CN105111280B - Template-fixed beta-hairpin peptidomimetics with protease inhibitory activity - Google Patents

Template-fixed beta-hairpin peptidomimetics with protease inhibitory activity Download PDF

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CN105111280B
CN105111280B CN201510320667.4A CN201510320667A CN105111280B CN 105111280 B CN105111280 B CN 105111280B CN 201510320667 A CN201510320667 A CN 201510320667A CN 105111280 B CN105111280 B CN 105111280B
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CN105111280A (en
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S·J·德马科
K·默勒
H·亨策
O·塞利耶
F·荣格
F·贡贝特
D·奥伯莱希特
C·卢丁
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Universitaet Zuerich
Spexis AG
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Polyphor AG
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Abstract

The invention provides template-fixed beta-hairpin peptidomimetics of the general formula (I) or salts thereof

Description

Template-fixed beta-hairpin peptidomimetics with protease inhibitory activity
The present application is a divisional application of the patent application having the title "template-fixed β -hairpin peptidomimetic having protease inhibitory activity" filed as 200580049141.1 filed as 17.2.2005.
Technical Field
The present invention provides template-fixed β -hairpin peptidomimetics comprising a template-fixed chain of 11 α -amino acid residues, generalized in that their position in the chain is Gly or Pro (4NHCOPhe), or of some kind as defined below. These template-fixed β -hairpin peptidomimetics are useful as inhibitors of proteases. They are particularly useful as inhibitors of different serine proteases, e.g., human cathepsin G, elastase or tryptase. Furthermore, the present invention provides a highly efficient method by which these compounds can be produced in a library format, if desired.
The β -hairpin peptidomimetics of the invention exhibit improved potency, oral bioavailability, improved half-life, and most importantly, high selectivity rates between different serine proteases, depending on the appropriate choice of certain types of α -amino acid residues and their position in the chain. Furthermore, these β -hairpin peptidomimetics exhibit low hemolysis and low cytotoxicity towards erythrocytes.
Background
Inhibitors of proteases are showing promising therapeutic utility in the treatment of diseases such as cancer (R.P.Beckert, A.Davidson, A.H.Drummond, M.Whittaker, Drug Disc.Today 1996,1, 16-26; L.L.Johnson, R.Dyer, D.J.Hupe, curr.Opin.chem.biol.1998,2,466-71; D.Leung, G.Abbenantete, and D.P.Fairlie, J.Med.chem.2000,43, 305-; c.albicans (C.Abad-Zappelero, R.Goldman, S.W.Muchmore, C.Hutchins, K.Stewart, J.Navaza, C.D.Payne, T.L.ray, protein Sci.1996,5,640-52), HIV (A.Wlodawer, J.W.Erickson, Annu.Rev.biochem.1993,62,543-85; P.L.Darke, J.R.Huff, adv.Pharmacol.1994,5,399-, hepatitis (J.L.Kim, K.A.Morgenster, C.Lin, T.Fox, M.D.Dwyer, J.A.Landro, S.P.Chemers, W.Markked, C.A.Lin, T.Fox, M.D.Dwy, C.Landso, S.S.P.P.J.P.J.Ser. Checker, C.J.J.J.J.Schaft.31, C.J.J.J.J.J.J.Shi, C.S.J.J.J.J.Schaft, C.D.D.D.D.D.D.D.D.D.D.D.D.D.D.D.D.D.A.A.A.D.A.D.D.A.A.Landson, C.S.S.S.S.S.S.S.S.S.S.S.P.S.S.A.S.S.S.H.H.A.A.A.A.A.A.A.A.A.A.A.A.A.A.A.A.A.A.A.A.A.A.A.A.A.D.D.A.D.D.D.A.D.A.A.D.D.H.D.D.A.D.H.D.D.D.H.D.D.A.H.A.A.A.D.D.D.A.A.A.A.D.D.A.D.H.A.A.D.A.A.A.A.A.A.A.A.A.A.D.A.A.A.A.A.A.A.A.A.A.A.A.A.A.A.A.D.A.D.D.D.A.A.A.A.A.A.A.A.A.A.D.D.A., S.Kahn, E.A.Mendriaz, Science,1999,286,735-41), angiogenesis (Kaatinen M et al, Atherosylrosisis 1996, 1231-.
Since most proteases bind to their substrates in an extended or β -chain conformation, good inhibitors must be able to mimic this conformation. Thus, ideally, the β -hairpin mimetic is adapted to lock the peptide sequence in an extended conformation.
Among proteases, serine proteases constitute important therapeutic targets. Serine proteases are classified by their substrate specificity (particularly the type of residue found at P1) into: trypsin-like (positively charged residues Lys/Arg at position P1 are preferred), elastase-like (residues P1 areSmall hydrophobic residues Ala/Val) or chymotrypsin-like (large hydrophobic residues Phe/Tyr/Leu at P1). It is already available in the PDB database (PDB:www.rcsb.org/pdb) Those serine proteases for which protease inhibitor X-ray crystal data were obtained include trypsin, alpha-chymotrypsin, gamma-chymotrypsin, human neutrophil elastase, thrombin, subtilisin, human cytomegalovirus, protease a, achromobacter (achromobacter), human cathepsin G, glutamate specific protease, carboxypeptidase D, coagulation factor VIIa, porcine coagulation factor 1XA, mesenterico peptidase, HCV protease and thermitase. Other serine proteases of therapeutic benefit include tryptase, complement convertase, hepatitis C-NS3 protease. Inhibitors of thrombin (e.g. j.l.metha, l.y.chen, w.w.nichols, c.mattsson, d.gustaffson, t.g.p.saldeen, j.cardiovasec.pharmacol.1998, 31,345-51; c.lila, p.gloanec, l.cadet, y.herve, j.fournier, f.leborgne, t.j.verbeuren, g.denanteuil, synth.comm.1998,28,4419-29) and factor Xa (e.g. j.p.vacca, annu.med.chem.1998, 33,81-90) are being used as anticoagulants in clinical assessments, inhibitors of elastase (j.r.williams, r.c.falcon, c.knee, r.l.in, r.l.chem.1997, r.r.r.t.21-51, r.t.t.t. ghem.r.r.r.r.t.t.r.t.t.t.t.t.t.t.t.r.r.t. lung, r.r.r.r.t. t. r.r.t. r.r.r.t. r. 7, r.r. r. r.r.r. r. r.r.r.r.r.r. r.. Finally, cathepsin G and elastase are intimately involved in the regulation of the activity of cytokines and their receptors. In particular at the site of inflammation, cathepsin G, elastase and protease 3 are released from infiltrating polymorphonuclear cells that have a close temporal correlation with elevated levels of inflammatory cytokines, which strongly suggests that these proteases are involved in the control of cytokine bioactivity and availability (u.bank, s.ansorge, j.leukc.biol.2001, 69,177-90). Therefore, inhibitors of elastase and cathepsin G constitute valuable targets for new drug candidates, in particular for chronic obstructive pulmonary disease (Ohbayashi H, ephert Opin.Investig.Drugs 2002,11,965-980)。
Among the various proteinaceous serine protease inhibitors that exist, one is a 14 amino acid cyclic peptide from sunflower seed, known as sunflower trypsin inhibitor (SFTI-1) (S.Luckett, R.Santiago Garcia, J.J.Barker, A.V.Konarev, P.R.Shewry, A.R.Clarke, R.L.Brady, J.mol.biol.1999,290, 525-533; Y.Q.Long, S.L.Lee, C.Y.Lin, I.J.Enyedy, S.Wang, P.Li, R.B.Dickson, P.P.roller, Biorg.&Med, chem, lett, 2001,11, 2515-containing 2519) which show sequence and conformational similarity to the trypsin-reactive loops of serine protease inhibitors of the Bowman-Birk family. The inhibitor adopts a beta-hairpin conformation when bound to the active site of bovine beta-trypsin. SFTI-1 inhibits beta-trypsin (K)i<0.1nM), cathepsin G (K)i0.15nM), elastase (K)i105. mu.M), chymotrypsin (K)i7.4. mu.M) and Thrombin (K)i～136mM)。
We illustrate here the approach of inhibitor design, which involves the transplantation of a β -hairpin loop from a naturally occurring peptide onto a hairpin-inducing template. Based on the well-defined 3D structure of the β -hairpin mimetics, libraries of compounds can be designed, which ultimately lead to novel inhibitors that exhibit different specificity characteristics for several classes of proteases.
Template-bound hairpin mimetics have been described in the literature (D, Obrecht, M.Altorfer, J.A.Robinson, adv.Med.Chem.1999,4, 1-68; J.A.Robinson, Syn.Lett.2000,4,429-441), template-fixed peptidomimetics which inhibit serine proteases and methods for their synthesis have been described in International patent application WO2003/054000A1 and Descours A, Moehle K., Renard A, RobinJ.ChemBioChem 2002,3,318-323, but these previously disclosed molecules have not exhibited high selectivity and particularly high potency. However, the ability to generate β -hairpin peptidomimetics using combinatorial and parallel synthesis methods has now been established (L.Jiang, K.Moehle, B.Dhanapaal, D.Obrecht, J.A.Robinson, Helv.Chim.acta.2000,83, 3097-.
The above-described methods allow the synthesis and screening of large hairpin mimetic libraries, which in turn facilitates structure-activity studies to a considerable extent, and thus the discovery of new molecules with highly potent and selective serine protease inhibitory activity, oral bioavailability, low hemolytic activity on human erythrocytes, and low cytotoxicity.
Disclosure of Invention
The beta-hairpin peptidomimetics of the invention are compounds of the general formula, and their pharmaceutically acceptable salts,
Figure BDA0000736457340000051
wherein
Figure BDA0000736457340000052
Is a group of one of the formulae:
Figure BDA0000736457340000061
Figure BDA0000736457340000071
wherein,
Figure BDA0000736457340000072
is Gly, or is the residue of an L-alpha-amino acid, wherein B is of the formula-NR20CH(R71) -or an enantiomer of one of the groups a1 to a69 as defined below;
is a group of one of the formulae:
Figure BDA0000736457340000074
Figure BDA0000736457340000081
Figure BDA0000736457340000091
Figure BDA0000736457340000101
R1is H, lower alkyl or aryl-lower alkyl;
R2is H, alkyl, alkenyl, - (CH)2)m(CHR61)sOR55、-(CH2)m(CHR61)sSR56、-(CH2)m(CHR61)sNR33R34、-(CH2)m(CHR61)sOCONR33R75、-(CH2)m(CHR61)sNR20CONR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R3Is H, alkyl, alkenyl, - (CH)2)m(CHR61)sOR55、-(CH2)m(CHR61)sSR56、-(CH2)m(CHR61)sNR33R34、-(CH2)m(CHR61)sOCONR33R75、-(CH2)m(CHR61)sNR20CONR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R4Is H, alkyl, alkenyl, - (CH)2)m(CHR61)sOR55、-(CH2)m(CHR61)sSR56、-(CH2)m(CHR61)sNR33R34、-(CH2)m(CHR61)sOCONR33R75、-(CH2)m(CHR61)sNR20CONR33R82、-(CH2)p(CHR61)sCOOR57、-(CH2)p(CHR61)sCONR58R59、-(CH2)p(CHR61)sPO(OR60)2、-(CH2)p(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R5Is alkyl, alkenyl, - (CH)2)o(CHR61)sOR55、-(CH2)o(CHR61)sSR56、-(CH2)o(CHR61)sNR33R34、-(CH2)o(CHR61)sOCONR33R75、-(CH2)o(CHR61)sN R20CONR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R6Is H, alkyl, alkenyl, - (CH)2)o(CHR61)sOR55、-(CH2)o(CHR61)sSR56、-(CH2)o(CHR61)sNR33R34、-(CH2)o(CHR61)sOCONR33R75、-(CH2)o(CHR61)sNR20CON R33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R7Is alkyl, alkenyl, - (CH)2)q(CHR61)sOR55、-(CH2)q(CHR61)sNR33R34、-(CH2)q(CHR61)sOCONR33R75、-(CH2)q(CHR61)sNR20CONR33R82、-(CH2)r(CHR61)sCO OR57、-(CH2)r(CHR61)sCONR58R59、-(CH2)r(CHR61)sPO(OR60)2、-(CH2)r(CHR61)sSO2R62Or- (CH)2)r(CHR61)sC6H4R8；
R8Is H, Cl, F, CF3、NO2Lower alkyl, lower alkenyl, aryl-lower alkyl, - (CH)2)o(CHR61)sOR55、-(CH2)o(CHR61)sSR56、-(CH2)o(CHR61)NR33R34、-(CH2)o(CHR61)sOCONR33R75、-(CH2)o(CHR61)sNR20CONR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sCOR64；
R9Is alkyl, alkenyl, - (CH)2)o(CHR61)sOR55、-(CH2)o(CHR61)sSR56、-(CH2)o(CHR61)sNR33R34、-(CH2)o(CHR61)sOCONR33R75、-(CH2)o(CHR61)sNR20CON R33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R10Is alkyl, alkenyl, - (CH)2)o(CHR61)sOR55、-(CH2)o(CHR61)sSR56、-(CH2)o(CHR61)sNR33R34、-(CH2)o(CHR61)sOCONR33R75、-(CH2)o(CHR61)sNR20CON R33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R11Is H, alkyl, alkenyl, - (CH)2)m(CHR61)sOR55、-(CH2)m(CHR61)sNR33R34、-(CH2)m(CHR61)sOCONR33R75、-(CH2)m(CHR61)sNR20CONR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R12Is H, alkyl, alkenyl, - (CH)2)m(CHR61)sOR55、-(CH2)m(CHR61)sSR56、-(CH2)m(CHR61)sNR33R34、-(CH2)m(CHR61)sOCONR33R75、-(CH2)m(CHR61)sNR20CONR33R82、-(CH2)r(CHR61)sCOOR57、-(CH2)r(CHR61)sCONR58R59、-(CH2)r(CHR61)sPO(OR60)2、-(CH2)r(CHR61)sSO2R62Or- (CH)2)r(CHR61)sC6H4R8；
R13Is alkyl, alkenyl, - (CH)2)q(CHR61)sOR55、-(CH2)q(CHR61)sSR56、-(CH2)q(CHR61)sNR33R34、-(CH2)q(CHR61)sOCONR33R75、-(CH2)q(CHR61)sNR20CONR33R82、-(CH2)q(CHR61)sCOOR57、-(CH2)q(CHR61)sCONR58R59、-(CH2)q(CHR61)sPO(OR60)2、-(CH2)q(CHR61)sSO2R62Or- (CH)2)q(CHR61)sC6H4R8；
R14Is H, alkyl, alkenyl, - (CH)2)m(CHR61)sOR55、-(CH2)m(CHR61)sNR33R34、-(CH2)m(CHR61)sOCONR33R75、-(CH2)m(CHR61)sNR20CONR33R82、-(CH2)q(CHR61)sCOOR57、-(CH2)q(CHR61)sCONR58R59、-(CH2)q(CHR61)sPO(OR60)2、-(CH2)q(CHR61)sSOR62Or- (CH)2)q(CHR61)sC6H4R8；
R15Is alkyl, alkenyl, - (CH)2)o(CHR61)sOR55、-(CH2)o(CHR61)sSR56、-(CH2)o(CHR61)sNR33R34、-(CH2)o(CHR61)sOCONR33R75、-(CH2)o(CHR61)sNR20CON R33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R16Is alkyl, alkenyl, - (CH)2)o(CHR61)sOR55、-(CH2)o(CHR61)sSR56、-(CH2)o(CHR61)sNR33R34、-(CH2)o(CHR61)sOCONR33R75、-(CH2)o(CHR61)sNR20CONR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R17Is alkyl, alkenyl, - (CH)2)q(CHR61)sOR55、-(CH2)q(CHR61)sSR56、-(CH2)q(CHR61)sNR33R34、-(CH2)q(CHR61)sOCONR33R75、-(CH2)q(CHR61)sNR20CONR33R82、-(CH2)q(CHR61)sCOOR57、-(CH2)q(CHR61)sCONR58R59、-(CH2)q(CHR61)sPO(OR60)2、-(CH2)q(CHR61)sSO2R62Or- (CH)2)q(CHR61)sC6H4R8；
R18Is alkyl, alkenyl, - (CH)2)p(CHR61)sOR55、-(CH2)p(CHR61)sSR56、-(CH2)p(CHR61)sNR33R34、-(CH2)p(CHR61)sOCONR33R75、-(CH2)p(CHR61)sNR20CONR33R82、-(CH2)p(CHR61)sCOOR57、-(CH2)p(CHR61)sCONR58R59、-(CH2)p(CHR61)sPO(OR60)2、-(CH2)p(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R19Is lower alkyl, - (CH)2)p(CHR61)sOR55、-(CH2)p(CHR61)sSR56、-(CH2)p(CHR61)sNR33R34、-(CH2)p(CHR61)sOCONR33R75、-(CH2)p(CHR61)sNR20CONR33R82、-(CH2)p(CHR61)sCOOR57、-(CH2)p(CHR61)sCONR58R59、-(CH2)p(CHR61)sPO(OR60)2、-(CH2)p(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8Or
R18And R19Together may form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-；
R20Is H, alkyl, alkenyl or aryl-lower alkyl;
R21is H, alkyl, alkenyl, - (CH)2)o(CHR61)sOR55、-(CH2)o(CHR61)sSR56、-(CH2)o(CHR61)sNR33R34、-(CH2)o(CHR61)sOCONR33R75、-(CH2)o(CHR61)sNR20CONR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R22Is H, alkyl, alkenyl, - (CH)2)o(CHR61)sOR55、-(CH2)o(CHR61)sSR56、-(CH2)o(CHR61)sNR33R34、-(CH2)o(CHR61)sOCONR33R75、-(CH2)o(CHR61)sNR20CONR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R23Is alkyl, alkenyl, - (CH)2)o(CHR61)sOR55、-(CH2)o(CHR61)sSR56、-(CH2)o(CHR61)sNR33R34、-(CH2)o(CHR61)sOCONR33R75、-(CH2)o(CHR61)sNR20CONR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R24Is alkyl, alkenyl, - (CH)2)o(CHR61)sOR55、-(CH2)o(CHR61)sSR56、-(CH2)o(CHR61)sNR33R34、-(CH2)o(CHR61)sOCONR33R75、-(CH2)o(CHR61)sNR20CON R33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R25Is H, alkyl, alkenyl, - (CH)2)m(CHR61)sOR55、-(CH2)m(CHR61)sSR56、-(CH2)m(CHR61)sNR33R34、-(CH2)m(CHR61)sOCONR33R75、-(CH2)m(CHR61)sNR20CO NR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sP O(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R26Is H, alkyl, alkenyl, - (CH)2)m(CHR61)sOR55、-(CH2)m(CHR61)sSR56、-(CH2)m(CHR61)sNR33R34、-(CH2)m(CHR61)sOCONR33R75、-(CH2)m(CHR61)sNR20CO NR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sP O(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8Or
R25And R26Together may form: - (CH)2)2-6-、-(CH2)rO(CH2)r-、-(CH2)rS(CH2)r-or- (CH)2)rNR57(CH2)r-；
R27Is H, alkyl, alkenyl, - (CH)2)o(CHR61)sOR55、-(CH2)o(CHR61)sSR56、-(CH2)o(CHR61)sNR33R34、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sOCONR33R75、-(CH2)o(CHR61)sNR20CONR33R82、-(CH2)o(CHR61)sP O(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R28Is alkyl, alkenyl, - (CH)2)o(CHR61)s-OR55、-(CH2)o(CHR61)sSR56、-(CH2)o(CHR61)sNR33R34、-(CH2)o(CHR61)sOCONR33R75、-(CH2)o(CHR61)sNR20CO NR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R29Is alkyl, alkenyl, - (CH)2)o(CHR61)sOR55、-(CH2)o(CHR61)sSR56、-(CH2)o(CHR61)sNR33R34、-(CH2)o(CHR61)sOCONR33R75、-(CH2)o(CHR61)sNR20CON R33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R30Is H, alkyl, alkenyl or aryl-lower alkyl.
R31Is H, alkyl, alkenyl, - (CH)2)p(CHR61)sOR55、-(CH2)p(CHR61)sNR33R34、-(CH2)p(CHR61)sOCONR33R75、-(CH2)p(CHR61)sNR20CONR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R32Is H, lower alkyl or aryl-lower alkyl;
R33is H, alkyl, alkenyl, - (CH)2)m(CHR61)sOR55、-(CH2)m(CHR61)sNR34R63、-(CH2)m(CHR61)sOCONR75R82、-(CH2)m(CHR61)sNR20CONR78R82、-(CH2)o(CHR61)sCOR64、-(CH2)o(CHR61)s-CONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R34Is H, lower alkyl, aryl or aryl-lower alkyl;
R33and R34Together may form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-；
R35Is H, alkyl, alkenyl, - (CH)2)m(CHR61)sOR55、-(CH2)m(CHR61)sNR33R34、-(CH2)m(CHR61)sOCONR33R75、-(CH2)m(CHR61)sNR20CONR33R82、-(CH2)p(CHR61)sCOOR57、-(CH2)p(CHR61)sCONR58R59、-(CH2)p(CHR61)sPO(OR60)2、-(CH2)p(CHR61)sSO2R62Or- (CH)2)p(CHR61)sC6H4R8；
R36Is H, alkyl, alkenyl, - (CH)2)o(CHR61)sOR55、-(CH2)p(CHR61)sNR33R34、-(CH2)p(CHR61)sOCONR33R75、-(CH2)p(CHR61)sNR20CONR33R82、-(CH2)p(CHR61)sCOOR57、-(CH2)p(CHR61)sCONR58R59、-(CH2)p(CHR61)sPO(OR60)2、-(CH2)p(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R37Is H, F, Br, Cl, NO2、CF3Lower alkyl, - (CH)2)p(CHR61)sOR55、-(CH2)p(CHR61)sNR33R34、-(CH2)p(CHR61)sOCONR33R75、-(CH2)p(CHR61)sNR20CO NR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sP O(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R38Is H, F, Br, Cl, NO2、CF3Alkyl, alkenyl, - (CH)2)p(CHR61)sOR55、-(CH2)p(CHR61)sNR33R34、-(CH2)p(CHR61)sOCONR33R75、-(CH2)p(CHR61)sNR20CO NR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sP O(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R39Is H, alkyl, alkenyl or aryl-lower alkyl;
R40is H, alkyl, alkenyl or aryl-lower alkyl;
R41is H, F, Br, Cl, NO2、CF3Alkyl, alkenyl, - (CH)2)p(CHR61)sOR55、-(CH2)p(CHR61)sNR33R34、-(CH2)p(CHR61)sOCONR33R75、-(CH2)p(CHR61)sNR20CO NR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sP O(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R42Is H, F, Br, Cl, NO2、CF3Alkyl, alkenyl, - (CH)2)p(CHR61)sOR55、-(CH2)p(CHR61)sNR33R34、-(CH2)p(CHR61)sOCONR33R75、-(CH2)p(CHR61)sNR20CO NR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sP O(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R43Is H, alkyl, alkenyl, - (CH)2)m(CHR61)sOR55、-(CH2)m(CHR61)sNR33R34、-(CH2)m(CHR61)sOCONR33R75、-(CH2)m(CHR61)sNR20CONR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)sPO(OR60)2、-(CH2)o(CHR61)sSO2R62Or- (CH)2)o(CHR61)sC6H4R8；
R44Is alkyl, alkenyl, - (CH)2)r(CHR61)sOR55、-(CH2)r(CHR61)sSR56、-(CH2)r(CHR61)sNR33R34、-(CH2)r(CHR61)sOCONR33R75、-(CH2)r(CHR61)sNR20CON R33R82、-(CH2)r(CHR61)sCOOR57、-(CH2)r(CHR61)sCONR58R59、-(CH2)r(CHR61)sPO(OR60)2、-(CH2)r(CHR61)sSO2R62Or- (CH)2)r(CHR61)sC6H4R8；
R45Is H, alkyl, alkenyl, - (CH)2)o(CHR61)sOR55、-(CH2)o(CHR61)sSR56、-(CH2)o(CHR61)sNR33R34、-(CH2)o(CHR61)sOCONR33R75、-(CH2)o(CHR61)sNR20CON R33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)s(CHR61)sCONR58R59、-(CH2)s(CHR61)sPO(OR60)2、-(CH2)s(CHR61)sSO2R62Or- (CH)2)s(CHR61)sC6H4R8；
R46Is H, alkyl, alkenyl or- (CH)2)o(CHR61)pC6H4R8；
R47Is H, alkyl, alkenyl or- (CH)2)o(CHR61)sOR55；
R48Is H, lower alkyl, lower alkenyl or aryl-lower alkyl;
R49is H, alkyl, alkenyl, - (CHR)61)sCOOR57、(CHR61)sCONR58R59、(CHR61)sPO(OR60)2、-(CHR61)sSOR62Or- (CHR)61)sC6H4R8；
R50Is H, lower alkyl or aryl-lower alkyl;
R51is H, alkyl, alkenyl, - (CH)2)m(CHR61)sOR55、-(CH2)m(CHR61)sSR56、-(CH2)m(CHR61)sNR33R34、-(CH2)m(CHR61)sOCONR33R75、-(CH2)m(CHR61)sNR20CO NR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)pP O(OR60)2、-(CH2)p(CHR61)sSO2R62Or- (CH)2)p(CHR61)sC6H4R8；
R52Is H, alkyl, alkenyl, - (CH)2)m(CHR61)sOR55、-(CH2)m(CHR61)sSR56、-(CH2)m(CHR61)sNR33R34、-(CH2)m(CHR61)sOCONR33R75、-(CH2)m(CHR61)sNR20CO NR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)pP O(OR60)2、-(CH2)p(CHR61)sSO2R62Or- (CH)2)p(CHR61)sC6H4R8；
R53Is H, alkyl, alkenyl, - (CH)2)m(CHR61)sOR55、-(CH2)m(CHR61)sSR56、-(CH2)m(CHR61)sNR33R34、-(CH2)m(CHR61)sOCONR33R75、-(CH2)m(CHR61)sNR20CO NR33R82、-(CH2)o(CHR61)sCOOR57、-(CH2)o(CHR61)sCONR58R59、-(CH2)o(CHR61)pP O(OR60)2、-(CH2)p(CHR61)sSO2R62Or- (CH)2)p(CHR61)sC6H4R8；
R54Is H, alkyl, alkenyl, - (CH)2)m(CHR61)sOR55、-(CH2)m(CHR61)sNR33R34、-(CH2)m(CHR61)sOCONR33R75、-(CH2)m(CHR61)sNR20CONR33R82、-(CH2)o(CHR61)COOR57、-(CH2)o(CHR61)sCONR58R59Or- (CH)2)o(CHR61)sC6H4R8；
R55Is H, lower alkyl, lower alkenyl, aryl-lower alkyl, - (CH)2)m(CHR61)sOR57、-(CH2)m(CHR61)sNR34R63、-(CH2)m(CHR61)sOCONR75R82、-(CH2)m(CHR61)sNR20CO NR78R82、-(CH2)o(CHR61)s-COR64、-(CH2)o(CHR61)COOR57Or- (CH)2)o(CHR61)sCON R58R59；
R56Is H, lower alkyl, lower alkenyl, aryl-lower alkyl, - (CH)2)m(CHR61)sOR57、-(CH2)m(CHR61)sNR34R63、-(CH2)m(CHR61)sOCONR75R82、-(CH2)m(CHR61)sNR20CO NR78R82、-(CH2)o(CHR61)s-COR64Or- (CH)2)o(CHR61)sCONR58R59；
R57Is H, lower alkyl, lower alkenyl, aryl lower alkyl or heteroaryl lower alkyl;
R58is H, lower alkyl, lower alkenyl, aryl, heteroaryl, aryl-lower alkyl or heteroaryl-lower alkyl;
R59is H, lower alkyl, lower alkenyl, aryl, heteroaryl, aryl-lower alkylOr heteroaryl-lower alkyl; or
R58And R59Together may form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-；
R60Is H, lower alkyl, lower alkenyl, aryl or aryl-lower alkyl;
R61is alkyl, alkenyl, aryl, heteroaryl, aryl-lower alkyl, heteroaryl-lower alkyl, - (CH)2)mOR55、-(CH2)mNR33R34、-(CH2)mOCONR75R82、-(CH2)mNR20CONR78R82、-(CH2)oCOOR37、-(CH2)oNR58R59Or- (CH)2)oPO(COR60)2；
R62Is lower alkyl, lower alkenyl, aryl, heteroaryl or aryl-lower alkyl;
R63is H, lower alkyl, lower alkenyl, aryl, heteroaryl, aryl-lower alkyl, heteroaryl-lower alkyl, -COR64、-COOR57、-CONR58R59、-SO2R62OR-PO (OR)60)2；
R34And R63Together can form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-；
R64Is H, lower alkyl, lower alkenyl, aryl, heteroaryl, aryl-lower alkyl, heteroaryl-lower alkyl, - (CH)2)p(CHR61)sOR65、-(CH2)p(CHR61)sSR66Or- (CH)2)p(CHR61)sNR34R63、-(CH2)P(CHR61)sOCONR75R82、-(CH2)P(CHR61)sNR20CONR78R82；
R65Is H, lower alkyl, lower alkenyl, aryl-lower alkyl, heteroaryl-lower alkyl, -COR57、-COOR57or-CONR58R59；
R66Is H, lower alkyl, lower alkenyl, aryl-lower alkyl, heteroaryl-lower alkyl or-CONR58R59；
m is 2 to 4; o is 0 to 4; p is 1 to 4; q is 0 to 2; r is 1 or 2; s is 0 or 1;
z is a chain of 11 α -amino acid residues, the positions of which in the chain are counted starting from the N-terminal amino acid, whereby, depending on their position in the chain, these amino acid residues are Gly, Pro (4NHCOPhe) or of the formula-A-CO-, or of the formula-B-CO-, or of one of the following types:
C:-NR20CH(R72)CO-；
D:-NR20CH(R73)CO-；
E:-NR20CH(R74)CO-；
F:-NR20CH(R84) CO-; and
H:-NR20-CH(CO-)-(CH2)4-7-CH(CO-)-NR20-、-NR20-CH(CO-)-(CH2)pSS(CH2)p-CH(CO-)-NR20-、-NR20-CH(CO-)-(-(CH2)pNR20CO(CH2)p-CH(CO-)-NR20-and-NR20-CH(CO-)-(-(CH2)pNR20CONR20(CH2)p-CH(CO-)-NR20-；
R71Is a lower alkyl group,Lower alkenyl, - (CH)2)p(CHR61)sOR75、-(CH2)p(CHR61)sSR75、-(CH2)p(CHR61)sNR33R34、-(CH2)p(CHR61)sOCONR33R75、-(CH2)p(CHR61)sNR20CON R33R82、-(CH2)o(CHR61)sCOOR75、-(CH2)pCONR58R59、-(CH2)pPO(OR62)2、-(CH2)pSO2R62Or- (CH)2)o-C6R67R68R69R70R76；
R72Is H, lower alkyl, lower alkenyl, - (CH)2)p(CHR61)sOR85Or- (CH)2)p(CHR61)sSR85；
R73Is- (CR)86R87)oR77、-(CH2)rO(CH2)oR77、-(CH2)rS(CH2)oR77Or- (CH)2)rNR20(CH2)oR77；
R74Is- (CH)2)pNR78R79、-(CH2)pNR77R80、-(CH2)pC(＝NR80)NR78R79、-(CH2)pC(＝NOR50)NR78R79、-(CH2)pC(＝NNR78R79)NR78R79、-(CH2)pNR80C(＝NR80)NR78R79、-(CH2)pN＝C(NR78R80)NR79R80、-(CH2)pC6H4NR78R79、-(CH2)pC6H4NR77R80、-(CH2)pC6H4C(＝NR80)NR78R79、-(CH2)pC6H4C(＝NOR50)NR78R79、-(CH2)pC6H4C(＝NNR78R79)NR78R79、-(CH2)pC6H4NR80C(＝NR80)NR78R79、-(CH2)pC6H4N＝C(NR78R80)NR79R80、-(CH2)rO(CH2)mNR78R79、-(CH2)rO(CH2)mNR77R80、-(CH2)rO(CH2)pC(＝NR80)NR78R79、-(CH2)rO(CH2)pC(＝NOR50)NR78R79、-(CH2)rO(CH2)pC(＝NNR78R79)NR78R79、-(CH2)rO(CH2)mNR80C(＝NR80)NR78R79、-(CH2)rO(CH2)mN＝C(NR78R80)NR79R80、-(CH2)rO(CH2)pC6H4CNR78R79、-(CH2)rO(CH2)pC6H4C(＝NR80)NR78R79、-(CH2)rO(CH2)pC6H4C(＝NOR50)NR78R79、-(CH2)rO(CH2)pC6H4C(＝NNR78R79)NR78R79、-(CH2)rO(CH2)pC6H4NR80C(＝NR80)NR78R79、-(CH2)rS(CH2)mN R78R79、-(CH2)rS(CH2)mNR77R80、-(CH2)rS(CH2)pC(＝NR80)NR78R79、-(CH2)rS(CH2)pC(＝NOR50)NR78R79、-(CH2)rS(CH2)pC(＝NNR78R79)NR78R79、-(CH2)rS(CH2)mNR80C(＝NR80)NR78R79、-(CH2)rS(CH2)mN＝C(NR78R80)NR79R80、-(CH2)rS(CH2)pC6H4CNR78R79、-(CH2)rS(CH2)pC6H4C(＝NR80)NR78R79、-(CH2)rS(CH2)pC6H4C(＝NOR50)NR78R79、-(CH2)rS(CH2)pC6H4C(＝NNR78R79)NR78R79、-(CH2)rS(CH2)pC6H4NR80C(＝NR80)NR78R79、-(CH2)pNR80COR64、-(CH2)pNR80COR77、-(CH2)pNR80CONR78R79、-(CH2)pC6H4NR80CONR78R79Or- (CH)2)pNR20CO-[(CH2)u-X]t-CH3Wherein X is-O-, -NR20-or-S-; u is 1 to 3 and t is 1 to 6;
R75is lower alkyl, lower alkenyl or aryl-lower alkyl;
R33and R75Together may form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-；
R75And R82Together may form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-；
R76Is H, lower alkyl, lower alkenyl, aryl-lower alkyl, - (CH)2)oOR72、-(CH2)oSR72、-(CH2)oNR33R34、-(CH2)oOCONR33R75、-(CH2)oNR20CONR33R82、-(CH2)oCOOR75、-(CH2)oCONR58R59、-(CH2)oPO(OR60)2、-(CH2)pSO2R62Or- (CH)2)oCOR64；
R77is-C6R67R68R69R70R76Or a heteroaryl group of one of the formulae
Figure BDA0000736457340000201
Figure BDA0000736457340000211
R78Is H, lower alkyl, aryl or aryl-lower alkyl;
R78and R82Together may form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-；
R79Is H, lower alkyl, aryl or aryl-lower alkyl, or
R78And R79Together may be: - (CH)2)2-7-、-(CH2)2O(CH2)2-or- (CH)2)2NR57(CH2)2-；
R80Is H or lower alkyl;
R81is H, lower alkyl or aryl-lower alkyl;
R82is H, lower alkyl, aryl, heteroaryl or aryl-lower alkyl;
R33and R82Together may form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-；
R83Is H, lower alkyl, aryl or-NR78R79；
R84Is- (CH)2)m(CHR61)sOH、-(CR86R87)pOR80、-(CR86R87)pCOOR80、-(CH2)m(CHR61)sSH、-(CR86R87)pSR80、-(CH2)pCONR78R79、-(CH2)pNR80CONR78R79、-(CH2)pC6H4CONR78R79、-(CH2)pC6H4NR80CONR78R79、-(CR86R87)oPO(OR60)2、-(CR86R87)pSO2R60、-(CR86R87)pSOR60、-(CH2)m(CHR61)sOPO(OR60)2Or- (CH)2)m(CHR61)sOSO2R60；
R85Is lower alkyl or lower alkenyl;
R86h, H is lower alkyl which may be substituted by halogen or halogen;
R87h, H is lower alkyl which may be substituted by halogen or halogen;
with the following conditions: in said chain of 11 a-amino acid residues Z,
-if n is 11, then the amino acid residues at positions 1 to 11 are:
-P1 type C or D or E or F;
-P2 type C or D or E or F;
-P3, type C or F, or the residue is Gly;
-P4 type C or D or F or E, or the residue is Gly or Pro;
-P5, type E or C or F, or the residue is Gly or Pro;
-P6, D or F or E or C, or the residue is Gly or Pro;
-P7, type C or E or F, or having the formula-A-CO-, or the residue is Gly or Pro;
-P8, type D or C or F, or of formula-A-CO, or the residue is Gly or Pro (4 NHCOPhe);
-P9 type C or D or E or F;
p10 type D or C or F or E, and
p11 type C or D or E or F, or
-P2 and P10 together mayForming a group of the H type, with the further proviso that: if the template isDProLPro, amino acid residues from P1 to P11 are not as follows:
-P1:Arg
-P2 Cys linked to Cys at position P10 by a disulfide bond
-P3:Thr
-P4:Lys
-P5:Ser
-P6:Ile
-P7:Pro
-P8:Pro
-P9:Ile
-P10 Cys linked via a disulfide bond to Cys at position P10, and
-P11:Phe
according to the invention, these β -hairpin peptidomimetics can be prepared by a method comprising:
(a) coupling the appropriately functionalized solid support with an appropriately N-protected derivative of the amino acid in position 5,6 or 7 in the desired end product, any functional groups that may be present in said N-protected amino acid derivative being likewise appropriately protected;
(b) removing the N-protecting group from the product thus obtained;
(c) coupling the product thus obtained with a suitably N-protected derivative of that amino acid which is one position closer to the N-terminal amino acid residue in the desired end product, any functional groups which may be present in said N-protected amino acid derivative being likewise suitably protected;
(d) removing the N-protecting group from the product thus obtained;
(e) repeating steps (c) and (d) until the N-terminal amino acid residue has been introduced;
(f) coupling the product thus obtained with a compound of the general formula
Figure BDA0000736457340000231
Wherein
Figure BDA0000736457340000241
As defined above, and X is an N-protecting group, or, if
Figure BDA0000736457340000242
When it is the above (a1) or (a2) group,
(fa) coupling the product obtained in step (e) with a suitably N-protected derivative of an amino acid of the general formula:
HOOC-B-H III or HOOC-A-H IV
Wherein B and A are as defined above, any functional groups which may be present in the N-protected amino acid derivative are likewise suitably protected;
(fb) removing the N-protecting group from the product thus obtained; and
(fc) coupling the product thus obtained with a suitably N-protected derivative of an amino acid of formula IV and III above, respectively, in which any functional groups that may be present are likewise suitably protected; and, respectively, if
Figure BDA0000736457340000243
In the case of the above group (a3),
(fa') coupling the product obtained in step (e) with a suitably N-protected derivative of an amino acid of formula III as described above, any functional groups which may be present in said N-protected amino acid derivative also being suitably protected;
(fb') removing the N-protecting group from the product thus obtained; and
(fc') coupling the product thus obtained with a suitably N-protected derivative of an amino acid of formula III above, any functional groups which may be present in said N-protected amino acid derivative likewise being suitably protected;
(g) removing the N-protecting group from the product obtained in step (f) or (fc');
(h) coupling the product thus obtained with a suitably N-protected derivative of the amino acid at position 11 in the desired end product, any functional groups that may be present in said N-protected amino acid derivative likewise being suitably protected;
(i) removing the N-protecting group from the product thus obtained;
(j) coupling the product thus obtained with a suitable N-protected derivative of the amino acid which is further from position 11 in the desired end product, any functional groups which may be present in said N-protected amino acid derivative likewise being suitably protected;
(k) removing the N-protecting group from the product thus obtained;
(l) Repeating steps (j) and (k) until all amino acid residues have been introduced;
(m) if desired, selectively deprotecting one or more protected functional groups present in the molecule and appropriately substituting the reactive groups thus released;
(n) if desired, forming an interchain linkage between the side chains of the appropriate amino acid residues at positions 2 and 10;
(o) detaching the product thus obtained from the solid support;
(p) cyclizing the cleaved product from the solid support;
(q) removing any protecting groups present on the functional groups of any members of the chain of amino acid residues and, if desired, any other protecting groups that may be present in the molecule; and
(r) if desired, converting the product thus obtained into a pharmaceutically acceptable salt thereof, or converting the pharmaceutically acceptable or unacceptable salt thus obtained into the corresponding free compound of formula I, or into a different, pharmaceutically acceptable salt.
Alternatively, the peptidomimetics of the present invention may be prepared by:
(a') coupling a suitably functionalized solid support with a compound of the general formula
Figure BDA0000736457340000261
Wherein
Figure BDA0000736457340000262
As defined above, and X is an N-protecting group, or, if
Figure BDA0000736457340000263
When it is the above group (a1) or (a2),
(a' a) coupling the suitably functionalized solid support with a suitably N-protected derivative of an amino acid of the general formula
HOOC-B-H III or HOOC-A-H IV
Wherein B and A are as defined above, any functional groups which may be present in the N-protected amino acid derivative are likewise suitably protected;
(a' b) removing the N-protecting group from the product thus obtained; and
(a' c) coupling the product thus obtained with a suitably N-protected derivative of an amino acid of the above general formulae IV and III, respectively, any functional groups which may be present in said N-protected amino acid derivative being likewise suitably protected; and, respectively, if
Figure BDA0000736457340000264
In the case of the above group (a3),
(a 'a') coupling said suitably functionalized solid support with a suitably N-protected derivative of an amino acid of formula III above, any functional groups that may be present in said N-protected amino acid derivative being likewise suitably protected;
(a 'b') removing the N-protecting group from the product thus obtained; and
(a 'c') coupling the product thus obtained with a suitably N-protected derivative of an amino acid of formula III above, any functional groups that may be present in said N-protected amino acid derivative likewise being suitably protected;
(b ') removing the N-protecting group from the product obtained in step (a '), (a ' c) or (a ' c ');
(c') coupling the product thus obtained with a suitably N-protected derivative of the amino acid at position 11 in the desired end product, any functional groups which may be present in said N-protected amino acid derivative likewise being suitably protected;
(d') removing the N-protecting group from the product thus obtained;
(e') coupling the product thus obtained with a suitably N-protected derivative of the amino acid which is further from position 11 in the desired end product, any functional groups which may be present in said N-protected amino acid derivative being likewise suitably protected;
(f') removing the N-protecting group from the product thus obtained;
(g ') repeating steps (e ') and (f ') until all amino acid residues have been introduced;
(h') if desired, selectively deprotecting one or more protected functional groups present in the molecule and appropriately substituting the reactive groups thus released;
(i') if desired, forming an interchain linkage between the side chains of the appropriate amino acid residues at positions 2 and 10;
(j') detaching the product thus obtained from the solid support;
(k') cyclizing the cleaved product from the solid support;
(l') removing any protecting groups present on the functional groups of any members of the chain of amino acid residues and, if desired, any other protecting groups that may be present in the molecule; and
(m') if desired, converting the product thus obtained into a pharmaceutically acceptable salt thereof, or converting a pharmaceutically acceptable or unacceptable salt thus obtained into the corresponding free compound of formula I, or into a different, pharmaceutically acceptable salt.
The peptidomimetics of the present invention may also be enantiomers of the compounds of formula I. These enantiomers can be prepared by modifications of the above process in which all enantiomers of chiral starting materials are used.
The term "alkyl", as used in this specification, alone or in combination, refers to a straight or branched chain saturated hydrocarbon group having up to 24 (preferably up to 12) carbon atoms. Similarly, the term "alkenyl" refers to a straight or branched chain hydrocarbon group having up to 24 (preferably up to 12) carbon atoms containing at least one or up to four olefinic double bonds depending on chain length. The term "lower" refers to groups and compounds having up to 6 carbon atoms. Thus, for example, the term "lower alkyl" refers to a straight or branched chain saturated hydrocarbon group having up to 6 carbon atoms, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, tert-butyl, and the like. The term "aryl" refers to an aromatic carbocyclic hydrocarbon radical containing one or two six-membered rings, such as phenyl or naphthyl, which may be substituted with up to three substituents, such as Br, Cl, F, CF3、NO2Lower alkyl or lower alkenyl. The term "heteroaryl" refers to an aromatic heterocyclic group containing one or two five-membered and/or six-membered rings, at least one of which contains up to three heteroatoms selected from the group consisting of O, S and N, and which rings are substituted or unsubstituted; representative examples of such optionally substituted heteroaryl groups are as hereinbefore described for R77The definition section of (a).
The structural element-A-CO-refers to the amino acid building block which, in combination with the structural element-B-CO-, forms the templates (a1) and (a 2). The structural element-B-CO-is combined with another structural element-B-CO-to form a template (a 3). Template (a3) is less preferred in formula I. Templates (a) to (p) constitute building blocks having N-and C-termini positioned at intervals in such a way that the distance between these two groups may be between 4.0 and 5.5A. The peptide chain Z is linked to the C-and N-termini of the templates (a) to (p) via the respective N-and C-termini, such that the templates and chain form a cyclic structure, for example as shown in formula I. In the case where the distance between the N-and C-terminus of the template is between 4.0 and 5.5A as used herein, the template will induce an H-linked network which is necessary for the formation of a beta-hairpin conformation in the peptide chain Z. Thus, the template and peptide chain form a β -hairpin mimetic.
The beta-hairpin conformation is highly correlated with serine protease inhibitory activity of the beta-hairpin mimetics of the invention. The conformational nature of the stable β -hairpin of templates (a) to (p) not only plays a key role in selective inhibitory activity but also has a key role in the synthetic process as defined above, since incorporation of the template at the beginning or in the vicinity of the middle of the linear protected peptide precursor can significantly enhance cyclization yields.
The structural units A1-A69 belong to the class of amino acids in which the N-terminus is a secondary amine forming part of a ring. Among the amino acids encoded by the gene, only proline falls into this class. The configuration of the structural units A1 to A69 is (D), they are combined with the structural unit of the (L) -configuration-B-CO-. The preferred combination with respect to template (a1) is-DA1-CO-LB-CO-toDA69-CO-LB-CO-. Thus, for example,DPro-Lpro constitutes a prototype of template (a 1). Less preferred but still feasible are also combinations of forming the template (a 2): -LA1-CO-DB-CO-di-carboxylic acidLA69-CO-DB-CO-combination. Thus, for example,LPro-Dpro constitutes a prototype of template (a 2).
It will be appreciated that the structural unit-A1-CO-to-A69-CO-wherein A has the (D) -configuration carries a group R in the alpha-position to the N-terminus1。R1Preferably H and lower alkyl, R1Most preferred are H and methyl. One skilled in the art will appreciate that A1-A69 is shown in the (D) -configuration for R1Is the case for H and methyl, which corresponds to the (R) -configuration. Generalizing R according to Cahn, Ingold and Prelog rules1The configuration may also be represented as (S).
Except that R1In addition, the structural units-A1-CO-to-A69-CO-may carry the name R2To R17Of (2)And (4) a substituent. The other substituent may be H, which if not H is preferably a small to medium size aliphatic or aromatic group. R2To R17Examples of preferred values of (c) are:
-R2h, lower alkyl, lower alkenyl, (CH)2)mOR55(wherein R is55Lower alkyl or lower alkenyl), (CH)2)mSR56(wherein R is56Lower alkyl or lower alkenyl), (CH)2)mNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; r33And R34Together form- (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-；R57H or lower alkyl), (CH)2)mOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H or lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R3H, lower alkyl, lower alkenyl, - (CH)2)mOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)mSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)mNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R4H, lower alkyl, lower alkenyl, - (CH)2)mOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)mSR56(it isIn R56Lower alkyl or lower alkenyl), - (CH)2)mNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H or lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R5Lower alkyl, lower alkenyl, - (CH)2)oOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)oSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)oNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), (CH)2)oNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), (CH)2)oN(R20)COR64(wherein R is20H or lower alkyl; r64Alkyl, alkenyl, aryl and aryl-lower alkyl, heteroaryl-lower alkyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H or lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R6H, lower alkyl, lower alkenyl, - (CH)2)oOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)oSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)oNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H or lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R7Lower alkyl, lower alkenyl, - (CH)2)qOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)qSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)qNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)qOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), (CH)2)qNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)qN(R20)COR64(it isIn R20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)rCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)qCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H or lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)rPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), (CH)2)rSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R8:H、F、Cl、CF3Lower alkyl, lower alkenyl, - (CH)2)oOR55(wherein R is55Lower alkyl or lower alkenyl), (CH)2)oSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)oNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H or lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R9Lower alkyl, lower alkenyl, - (CH)2)oOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)oSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)oNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H or lower alkyl; or R58And R59Together areForming: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R10Lower alkyl, lower alkenyl, - (CH)2)oOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)oSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)oNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl orLower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R11H, lower alkyl, lower alkenyl, - (CH)2)mOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)mSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)mNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R12H, lower alkyl, lower alkenyl, - (CH)2)mOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)mSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)mNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or; r33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)rCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)rCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H or lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)rPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R13Lower alkyl, lower alkenyl, - (CH)2)qOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)qSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)qNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)qOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form:-(CH2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)qNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)qN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)rCOO57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)qCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H or lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)rPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)rSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R14H, lower alkyl, lower alkenyl, - (CH)2)mOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)mSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)mNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mN(R20)COR64(wherein R is20H, lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H or lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl), - (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R15Lower alkyl, lower alkenyl, - (CH)2)oOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)oSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)oNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), (CH)2)oN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), particularly advantageous NR20CO lower alkyl (R)20H or lower alkyl), -CH (e.g. methyl-ethyl-methyl-), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R16Lower alkyl, lower alkenyl, - (CH)2)oOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)oSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)oNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H or lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R17Lower alkyl, lower alkenyl, - (CH)2)qOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)qSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)qNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)qOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)qNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)qN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)rCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)qCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)rPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)rSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
Among the structural units A1 to A69, the following are preferred: r2A5, A8, A22, A25, R being H2A38, A42, A47 and A50 for H. Most preferred are structural units of type A8':
Figure BDA0000736457340000401
wherein R is20Is H or lower alkyl; and, R64Is alkyl, alkenyl, [ (CH)2)u-X]t-CH3(wherein X is-O-, -NR-20or-S-, u is 1-3 and t is 1-6), aryl-lower alkyl or heteroaryl-lower alkyl; in particular wherein R64Is n-hexyl (A8' -1), n-heptyl (A8' -2), 4- (phenyl) benzyl (A8' -3), diphenylmethyl (A8' -4), 3-amino-propyl (A8' -5), 5-amino-pentyl (a8' -6), methyl (a8' -7), ethyl (a8' -8), isopropyl (a8' -9), isobutyl (a8' -10), n-propyl (a8' -11), cyclohexyl (a8' -12), cyclohexylmethyl (a8' -13), n-butyl (a8' -14), phenyl (a8' -15), benzyl (a8' -16), (3-indolyl) methyl (a8' -17), 2- (3-indolyl) ethyl (a8' -18), (4-phenyl) phenyl (a8' -19), n-nonyl (a8' -20), CH (a8' -7), n-nonyl (a8' -20)3-OCH2CH2-OCH2-and CH3-(OCH2CH2)2-OCH2-those of (a).
The structural unit A70 belongs to the class of open-chain alpha-substituted alpha-amino acids, the structural units A71 and A72 belong to the class of corresponding beta-amino acid analogs, and the structural units A73-A104 belong to the class of cyclic analogs of A70. Such amino acid derivatives have been shown to bind small peptides in a well-defined reverse-turn or U-shaped conformation (C.M. Venkatachalam, Biopolymers,1968,6, 1425-1434; W.Kabsch, C Sander, Biopolymers 1983,22, 2577). Ideally, such Building Blocks or templates are suitable for stabilizing the beta-hairpin conformation in the peptide loop (D.Obrecht, M.Altorfer, J.A.Robinson, "Novelpeptide Mimetic Building Blocks and strands for effective Lead Finding", adv.Med chem.1999, Vol.4, 1-68; P.Balarlaram, "Non-standing amino acids in peptides and proteins engineering", Current.Opin.Struct.1992.2, 845, M.Crisma, G.Valle, C.Toniolo, S.Prubidad, R.B.Rao, P.Hbaltam, "beta-transition structures for structures of peptides of proteins, R.B.Rao, P.Hbaltam," beta-hairpin structures of proteins, proteins of proteins, J.1990, Al.35-J.1990, Biocoding. J.35, J.1990, J.12, J.Biocoding. J.249, et al., Biocoding. J.J.35, et al., Biocoding. J.J.249, et al., Biocoding. J.J.J.S.12, J.J.J.J.Pat..
It has been shown that: two enantiomers of the Building block-A70-CO-to A104-CO-of the combined L-configuration Building block-B-CO-are capable of stabilizing and inducing beta-hairpin conformations with high efficiency (D.Obrecht, M.Altorfer, J.A.Robinson, "Novel Peptide Mimetic combinations Blocks and Strategies for Efficient lead filing", adv.Med chem.1999, Vol.4, 1-68; D.Obrecht, C.Spiegler, P).
Figure BDA0000736457340000411
K.Müller,H.Heimgartner,F.Stierli,Helv.Chim.Acta 1992,75,1666-1696；D.Obrecht,U.Bohdal,J.Daly,C.Lehmann,P.
Figure BDA0000736457340000412
K.Müller,Tetrahedron 1995,51,10883-10900；D.Obrecht,C.Lehmann,C.Ruffieux,P.
Figure BDA0000736457340000413
K.Müller,Helv.Chim.Acta 1995,78,1567-1587；D.Obrecht,U.Bohdal,C.Broger,D.Bur,C.Lehmann,R.Ruffieux,P.C.Spiegler,Helv.Chim.Acta 1995,78,563-580；D.Obrecht,H.Karajiannis,C.Lehmann,P.
Figure BDA0000736457340000422
C.Spiegler,Helv.Chim.Acta1995,78,703-714)。
Thus, for the purposes of the present invention, template (A1) may also consist of-A70-CO-to A104-CO-, where the structural units A70 to A104 are structural units-B-CO-having either the (D) -or the (L) -configuration in combination with the (L) -configuration.
R in A70-A10420Preferably H or lower alkyl, with methyl being most preferred. R in structural units A70 to A10418、R19And R21To R29Preferred values of (a) are as follows:
-R18lower alkyl.
-R19Lower alkyl, lower alkenyl, - (CH)2)pOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)pSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)pNR33R34(wherein R is33Lower alkyl or lower alkenyl, R34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)pCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)pCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H or lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)pSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)oC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R21H, lower alkyl, lower alkenyl, - (CH)2)oOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)oSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)oNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R22Lower alkyl, lower alkenyl, - (CH)2)oOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)oSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)oNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8H, F, Cl, CF, lower alkyl, lower alkenyl or lower alkoxy).
-R23H, lower alkyl, lower alkenyl, - (CH)2)oOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)oSR56(wherein R is56Lower alkyl or lower alkenyl), - (C)CH2)oNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-, wherein R57H or lower alkyl), - (CH)2)oN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), particularly advantageously NR20CO lower alkyl (R)20H or lower alkyl), -CH (e.g. methyl-ethyl-methyl-), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy);
-R24lower alkyl, lower alkenyl, - (CH)2)oOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)oSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)oNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl;or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), particularly advantageously NR20CO lower alkyl (R)20H or lower alkyl), -CH (e.g. methyl-ethyl-methyl-), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl, or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy);
-R25h, lower alkyl, lower alkenyl, - (CH)2)mOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)mNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(whereinR62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R26H, lower alkyl, lower alkenyl, - (CH)2)mOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)mNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or, R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-or, R25And R26May be taken together to form- (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl).
-R27H, lower alkyl, lower alkenyl, - (CH)2)oOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)oSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)oNR33R34(wherein R is33Lower alkyl or lower alkenyl, R34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(whereinR8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R28Lower alkyl, lower alkenyl, - (CH)2)oOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)oSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)oNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59：-(CH2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R29Lower alkyl, lower alkenyl, - (CH)2)oOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)oSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)oNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), particularly advantageously NR20CO lower alkyl (R)20H or lower alkyl), -CH (e.g. methyl-ethyl-methyl-), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
R23、R24And R29Is preferably-NR20-CO-lower alkyl, wherein R20Is H or lower alkyl.
For templates (b) through (p), e.g., (b1) and (l), preferred values for the plurality of symbols are as follows:
-R1h or lower alkyl;
-R8:H、F、Cl、CF3lower alkyl, lower alkenyl, - (CH)2)oOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)oSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)oNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H or lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R20H or lower alkyl.
-R30H and methyl.
-R31H, lower alkyl, lower alkenyl, - (CH)2)pOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)pNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), (-CH2)oCONR58R59(wherein R is58Lower alkyl, or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)rC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy); most preferred is-CH2CONR58R59(R58H or lower alkyl; r59Lower alkyl or lower alkenyl).
-R32H and methyl.
-R33Lower alkyl, lower alkenyl, - (CH)2)mOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)mNR34R63(wherein R is34Lower alkyl or lower alkenyl; r63H or lower alkyl; or R34And R63Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), (CH)2)mOCONR75R82(wherein R is75Lower alkyl or lower alkenyl; r82H or lower alkyl; or R75And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mNR20CONR78R82(wherein R is20H or lower alkyl; r78H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R78And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl).
-R34H or lower alkyl.
-R35H, lower alkyl, lower alkenyl, - (CH)2)mOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)mNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl),-(CH2)mN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl).
-R36Lower alkyl, lower alkenyl or aryl-lower alkyl.
-R37H, lower alkyl, lower alkenyl, - (CH)2)pOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)pNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R38H, lower alkyl, lower alkenyl, - (CH)2)pOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)pNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R78Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R39H, lower alkyl, lower alkenyl, - (CH)2)mOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)mN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl).
-R40Lower alkyl, lower alkenyl or aryl-lower alkyl.
-R41H, lower alkyl, lower alkenyl, - (CH)2)pOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)pNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R42H, lowLower alkyl, lower alkenyl, - (CH)2)pOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)pNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form:-(CH2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R43H, lower alkyl, lower alkenyl, - (CH)2)mOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)mSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)mNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lowerAn alkenyl group; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oPO(OR60)2(wherein R is60Lower alkyl or lower alkenyl), - (CH)2)oSO2R62(wherein R is62Lower alkyl or lower alkenyl) or- (CH)2)qC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R44Lower alkyl, lower alkenyl, - (CH)2)pOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)pSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)pNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R78Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)pN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)pCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)pCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl) or- (CH)2)oC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower,Lower alkenyl or lower alkoxy).
-R45H, lower alkyl, lower alkenyl, - (CH)2)oOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)oSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)oNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)sOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)oN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl) or- (CH)2)sC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R46H, lower alkyl, lower alkenyl, - (CH)2)sOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)sSR56(wherein R is56Lower alkyl or lower alkenyl), - (CH)2)sNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)sOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)sNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkylA group; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)sN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl) or- (CH)2)sC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R47H OR OR55(wherein R is55Lower alkyl or lower alkenyl).
-R48H or lower alkyl.
-R49H; lower alkyl, - (CH)2)oCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)oCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl) or (CH)2)sC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R50H and methyl.
-R51H, lower alkyl, lower alkenyl, - (CH)2)mOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)mNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), (CH)2)mOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lowerAlkenyl), - (CH)2)pCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)pCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl) or- (CH)2)rC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R52H, lower alkyl, lower alkenyl, - (CH)2)mOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)mNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Lower alkyl; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl；R82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-；R57H or lower alkyl), - (CH)2)mN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)pCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)pCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl) or- (CH)2)rC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R53H, lower alkyl, lower alkenyl, - (CH)2)mOR55(wherein R is55Lower alkyl or lower alkenyl), - (CH)2)mNR33R34(wherein R is33Lower alkyl or lower alkenyl; r34H or lower alkyl; or R33And R34Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mOCONR33R75(wherein R is33H or lower alkyl or lower alkenyl; r75Is low gradeAn alkyl group; or R33And R75Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mNR20CONR33R82(wherein R is20H or lower alkyl; r33H or lower alkyl or lower alkenyl; r82H or lower alkyl; or R33And R82Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl), - (CH)2)mN(R20)COR64(wherein R is20H or lower alkyl; r64Lower alkyl or lower alkenyl), - (CH)2)pCOOR57(wherein R is57Lower alkyl or lower alkenyl), - (CH)2)pCONR58R59(wherein R is58Lower alkyl or lower alkenyl; and R59H, lower alkyl; or R58And R59Together form: - (CH)2)2-6-、-(CH2)2O(CH2)2-、-(CH2)2S(CH2)2-or- (CH)2)2NR57(CH2)2-; wherein R is57H or lower alkyl) or- (CH)2)rC6H4R8(wherein R is8:H、F、Cl、CF3Lower alkyl, lower alkenyl or lower alkoxy).
-R54Lower alkyl, lower alkenyl or aryl-lower alkyl.
Most preferably, R1Is H; r20Is H; r30Is H; r31Is carboxymethyl or lower alkoxycarbonylmethyl; r32Is H; r35Is methyl; r36Is a methoxy group; r37Is H and R38Is H.
Among the structural units a70 to a104, the following are preferred: r22A74, A75, A76, R being H22A77, a78 and a79 for H.
The structural units-B-CO-in templates (a1), (a2) and (a3) refer to L-amino acid residues. A preferred value for B is-NR20CH(R71) -and R2A5, A8, A22, A25, R being H2Is an enantiomer of the a38, a42, a47, and a50 groups of H, most preferred are:
Figure BDA0000736457340000601
Figure BDA0000736457340000621
furthermore, the most preferred values of B also include groups of the type A8 "in the (L) -configuration:
Figure BDA0000736457340000622
wherein R is20Is H or lower alkyl; r64Is alkyl, alkenyl, - [ (CH)2)u-X]t-CH3(wherein X is-O-, -NR-20-or-S-, u is 1-3 and t is 1-6), aryl-lower alkyl or heteroaryl-lower alkyl; especially wherein R64Is n-hexyl (A8' -21), n-heptyl (A8' -22), 4- (phenyl) benzyl (A8' -23), diphenylmethyl (A8' -24), 3-amino-propyl (A8' -25), 5-amino-pentyl (A8' -26), methyl (A8' -27), ethyl (A8' -28), iso-pentyl (A8' -28)Propyl (A8' -29), isobutyl (A8' -30), n-propyl (A8' -31), cyclohexyl (A8' -32), cyclohexylmethyl (A8' -33), n-butyl (A8' -34), phenyl (A8' -35), benzyl (A8' -36), (3-indolyl) methyl (A8' -37), 2- (3-indolyl) ethyl (A8' -38), (4-phenyl) phenyl (A8' -39), n-nonyl (A8' -40), CH (CH) 8' -37)3-OCH2CH2-OCH2- (A8 "-41) and CH3-(OCH2CH2)2-OCH2Those of (A8 "-42).
Typically, the peptide chain Z of the β -hairpin mimetic described herein is defined by an amino acid residue belonging to one of the following groups:
the radical C-NR20CH(R72) CO-; "hydrophobic: small to medium sized "
The radical D-NR20CH(R73) CO-; "hydrophobic: large aromatic or heteroaromatic compounds "
The radicals E-NR20CH(R74) CO-; "polar cationic" and "urea-derived"
The group F-NR20CH(R84) CO-; polar uncharged or anionic "
The radical H-NR20-CH(CO-)-(CH2)4-7-CH(CO-)-NR20-、-NR20-CH(CO-)-(CH2)pSS(CH2)p-CH(CO-)-NR20-、-NR20-CH(CO-)-(-(CH2)pNR20CO(CH2)p-CH(CO-)-NR20-and-NR20-CH(CO-)-(-(CH2)pN R20CONR20(CH2)p-CH(CO-)-NR20-；
"interchain bond"
Furthermore, the amino acid residues in the chain Z may also have the formula-A-CO-or the formula-B-CO-, wherein A and B are as defined above. Finally, Gly can also be an amino acid residue in chain Z, and Pro (4-NHCOPhe) can also be amino acid residues in chain Z, except possibly at the position of the interchain bond (H).
The group C comprises a compound having a small atomSize to medium size hydrophobic side chain groups (according to R for substituent72General definition of (d) amino acid residues. Hydrophobic residues refer to amino acid side chains that are uncharged at physiological pH and are rejected by aqueous solutions. In addition, these side chains are typically free of hydrogen bond donor groups such as, but not limited to, primary and secondary amides, primary and secondary amines and their corresponding protonated salts, thiols, alcohols, phosphonates, phosphates, ureas, or thioureas. However, they may contain hydrogen bond acceptor groups such as ethers, thioethers, esters, tertiary amides, alkyl or aryl phosphonates and phosphates or tertiary amines. The small to medium size amino acids encoded by the gene include alanine, isoleucine, leucine, methionine and valine.
The radical D comprises radicals having aromatic and heteroaromatic side chains (according to the formula for the substituent R)73General definition of (d) amino acid residues. Aromatic amino acid residues refer to hydrophobic amino acids having a side chain containing at least one ring with a conjugated pi-electron system (aromatic group). In addition, they may contain hydrogen bond donor groups (such as, but not limited to, primary and secondary amides, primary and secondary amines and their corresponding protonated salts, thiols, alcohols, phosphonates, phosphates, ureas, or thioureas) and hydrogen bond acceptor groups (such as, but not limited to, ethers, thioethers, esters, tertiary amides, alkyl or aryl phosphonates and phosphates or tertiary amines). Aromatic amino acids encoded by a gene include phenylalanine and tyrosine.
A heteroaromatic amino acid residue refers to a hydrophobic amino acid residue having a side chain with at least one ring with a conjugated pi-electron system including at least one heteroatom, such as, but not limited to O, S and N (depending on the R for substituent)77General definition of (ii). In addition, such residues may contain hydrogen bond donor groups (such as, but not limited to, primary and secondary amides, primary and secondary amines and their corresponding protonated salts, thiols, alcohols, phosphonates, phosphates, ureas, or thioureas) and hydrogen bond acceptor groups (such as, but not limited to, ethers, thioethers, esters, tertiary amides, alkyl or aryl phosphonates and phosphates or tertiary amines). Gene-encoded heteroaromatic amino acidsIncluding tryptophan and histidine.
Group E comprises amino acids having side chains with polar cations, amido groups, and urea-derived residues (as per for substituent R)74General definition of (ii). Polar cations refer to basic side chains that are protonated at physiological pH. The polar cationic amino acids encoded by the gene include arginine, lysine and histidine. Citrulline is an example of a urea-derived amino acid residue.
Group F comprises amino acids having a side chain with a polar uncharged or anionic residue (as for substituent R)84General definition of (ii). Polar uncharged or anionic residues refer to hydrophilic side chains which are uncharged and anionic (carboxylic acids included) respectively at physiological pH, but which are not repelled by aqueous solutions. Typically, such side chains contain hydrogen bond donor groups such as, but not limited to, primary and secondary amides, carboxylic acids and esters, primary and secondary amines, thiols, alcohols, phosphonates, phosphates, ureas, or thioureas. These groups can form a hydrogen bonding network with water molecules. In addition, they may also contain hydrogen bond acceptor groups such as, but not limited to, ethers, thioethers, esters, tertiary amides, carboxylic acids and esters (salts) of carboxylic acids, alkyl or aryl phosphonates and phosphates or tertiary amines. The polar uncharged amino acids encoded by the gene include asparagine, cysteine, glutamine, serine and threonine, and also include aspartic acid and glutamic acid.
The group H comprises side chains which are preferably (L) -amino acids in opposite positions in the beta-chain region, which may form inter-chain bonds. The most well known bond is the disulfide bond formed by cysteine and homo-cysteine located opposite the β -chain. Various methods are known for disulfide bond formation, including J.P.Tam et al Synthesis 1979, 955-957; stewart et al, Solidphase Peptide Synthesis,2d Ed., Pierce Chemical Company, III., 1984; ahmed et al J.biol.chem.1975,250, 8477-8482; and those described by Pennington et al, Peptides, page 164-166, Giralt and Andreu, eds., ESCOM Leiden, the Netherlands, 1990. Most advantageously, for the scope of the present invention, a disulfide bond can be made using an acetamidomethyl (Acm) -protecting group for cysteine. A well established interchain bond consists in linking ornithine and lysine, respectively, to glutamic and aspartic acid residues located at opposite beta-strand positions through the formation of an amide bond. The preferred protecting group is allyloxycarbonyl (Alloc) for the side chain amino groups of ornithine and lysine and allyl ester for aspartic acid and glutamic acid. Finally, interchain linkages may also be established by linking the amino groups of lysine and ornithine in opposite β -strand positions with reagents such as N, N-carbonylimidazole to form cyclic ureas.
As previously described, the positions for the interchain bonds are positions P2 and 10 taken together. Such interchain linkages are known to stabilize the β -hairpin conformation and thus constitute important structural elements for the design of β -hairpin mimetics.
The most preferred amino acid residues in the chain Z are those derived from natural alpha-amino acids. Suitable amino acids (or their residues) for the purposes of the present invention are listed below, the abbreviations corresponding to the usual practice generally adopted:
three-letter code and single-letter code
Ala L-alanine A
Arg L-arginine R
Asn L-asparagine N
Asp L-aspartic acid D
Cys L-cysteine C
Glu L-glutamic acid E
Gln L-Glutamine Q
Gly glycine G
His L-histidine H
Ile L-isoleucine I
Leu L-leucine L
Lys L-lysine K
Met L-methionine M
Phe L-phenylalanine F
Pro L-proline P
DPro D-prolineDP
Ser L-serine S
Thr L-threonine T
Trp L-Tryptophan W
Tyr L-tyrosine Y
Val L-valine V
Other α -amino acids (or residues thereof) suitable for the purposes of the present invention include:
Figure BDA0000736457340000661
Figure BDA0000736457340000681
Figure BDA0000736457340000691
particularly preferred residues for the group C are:
Figure BDA0000736457340000692
Figure BDA0000736457340000701
particularly preferred residues for the group D are:
Figure BDA0000736457340000702
Figure BDA0000736457340000711
particularly preferred residues for the group E are:
Figure BDA0000736457340000721
particularly preferred residues for the group F are:
typically, the peptide chain Z in the β -hairpin mimetic of the invention comprises 11 amino acid residues. Positions P1 to P11 of each amino acid residue in the chain Z are explicitly defined as follows: p1 represents the first amino acid in chain Z which is coupled with its N-terminus to the C-terminus of templates (B) - (P) or to the C-terminus of the-B-CO-group in template (a1), the C-terminus of the-A-CO-group in template (a2), the C-terminus of the-B-CO-group forming the C-terminus of template (a 3); p11 represents the last amino acid in chain Z which is coupled with its C-terminus to the N-terminus of templates (B) - (P) or to the N-terminus of the-A-CO-group in template (a1), the N-terminus of the-B-CO-group in template (a2), the N-terminus of the-B-CO-group forming the N-terminus of template (a 3). Each of positions P1 to P11 preferably contains an amino acid residue belonging to one of the C, D, E, F, H types described above, or having the formula-a-CO-or having the formula-B-CO-, or, alternatively, Gly, Pro, or Pro (4NHCOPhe), as follows:
in general, the α -amino acid residues in positions 1 to 11 of the chain Z are preferably:
-P1 type C or D or E or F;
-P2 type E or F or C;
-P3, type C or F, or the residue is Gly;
-P4 type C or E or F, or the residue is Gly or Pro;
-P5, type E or F, or the residue is Gly or Pro;
-P6 type C or D or F, or the residue is Gly or Pro;
-P7, form F or of formula-A-CO-, or the residue is Gly or Pro;
-P8, D or C or of formula-A-CO-, or the residue is Gly or Pro (4 NHCOPhe);
-P9 type C or D or E or F;
-P10 type F or C or E:
-P11 type E or F or C or D; or
-P2 and P10 together form a group of H type;
with the following conditions: if the template isDPro-LPro, then the amino acid residues from P1 to P11 are not:
-P1:Arg
cys linked to Cys at position P10 by a disulfide bond-P2
-P3:Thr
-P4Lys
-P5Ser
-P6Ile
-P7Pro
-P8Pro
-P9Ile
-Cys of P10 linked to Cys at P10 by a disulfide bond; and
-P11Phe。
the α -amino acid residues in positions 1 to 11 are most preferably:
-P1: Nle, Ile, Aoc, hLeu, Chg, OctG, hPhe, 4AmPhe, Cha, Phe, Tyr, 2Cl-Phe, Trp, 1-Nal, Leu, Cha, or Arg;
-P2 Cys, Glu, Nle, Thr or Gln;
-P3 Thr, Ala or Abu;
-P4 Lys, Nle, Ala, Abu or Thr;
-P5 Ser, AlloThr or Dpr;
-P6 Ile, c5al, Leu, Nle, Aoc, OctG, Cha, hLeu, hPhe, Chg, t-BuA, Glu or Asp;
-P7:Pro；
p8 Pro, Ala or Pro (4 NHCOPhe);
-P9 Tyr, Phe, Ile, Nle, Cha, Gln, Arg, Lys, His, Thr, or Ala;
-P10 Cys, Arg, Nle, Gln, Lys, Met, Thr or Ser;
-P11 Tyr, Gln, Arg, Ser, Nle, 2-Nal, 2Cl-Phe, Cha, Phg, Tyr, Phe, Asp, Asn or Thr; and
cys, which can form a disulfide bond if present at P2 and P10.
In the case of inhibitors of cathepsin G, the α -amino acid residues in positions 1 to 11 of chain Z are preferably:
-P1, form C or D or E;
-P2 type F or C;
-P3: form F;
-P4, type C or E;
-P5, form E or F;
-P6: form F;
-P7, form F or of formula-A-CO-, or the residue is Gly or Pro;
-P8, type C or of formula-A-CO-, or the residue is Gly or Pro (4 NHCOPhe);
-P9 type C or D or F;
-P10 type F or C or E;
-P11, form E or D or F; or
-P2 and P10 together form a group of the H type.
For inhibitors of cathepsin G, the α -amino acid residues in positions 1 to 11 are most preferably:
-P1 Phe, hPhe, 4AmPhe, Nle, Chg, Ile, Tyr, Arg, Trp, 2Cl-Phe, Arg, 1-Nal or Cha;
-P2 Cys, Glu or Nle;
-P3:Thr；
-P4: Lys or Nle;
-P5 Ser, AlloThr or Dpr;
-P6 Asp or Glu;
-P7:Pro；
-P8:Pro；
-P9 Ile, Nle, Cha, Gln, Tyr or Ala;
-P10 Cys, Arg or Nle;
-P11 Thr, Asp, Ser, Tyr, Phe, Asn or Arg; and
cys, which can form a disulfide bond if present at P2 and P10.
In the case of inhibitors of elastase, the α -amino acid residues in positions 1 to 11 of chain Z are preferably:
-P1, form C or D;
-P2: form F;
-P3 type F or C;
-P4, type C or F;
-P5: form F;
-P6: type C;
-P7 of formula-A-CO-, or the residue is Gly or Pro;
-P8 of formula-A-CO-, or the residue is Gly or Pro (4 NHCOPhe);
-P9, form D or F or C;
-P10 type F or C or E;
-P11, form E or F or D; or
-P2 and P10 together form a group of the H type.
For inhibitors of elastase, the α -amino acid residues at positions 1 to 11 are most preferably:
-P1 Ile, Nle, Aoc, hLeu, Chg, OctG or hPhe;
-P2 Cys, Glu, Thr or Gln;
-P3 Thr, Ala or Abu;
-P4: Ala, Thr or Abu;
-P5:Ser；
-P6 OctG, Ile, Cha, Leu, c5al, Nle, Aoc, Chg, tBuA or hLeu;
-P7:Pro；
pro or Pro (4NHCOPhe) P8;
-P9 Gln, Tyr, ILe or Phe;
-P10 Cys, Lys, Gln, Thr, Met or Arg;
-P11 Tyr, Ser, Arg, Gln, Nle, 2-Nal, 2Cl-Phe, Cha, or Phg; and
cys, which can form a disulfide bond if present at P2 and P10.
In the case of tryptase inhibitors, the α -amino acid residues in positions 1 to 11 of chain Z are preferably:
-P1, form C or D or E;
-P2: form F;
-P3: form F;
-P4: form E;
-P5: form F;
-P6, form C or D;
-P7, form F or of formula-A-CO-, or the residue is Gly or Pro;
-P8, form C or of formula-A-CO-, or the residue is Gly or Pro;
-P9 type C or E or F;
-P10: form F;
-P11, form E or D; or
-P2 and P10 together form a group of H type;
with the following conditions: if the template isDPro-LPro, then the amino acid residues from P1 to P11 are not:
-P1:Arg
cys linked to Cys at position P10 by a disulfide bond-P2
-P3:Thr
-P4 Lys
-P5 Ser
-P6 Ile
-P7 Pro
-P8 Pro
-P9 Ile
-Cys of P10 linked to Cys at P10 by a disulfide bond; and
-P11 Phe。
for tryptase inhibitors, the α -amino acid residues in positions 1 to 11 of chain Z are most preferably:
cha, Tyr or Trp, P1
-P2:Cys
-P3:Thr
-P4:Lys
-P5:Ser
-P6:Leu
-P7:Pro
-P8:Pro
-P9:Lys
-P10:Cys
-P11: Arg; and
the Cys residues present at positions P2 and P10 may form a disulfide bond.
Particularly preferred β -peptide mimetics of the invention include those disclosed in examples 5, 19, 20, 22, 23, 38, 39, 40 and 75 as cathepsin G inhibitors; those disclosed as elastase inhibitors in examples 91, 121, 153, 154, 155, 156, 157, 158, 159, 160, 161177 and 178; and those disclosed as tryptase inhibitors in examples 193, 194 and 195.
The methods of the invention can be advantageously practiced as a parallel array synthesis to generate a library of template-fixed β -hairpin peptidomimetics of formula I above. Such parallel syntheses allow large arrays (typically 24 to 192, preferably 96) of compounds of formula I to be obtained in high yields and with defined purity, minimizing the formation of dimer and polymer by-products. Thus, proper selection of the functionalized solid support (i.e., solid support plus linker molecule), template, and cyclization sites plays an important role.
Can be prepared from polystyrene cross-linked with preferably 1-5% divinylbenzene, polystyrene coated with polyethylene glycol spacers (spacers)R) And polyacrylamide resins are conveniently obtained as a functionalized solid support (see also Obrecht d; villalgordo, J. -M, "Solid-Supported Combinatorial and parallel Synthesis of Small-Molecular-Weight Compound Libraries", Tetrahedron organic chemistry Series, Vol.17, Pergamon, Elsevier Science, 1998).
The solid support may be functionalized by a linker, i.e. a bifunctional spacer molecule containing an anchor group at one end for attachment to the solid support and a selectively cleavable functional group at the other end for subsequent chemical transformation and cleavage procedures. For the purposes of the present invention, two types of linker are used:
type 1 linkers are designed to release the amide group under acidic conditions (Rink H, Tetrahedron Lett.1987,28, 3783-. This type of linker forms an amide of the amino acid carboxyl group; examples of resins functionalized by such linker structures include 4- [ ((((2, 4-dimethoxyphenyl) Fmoc-aminomethyl) phenoxyacetamido) aminomethyl ] PS resin, 4- [ (((2, 4-dimethoxyphenyl) Fmoc-aminomethyl) phenoxyacetamido) aminomethyl ] -4-methylbenzhydrylamine PS resin (Rink amide MBHA PS resin), and 4- [ ((((2, 4-dimethoxyphenyl) Fmoc-aminomethyl) phenoxyacetamido) aminomethyl ] benzhydrylamine PS-resin (Rink amide BHA PS resin). Preferably, the support is obtained from polystyrene cross-linked with most preferably 1-5% divinylbenzene and functionalized with a 4- (((2, 4-dimethoxyphenyl) Fmoc-aminomethyl) phenoxyacetamido) linker.
Type 2 linkers are designed to eventually release the carboxyl group under acidic conditions. This type of linker forms acid labile esters with the carboxyl group of amino acids, typically acid labile benzyl, benzhydryl and trityl esters; examples of such linker structures include 2-methoxy-4-hydroxymethylphenoxy (Sasrin)RLinker), 4- (2, 4-dimethoxyphenyl-hydroxymethyl) -phenoxy (Rink linker), 4- (4-hydroxymethyl-3-methoxyphenoxy) butyric acid (HMPB linker), trityl and 2-chlorotrityl. Preferably, the support is obtained from polystyrene cross-linked with most preferably 1-5% divinylbenzene, and functionalized with a 2-chlorotrityl linker.
When the process of the invention is carried out as a parallel array synthesis, it may be advantageously carried out as described below, but it will be immediately apparent to the skilled person how to vary these procedures when one wishes to synthesize a single compound of formula I as described above.
A large number of reaction vessels (generally 24 to 192, typically 96) equal to the total number of compounds to be synthesized by the parallel method are loaded with 25 to 1000mg, preferably 100mg, of a suitable functionalized solid support, preferably obtained from polystyrene cross-linked with 1 to 3% divinylbenzene, or from Tentagel resin.
Solvents to be used must swell the resin and include, but are not limited to, Dichloromethane (DCM), Dimethylformamide (DMF), N-methylpyrrolidinone (NMP), dioxane, toluene, Tetrahydrofuran (THF), ethanol (EtOH), Trifluoroethanol (TFE), isopropanol, and the like. Solvent mixtures containing at least one component polar solvent (e.g., 20% TFE/DCM, 35% THF/NMP) are advantageous for ensuring and dissolving high reactivity and solvation of the peptide chain bound to the resin (Fields, g.b., Fields, c.g., j.am. chem. soc.1991,113, 4202-4207).
Considerable progress has been made in the synthesis of protected peptide fragments with the development of various linkers that release a C-terminal carboxylic acid group under mildly acidic conditions and do not affect the acid labile groups that protect functional groups in the side chains. 2-methoxy-4-hydroxybenzyl alcohol-derived linker (Sasrin)RThe linker, Mergler et al, Tetrahedron Lett.1988,294005-4008) was cleavable with diluted trifluoroacetic acid (0.5-1% TFA in DCM), and was stable to Fmoc deprotection conditions during peptide synthesis, with additional protecting groups based on Boc/tBu being compatible with this protection scheme. Other linkers suitable for use in the methods of the invention include: super acid-labile 4- (2, 4-dimethoxyphenyl-hydroxymethyl) -phenoxy linker (Rink linker, Rink, h.tetrahedron lett.1987,28, 3787-sec 3790) where removal of the peptide requires 10% acetic acid in DCM or 0.2% trifluoroacetic acid in DCM; 4- (4-hydroxymethyl-3-methoxyphenoxy) butanoic acid derived linker (HMPB-linker,&riniker, Peptides1991,1990131) which can also be cleaved by 1% TFA/DCM to generate peptide fragments containing all acid labile side chain protecting groups; and also, a 2-chlorotrityl chloride linker (Barlos et al, Tetrahedron Lett.1989,30, 3943-.
Suitable protecting groups for amino acids and their residues, respectively, are, for example:
for amino groups (as present, e.g. also in lysine side chains)
Figure BDA0000736457340000811
For carboxyl groups (if present, for example also in the aspartic and glutamic acid side chains), by conversion into esters with components of alcohols
Figure BDA0000736457340000812
Figure BDA0000736457340000821
For guanidino groups (as present, e.g. in arginine side chains)
Figure BDA0000736457340000822
For hydroxy groups (as present, e.g. in threonine and serine side chains)
tBu tert-butyl
Bn benzyl group
Trt trityl radical
And for mercapto groups (as present, e.g. in cysteine side chains)
Figure BDA0000736457340000823
Preferably, 9-fluorenylmethyloxycarbonyl (Fmoc) -protected amino acid derivatives are used as building blocks for the construction of template-fixed beta-hairpin loop mimetics of formula I. For deprotection, i.e. cleavage of the Fmoc group, 20% piperidine in DMF or 2% DBU/2% piperidine in DMF may be used.
The amount of reactant (i.e., amino acid derivative) is typically 1 to 20 equivalents based on milliequivalents (meq/g) of functionalized solid support per gram of loading initially weighed into the reaction tube (typically 0.1 to 2.85meq/g for polystyrene resins). Additional equivalents of reactants may be used, if desired, to drive the reaction to completion in a reasonable time. The reaction cuvette was reinserted into the storage section (reservoir block) in combination with the tube rack section (holder block) and manifold (maniffold), and the device was secured together. The gas flow through the manifold is initialized to provide a controlled environment, such as nitrogen, argon, air, etc. The gas stream may also be heated or cooled before flowing through the manifold. Heating or cooling of the reaction well is performed by heating the reaction table or cooling with isopropyl alcohol/dry ice or the like to cause the desired synthesis reaction to occur. Agitation is achieved by shaking or magnetic stirring (in a reaction tube). Preferred workstations, but not limited to, are Labsource's Combi-chem workstation and the MultiSyn Tech's-Syro synthesizer.
Amide bond formation requires activation of the alpha-carboxyl group for the acylation step. When passing through the usual carbodiimides, e.g. dicyclohexylcarbodiimide (DCC, Sheehan)&When Hess, J.Am.chem.Soc.1955,77,1067-1068) or diisopropylcarbodiimide (DIC, Saratakis et al biochem.Biophys.Res.Commun.1976,73,336-342) are subjected to such activation, dicyclohexylurea and diisopropylurea, respectively, are insoluble and soluble in the solvents conventionally used. In one variation of the carbodiimide process, a process involving 1-hydroxybenzotriazole (HOBt,
Figure BDA0000736457340000831
&geiger, chem. Ber 1970,103,788-798) as an additive to the coupling mixture. HOBt prevents dehydration, inhibits racemization of activated amino acids, and acts as a catalyst to advance slow coupling reactions. Certain phosphorus reagents have been tried as direct coupling reagents, such as benzotriazol-1-yl-oxy-tris- (dimethylamino) -phosphonium hexafluorophosphate (BOP, Castro et al, Tetrahedron Lett.1975,14, 1219; Synthesis,1976,751-752) or benzotriazol-1-yl-oxy-tris-pyrrolidinyl-phosphonium hexafluorophosphate (Py-BOP, Coste et al, TetraDron Lett.1990,31, 205-; these phosphorus reagents are also suitable for in situ formation of HOBt esters with protected amino acid derivatives. More recently, Diphenoxyphosphorylazide (DPPA) or O- (7-aza-benzotriazol-1-yl) -N, N, N ', N' -tetramethyluronium tetrafluoroborate (TATU) or O- (7-aza-benzotriazol-1-yl) -N, N, N ', N' -tetramethyluronium Hexafluorophosphate (HATU)/7-aza-1-hydroxybenzotriazole (HOAt, Carpino et al, Tetrahedron Lett.1994,35, 2279-.
Since it is necessary to perform a near quantitative coupling reaction, experimental evidence of reaction completion is required. Ninhydrin tests (Kaiser et al, anal. biochemistry 1970,34,595) can be performed quickly and easily after each coupling step, where a positive colorimetric reaction against an aliquot of the resin-bound peptide quantitatively indicates the presence of a primary amine. The Fmoc chemistry method allows spectrophotometric detection of Fmoc chromophores when released with bases (Meienhofer et al, int.J. peptide Protein Res.1979,13, 35-42).
The resin bound intermediate in each reaction tube is washed free of excess residual reagents, solvents and by-products by repeated exposure to neat solvent by either:
1) loading a solvent (preferably, 5ml) into the reaction wells, immersing the reaction tubes and the tube rack components and manifold, agitating for 5 to 300 minutes, preferably, 15 minutes, and draining by gravity, followed by draining the solvent by applying air pressure through the manifold inlet (while closing the outlet);
2) the manifold is removed from the tube rack assembly, an aliquot (preferably 5ml) of solvent is dispersed through the top of the reaction tube, and drained by gravity through a filter into a receiving container (e.g., a tube or vial).
The above two washing procedures are repeated up to about 50 times (preferably, about 10 times), and the efficiency of the removal of the reagents, solvents and by-products is monitored by TLC, GC or the like or by observing the washings.
For each successive conversion, the procedure described above for reacting the resin bound compound with the reagent in the reaction well followed by removal of excess reagent, by-products and solvent is repeated until the final resin bound, fully protected linear peptide is obtained.
Prior to dissociation of such fully protected linear peptides from the solid support, one or more protected functional groups present in the molecule may be selectively deprotected, if desired, and the reactive groups released thereby suitably replaced. To achieve this effect, the functional group to be treated must initially be protected with a protecting group which can be selectively removed and which does not interfere with the remaining protecting groups present. Alloc (allyloxycarbonyl) is selectively removed (e.g., via CH)2Cl2Phenyl silane and Pd deg.) and does not affect the remaining protecting groups (e.g., Fmoc) present in the molecule. The reactive groups thus released may then be treated with a reagent suitable for introducing the desired substituent. Thus, for example, an amino group can be acylated by an acylating agent corresponding to the acyl substituent to be introduced. To form the pegylated amino acid, e.g., IPegK or SPegK, preferably 5 equivalents of HATU (N- [ (dimethylamino) -1H-1,2, 3-triazolo [4,5-b ] hexafluorophosphate in dry DMF]Pyridin-1-ylmethylene]-N-methylmethanamine N-oxide) and a solution of 10 equivalents of DIPEA (diisopropylethylamine) in anhydrous DMF, and, respectively, 5 equivalents of 2- [2- (2-methoxyethoxy) ethoxy ] ethoxy]Acetic acid (lPeg) and 2- (2-methoxyethoxy) acetic acid (sPeg) were applied to the liberated amino group of the appropriate amino acid side chain for 3 hours. After this time the resin was filtered and washed, the procedure was repeated with a fresh solution of the reagents for another 3 hours.
Prior to dissociation of the fully protected linear peptide from the solid support, interchain linkages may also be formed between the side chains of the appropriate amino acid residues at positions 2 and 10, if desired.
Interchain bonds and their formation have been discussed above in connection with the explanations for the group of the H type, which may be, for example, a disulfide bond formed between cysteine and homocysteine residues at opposite positions of the β -chain, or a lactam bridge formed by glutamic and aspartic acid residues linked to ornithine and lysine residues, respectively, at opposite β -chain positions, or a glutamic acid residue linked to a2, 4-diaminobutyric acid residue, via an amide bond. The formation of such interchain bonds can be accomplished by methods well known in the art.
To form disulfide bonds, preferably 10 equivalents of iodine solution is applied to DMF, or to CH2Cl21.5 hours in the/MeOH mixture, after filtration of the iodine solution, repeated for another 3 hours with fresh iodine solution, or else 10 equivalents of iodine solution were applied to the mixture of DMSO and acetic acid solution (with 5% NaHCO)3Buffered to pH 5-6) for 4 hours, or applied by stirring for 24 hours to water adjusted to pH 8 with ammonium hydroxide solution, or to ammonium acetate buffer adjusted to pH 8 in the presence of air, or to NMP and tri-n-butylphosphine (preferably, 50 equivalents).
Dissociation of the fully protected linear peptide from the solid support is achieved by dipping the reaction tube and tube rack components and manifold into a reaction well containing a lysis reagent solution (preferably 3 to 5 ml). Gas flow, temperature control, agitation, and reaction monitoring are performed as described above and as needed to achieve dissociation reactions. The reaction tubes, as well as the tube holder assembly and manifold, are removed from the storage section, lifted above the solution level but below the upper edge of the reaction wells, and air pressure is applied via the manifold inlet (while the outlet is closed) to efficiently discharge the final product solution into the storage wells. The remaining resin in the reaction tube is then washed 2 to 5 times with 3 to 5ml of a suitable solvent as described above to extract (wash out) as much of the cleavage product as possible. The product solutions thus obtained were combined and carefully carried out to avoid cross-mixing. The various solutions/extracts are then manipulated as required to isolate the final compound. Typical operations include, but are not limited to: evaporation, concentration, liquid/liquid extraction, acidification, basification, neutralization or other reactions in solution.
The solution containing the fully protected linear peptide derivative which has been cleaved from the solid support and neutralized with a base is subjected to evaporation. The cyclization is then carried out in solution using solvents such as DCM, DMF, dioxane, THF, etc. A variety of coupling reagents as previously mentioned can be used for the cyclization. The cyclization is continued for about 6 to 48 hours, preferably about 16 hours. The progress of the reaction is followed, for example, by RP-HPLC (reverse phase high performance liquid chromatography). The solvent is then removed by evaporation, the fully protected cyclic peptide derivative is dissolved in a water-immiscible solvent (e.g. DCM) and the solution is extracted with water or a mixture of water-miscible solvents to remove any excess coupling reagent.
Finally, with 95% TFA, 2.5% H2O, 2.5% TIS or other combinations of scavengers to effect cleavage of the protecting group. The cleavage reaction time is usually 30 minutes to 12 hours, preferably, about 2.5 hours. The volatiles were evaporated to dryness, the crude peptide was dissolved in 20% AcOH in water and extracted with isopropyl ether or other solvent suitable for this. The aqueous layer was collected and evaporated to dryness to obtain the fully deprotected cyclic peptide derivative of formula I as the final product.
Alternatively, dissociation from the solid support, cyclization and complete deprotection of the fully protected peptide can be achieved manually in glass containers.
Depending on its purity, the peptide derivative can be used directly in the biological assay, or it must be further purified, for example by preparative HPLC.
As mentioned before, the fully deprotected product of formula I thus obtained may then be converted into a pharmaceutically acceptable salt, or a pharmaceutically acceptable or unacceptable salt thus obtained may be converted into the corresponding free compound of formula I, or into a different, pharmaceutically acceptable salt, if desired. Any of these operations may be performed by methods well known in the art.
The template material of formula II used in the process of the present invention, the pre-starting materials (pre-starting materials) material therefor and the preparation of these materials and the pre-starting materials are described in the international application PCT/EP02/01711 of the same applicant, which is disclosed as WO02/070547 a 1.
The β -hairpin peptidomimetics of the invention are useful in a wide range of applications where inflammatory or pulmonary diseases or infections or immune diseases or cardiovascular diseases or neurodegenerative diseases are mediated or caused by serine protease activity or where cancer is mediated or caused by serine protease activity. For the control or prevention of a given condition or disease that can be treated with a protease inhibitor, the β -hairpin peptidomimetic can be administered per se or as a suitable formulation together with carriers, diluents or excipients well known in the art.
When β -hairpin peptidomimetics are used to treat or prevent emphysema, rheumatoid arthritis, osteoarthritis, arteriosclerosis, psoriasis, cystic fibrosis, multiple sclerosis, adult respiratory distress syndrome, pancreatitis, asthma, allergic rhinitis, inflammatory skin diseases, post-angioplasty restenosis, cardiac hypertrophy, heart failure, or cancer (such as, but not limited to, breast cancer or cancers associated with angiogenesis or metastasis), the β -hairpin peptidomimetics may be administered alone, as a mixture of multiple β -hairpin peptidomimetics, in combination with other anti-inflammatory or antimicrobial or anti-cancer agents, and/or in combination with other pharmaceutically active agents. The β -hairpin peptidomimetics can be administered as such or as a pharmaceutical composition.
Pharmaceutical compositions comprising the β -hairpin peptidomimetics of the invention may be produced by conventional mixing, dissolving, granulating, manufacturing coated tablets, finely grinding, emulsifying, encapsulating, entrapping (entrap) or lyophilizing processes. Pharmaceutical compositions may be formulated by conventional means using one or more physiologically acceptable carriers, diluents, excipients or adjuvants which facilitate processing of the active β -hairpin peptidomimetic into a pharmaceutically acceptable formulation. The appropriate formulation depends on the method of administration selected.
For topical administration, the beta-hairpin peptidomimetics of the present invention can be formulated as solutions, gels, ointments, creams, suspensions, and the like, as is well known in the art.
Systemic formulations include those designed for administration by injection (e.g., subcutaneous, intravenous, intramuscular, intrathecal, or intraperitoneal injection), as well as those designed for transdermal, transmucosal, oral, or pulmonary administration.
For injection, the β -hairpin peptidomimetics of the invention can be formulated in a suitable solution, preferably in a physiologically compatible buffer, for example, a Hink's solution, ringer's solution or physiological saline buffer. The solution may contain formulating substances such as suspending, stabilizing and/or dispersing agents. Alternatively, the β -hairpin peptidomimetics of the invention may be in powder form for combination with a suitable vehicle (e.g., sterilized pyrogen-free water) prior to use.
For transmucosal administration, penetrants appropriate to the barrier to be permeated are used in the formulation, as is known in the art.
For oral administration, the β -hairpin peptidomimetics of the invention can be readily formulated by combining them with pharmaceutically acceptable carriers known in the art. Such carriers enable the beta-hairpin peptidomimetics of the invention to be formulated as tablets, pills, dragees, capsules, liquids, gels, syrups, slurries, suspensions and the like, for oral formulation by a patient to be treated. For oral formulations (e.g., powders, capsules, and tablets), suitable excipients include fillers (e.g., sugars such as lactose, sucrose, mannitol, and sorbitol; cellulose preparations such as corn starch, wheat starch, rice starch, potato starch, gelatin, tragacanth, methyl cellulose, hydroxypropylmethyl cellulose, sodium carboxymethyl cellulose, and/or polyvinylpyrrolidone (PVP)), granulating agents, and binding agents. If desired, disintegrating agents can be added, for example crosslinked polyvinylpyrrolidone, agar or alginic acid or a salt thereof (e.g. sodium alginate). The solid dosage forms may be sugar coated or enteric coated, if desired, using standard techniques.
For oral liquid preparations (e.g., suspensions, elixirs and solutions), suitable carriers, excipients or diluents include water, glycols, oils, alcohols and the like. In addition, flavoring agent, antiseptic, colorant, etc. can be added.
For buccal administration, the compositions may take the form of tablets, lozenges, and the like, which are generally formulated.
For administration by inhalation, the β -hairpin peptidomimetics of the present invention may be conveniently delivered in the form of an aerosol spray from a pressurized pack or a nebulizer, using a suitable propellant, e.g., dichlorodifluoromethane, trichlorofluoromethane, carbon dioxide or other suitable gas. In the case of a pressurized aerosol, the dosage unit may be determined by providing a valve to deliver a metered amount. Capsules and cartridges of, for example, gelatin for use in an inhaler or insufflator may be formulated containing a powder mix of the beta-hairpin peptidomimetic of the invention and a suitable powder base such as lactose or starch.
The compounds may also be formulated in rectal or vaginal compositions such as suppositories with suitable suppository bases such as cocoa butter or other glycerides.
In addition to the formulations described above, the β -hairpin peptidomimetics of the invention can also be formulated as depot preparations. Such long acting formulations may be administered by implantation (e.g. subcutaneously or intramuscularly) or by intramuscular injection. To produce such depot formulations, the β -hairpin peptidomimetics of the invention can be formulated with suitable polymeric or hydrophobic materials (e.g., as an emulsion in an acceptable oil) or ion exchange resins, or as sparingly soluble salts.
In addition, other drug delivery systems may be used, such as liposomes and emulsions, as are well known in the art. Certain organic solvents, such as dimethyl sulfoxide, may also be used. In addition, sustained release systems, such as semipermeable matrices of solid polymers containing the therapeutic agent, can be used to deliver the β -hairpin peptidomimetics of the invention. A variety of sustained release materials have been identified and are well known to those skilled in the art. Sustained release capsules may release compounds for weeks up to over 100 days, depending on their chemical nature. Depending on the chemical nature and biological stability of the therapeutic agent, additional strategies may be used for protein stabilization.
Because the β -hairpin peptidomimetics of the invention may contain charged residues, they may be included as such in any of the above formulations, or as a pharmaceutically acceptable salt. The solubility of pharmaceutically acceptable salts in aqueous or other protic solvents tends to be greater than that of the corresponding free forms.
The β -hairpin peptidomimetics of the invention or compositions thereof will generally be used in an amount effective to achieve the intended purpose. It will be appreciated that the amount used will depend on the particular application.
For topical administration to treat or prevent a disease treatable with a β -hairpin mimetic, a therapeutically effective dose can be determined using, for example, the in vitro assay provided in the examples. Treatment may be applied when the disease is visible or even when it is not yet visible. One of ordinary skill in the art will be able to determine, without undue experimentation, a therapeutically effective amount for treating a localized disease.
For systemic administration, the therapeutically effective amount can be estimated initially from in vitro tests. For example, the dose can be formulated in animal models to achieve a circulating β -hairpin peptidomimetic concentration range that includes the IC determined in cell culture50. Such information can be used to more accurately determine effective dosages in humans.
Initial dosages can be determined from in vivo data, such as animal models, using techniques well known in the art. One of ordinary skill in the art can readily optimize administration to humans based on animal data.
The dosage for use as a serpin can be individually adjusted to provide plasma levels of the beta-hairpin peptidomimetics of the invention sufficient to maintain a therapeutic effect. Therapeutically effective serum levels can be obtained by administering multiple doses per day.
In the case of local administration or selective absorption, the effective local concentration of the β -hairpin peptidomimetics of the invention may be independent of the plasma concentration. One of ordinary skill in the art will be able to optimize therapeutically effective topical dosages without undue experimentation.
The amount of β -hairpin peptidomimetic administered will, of course, depend on the subject being treated, the weight of the subject, the severity of the patient, the mode of administration, and the judgment of the prescribing physician.
Generally, a therapeutically effective dose of the beta-hairpin peptidomimetics described herein will provide therapeutic benefit without causing substantial toxicity.
Toxicity of the β -hairpin peptidomimetics of the invention can be determined by standard pharmaceutical procedures in cell cultures or experimental animals, e.g., by determining LD50(dose leading to 50% death in the population) or LD100(dose that resulted in 100% mortality of the population). The dose ratio between toxic and therapeutic effects is the therapeutic index. Compounds that exhibit high therapeutic indices are preferred. The data obtained from these cell culture assays and animal studies can be used to formulate dosage ranges that are non-toxic for use in humans. The dosage of the β -hairpin peptidomimetics of the invention is preferably in the range of circulating concentrations that include an effective dose with little or no toxicity. The dosage may vary within this range depending upon the dosage form employed and the route of administration utilized. The exact formulation, route of administration, and dosage can be selected by the individual physician taking into account the patient's condition. (see, e.g., Fingl et al 1975 in: The Pharmacological Basis of Therapeutics, Ch.1, p.1).
Detailed Description
The following examples illustrate the invention in more detail, but are not intended to limit the scope of the invention in any way. The following abbreviations are used in these examples:
HBTU 1-benzotriazol-1-yl-tetramethyluronium hexafluorophosphate (Knorr et al tetrahedron Lett.1989,30, 1927-;
HOBt 1-hydroxybenzotriazole;
DIEA is diisopropylethylamine;
HOAT 7-aza-1-hydroxybenzotriazole;
HATU O- (7-aza-benzotriazol-1-yl) -N, N, N ', N' -tetramethyluronium hexafluorophosphate (Carpino et al Tetrahedron Lett.1994,35, 2279-.
Examples
1. Peptide synthesis
Coupling the first protected amino acid residue to a resin
0.5g of 2-chlorotrityl chloride resin (Barlos et al Tetrahedron Lett.1989,30, 3943-. Suspending the resin in CH2Cl2(2.5ml) and stirring was continued for 30 minutes and allowed to swell at room temperature. With 0.415mMol (1 equivalent) of the first appropriately protected amino acid residue (see below) and CH2Cl2The resin was treated with 284. mu.l (4 equivalents) of Diisopropylethylamine (DIEA) in (2.5ml) and the mixture was shaken for 4 h at 25 ℃. The resin color changed to purple and the solution remained pale yellow. Oscillatory resin (CH)2Cl217/2/1/MeOH/DIEA), 30ml for 30 minutes; then washed in the following order: by CH2Cl2(1x)、DMF(1x)、CH2Cl2(1x)、MeOH(1x)、CH2Cl2(1x)、MeOH(1x)、CH2Cl2(2x)、Et2Washed O (2 ×) and dried under vacuum for 6 hours.
Typically, the loading is 0.6-0.7 mMol/g.
The following pre-loaded resins were prepared: Fmoc-Pro-2-chlorotrityl resin, Fmoc-Asp (OtBu) -2-chlorotrityl resin, Fmoc-Pro (5RPhe) -2-chlorotrityl resin, Fmoc-Leu-2-chlorotrityl resin, Fmoc-Glu (OtBu) -2-chlorotrityl resin, Fmoc-Asp (OtBu) -2-chlorotrityl resin, Fmoc-Phe-2-chlorotrityl resin, Fmoc-Gln (Trt) -2-chlorotrityl resin, Fmoc-Ser (OtBu) -2-chlorotrityl resin, Fmoc-Val-2-chlorotrityl resin, Fmoc-Thr (OtBu) -2-chlorotrityl resin, and Fmoc-Ile-2-chlorotrityl resin.
Synthesis of fully protected peptide fragments
The synthesis was performed using a Syro-peptide synthesizer (Multisyntech) using 24 to 96 reaction vessels. In each container 60mg (weight of resin before loading) of the above resin was placed. The following reaction cycles were set and allowed to proceed:
Figure BDA0000736457340000941
steps 3 to 6 are repeated to add each amino acid.
After synthesis of the fully protected peptide fragment has been terminated, either program A or program B, described below, is then employed, depending on whether an interchain linkage (i.e., disulfide β -chain linkage) is to be formed.
Procedure A cyclization and work-up of backbone-cyclized peptide
Cleaving the fully protected peptide fragment
After completion of the synthesis, the resin was suspended in CH2Cl21ml (0.39mMol) of 1% TFA (v/v), suspended for 3 minutes, filtered, washed with CH2Cl21ml (1.17mMol,3 equivalents) of 20% DIEA (v/v) neutralised the filtrate. This procedure was repeated twice to ensure that lysis was complete. An aliquot (200 μ L) of the filtrate was completely deprotected with 0.5ml of cleavage mixture containing 95% trifluoroacetic acid (TFA), 2.5% water and 2.5% Triisopropylsilane (TIS), and the filtrate was analyzed by reverse phase-LC MS to monitor the efficiency of linear peptide synthesis.
Cyclization of linear peptides
The fully protected linear peptide was dissolved in DMF (8ml, 10 mg/ml). 2 equivalents of HATU (0.72mMol) in 1ml of DMF and 4 equivalents of DIEA (1.44mMol) in 1ml of DMF were added and the mixture was stirred at room temperature for 16 hours. The volatiles were evaporated to dryness. The crude cyclized peptide was dissolved in 7ml of CH2Cl2In (1), the mixture was extracted three times with 10% acetonitrile in water (4.5 ml). Will CH2Cl2The layer was evaporated to dryness.
Deprotection and purification of cyclic peptides
The cyclic peptide obtained was dissolved in 3ml of a cleavage mixture containing 95% trifluoroacetic acid (TFA), 2.5% water and 2.5% Triisopropylsilane (TIS). The mixture was left at 20 ℃ for 2.5 hours and then concentrated under vacuum. The crude peptide was dissolved in 20% AcOH in water (7ml) and extracted three times with diisopropyl ether (4 ml). The aqueous layer was collected, evaporated to dryness and the residue was purified by preparative reverse phase LC-MS.
After lyophilization, the product was obtained as a white powder and analyzed by LC-MS. Analytical data obtained after preparative HPLC and ESI-MS including purity are shown in table 1.
The analysis method comprises the following steps:
analytical HPLC retention times (RT in minutes) were determined using Jupiter protein 90A,150x 2.0mm, (code 00F4396-B0-Phenomenex) using the following solvents: solvent A (H)2O + 0.1% TFA) and solvent B (CH)3CN + 0.1% TFA), and the following gradient was used: and (3) 0 minute: 95% of A and 5% of B; 20 minutes: 40% A, 60% B; 21-23 minutes: 0% A, 100% B; 23.1-30 minutes: 95% of A and 5% of B.
Procedure B: cyclization and work-up of backbone cyclized peptides having disulfide beta-chain linkages
Formation of disulfide beta-chain bonds
After completion of the synthesis, the resin was expanded in 3ml of anhydrous DMF for 1 hour. Then a solution of 10 equivalents of iodine in DMF (6ml) was added to the reactor followed by stirring for 1.5 hours. The resin was filtered and a solution of fresh iodine (10 equivalents) in DMF (6ml) was added followed by stirring for a further 3 hours. The resin was filtered with DMF (3X) and CH2Cl2(3x) washing.
Backbone cyclization, cleavage and purification of peptides
After disulfide beta-chain bond formation, the resin was suspended in CH2Cl21ml (0.39mMol) of (1%) in 1% TFA (v/v), suspended for 3 minutes, filtered, washed with CH2Cl21ml (1.17mMol,3 equivalents) of 20% DIEA (v/v) neutralises the filtrate. This procedure was repeated twice to ensure complete lysis. With 2ml CH2Cl2The resin was washed. Evaporation of CH2Cl2And (4) layering to dryness.
The fully protected linear peptide was dissolved in 8ml of anhydrous DMF. To the peptide was then added 2 equivalents of HATU in anhydrous DMF (1ml) and 4 equivalents of DIPEA in anhydrous DMF (1ml), followed by stirring for 16 hours. The volatiles were evaporated to dryness. The crude cyclized peptide was dissolved in 7ml CH2Cl2In (1), the mixture was extracted three times with 10% acetonitrile in water (4.5 ml). Evaporation of CH2Cl2And (4) layering to dryness. To completely deprotect the peptide, 3ml of cleavage cocktail TFA TIS H was added2O (95:2.5:2.5), and the mixture was left for 2.5 hours. The volatiles were evaporated to dryness and the crude peptide was dissolved in 20% AcOH in water (7ml) and extracted three times with diisopropyl ether (4 ml). The water layer was collected, evaporated to dryness and the residue was purified by preparative reverse phase LC-MS.
After lyophilization, the product was obtained as a white powder and analyzed by ESI-MS analysis as described above. Analytical data obtained after preparative HPLC and ESI-MS including purity are shown in table 1.
Examples 1-45, 52-63, 65-67, 70-71, 75-114, 129, 131-. The peptide was synthesized starting from amino acid Pro grafted with resin. The starting resin was Fmoc-Pro-2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Pro-DPro-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Examples 1-6, 9-45, 52-63, 65-67, 70-71, 75-103, 112-114, 129, 131,133,136-138,140-141,143-146,148-153,155, 157-162 and 179-196 were cleaved from the resin as described in procedure B for disulfide bond formation, cyclization, deprotection and purification. Examples 82, 123, 149, 159, 161 and 178 were cleaved from the resin as described in procedure B. The following procedure was used to form disulfide bonds:
the crude product was dissolved in 0.1M ammonium acetate buffer (pH adjusted to 8) (concentration: 1mg of crude product per ml). The mixture was stirred at room temperature in the presence of air. The reaction was monitored by reverse phase LC-MS. After the reaction was complete, the solution was evaporated to dryness and the residue was purified by preparative reverse phase LC-MS.
Cyclization of the backbone was performed as described in procedure a. Deprotection was performed using the following procedure:
for complete deprotection of the peptide, 5ml of cleavage cocktail TFA: H was added2Phenol, thioanisole, ethanedithiol (82.5:5:5:5:2.5), and the mixture was left at room temperature for 5 hours. The peptide was precipitated by addition of cold diethyl ether (10 ml). After the centrifugation, the slurry is subjected to a centrifugal separation,the supernatant phase was removed. The precipitate was washed three times with 5ml diethyl ether and purified by preparative reverse phase LC-MS.
After lyophilization, the product was obtained as a white powder and analyzed by ESI-MS analysis as described above.
Examples 7, 8, 104, 111, 132, 134, 135, 139, 142, 147, 154 and 156 were cleaved from the resin as described in procedure A, and were cyclized, deprotected and purified.
HPLC-retention time (min) was determined using the analytical method described above:
example 1(15.37), example 2(11.54), example 3(7.82), example 4(8.62), example 5(16.51), example 6(13.67), example 7(3.61), example 8(4.11), example 9(5.82), example 10(7.98), example 11(8.38), example 12(6.80), example 13(7.41), example 14(6.20), example 15(8.68), example 16(9.82), example 17(5.59), example 20(7.32), example 21(8.66), example 22(8.68), example 23(12.66), example 24(8.67), example 25(7.53), example 26(9.02), example 27(8.06), example 28(9.62), example 29(8.78), example 30 (10.32), example 31 (7.35), example 31.34), example 31.31 (33.34), example 31.31.31), example 31.31.32, example 31.31, example 31.32, and example 9.32, Example 36(10.98), example 37(7.56), example 38(9.29), example 39(8.32), example 40(10.11), example 41(7.23), example 42(8.83), example 43(7.92), example 44(9.87), example 45(8.26), example 52(6.20), example 53(8.68), Ex 54(9.82), example 55(5.59), example 56(6.06), example 57(6.47), example 58(7.32), example 59(8.68), example 60(10.66), example 61(8.54), example 62(9.83), example 63(16.54), example 65(15.71), example 66(17.50), example 67(15.87), example 70(12.87), example 71(13.48), example 22.76), example 10 (14.75), example 10.79 (10.75), example 10.79 (5.75), example 44 (7.32), example 59 (7.68), example 60 (10.68), example 60(10.66), example 70.66), example 70 (8.54), example 71, example 80 (14.75), example 80), example 10.75 (10.75), example 10.75, example, Example 81(11.63), example 82(5.71), example 83(5.45), example 84(11.14), example 85(10.90), example 86(13.78), example 87(13.98), example 88(14.35), example 89(15.21), example 90(14.72), example 91(11.97), example 92(11.77), example 93(15.25), example 94(14.61), example 95(20.46), example 96(15.08), example 97(20.78), example 98(18.28), example 99(14.62), example 100(13.90), example 101(13.76), example 102(20.53), example 103(14.14), example 104(11.60), example 105(11.90), example 106(11.63), example 107(11.78), example 108 (108), example 102 (20.03), example 103 (15.67), example 110.112), example 5.67), example 11.11.11 (11.13), example 11.15.15.15.49), example 89 (15.7), example 90), example 97(20.78), example 98(18.28), example 99(14.62), example 100(13.90), example 107), example 110.78), example 110.15.15.15.15.7), example 15.15.15.15.15., Example 114(5.55), example 129(17.22), example 131(11.97), example 132(13.56), example 133(14.57), example 134(14.72), example 135(17.53), example 136(18.28), example 137(14.72), example 138(14.35), example 139(15.40), example 140(11.14), example 141(5.71), example 142(13.97), example 143(13.94), example 144(15.08), example 145(20.87), example 146(17.91), example 147(17.11), example 148(7.83), example 149(16.22), example 150(20.09), example 151(20.72), example 152(21.38), example 153(17.97), example 154(16.58), example 155(19.46), example 160.156 (15.156), example 160.65 (15.161), example 159 (15.65), example 137 (15.65), example 159 (15.65), example 137 (15.30.30.91), example 138, 17.91, 17.11), example 48(7.83), example 149.22), example 150 (16.22), example 150.65), example 151 (15.65), example 161, and example 161 Example 162(17.79), example 179(4.25), example 180(11.43), example 181(12.30), example 182(12.83), example 183(10.51), example 184(12.12), example 185(10.14), example 186(10.09), example 187(10.14), example 188(10.65), example 189(10.73), example 190(10.10), example 191(10.17), example 192(10.19), example 193(11.02), example 194(9.92), example 195(10.74), example 196 (9.94).
Example 46 is shown in table 1. The peptide was synthesized starting from amino acid Pro grafted with resin. The starting resin was Fmoc-Pro-2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Pro-DAsp (OtBu) -P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. In-line with the aboveThereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 46 (8.94).
Example 47 is shown in table 1. The synthesis of the peptide starts with the amino acid Asp grafted with the resin. The starting resin was Fmoc-Asp (OtBu) -2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Asp (OtBu) -DPro-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 47 (7.29).
Example 48 is shown in table 1. The synthesis of the peptide starts with the amino acid Pro (5RPhe) grafted with the resin. The starting resin was Fmoc-Pro (5RPhe) -2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Pro (5RPhe) -DPr o-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 48 (10.07).
Example 49 is shown in table 1. The synthesis of the peptide starts with the amino acid Pro grafted with the resin. The starting resin was Fmoc-Pro-2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Pro-DAla-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 49 (8.09);
example 50 is shown in table 1. The synthesis of the peptide starts with the amino acid Pro grafted with the resin. The starting resin was Fmoc-Pro-2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Pro-DIle-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 50 (9.78).
Example 51 is shown in table 1. The synthesis of the peptide starts with the amino acid Leu grafted with the resin. The starting resin was Fmoc-Leu-2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Leu-DPro-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 51 (8.94);
example 64 is shown in table 1. The synthesis of the peptide starts with the amino acid Glu grafted with the resin. The starting resin was Fmoc-Glu (OtButt) -2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Glu (OtBu) -DPro-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 64 (13.17).
Example 68 is shown in table 1. The synthesis of the peptide starts with the amino acid Asp grafted with the resin. The starting resin was Fmoc-Asp (OtBu) -2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Asp (OtBu) -DAla-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 68 (12.44).
Example 69 is shown in table 1. The synthesis of the peptide starts with the amino acid Pro grafted with the resin. The starting resin was Fmoc-Pro-2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Pro-DAsn (Trt) -P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 69 (12.97).
Example 72 is shown in table 1. The synthesis of the peptide starts with the amino acid Pro grafted with the resin. The starting resin was Fmoc-Pro-2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Pro-DThr (OtBu) -P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 72 (13.34).
Example 73 is shown in table 1. The synthesis of the peptide starts with the amino acid Pro grafted with the resin. Initiation ofThe resin was Fmoc-Pro-2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Pro-DIle-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 73 (9.78).
Example 74 is shown in table 1. The synthesis of the peptide starts with the amino acid Leu grafted with the resin. The starting resin was Fmoc-Leu-2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Leu-DPro-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 74 (8.94).
Example 115 is shown in table 1. The synthesis of the peptide starts with the amino acid Pro grafted with the resin. The starting resin was Fmoc-Pro-2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Pro-DAsp (OtBu) -P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 115 (4.82).
Example 116 is shown in table 1. The synthesis of the peptide starts with the amino acid Pro grafted with the resin. The starting resin was Fmoc-Pro-2-chlorotrityl resin, which was prepared as described above. Synthesis of Linear on solid support according to the procedure described previouslyA peptide having the sequence: resin-Pro-DPhe-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 116 (5.98).
Example 117 is shown in table 1. The synthesis of the peptide starts with the amino acid Pro grafted with the resin. The starting resin was Fmoc-Pro-2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Pro-DArg (Trt) -P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 117 (4.48).
Example 118 is shown in table 1. The synthesis of the peptide starts with the amino acid Pro grafted with the resin. The starting resin was Fmoc-Pro-2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Pro-DSer (OtBu) -P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 118 (4.73).
Example 119 is shown in table 1. The synthesis of the peptide starts with the amino acid Pro grafted with the resin. The starting resin was Fmoc-Pro-2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Pro-DVal-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, disulfide bond formation as described in procedure B,the peptide is cleaved from the resin, cyclized, deprotected and purified.
HPLC-retention time (min) was determined using the analytical method described above:
example 119 (5.47).
Example 120 is shown in table 1. The synthesis of the peptide starts with the amino acid Pro grafted with the resin. The starting resin was Fmoc-Pro-2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Pro-DPic-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 120 (5.48).
Example 121 is shown in table 1. The synthesis of the peptide starts with the amino acid Asp grafted with the resin. The starting resin was Fmoc-Asp (OtBu) -2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Asp (OtBu) -DPro-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 121 (4.56).
Examples 122 and 167 are shown in table 1. The synthesis of the peptide starts with the amino acid Phe grafted with a resin. The starting resin was Fmoc-Phe-2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Phe-DPro-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 122 (5.75); 167(5.75).
Examples 123, 164, 169, 170, 172, 173, 175, 177 and 178 are shown in Table 1. The synthesis of the peptide starts with the amino acid Gln grafted with the resin. The starting resin was Fmoc-Gln (Trt) -2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Gln (Trt) -DPro-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
examples 123(4.35), 164(13.20), 169(16.81), 170(14.57), 172(16.78), 173(13.57), 175(15.94), 177(16.78), 178 (17.45).
Example 124 is shown in table 1. The synthesis of the peptide starts with the amino acid Ser grafted with the resin. The starting resin was Fmoc-Ser (OtBu) -2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Ser (OtBu) -DPro-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 124 (4.46).
Example 125 is shown in table 1. The synthesis of the peptide starts with the amino acid Val grafted with the resin. The starting resin was Fmoc-Val-2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Val-DPro-P11-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 125 (18.42).
Example 126 is shown in table 1. The synthesis of the peptide starts with the amino acid Thr grafted with the resin. The starting resin was Fmoc-Thr (OtBu) -2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Thr (OtBu) -DThr (OtBu) -P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 126 (4.35).
Examples 127, 163, 165 and 174 are shown in Table 1. The synthesis of the peptide starts with the amino acid Glu grafted with the resin. The starting resin was Fmoc-Glu (OtBu) -2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Glu (OtBu) -DLys (Boc) -P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
examples 127(4.11), 163(14.93), 165(14.40) and 174 (12.73).
Example 128 is shown in table 1. The synthesis of the peptide starts with the amino acid Thr grafted with the resin. The starting resin was Fmoc-Thr (OtBu) -2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Thr (OtBu) -DPhe-P11-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 128 (5.26).
Example 130 is shown in table 1. The synthesis of the peptide starts with the amino acid Pro grafted with the resin. The starting resin was Fmoc-Pro-2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Pro-DAla-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 130 (14.79).
Example 166 is shown in table 1. The synthesis of the peptide starts with the amino acid Ile grafted with the resin. The starting resin was Fmoc-Ile-2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Ile-DPhe-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 166 (16.80).
Example 168 is shown in table 1. The synthesis of the peptide starts with the amino acid Asp grafted with the resin. The starting resin was Fmoc-Asp (OtBu) -2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Asp (OtBu) -DPro-P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
example 168 (4.56).
Examples171, and 176 are shown in table 1. The synthesis of the peptide starts with the amino acid Gln grafted with the resin. The starting resin was Fmoc-Gln (Trt) -2-chlorotrityl resin, which was prepared as described above. The linear peptide was synthesized on a solid support according to the procedure described previously, which was the following sequence: resin-Gln (Trt) -DGln (Trt) -P11-P10-P9-P8-P7-P6-P5-P4-P3-P2-P1. Thereafter, the peptide was cleaved from the resin following disulfide bond formation as described in procedure B, and cyclization, deprotection and purification were performed.
HPLC-retention time (min) was determined using the analytical method described above:
examples 171(15.40) and 176 (13.67).
Figure BDA0000736457340001091
Figure BDA0000736457340001101
Figure BDA0000736457340001111
Figure BDA0000736457340001121
Figure BDA0000736457340001131
Figure BDA0000736457340001141
Figure BDA0000736457340001151
The Cys at the 2-and 10-positions in examples 1-6,9-103,112-131,133,136-138,140-141,143-146,148-153,155,157-196 formed disulfide bonds.
2. Biological method
2.1. Preparation of peptide samples
The lyophilized peptide was weighed on a microbalance (Mettler MT5) and dissolved in sterile water to a final concentration of 1mM, unless otherwise stated. The stock solution was stored at +4 ℃ in the dark.
2.2. Enzyme assay
Enzyme and substrate conditions are shown in table 2.
Kinetic measurements were performed in 96-well flat-bottom plates (Greiner) on a Genios plate reader (Tecan) in a total reaction volume of 100 μ Ι. The enzyme was combined with the peptide (inhibitor) in a buffer containing 100mM HEPES (pH7.5), 50mM CaCl20.025% tween-20, 5% DMSO and 1mM substrate. The rate of substrate hydrolysis was measured by monitoring the change in absorbance at 405nm for 30 minutes to verify the linearity of the reaction curve. The average rate from minute 1 to minute 10 was used for all calculations. Initial calculations of background subtraction, average rate, mean in duplicate, and% inhibition were performed using Magellan software from Tecan (5 th edition). IC 50% was calculated using Grafit (version 5.0.10) from Erithacus Software by fitting inhibition data from 6 different inhibitor concentrations to the following 4-parameter equation:
Figure BDA0000736457340001161
in this equation, s is the slope coefficient, x is the inhibitor concentration, and y is the% inhibition of the given concentration of inhibitor.
Km/KiMeasurement of
K determination of serine protease substrates from the Lineweaver-Burke plot (Grafit v5)m. Using formula KiIC 50%/(1 + ([ substrate ]]/Km) To calculate the Ki value of the inhibitor.
Increasing concentrations of substrate were reacted with the enzyme and the rate per reaction (ABS/mSec) was plotted against substrate concentration. A reciprocal plot (Lineweaver-Burke) is also plotted to provide KmAnd Vmax(see reference 1 described below).
TABLE 2
Figure BDA0000736457340001171
Figure BDA0000736457340001181
2.3. Cytotoxicity assays
Reduction Using MTT [ see references 2 and 3, infra]The cytotoxicity of the peptides on HELA cells (Acc57) and COS-7 cells (CRL-1651) was determined. The method is briefly described as follows: respectively at 7.0x10 per hole3Individual cell and 4.5x103The individual cells were seeded with HELA and COS-7 cells and allowed to grow in 96 well microtiter plates at 37 ℃ in 5% CO2And growing for 24 hours. At this point, time zero (Tz) was determined by MTT reduction (see below). The supernatant in the remaining wells was discarded and fresh medium and 12.5, 25 and 50 μ M serial dilutions of peptides were pipetted into the wells. Each peptide concentration was determined in triplicate. At 37 deg.C, 5% CO2Incubation of the cells was continued for 48 hours. The wells were then washed with Phosphate Buffered Saline (PBS), and then 100. mu.l of MTT reagent (0.5mg/ml in RPMI1640 and DMEM medium, respectively) was added to the wells. This was incubated at 37 ℃ for 2 hours, followed by aspiration of the medium and addition of 100. mu.l of isopropanol to each well. The absorbance (OD) at 595nm of the dissolved product was measured595Peptides). For each concentration, the average was calculated from triplicate measurements. The percentage of growth was calculated as follows: (OD)595peptide-OD595Tz-OD595void)/(OD595Tz-OD595Empty wells) x 100%, which is plotted against each peptide concentration.
LC50 values (lethal concentration, defined as the concentration at which 50% of cells are killed) were determined for each peptide using the TREND line function of EXCEL (microsoft office 2000) (═ TREND (C50: C0,% 50:% 0, -50)), at (50, 25, 12.5 and 0 μm), the corresponding percentage of growth and the value-50.
By using concentration (50, 25, 12.5 and 0. mu.g/ml), corresponding percentage and value 50 TREND line function, (TREND (C)50:C0,％50Thousandths, 50)), the GI50 (growth inhibition) concentration was calculated for each peptide.
2.4. Hemolysis of blood
Peptides were tested for their hemolytic activity against human red blood cells (hRBC). Fresh hRBC were washed three times with Phosphate Buffered Saline (PBS) by centrifugation at 2000x g for 10 minutes. Peptides were incubated with 20% v/v hRBC at 37 ℃ for 1 hour at a concentration of 100. mu.M. The final red blood cell concentration was approximately 0.9x10 per ml9And (4) cells. By respectively in the presence of H in only PBS and in the presence of H2hRBC were incubated at 0.1% Triton X-100 in O and values for 0% and 100% cell lysis were determined, respectively. The samples were centrifuged, the supernatants were diluted 20-fold in PBS buffer, and the Optical Density (OD) of the samples at 540nM was measured. 100% lysis value (OD)540H2O) gives an OD of about 1.3 to 1.8540. The percent hemolysis was calculated as: (OD)540peptide/OD540H2O)x100％.
2.5 plasma stability
Mu.l of plasma/albumin solution was placed in a polypropylene (PP) tube and blended with 45. mu.l of compound from 100mM solution B (obtained from 135. mu.l PBS and 15. mu.l peptide in 1mM PBS pH 7.4). A150. mu.l aliquot was transferred to each well of a10 kDa filter plate (Millipore MAPPB 1010Biomax membrane). For the "0 min control", 270. mu.l PBS was placed in a PP tube, 30. mu.l stock B was added, and mixed with shaking. Mu.l of the control solution was placed in one well of the filter plate and used as "filtered control".
Another 150. mu.l of control solution was taken and placed directly into a receiving well (reserved for filtrate) for use as "unfiltered control". the entire plate including the evaporation lid was incubated at 37 ℃ for 60 minutes. centrifugation at 4300rpm (3500g) and 15 ℃ for at least 2 hours on a plasma sample (rat plasma: Harlan Sera laboratory, UK; human plasma: Blutspendeszentrum Z ü rich) to produce 100. mu.l of filtrate approximately 1 hour of centrifugation was sufficient for a "plasmaphanin" sample (freshly prepared human albumin: Sigma A-4327; rat albumin: Sigma A-6272, both at a concentration of 40mg/ml in PBS.) the filtrate in the PP plate received was analyzed by LC/MS in the column: Jupiter C18 (Phennex), mobile phase (A) 0.1% formic acid in water and (B) acetonitrile,gradient: 5% -100% (B) in 2 minutes, electrospray ionization, MRM detection (triple quadrupole). The area of the peak was determined and the values determined in triplicate were averaged. Control 1 and control 2:100- (100x T) in combination (filtered and unfiltered time points 0 min)60/T0) Is expressed in percentage. The average of these values is then calculated.
2.6. Pharmacokinetic studies (PK)
Pharmacokinetic studies in rats after a single oral (gavage) and intravenous administration
A pharmacokinetic study was performed on the compound of example 75("Ex.75") after a single intravenous (i.v.) and oral (p.o., gavage) administration using 332g (+ -10 g) male Wistar mice obtained from RCC Ltd, laboratory animals, CH-4414F ü llisdorf, Switzerland, plus vehicle saline to give a final concentration of 2.5mg/ml for intravenous administration and a volume of 2ml/kg for oral administration and a volume of 10ml/kg for oral administration, injecting the peptide of example 75 to provide a final intravenous dose of 5mg/kg and an oral dose of 50mg/kg according to the protocol described below at different time points, using DiLab Saumpler by automated blood sample collection, a blood sample (approximately 0.24ml) was taken into heparinized tubes at time points of 0,5 minutes (intravenous only), 15, 30 minutes and 1,2, 4, 24 and 36 hours and 16-8 hours (frozen plasma was analyzed by HPLC at 80 ℃ before addition to heparinized tubes.
Preparation of plasma calibration samples
"blank" rat plasma from untreated animals was used. 0.1ml aliquots of plasma were mixed with 50ng propranolol (internal standard, IS) (by mixing in
Figure BDA0000736457340001201
Sample preparation by solid phase extraction on HLB cartridges (Waters) and a known amount of the peptide of example 75 were mixed to obtain 9 plasma calibration samples ranging between 5-2000 ng/ml. With 1ml of methanol and then with 1ml of 1% NH in water3Regulating
Figure BDA0000736457340001202
Upper HLB cartridge. Then 400. mu.l of 1% NH in water3Samples were diluted and loaded. With 1ml of methanol/1% NH in water3(5/95) washing the board. Elution was performed using 1ml of 0.1% TFA in methanol.
The plate containing the eluate was introduced into a concentrator system and allowed to dry. The residue was dissolved in 100. mu.l formic acid 0.1%/acetonitrile 95/5(v/v) and analyzed in HPLC/MS on a reverse phase analytical column (Jupiter C18,50X2.0mm,5 μm, Phenomenex) using gradient elution (mobile phase A: 0.1% formic acid in water; B: acetonitrile, from 5% B to 100% B over 2 minutes).
Preparation of plasma samples
From each sample 100 μ l of plasma was taken for extraction. If the volume is less than 100. mu.l, then an appropriate amount of "blank" mouse plasma is added to keep the matrix the same as the calibration curve. The samples were then blended with IS and processed as described for the calibration curve.
Pharmacokinetic evaluation
Using software PK solution 2.0TM(Summit Research Service, Montrose, CO 81401USA), PK analysis was performed on the collected data (typically n ═ 2 or 3). The area under the curve AUC was calculated by linear trapezoidal law. The AUC(t-∞)Estimated as Ct/b (b: elimination rate constant). AUC(t-∞)Is AUC(0-t)And AUC(t-∞)The sum of (1). The elimination half-life is calculated by linear regression over at least three data points of the elimination period. By a correlation coefficient (r)2) To evaluate the time interval chosen for half-life determination, the correlation coefficient should be at least greater than 0.85 and most preferably greater than 0.96. In the case of intravenous administration, t is determined by extrapolating the curve from the first two time pointsZeroThe initial concentration of (c). Finally, by AUC after intraperitoneal administration normalized to intravenous administration(0-∞)In contrast, bioavailability after intraperitoneal administration was calculated.
3.0 results
The results of the experiments described above in 2.2-2.5 are shown in table 3 below.
Figure BDA0000736457340001221
Figure BDA0000736457340001241
Figure BDA0000736457340001251
Figure BDA0000736457340001271
Figure BDA0000736457340001291
Figure BDA0000736457340001301
Figure BDA0000736457340001311
The results of the experiment described in 2.5 above are shown in table 4 below.
TABLE 4
Figure BDA0000736457340001321
The results of the experiment described in 2.6(PK) above are shown in table 5 below.
TABLE 5
The large inter-individual differences in plasma concentrations of the peptides of example 75 were most significant after a single oral administration (intravenous% C.V-6-68%, except for one value 173% at the lowest measured concentration; oral% c.v.: 113-.
Intravenous administration
After intravenous administration of the peptide of example 75 at a dose level of 5mg/kg body weight, the peptide of example 75 follows intravenous kinetics. The peptide of example 75 showed, after PK analysis, extrapolated CInitiation of14069ng/ml, C observed at 5 minutes (0.083 hours)max10762 ng/ml. Plasma levels dropped rapidly to 5774 and 3455ng/ml at 15 and 30 minutes, respectively. From 1 to 2 hours, plasma levels at 0.46 hour end t1/2Reduced to 18ng/ml at 4 hours. AUC0.tAnd AUC0-∞6044 and 6047ng x h/ml, respectively, with an initial distribution volume of 355 ml/kg. The apparent volume of distribution was 547 ml/kg.
Oral administration
After oral administration of the peptide of example 75 at a dosage level of 50mg/kg body weight, the plasma levels of the peptide of example 75 follow the kinetics of oral administration. After PK analysis, the peptide of example 75 showed C observed at 0.25 hr (15 min)maxIs 464 ng/ml. From 0.25 hours, plasma levels at 0.87 hours end t1/2Reduced to 24ng/ml at 4 hours. AUC0-tAnd AUC0-∞782 and 813ng x h/ml, respectively. The apparent distribution volume was 1008ml/kg, taking into account an absorption of 1.3%.
Comparison of oral administration and intravenous administration
Since the dose levels were different in the orally administered group and the intravenously administered group, the values were compared after dose normalization.
Compared to the normalized AUC after intravenous administration of the peptide of example 75 (100%: 6047ng-h/ml)0-∞Values, percentage of absorbed example 75 peptide after oral administration (F) 1.3% (81ng x h/ml), normalized C after oral administrationmaxThe value was approximately 234 times lower (46 vs 10762 ng/ml; Table 3). The apparent volume of distribution after oral administration was approximately 1.8 times higher than after intravenous administration (1008 vs 547 ml/kg).
Reference to the literature
1.Barrtt,A.J.Methods in Enzymology 1981,80,561-565；Leatherbarrow,R.J.1992,GraFit,Erithacus Software Ltd.,Staines,U.K.
2.Mossman T.J.Immunol.Meth.1983,65:55-63
3.Berridge MV,Tan AS.Arch.Biochem.Biophys.1993,303:474-482

Claims (66)

1. A compound of the general formula,
Figure FDA0002210548630000011
wherein
Figure FDA0002210548630000012
Is a dipeptide consisting of two different amino acid building blocksDPro-LPro, and Z is a chain of 11 amino acid residues, the positions of which in the chain are counted starting from the N-terminal amino acid, wherein the amino acid residues in positions 1 to 11 of the Z chain are:
-P1: nle, Ile, Chg, OctG, hPhe, Cha, Phe, Tyr, Trp, or Arg;
-P2: cys, Glu or Gln;
-P3：Thr；
-P4: lys, Nle, or Ala;
-P5：Ser；
-P6: ile, Leu, OctG, Cha, or Asp;
-P7：Pro；
-P8: pro or Pro (4 NHCOPhe);
-P9: ile, Nle, Cha, Gln or Lys;
-P10: cys, Lys or Thr;
-P11: tyr, Gln, Arg, Phe, Asp, or Thr; and
when Cys is present in P2 and P10, it forms a disulfide bond,
with the proviso that the amino acid chain Z does not have the amino acid sequence of the following amino acid residues:
-P1 is Arg,
-P2 is Cys, which is linked by a disulfide bond to the Cys at position P10,
-P3 is Thr,
-P4 is Lys and is a lysine,
-P5 is Ser,
-P6 is the number Ile,
-P7 is Pro,
-P8 is Pro,
-P9 is the number Ile,
-P10 is Cys, which is linked to Cys in position P2 by a disulfide bond, and
-P11 is Phe;
the compounds are in free form or in pharmaceutically acceptable salt form.
2. A compound according to claim 1, wherein the amino acid residues in positions 1 to 11 of the Z chain are:
-P1: phe, hPhe, Nle, Chg, Ile, Tyr, Arg, Trp, or Cha;
-P2: cys or Glu;
-P3：Thr；
-P4: lys or Nle;
-P5：Ser；
-P6：Asp；
-P7：Pro；
-P8：Pro；
-P9: ile, Nle, Cha, or Gln;
-P10：Cys；
-P11: thr, Asp, Tyr, Phe or Arg; and
-when Cys is present at P2 and P10, it forms a disulfide bond.
3. A compound according to claim 1, wherein the amino acid residues in positions 1 to 11 of the Z chain are:
-P1: ile, Nle, Chg, OctG or hPhe;
-P2: cys, Glu or Gln;
-P3：Thr；
-P4：Ala；
-P5：Ser；
-P6: OctG, Ile, Cha, or Leu;
-P7：Pro；
-P8: pro or Pro (4 NHCOPhe);
-P9: gln or Ile;
-P10: cys, Lys or Thr;
-P11: tyr, Arg, Gln, or Phe; and
-when Cys is present at P2 and P10, it forms a disulfide bond.
4. A compound according to claim 1, wherein the amino acid residues in positions 1 to 11 of the Z chain are:
-P1: cha, Tyr or Trp
-P2：Cys
-P3：Thr
-P4：Lys
-P5：Ser
-P6：Leu
-P7：Pro
-P8：Pro
-P9：Lys
-P10：Cys
-P11: arg; and
disulfide bonds are formed between P2 and the Cys residues present at position P10.
5. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：Phe；
-P2：Cys；
-P3：Thr；
-P4：Lys；
-P5：Ser；
-P6：Asp；
-P7：Pro；
-P8：Pro；
-P9：Ile；
-P10：Cys；
-P11：Tyr；
disulfide bonds are formed between P2 and Cys at position P10.
6. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：Ile；
-P2：Cys；
-P3：Thr；
-P4：Lys；
-P5：Ser；
-P6：Asp；
-P7：Pro；
-P8：Pro；
-P9：Nle；
-P10：Cys；
-P11：Arg；
disulfide bonds are formed between P2 and Cys at position P10.
7. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：Ile；
-P2：Cys；
-P3：Thr；
-P4：Lys；
-P5：Ser；
-P6：Asp；
-P7：Pro；
-P8：Pro；
-P9：Cha；
-P10：Cys；
-P11：Arg；
disulfide bonds are formed between P2 and Cys at position P10.
8. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：Ile；
-P2：Cys；
-P3：Thr；
-P4：Nle；
-P5：Ser；
-P6：Asp；
-P7：Pro；
-P8：Pro；
-P9：Ile；
-P10：Cys；
-P11：Arg
disulfide bonds are formed between P2 and Cys at position P10.
9. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：Phe；
-P2：Cys；
-P3：Thr；
-P4：Nle；
-P5：Ser；
-P6：Asp；
-P7：Pro；
-P8：Pro；
-P9：Ile；
-P10：Cys；
-P11：Tyr；
disulfide bonds are formed between P2 and Cys at position P10.
10. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：Chg；
-P2：Cys；
-P3：Thr；
-P4：Lys；
-P5：Ser；
-P6：Asp；
-P7：Pro；
-P8：Pro；
-P9：Ile；
-P10：Cys；
-P11：Tyr；
disulfide bonds are formed between P2 and Cys at position P10.
11. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：Arg；
-P2：Cys；
-P3：Thr；
-P4：Lys；
-P5：Ser；
-P6：Asp；
-P7：Pro；
-P8：Pro；
-P9：Ile；
-P10：Cys；
-P11：Phe；
disulfide bonds are formed between P2 and Cys at position P10.
12. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：Nle；
-P2：Cys；
-P3：Thr；
-P4：Ala；
-P5：Ser；
-P6：Ile；
-P7：Pro；
-P8：Pro；
-P9：Gln；
-P10：Cys；
-P11：Gln；
disulfide bonds are formed between P2 and Cys at position P10.
13. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：Nle；
-P2：Cys；
-P3：Thr；
-P4：Ala；
-P5：Ser；
-P6：Ile；
-P7：Pro；
-P8：Pro；
-P9：Gln；
-P10：Cys；
-P11：Tyr；
disulfide bonds are formed between P2 and Cys at position P10.
14. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：hPhe；
-P2：Cys；
-P3：Thr；
-P4：Ala；
-P5：Ser；
-P6：OctG；
-P7：Pro；
-P8：Pro；
-P9：Gln；
-P10：Cys；
-P11：Gln
disulfide bonds are formed between P2 and Cys at position P10.
15. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：OctG；
-P2：Gln；
-P3：Thr；
-P4：Ala；
-P5：Ser；
-P6：Ile；
-P7：Pro；
-P8：Pro；
-P9：Gln；
-P10：Thr；
-P11：Tyr。
16. a compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：hPhe；
-P2：Cys；
-P3：Thr；
-P4：Ala；
-P5：Ser；
-P6：Cha；
-P7：Pro；
-P8：Pro；
-P9：Gln；
-P10：Cys；
-P11：Phe；
disulfide bonds are formed between P2 and Cys at position P10.
17. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：OctG；
-P2：Glu；
-P3：Thr；
-P4：Ala；
-P5：Ser；
-P6：Ile；
-P7：Pro；
-P8：Pro；
-P9：Gln；
-P10：Lys；
-P11：Tyr。
18. a compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：OctG；
-P2：Cys；
-P3：Thr；
-P4：Ala；
-P5：Ser；
-P6：Cha；
-P7：Pro；
-P8：Pro；
-P9：Gln；
-P10：Cys；
-P11：Phe；
disulfide bonds are formed between P2 and Cys at position P10.
19. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：hPhe；
-P2：Cys；
-P3：Thr；
-P4：Ala；
-P5：Ser；
-P6：Cha；
-P7：Pro；
-P8：Pro；
-P9：Gln；
-P10：Cys；
-P11：Gln；
disulfide bonds are formed between P2 and Cys at position P10.
20. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：OctG；
-P2：Cys；
-P3：Thr；
-P4：Ala；
-P5：Ser；
-P6：Cha；
-P7：Pro；
-P8：Pro(4NHCOPhe)；
-P9：Gln；
-P10：Cys；
-P11：Gln；
disulfide bonds are formed between P2 and Cys at position P10.
21. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：hPhe；
-P2：Cys；
-P3：Thr；
-P4：Ala；
-P5：Ser；
-P6：OctG；
-P7：Pro；
-P8：Pro；
-P9：Gln；
-P10：Cys；
-P11：Tyr；
disulfide bonds are formed between P2 and Cys at position P10.
22. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：Chg；
-P2：Cys；
-P3：Thr；
-P4：Lys；
-P5：Ser；
-P6：Asp；
-P7：Pro；
-P8：Pro；
-P9：Ile；
-P10：Cys；
-P11：Thr；
disulfide bonds are formed between P2 and Cys at position P10.
23. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：Chg；
-P2：Cys；
-P3：Thr；
-P4：Lys；
-P5：Ser；
-P6：Asp；
-P7：Pro；
-P8：Pro；
-P9：Ile；
-P10：Cys；
-P11：Asp；
disulfide bonds are formed between P2 and Cys at position P10.
24. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：OctG；
-P2：Cys；
-P3：Thr；
-P4：Ala；
-P5：Ser；
-P6：Cha；
-P7：Pro；
-P8：Pro；
-P9：Gln；
-P10：Cys；
-P11：Gln；
disulfide bonds are formed between P2 and Cys at position P10.
25. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：OctG；
-P2：Cys；
-P3：Thr；
-P4：Ala；
-P5：Ser；
-P6：Ile；
-P7：Pro；
-P8：Pro；
-P9：Gln；
-P10：Cys；
-P11：Tyr；
disulfide bonds are formed between P2 and Cys at position P10.
26. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：OctG；
-P2：Cys；
-P3：Thr；
-P4：Ala；
-P5：Ser；
-P6：OctG；
-P7：Pro；
-P8：Pro；
-P9：Gln；
-P10：Cys；
-P11：Gln；
disulfide bonds are formed between P2 and Cys at position P10.
27. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：Cha；
-P2：Cys；
-P3：Thr；
-P4：Lys；
-P5：Ser；
-P6：Leu；
-P7：Pro；
-P8：Pro；
-P9：Lys；
-P10：Cys；
-P11：Arg；
disulfide bonds are formed between P2 and Cys at position P10.
28. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：Tyr；
-P2：Cys；
-P3：Thr；
-P4：Lys；
-P5：Ser；
-P6：Leu；
-P7：Pro；
-P8：Pro；
-P9：Lys；
-P10：Cys；
-P11：Arg；
disulfide bonds are formed between P2 and Cys at position P10.
29. A compound of formula I according to claim 1, wherein the amino acid residues in positions 1 to 11 of the chain Z are:
-P1：Trp；
-P2：Cys；
-P3：Thr；
-P4：Lys；
-P5：Ser；
-P6：Leu；
-P7：Pro；
-P8：Pro；
-P9：Lys；
-P10：Cys；
-P11：Arg；
disulfide bonds are formed between P2 and Cys at position P10.
30. Use of a compound according to any one of claims 1 to 29 in the manufacture of a medicament for the treatment of cardiovascular diseases and/or immunological diseases and/or neurodegenerative diseases.
31. A pharmaceutical composition comprising a compound according to any one of claims 1 to 29 and a pharmaceutically inert carrier.
32. A pharmaceutical composition according to claim 31 in a form suitable for topical, transmucosal or pulmonary administration, or for administration by injection or inhalation.
33. A pharmaceutical composition according to claim 31, in a form suitable for oral, transdermal or buccal administration.
34. The pharmaceutical composition according to any one of claims 31 to 33, which is in the form of a tablet, dragee, capsule, solution, gel, plaster, cream, ointment, syrup, slurry, suspension, nebulizer or suppository.
35. A pharmaceutical composition according to any one of claims 31 to 33, which is in the form of a liquid or a spray.
36. Use of a compound according to any one of claims 1 to 29 for the manufacture of a medicament for use as a protease inhibitor.
37. The use according to claim 36, wherein the protease inhibitor medicament is used to prevent infection in a healthy individual, or to slow infection in an infected patient, or for protease activity to mediate or cause cancer, or for protease activity to mediate or cause an immune disease, or for protease activity to mediate or cause inflammation, or for a situation in which protease activity mediates or causes an immune response.
38. Use of a compound according to any one of claims 2, 5-11 and 22-23 for the manufacture of a medicament for use as an inhibitor of cathepsin G.
39. Use of a compound according to any one of claims 3, 12-21 and 24-26 for the manufacture of a medicament for use as an inhibitor of elastase.
40. Use of a compound according to any one of claims 4 and 27-29 for the manufacture of a medicament for use as an inhibitor of tryptase.
41. A process for the production of a compound according to any one of claims 1 to 29, which process comprises:
(a) coupling the appropriately functionalized solid support with an appropriately N-protected derivative of the amino acid at position 5,6 or 7 in the desired end product, any functional groups that may be present in said N-protected amino acid derivative being likewise appropriately protected;
(b) removing the N-protecting group from the product thus obtained;
(c) coupling the product thus obtained with a suitably N-protected derivative of that amino acid which is in the desired end product further to the N-terminal amino acid residue position, any functional groups which may be present in said N-protected amino acid derivative being likewise suitably protected;
(d) removing the N-protecting group from the product thus obtained;
(e) repeating steps (c) and (d) until the N-terminal amino acid residue has been introduced;
(f) coupling the product thus obtained with a compound of the general formula
Figure FDA0002210548630000191
Wherein
Has the meaning as defined in claim 1 and X is an N-protecting group, or,
(fa) coupling the product obtained in step (e) with a suitably N-protected derivative of the amino acid L-proline;
(fb) removing the N-protecting group from the product thus obtained; and
(fc) coupling the product thus obtained with a suitably N-protected derivative of the amino acid D-proline;
(g) removing the N-protecting group from the product obtained in step (f) or (fc);
(h) coupling the product thus obtained with a suitably N-protected derivative of the amino acid at position 11 in the desired end product, any functional groups that may be present in said N-protected amino acid derivative likewise being suitably protected;
(i) removing the N-protecting group from the product thus obtained;
(j) coupling the product thus obtained with a suitably N-protected derivative of that amino acid which is further from position 11 in the desired end product, any functional groups which may be present in said N-protected amino acid derivative likewise being suitably protected;
(k) removing the N-protecting group from the product thus obtained;
(l) Repeating steps (j) and (k) until all amino acid residues have been introduced;
(m) selectively deprotecting one or more protected functional groups present in the molecule and appropriately substituting the reactive groups thus released;
(n) forming an interchain bond between the side chains of the appropriate amino acid residues at positions 2 and 10;
(o) detaching the product thus obtained from the solid support;
(p) cyclizing the cleaved product from the solid support; and
(q) removing any protecting groups present on the functional groups of any members of the chain of amino acid residues and any other protecting groups that may be present in the molecule.
42. A process for the production of a compound according to any one of claims 1 to 29, which process comprises:
(a) coupling the appropriately functionalized solid support with an appropriately N-protected derivative of the amino acid at position 5,6 or 7 in the desired end product, any functional groups that may be present in said N-protected amino acid derivative being likewise appropriately protected;
(b) removing the N-protecting group from the product thus obtained;
(c) coupling the product thus obtained with a suitably N-protected derivative of that amino acid which is in the desired end product further to the N-terminal amino acid residue position, any functional groups which may be present in said N-protected amino acid derivative being likewise suitably protected;
(d) removing the N-protecting group from the product thus obtained;
(e) repeating steps (c) and (d) until the N-terminal amino acid residue has been introduced;
(f) coupling the product thus obtained with a compound of the general formula
Wherein
Has the meaning as defined in claim 1 and X is an N-protecting group, or,
(fa) coupling the product obtained in step (e) with a suitably N-protected derivative of the amino acid L-proline;
(fb) removing the N-protecting group from the product thus obtained; and
(fc) coupling the product thus obtained with a suitably N-protected derivative of the amino acid D-proline;
(g) removing the N-protecting group from the product obtained in step (f) or (fc);
(h) coupling the product thus obtained with a suitably N-protected derivative of the amino acid at position 11 in the desired end product, any functional groups that may be present in said N-protected amino acid derivative likewise being suitably protected;
(i) removing the N-protecting group from the product thus obtained;
(j) coupling the product thus obtained with a suitably N-protected derivative of that amino acid which is further from position 11 in the desired end product, any functional groups which may be present in said N-protected amino acid derivative likewise being suitably protected;
(k) removing the N-protecting group from the product thus obtained;
(l) Repeating steps (j) and (k) until all amino acid residues have been introduced;
(n) forming an interchain bond between the side chains of the appropriate amino acid residues at positions 2 and 10;
(o) detaching the product thus obtained from the solid support;
(p) cyclizing the cleaved product from the solid support; and
(q) removing any protecting groups present on the functional groups of any members of the chain of amino acid residues and any other protecting groups that may be present in the molecule.
43. A method according to claim 41 or claim 42, further comprising the steps of:
(r) converting the obtained product into a pharmaceutically acceptable salt, or converting the obtained pharmaceutically acceptable or unacceptable salt into the corresponding free compound of formula I or into a different pharmaceutically acceptable salt.
44. A process for the production of a compound according to any one of claims 1 to 29, which process comprises:
(a') coupling a suitably functionalized solid support with a compound of the general formula
Figure FDA0002210548630000221
Wherein
Figure FDA0002210548630000222
Has the meaning as defined in claim 1 and X is an N-protecting group, or,
(a' a) coupling the suitably functionalized solid support with a suitable N-protected derivative of the amino acid L-proline;
(a' b) removing the N-protecting group from the product thus obtained; and
(a' c) coupling the product thus obtained with a suitably N-protected derivative of the amino acid D-proline;
(b ') removing the N-protecting group from the product obtained in step (a ') or (a ' c);
(c') coupling the product thus obtained with a suitably N-protected derivative of the amino acid at position 11 in the desired end product, any functional groups which may be present in said N-protected amino acid derivative likewise being suitably protected;
(d') removing the N-protecting group from the product thus obtained;
(e') coupling the product thus obtained with a suitably N-protected derivative of that amino acid which is further from position 11 in the desired end product, any functional groups which may be present in said N-protected amino acid derivative likewise being suitably protected;
(f') removing the N-protecting group from the product thus obtained;
(g ') repeating steps (e ') and (f ') until all amino acid residues have been introduced;
(h') selectively deprotecting one or more protected functional groups present in the molecule and appropriately substituting the reactive groups thus released;
(i') forming an interchain linkage between the side chains of the appropriate amino acid residues at positions 2 and 10;
(j') detaching the product thus obtained from the solid support;
(k') cyclizing the cleaved product from the solid support; and
(l') removing any protecting groups present on the functional groups of any members of the chain of amino acid residues and removing any other protecting groups that may be present in the molecule.
45. A process for the production of a compound according to any one of claims 1 to 29, which process comprises:
(a') coupling a suitably functionalized solid support with a compound of the general formula
Wherein
Figure FDA0002210548630000232
Has the meaning as defined in claim 1 and X is an N-protecting group, or,
(a' a) coupling the suitably functionalized solid support with a suitable N-protected derivative of the amino acid L-proline;
(a' b) removing the N-protecting group from the product thus obtained; and
(a' c) coupling the product thus obtained with a suitably N-protected derivative of the amino acid D-proline;
(b ') removing the N-protecting group from the product obtained in step (a ') or (a ' c);
(c') coupling the product thus obtained with a suitably N-protected derivative of the amino acid at position 11 in the desired end product, any functional groups which may be present in said N-protected amino acid derivative likewise being suitably protected;
(d') removing the N-protecting group from the product thus obtained;
(e') coupling the product thus obtained with a suitably N-protected derivative of that amino acid which is further from position 11 in the desired end product, any functional groups which may be present in said N-protected amino acid derivative likewise being suitably protected;
(f') removing the N-protecting group from the product thus obtained;
(g ') repeating steps (e ') and (f ') until all amino acid residues have been introduced;
(i') forming an interchain linkage between the side chains of the appropriate amino acid residues at positions 2 and 10;
(j') detaching the product thus obtained from the solid support;
(k') cyclizing the cleaved product from the solid support; and
(l') removing any protecting groups present on the functional groups of any members of the chain of amino acid residues and removing any other protecting groups that may be present in the molecule.
46. A method according to claim 44 or claim 45, further comprising the steps of:
(m') converting the obtained product into a pharmaceutically acceptable salt, or converting the obtained pharmaceutically acceptable or unacceptable salt into the corresponding free compound of formula I or into a different pharmaceutically acceptable salt.
47. A compound of formula (I) in free form or in pharmaceutically acceptable salt form,
Figure FDA0002210548630000241
wherein
Figure FDA0002210548630000242
Is a dipeptide consisting of two different amino acid building blocksDPro-LGln，
Z is a chain of 11 amino acid residues, the positions of which in the chain are counted starting from the N-terminal amino acid, wherein the chain of 11 amino acid residues is:
Nle-Cys-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Cys-Tyr；
Nle-Cys-Thr-Ala-Ser-Cha-Pro-Pro-Gln-Cys-Gln；
Nle-Cys-Thr-Ala-Ser-OctG-Pro-Pro-Gln-Cys-Tyr；
Nle-Cys-Thr-Ala-Ser-Cha-Pro-Pro-Gln-Cys-Tyr；
Nle-Cys-Thr-Ala-Ser-Cha-Pro-Pro-Gln-Cys-Cha；
hPhe-Cys-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Cys-Tyr；
Nle-Cys-Thr-Ala-Ser-Cha-Pro-Pro-Gln-Cys-2Cl-Phe；
OctG-Cys-Thr-Ala-Ser-Ile-Pro-Pro-Gln-Cys-Tyr; or
OctG-Cys-Thr-Ala-Ser-OctG-Pro-Pro-Gln-Cys-Gln，
And Cys present at positions P2 and P10 may form a disulfide bond.
48. Use of a compound according to claim 47 in the manufacture of a medicament for the treatment of cardiovascular disease and/or immunological disease and/or neurodegenerative disease.
49. A pharmaceutical composition comprising a compound according to claim 47 and a pharmaceutically inert carrier.
50. A pharmaceutical composition according to claim 49 in a form suitable for topical, transmucosal or pulmonary administration, or for administration by injection or inhalation.
51. A pharmaceutical composition according to claim 49, in a form suitable for oral, transdermal or buccal administration.
52. A pharmaceutical composition according to any one of claims 49 to 51 in the form of a tablet, dragee, capsule, solution, gel, plaster, cream, ointment, syrup, slurry, suspension, spray or suppository.
53. A pharmaceutical composition according to any one of claims 49 to 51 in the form of a liquid or spray.
54. Use of a compound according to claim 47 for the manufacture of a medicament for use as a protease inhibitor.
55. The use according to claim 54, wherein the protease inhibitor medicament is used to prevent infection in a healthy individual, or to slow infection in an infected patient, or for protease activity to mediate or cause cancer, or for protease activity to mediate or cause an immune disease, or for protease activity to mediate or cause inflammation, or for a situation in which protease activity mediates or causes an immune response.
56. Use of a compound according to claim 47 for the preparation of a medicament for use as an inhibitor of elastase.
57. A process for the production of a compound of formula I as defined in claim 47, which process comprises:
(a) coupling the appropriately functionalized solid support with an appropriately N-protected amino acid, which in the desired end product is at position 5,6 or 7, any functional group that may be present in the N-protected amino acid being likewise appropriately protected;
(b) removing the N-protecting group from the product thus obtained;
(c) coupling the product thus obtained with a suitably N-protected amino acid, which amino acid is in a position closer to the N-terminal amino acid residue in the desired end product, any functional groups that may be present in said N-protected amino acid being likewise suitably protected;
(d) removing the N-protecting group from the product thus obtained;
(e) repeating steps (c) and (d) until the N-terminal amino acid residue is introduced;
(fa) coupling the product obtained in step (e) with the appropriate N-protected amino acid L-glutamine;
(fb) removing the N-protecting group from the product thus obtained; and
(fc) coupling the product thus obtained with the appropriately N-protected amino acid D-proline;
(g) removing the N-protecting group from the product obtained in step (fc);
(h) coupling the product thus obtained with a suitably N-protected amino acid, which is in position 11 in the desired end product, any functional groups that may be present in said N-protected amino acid being likewise suitably protected;
(i) removing the N-protecting group from the product thus obtained;
(j) coupling the product thus obtained with said appropriately N-protected amino acid, which is further from position 11 in the desired end product, any functional group that may be present in said N-protected amino acid being likewise appropriately protected;
(k) removing the N-protecting group from the product thus obtained;
(l) Repeating steps (j) and (k) until all amino acid residues have been introduced;
(m) selectively deprotecting one or more protected functional groups present in the molecule and appropriately substituting the reactive groups thus released;
(n) forming an interchain bond between the side chains of the appropriate amino acid residues at positions 2 and 10;
(o) detaching the product thus obtained from the solid support;
(p) cyclizing the cleaved product from the solid support; and
(q) removing any protecting groups present on the functional groups of any amino acid residues in the cyclic product thus obtained and any protecting groups which may also be present in the molecule thus obtained.
58. A process for the production of a compound of formula I as defined in claim 47, which process comprises:
(a) coupling the appropriately functionalized solid support with an appropriately N-protected amino acid, which in the desired end product is at position 5,6 or 7, any functional group that may be present in the N-protected amino acid being likewise appropriately protected;
(b) removing the N-protecting group from the product thus obtained;
(c) coupling the product thus obtained with a suitably N-protected amino acid, which amino acid is in a position closer to the N-terminal amino acid residue in the desired end product, any functional groups that may be present in said N-protected amino acid being likewise suitably protected;
(d) removing the N-protecting group from the product thus obtained;
(e) repeating steps (c) and (d) until the N-terminal amino acid residue is introduced;
(fa) coupling the product obtained in step (e) with the appropriate N-protected amino acid L-glutamine;
(fb) removing the N-protecting group from the product thus obtained; and
(fc) coupling the product thus obtained with the appropriately N-protected amino acid D-proline;
(g) removing the N-protecting group from the product obtained in step (fc);
(h) coupling the product thus obtained with a suitably N-protected amino acid, which is in position 11 in the desired end product, any functional groups that may be present in said N-protected amino acid being likewise suitably protected;
(i) removing the N-protecting group from the product thus obtained;
(j) coupling the product thus obtained with said appropriately N-protected amino acid, which is further from position 11 in the desired end product, any functional group that may be present in said N-protected amino acid being likewise appropriately protected;
(k) removing the N-protecting group from the product thus obtained;
(l) Repeating steps (j) and (k) until all amino acid residues have been introduced;
(n) forming an interchain bond between the side chains of the appropriate amino acid residues at positions 2 and 10;
(o) detaching the product thus obtained from the solid support;
(p) cyclizing the cleaved product from the solid support; and
(q) removing any protecting groups present on the functional groups of any amino acid residues in the cyclic product thus obtained and any protecting groups which may also be present in the molecule thus obtained.
59. A process for the production of a compound of formula I as defined in claim 47, which process comprises:
(a) coupling the appropriately functionalized solid support with an appropriately N-protected amino acid, which in the desired end product is at position 5,6 or 7, any functional group that may be present in the N-protected amino acid being likewise appropriately protected;
(b) removing the N-protecting group from the product thus obtained;
(c) coupling the product thus obtained with a suitably N-protected amino acid, which amino acid is in a position closer to the N-terminal amino acid residue in the desired end product, any functional groups that may be present in said N-protected amino acid being likewise suitably protected;
(d) removing the N-protecting group from the product thus obtained;
(e) repeating steps (c) and (d) until the N-terminal amino acid residue is introduced;
(fa) coupling the product obtained in step (e) with the appropriate N-protected amino acid L-glutamine;
(fb) removing the N-protecting group from the product thus obtained; and
(fc) coupling the product thus obtained with the appropriately N-protected amino acid D-proline;
(g) removing the N-protecting group from the product obtained in step (fc);
(h) coupling the product thus obtained with a suitably N-protected amino acid, which is in position 11 in the desired end product, any functional groups that may be present in said N-protected amino acid being likewise suitably protected;
(i) removing the N-protecting group from the product thus obtained;
(j) coupling the product thus obtained with said appropriately N-protected amino acid, which is further from position 11 in the desired end product, any functional group that may be present in said N-protected amino acid being likewise appropriately protected;
(k) removing the N-protecting group from the product thus obtained;
(l) Repeating steps (j) and (k) until all amino acid residues have been introduced;
(m) selectively deprotecting one or more protected functional groups present in the molecule and appropriately substituting the reactive groups thus released;
(o) detaching the product thus obtained from the solid support;
(p) cyclizing the cleaved product from the solid support; and
(q) removing any protecting groups present on the functional groups of any amino acid residues in the cyclic product thus obtained and any protecting groups which may also be present in the molecule thus obtained.
60. A process for the production of a compound of formula I as defined in claim 47, which process comprises:
(a) coupling the appropriately functionalized solid support with an appropriately N-protected amino acid, which in the desired end product is at position 5,6 or 7, any functional group that may be present in the N-protected amino acid being likewise appropriately protected;
(b) removing the N-protecting group from the product thus obtained;
(c) coupling the product thus obtained with a suitably N-protected amino acid, which amino acid is in a position closer to the N-terminal amino acid residue in the desired end product, any functional groups that may be present in said N-protected amino acid being likewise suitably protected;
(d) removing the N-protecting group from the product thus obtained;
(e) repeating steps (c) and (d) until the N-terminal amino acid residue is introduced;
(fa) coupling the product obtained in step (e) with the appropriate N-protected amino acid L-glutamine;
(fb) removing the N-protecting group from the product thus obtained; and
(fc) coupling the product thus obtained with the appropriately N-protected amino acid D-proline;
(g) removing the N-protecting group from the product obtained in step (fc);
(h) coupling the product thus obtained with a suitably N-protected amino acid, which is in position 11 in the desired end product, any functional groups that may be present in said N-protected amino acid being likewise suitably protected;
(i) removing the N-protecting group from the product thus obtained;
(j) coupling the product thus obtained with said appropriately N-protected amino acid, which is further from position 11 in the desired end product, any functional group that may be present in said N-protected amino acid being likewise appropriately protected;
(k) removing the N-protecting group from the product thus obtained;
(l) Repeating steps (j) and (k) until all amino acid residues have been introduced;
(o) detaching the product thus obtained from the solid support;
(p) cyclizing the cleaved product from the solid support; and
(q) removing any protecting groups present on the functional groups of any amino acid residues in the cyclic product thus obtained and any protecting groups which may also be present in the molecule thus obtained.
61. A method according to any one of claims 57 to 60, further comprising the steps of:
(r) converting the obtained compound of formula I in free form into a pharmaceutically acceptable salt, or converting the obtained pharmaceutically acceptable or unacceptable salt into the corresponding compound of formula I in free form or into a different pharmaceutically acceptable salt.
62. A process for the production of a compound of formula I as defined in claim 47, which process comprises:
(a' a) coupling said suitably functionalized solid support with said suitably N-protected amino acid L-glutamine;
(a' b) removing the N-protecting group from the product thus obtained; and
(a' c) coupling the product thus obtained with the appropriately N-protected amino acid D-proline;
(b ') removing the N-protecting group from the product obtained in step (a' c);
(c') coupling the product thus obtained with said appropriately N-protected amino acid, which is in position 11 in the desired end product, any functional group that may be present in said N-protected amino acid being likewise appropriately protected;
(d') removing the N-protecting group from the product thus obtained;
(e') coupling the product thus obtained with said appropriately N-protected amino acid, which is further from position 11 in the desired end product, any functional group that may be present in said N-protected amino acid being likewise appropriately protected;
(f') removing the N-protecting group from the product thus obtained;
(g ') repeating steps (e ') and (f ') until all amino acid residues have been introduced;
(h') selectively deprotecting one or more protected functional groups present in the molecule and appropriately substituting the reactive groups thus released;
(i') forming an interchain linkage between the side chains of the appropriate amino acid residues at positions 2 and 10;
(j') detaching the product thus obtained from the solid support;
(k') cyclizing the cleaved product from the solid support; and
(l') removing any protecting groups present on the functional groups of any amino acid residues in the cyclic product thus obtained and any protecting groups which may also be present in the molecule thus obtained.
63. A process for the production of a compound of formula I as defined in claim 47, which process comprises:
(a' a) coupling said suitably functionalized solid support with said suitably N-protected amino acid L-glutamine;
(a' b) removing the N-protecting group from the product thus obtained; and
(a' c) coupling the product thus obtained with the appropriately N-protected amino acid D-proline;
(b ') removing the N-protecting group from the product obtained in step (a' c);
(c') coupling the product thus obtained with said appropriately N-protected amino acid, which is in position 11 in the desired end product, any functional group that may be present in said N-protected amino acid being likewise appropriately protected;
(d') removing the N-protecting group from the product thus obtained;
(e') coupling the product thus obtained with said appropriately N-protected amino acid, which is further from position 11 in the desired end product, any functional group that may be present in said N-protected amino acid being likewise appropriately protected;
(f') removing the N-protecting group from the product thus obtained;
(g ') repeating steps (e ') and (f ') until all amino acid residues have been introduced;
(i') forming an interchain linkage between the side chains of the appropriate amino acid residues at positions 2 and 10;
(j') detaching the product thus obtained from the solid support;
(k') cyclizing the cleaved product from the solid support; and
(l') removing any protecting groups present on the functional groups of any amino acid residues in the cyclic product thus obtained and any protecting groups which may also be present in the molecule thus obtained.
64. A process for the production of a compound of formula I as defined in claim 47, which process comprises:
(a' a) coupling said suitably functionalized solid support with said suitably N-protected amino acid L-glutamine;
(a' b) removing the N-protecting group from the product thus obtained; and
(a' c) coupling the product thus obtained with the appropriately N-protected amino acid D-proline;
(b ') removing the N-protecting group from the product obtained in step (a' c);
(c') coupling the product thus obtained with said appropriately N-protected amino acid, which is in position 11 in the desired end product, any functional group that may be present in said N-protected amino acid being likewise appropriately protected;
(d') removing the N-protecting group from the product thus obtained;
(e') coupling the product thus obtained with said appropriately N-protected amino acid, which is further from position 11 in the desired end product, any functional group that may be present in said N-protected amino acid being likewise appropriately protected;
(f') removing the N-protecting group from the product thus obtained;
(g ') repeating steps (e ') and (f ') until all amino acid residues have been introduced;
(h') selectively deprotecting one or more protected functional groups present in the molecule and appropriately substituting the reactive groups thus released;
(j') detaching the product thus obtained from the solid support;
(k') cyclizing the cleaved product from the solid support; and
(l') removing any protecting groups present on the functional groups of any amino acid residues in the cyclic product thus obtained and any protecting groups which may also be present in the molecule thus obtained.
65. A process for the production of a compound of formula I as defined in claim 47, which process comprises:
(a' a) coupling said suitably functionalized solid support with said suitably N-protected amino acid L-glutamine;
(a' b) removing the N-protecting group from the product thus obtained; and
(a' c) coupling the product thus obtained with the appropriately N-protected amino acid D-proline;
(b ') removing the N-protecting group from the product obtained in step (a' c);
(c') coupling the product thus obtained with said appropriately N-protected amino acid, which is in position 11 in the desired end product, any functional group that may be present in said N-protected amino acid being likewise appropriately protected;
(d') removing the N-protecting group from the product thus obtained;
(e') coupling the product thus obtained with said appropriately N-protected amino acid, which is further from position 11 in the desired end product, any functional group that may be present in said N-protected amino acid being likewise appropriately protected;
(f') removing the N-protecting group from the product thus obtained;
(g ') repeating steps (e ') and (f ') until all amino acid residues have been introduced;
(j') detaching the product thus obtained from the solid support;
(k') cyclizing the cleaved product from the solid support; and
(l') removing any protecting groups present on the functional groups of any amino acid residues in the cyclic product thus obtained and any protecting groups which may also be present in the molecule thus obtained.
66. A method according to any one of claims 62 to 65, further comprising the steps of:
(m') converting the obtained compound of formula I in free form into a pharmaceutically acceptable salt, or converting the obtained pharmaceutically acceptable or unacceptable salt into the corresponding compound of formula I in free form or into a different pharmaceutically acceptable salt.
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