2 , 4-SUBSTITUTED PYRIMIDINES AS CYSTEINE PROTEASE INHIBITORS
FIELD OF THE INVENTION
The invention is directed to certain substituted heteroaryl nitrile derivatives, which are protease inhibitors. More specifically, the compounds are inhibitors of cysteine proteases. In particular, the compounds inhibit cysteine proteases of the papain superfamily, more specifically those of the falcipain family, which are cysteine proteases found in the malaria parasite Plasmodium falciparum, and also cysteine proteases of the cathepsin family such as cathepsins K, L, S and B, particularly cathepsin K.
BACKGROUND OF THE INVENTION
Malaria is one of the major disease problems of the developing world. The most virulent malaria-causing parasite in humans is Plasmodium falciparum, which is the cause of hundreds of millions of cases of malaria per annum, and is thought to cause over 1 million deaths each year, Breman, J. G., et al., (2001) Am. Trap. Med. Hyg. 64, 1-11. One problem encountered in the treatment of malaria is the build-up of resistance by the parasite to available drugs. Thus there is a need to develop new antimalarial drugs.
One way of identifying a potential new drug with antimalarial activity is to study biological targets found in the Plasmodium falciparum parasite, in turn by investigating biological pathways in which particular targets might be identified. In Plasmodium falciparum, haemoglobin is transported to an acidic food vacuole, where it is degraded. It appears that multiple enzymes, including food vacuole cysteine, aspartic, and metalloproteases, and a cytosolic aminopeptidase, contribute to haemoglobin hydrolysis, Francis S. E. et al., (1997) Annu. Rev. Microbiol. 51 , 97-123; Rosenthal P.J. Protease inhibitors. In: Rosenthal PJ. , ed. Antimalarial Chemotherapy: Mechanisms of Action, Resistance, and New Directions in Drug Discovery, Totowa, N.J.: Humana Press, (2001) 325-345. Plasmodial haemoglobinases are therefore potential therapeutic targets.
Cysteine protease inhibitors were shown some years ago to block haemoglobin degradation by erythrocytic parasites, causing a characteristic morphological abnormality in which the food vacuole fills with undegraded haemoglobin and parasite development is blocked, Rosenthal P. J., et al., (1998) J. Clin. Invest. 82, 1560-6; Gamboa de Dominguez N.D. and Rosenthal P. J., (1996) Blood 87, 4448-54. Efforts to identify enzymes responsible for haemoglobin degradation led to the characterization of "falcipain" as a trophozoite food vacuole cysteine protease, Rosenthal PJ. and Nelson R.G., (1992) MoI Biochem Parasitol 51 , 143-52; Salas F. et al., (1995) Infect. Immun. 63 2120-5. It has more recently been found that "falcipain" actually constitutes three related papain-family cysteine proteases which share a number of unusual features, known as falcipain-1 , falcipain-2 and falcipain-3, Rosenthal, P. J., et al., (2002) Curr. Pharm. Des. 8, 1659-1672. Falcipain-2 is the principal cysteine protease of Plasmodium falciparum trophozoites,
Shenai B.R. et. al., (2000) J Biol Chem 275, 29000-10. Importantly, cysteine protease inhibitors that inhibit falcipain-2 consistently block haemoglobin hydrolysis and parasite development. These data suggest that falcipain-2 is a key target enzyme, but it is likely that the other two falcipains are also appropriate targets and that, in many cases, they are inhibited by the same compounds that are active against falcipain-2. Like falcipain-2, falcipain-3 readily hydrolyzes native haemoglobin under mildly reducing conditions that are similar to those found in physiological systems, Shenai B.R. et al., (2000) J. Biol. Chem. 275, 29000-10; Sijwali P.S. et al., (2001) Biochem. J. 360, 481-9; Shenai B.R. and Rosenthal P.J., (2002) MoI. Biochem. Parasitol. 122, 99-104. Falcipain-2 and falcipain-3 are similar in structure but falcipain-1 is a more distant relative; it is thought that this enzyme plays a key role in the invasion of erythrocytes by Plasmodium falciparum merozoites but that it is not essential for normal development during the erythrocytic stage, Sijwali, P. S., et al., Proceedings of the National Academy of Sciences of the United States of America 101 , 8721-8726. Whether falcipain-1 also plays a role in haemoglobin processing is unknown. Very recently, a fourth papain-family cysteine protease has been found, now known as falcipain-21. Falcipain-2' is nearly identical in sequence to falcipain-2, differing by only 3 amino acids, none of which are located at the active site. The structure of falcipain-2' is not known, but is likely to be very similar to that of falcipain-2. The biological role of falicipain-2' is also expected to be very similar, although probably not identical, to that of falcipain-2. In any event, cysteine protease inhibition, and in particular the inhibition of falcipain-2, blocks parasite development. Falcipain-2 and related plasmodial cysteine proteases are thus logical targets for antimalarial chemotherapy and therefore there is a need for compounds which are inhibitors of these targets.
P. vivax is the second most important human malaria parasite, after P. falciparum. Although less virulent than P. falciparum, P. vivax is the most widely distributed human malaria parasite, and it causes extensive morbidity (Mendis, K., Sina, B. J., Marchesini, P. and Carter, R. (2001 ) "The neglected burden of Plasmodium vivax malaria" Am. J. Trop. Med. Hyg. 64, 97-106). These two parasites are responsible for more than 90% of episodes of human malaria, totalling several hundred million cases annually. However, comprehensive studies of P. vivax have been limited due to technical shortcomings. Notably, unlike the case with P. falciparum, routine in vitro culture of P. vivax is not available, and animal models are limited to primates. Very recently (Na, B.K., Shenai, B. R., Sijwali, P. S., Choe, Y., Pandey, K. C, Singh, A., Craik, C. S., Rosenthal, P. J. (2004) identification and biochemical characterization of vivapains, cysteine proteases of the malaria parasite Plasmodium vivax. Biochem. J. 378, 529-538), two cysteine protease genes (vivapain-2 and vivapain-3) from P. vivax have been identified and cloned and the heterologously expressed gene products have been characterized biochemically. It was found that these cysteine proteases are apparent orthologues of falcipain-2 and falcipain- 3, but key differences in the biochemical properties of the plasmodial proteases warrant
attention to the inhibition of each enzyme in the evaluation of antimalarial protease inhibitors.
Cathepsins are a family of enzymes which are part of the papain superfamily of cysteine proteases. Certain cathepsins, for example cathepsins K, B, L, and S have been described in the literature. Cathepsin K polypeptide and the cDNA encoding such polypeptide were disclosed in U.S. Patent No. 5,501 ,969. Cathepsin K has also been variously denoted as cathepsin O or cathepsin 02 in the literature. The designation cathepsin K is considered to be the most appropriate and is used herein. Cathepsin K has been expressed, purified, and characterised, Bossard, M. J., et al., (1996) J. Biol.
Chem. 271 , 12517-12524; Drake, F.H., et al., (1996) J. Biol. Chem. 271 , 12511-12516;
Bromme, D., et al., (1996) J. Biol. Chem. 271, 2126-2132.
Cathepsins function in the normal physiological process of protein degradation in animals, including humans, e.g. in the degradation of connective tissue. However, elevated levels of these enzymes in the body can result in pathological conditions leading to disease. Thus, cathepsins have been implicated as causative agents in various disease states, including but not limited to, infections by Pneumocystis Carinii, Trypsanoma cruzi, Trypsanoma brucei, and Crithidia fusiculata; as well as in schistosomiasis, malaria, tumour invasion and tumour metastasis, metachromatic leukodystrophy, muscular dystrophy, amytrophy, inflammation, rheumatoid arthritis, osteoarthritis, osteoporosis, coronary disease, atherosclerosis, autoimmune diseases, respiratory diseases such as obstructive pulmonary disorder (COPD), immunologically mediated diseases (for example, transplant rejection), and other related diseases, see: International Publication Number WO 94/04172, published on March 3, 1994, and references cited therein; see also: European Patent Application EP 0 603 873 A1 , and references cited therein. Two bacterial cysteine proteases from P. gingivallis, called gingipains, have been implicated in the pathogenesis of gingivitis, Potempa, J., et al., (1994) Perspectives in Drug Discovery and Design 2, 445-458.
Cathepsin K is believed to play a causative role in diseases of excessive bone or cartilage loss. Bone is composed of a protein matrix in which spindle- or plate-shaped crystals of hydroxyapatite are incorporated. Type I collagen represents the major structural protein of bone comprising approximately 90% of the protein matrix. The remaining 10% of matrix is composed of a number of non-collagenous proteins, including osteocalcin, proteoglycans, osteopontin, osteonectin, thrombospondin, fibronectin, and bone sialoprotein. Skeletal bone undergoes remodeling at discrete foci throughout life. These foci, or remodeling
units, undergo a cycle consisting of a bone resorption phase followed by a phase of bone replacement.
Bone resorption is carried out by osteoclasts, which are multinuclear cells of haematopoietic lineage. In several disease states, such as osteoporosis and Paget's disease, the normal balance between bone resorption and formation is disrupted, and there is a net loss of bone at each cycle of resorption and formation. Ultimately, this leads to weakening of the bone and may result in increased fracture risk with minimal trauma. Several published studies have demonstrated that inhibitors of cysteine proteases are effective at inhibiting osteoclast-mediated bone resorption, thus indicating an essential role for cysteine proteases in bone resorption. For example, Delaisse, et al., (1980) Biochem. J., 192, 365, suggests that inhibitors of cysteine proteases (e.g., leupeptin, Z- Phe-Ala-CHN2) prevent bone resorption, while serine protease inhibitors were ineffective. Delaisse et. al., (1984) Biochem. Biophys. Res. Commun. 125, 441 , discloses that E-64 (L-trans-epoxysuccinyl-leucinamido-(4-guanidino)butane) and leupeptin are also effective at preventing bone resorption in vivo in rats. Lerner, et al,, (1992) J. Bone Min. Res. 7, 433, discloses that cystatin, an endogenous cysteine protease inhibitor, inhibits PTH stimulated bone resorption in mouse calvariae. Other studies report a correlation between inhibition of cysteine protease activity and bone resorption. Tezuka, et al., (1994) J. Biol. Chem. 269, 1106; Inaoka, et al., (1995) Biochem. Biophys. Res. Commun., 206, 89 and Shi, et al., (1995) FEBS Lett. 357, 129 disclose that under normal conditions cathepsin K is abundantly expressed in osteoclasts and may be the major cysteine protease present in these cells.
The abundant selective expression of cathepsin K in osteoclasts strongly suggests that this enzyme is essential for bone resorption. Thus, inhibition of cathepsin K may provide an effective treatment for diseases of excessive bone loss, including, but not limited to, osteoporosis, gingival diseases such as gingivitis and periodontitis, Paget's disease, hypercalcemia of malignancy, and metabolic bone disease. Cathepsin K levels have also been demonstrated to be elevated in chondroclasts of osteoarthritic synovium. Cathepsin K is also expressed in synovial giant cells taken from osteoarthritic patients (Dodds, et al., (1999) Arthritis & Rheumatism, 42, 1588, and Hou, et al., (2002), American Journal of Pathology 159, 2167). Cathepsin K staining is observed in osteoarthritic as well as rheumatoid arthritic samples (Hou, et al., (2002), American Journal of Pathology 159, 2167). The expression of cathepsin K has also been localized to cartilage tissue and a decrease in pH in cartilage correlated with severity of damage (Konttinen, et al., (2002),
Arthritis & Rheumatism, 46, 953). This observation, combined with the fact that cathepsin
K is an acidic lysosomal protease, strongly suggests a physiological role of cathepsin K in cartilage turnover in addition to bone resorption. These researchers also demonstrated that cathepsin K can degrade aggrecan and type Il collagen, the two major protein components of the cartilage matrix. Thus, inhibition of cathepsin K may also be useful for treating diseases of excessive cartilage or matrix degradation, including, but not limited to, osteoarthritis and rheumatoid arthritis. Cathepsin K has been shown to be abnormally or overexpressed in numerous tumors and in prostate cancer (Littlewood-Evans, et al., (1997), Cancer Res., 57, 5386 and Brubaker, et al., (2003), J. Bone Miner. Res., 18, 222). Furthermore, increased levels of bone resorption marker have been detected in bone metastases of prostate cancer suggesting that cathepsin K inhibitor may have utility in preventing metastasis of tumors to bone (Ishikawa, et al., (2001), MoI. Carcinog., 32, 84 and Brubaker, et al., (2003), J. Bone Miner. Res., 18, 222). Metastatic neoplastic cells also typically express high levels of other proteolytic enzymes such as cathepsin B, S and L that degrade the surrounding matrix. Thus, inhibition of cathepsin K may also be useful for treating certain tumors and neoplastic diseases.
Cathepsin L has been implicated in several diseases including osteoporosis, osteoarthritis, rheumatoid arthritis, lymphoproliferative diseases, cancer, metastasis, atherosclerosis (Lecaille, et al., (2002) Chem. Rev. 102, 4459 and Liu, et al., (2004), Arterioscler Throm Vase Biol. 24, 1359). Cathepsin L-deficient mice have also been shown to have increased resistance to osteoporosis following ovariectomy suggesting its potential for osteoporosis (Potts, et al., (2004) Int. J. Exp. Path. 85, 85). Cathepsin L is required for endothelial progenitor cell-induced neovascularization (Urbich, et al., (2005) Nat. Med. 11 , 206). Similarly, targeting cathepsin L by specific ribozymes decreases cathepsin L protein synthesis and cartilage destruction in rheumatoid arthritis (Schedel, et al., (2004) Gene Ther. 11 , 1040) suggesting its potential role in rheumatoid arthritis.
Cathepsin S has been implicated in several diseases including immune and auto-immune disorders, rheumatoid arthritis, inflammation, inflammatory bowel disease, myesthania gravis, atherosclerosis, lymphoproliferative diseases, cancer, metastasis (Lecaille, et al., (2002) Chem. Rev. 102, 4459 and Liu, et al., (2004), Arterioscler Throm Vase Biol. 24, 1359). 5. Cathepsin S is thought to play a role in invariant chain degradation and antigen presentation and cathepsin S null mice have been shown to have a diminished collagen- induced arthritis (Nakagawa, et al., (1999) Immunity, 10, 207) suggesting its potential role in rheumatoid arthritis.
Cathepsin B has been implicated in immune and auto-immune disorders, rheumatoid arthritis, inflammation, inflammatory bowel disease, myesthania gravis, osteoarthritis, lymphoproliferative diseases, cancer, metastasis (Lecaille, et al., (2002) Chem. Rev. 102, 4459 and Lang, et al., (2000), J. Rheumatol. 27, 1970). Cathepsin B has been implicated in the processing of invariant chain (Zhang, et al., (2000) Immunology, 100, 13) suggesting its role in immune disorders such as those listed above. Cathepsin B is one of the most highly expressed cysteine protease in cartilage and inhibitors of cathepsin B has been shown to inhibit cartilage degradation. Cat B may contribute to matrix degradation through cleavage of aggrecan and collagen, two components of cartilage matrix (Mort et al., (1998), Biochem. J., 335, 491). Additionally, cathepsin B could contribute to the mechanical loading component of osteoarthritis by cleaving lubricin, an abundant lubricating protein in synovial fluid. Cleavage of lubricin by cathepsin B has been shown to increase the coefficient of friction in synovial fluid and intact joints (Elsaid, K.A. et al. (2005), Transactions of the Orthopedic Research Society, 51st Annual Meeting, Abstract 924). These data suggest potential for cathepsin B inhibitors in osteoarthritis.
In view of the number of pathological responses and conditions that are mediated by cathepsins K, L, S and B and particularly by cathepsin K, there is a need for inhibitors of these cathepsins which can be used in the treatment of a variety of conditions.
European Patent Application No. EP 0604798 A1 discloses certain N-arylhydrazine derivatives useful for the control of insects or acarina.
SUMMARY OF THE INVENTION The invention is directed to novel heteroaryl nitrile derivatives and their use as protease inhibitors, more specifically inhibitors of cysteine protease, even more specifically inhibitors of cysteine proteases of the papain superfamily. In one aspect of the invention the cysteine proteases are those of the falcipain family, for example falcipain-2 and falcipain-3, which are examples of cysteine proteases indicated in malaria. In another aspect of the invention the cysteine proteases are those of the cathepsin family for example cathepsins K, L, S and B, particularly cathepsin K, which is a cysteine protease indicated for example in conditions characterised by excessive bone loss such as osteoporosis and bone metastasis, and other bone and joint diseases such as osteoarthritis. The compounds of the invention may also have utility as serine protease inhibitors.
The invention involves the compounds represented hereinbelow, pharmaceutical compositions comprising such compounds and use of the compounds as protease inhibitors.
DETAILED DESCRIPTION OF THE INVENTION
The present invention provides at least one chemical entity selected from a compound of Formula I:
wherein:
R1 represents C1-8alkyl, -C1-8alkyleneNRERF, -C1-8alkyleneNRGC(O)OC1-6alkyl, -C1-8alkyleneNRGC(O)C1-6alkyl or -C1-8alkylene-cycloalkyl;
R3 represents hydrogen or C1-6alkyl;
R4 represents hydrogen or C1-6alkyl;
and
a) X represents NR5 and A represents C(O) and i) R2 represents R2a or R2b wherein
R2a represents -NRH-aryl, -NRH-heteroaryl, -NRH-aryl-heteroaryl or -NRH- heteroaryl-aryl; and
R2b represents -C1-6alkyleneRA, aryl, biaryl, -aryl-heteroaryl, -heteroaryl-aryl, -aryl-heterocyclyl, -aryl-d-aalkylene-heterocyclyl, -aryl-O-d-salkylene-heterocyclyl, aryl-C-i-salkylene-heteroaryl, -aryl-heteraoaryl-d-salkylene-heterocyclyl,
-heteroaryl-aryl-C^salkylene-heterocyclyl, aryloxy, heteroaryl, cycloalkyl, -cycloalkyl-aryl, cycloalkyloxy, heterocyclyl, -NRH-aryl-heterocyclyl, -NRH-cycloalkyl, -NRBCi-6alkyleneRc, -Od-ealkyleneRD or -OC1-6alkenyl; -aryl-d- 3alkylene-heterocyclyl-RJ, -aryl-C1-3alkylene-heteroaryl-RK, C1-3alkyene(NH2)-aryl;
and
R5 represents hydrogen, C^alkyl, C1-6alkenyl, -C(O)R2a, -C-^alkylene-heterocyclyl, -C1-8alkyleneNRGC(O)Ci-6alkyl, -C1-8alkyleneNRGC(O)OCi-6alkyl,
-Ci-8alkyleneNRERF, N-phthalidimido-C-i-aalkylene- or -C(O)C1-6alkyl;
or
ii) R2 and R5 together with the carbon and nitrogen atoms to which they are respectively attached form a group selected from
or
b) X represents NR5 and A represents -SO2- and R2 represents Ci-6alkyl or C1-6aralkyl;
and
R5 represents hydrogen, C1-6alkyl, C1-6alkenyl, -C^salkylene-heterocyclyl, -C1-8alkyleneNRGC(O)C1-6alkylI -C1-8alkyleneNRGC(O)OC1-6alkyl,
-C1-8alkyleneNRERF, N-phthalidimido-Ci-8alkylene- or -C(O)C1-6alkyl;
or
c) X represents CH2 and A represents C(O) and
R2 represents -NRH-aryl, -NRH-heteroaryl, -NRHC1-6aralkyl,
-NRHCi-6alkylene-heteroaryl, -NRH-aryl-heteroaryl, -NRH-aryl-heterocyclyl or -OC1-6aralkyl;
wherein
RA, Rc and RD independently represent hydrogen, halogen, -NRERF, cyano, CCI3, -C(O)C1-6alkyl, C1-3alkyl, cycloalkyl, heterocyclyl, aryl, biaryl, -aryl-heteroaryl, -aryl-C^ 3alkylene-heterocyclyl, -aryl-O-C^alkylene-heterocyclyl, -C1-3alkenylaryl, heteroaryl, C1-6aralkyl, -NHC(O)C1-6alkyl, -NHC(O)OC1-6alkyl, -NHC(O)C1-6aralkyl or -NHC(O)OC1-6aralkyl;
RB represents hydrogen or Ci-8alkyl;
RE and RF independently represent hydrogen or C1-3alkyl; or RE represents cycloalkyl and RF represents hydrogen; or RE and RF together with the nitrogen atom to which they are attached form a 5- or 6-membered heterocyclic ring;
RG represents hydrogen or C1-3alkyl;
RH represents hydrogen, d-6alkyl, -CLealkyleneNHC^C^alkyl, -C1-6alkyleneNHC(O)OC1-4alkyl, or -C1-6alkyleneNRERF;
RJ represents aryl, heteroaryl, heterocyclyl, -C1-3alkylene(aryl)2, -C^alkylene-heteroaryl, -C1-3aralkyl, -Ci^alkylene-oxo-heterocyclyl, -O-C(O)C1-3alkylene-aryl,
Rκ represents one or two aryl substituents;
and salts and solvates thereof.
Terms and Definitions
As used herein, the term "alkyl" as a group or a part of a group refers to a linear or branched alkyl group containing the indicated number of carbon atoms. Examples of such groups include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n- pentyl, isopentyl, neopentyl or hexyl and the like. In another embodiment, the term "alkyl" as a group or a part of a group refers to a linear or branched optionally substituted alkyl group, containing from 1-6 carbon atoms; examples of such groups include methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, tert-butyl, pentyl, neopentyl, or hexyl. Preferred alkyl moieties are C1-4alkyl. In one embodiment alkyl is unsubstituted or substituted at one, two or three positions. Optional substituents include halogen, alkoxy, NR7R8, SOR9 and SO2R9, wherein R7 and R8 independently represent hydrogen or alkyl or R7 and R8 together with the N-atom to which they are attached form a heteroaryl or heterocyclyl ring, and R9 represents alkyl.
As used herein, the term "alkylene" as a group or a part of a group refers to a linear or branched saturated hydrocarbon linker group containing the indicated number of carbon atoms. Examples of such groups include methylene, ethylene and the like.
As used herein, the term "alkenyl" as a group or a part of a group refers to a linear or branched hydrocarbon group containing one or more carbon-carbon double bonds and containing the indicated number of carbon atoms. Examples of such groups include ethenyl, propenyl, buteηyl, pentenyl or hexenyl and the like.
As used herein, the term "alkoxy" as a group or a part of a group refers to an -O-alkyl group wherein alkyl is as herein defined. Examples of such groups include methoxy, ethoxy, propoxy, prop-2-oxy, butoxy, but-2-oxy or methylprop-2-oxy, pentoxy, hexoxy and the like.
In another embodiment, the term "alkoxy"' refers to a linear or branched optionally substituted alkoxy group, containing from 1-6 carbon atoms; examples of such groups include methoxy, ethoxy, propoxy, prop-2-oxy, butoxy, but-2-oxy or methylprop-2-oxy and the like. In one embodiment alkoxy moieties are C^alkoxy. In another embodiment alkoxy is unsubstituted or substituted at one, two or three positions. Optional substituents include halogen, for example fluorine.
As used herein, the term "aralkyl" as a group or a part of a group refers to an alkyl group as herein defined which contains the indicated number of carbon atoms, the alkyl group being substituted with an aryl group as herein defined.
As used herein, the term "aryl" as a group or a part of a group refers to an optionally substituted hydrocarbon aromatic group containing one, two or three conjugated or fused rings with at least one ring having a conjugated pi-electron system. Examples of such groups include optionally substituted phenyl, naphthyl or tetrahydronaphthalenyl and the like. In one embodiment aryl represents phenyl. In one embodiment aryl moieties are unsubstituted. In another embodiment aryl moieties are monosubstituted, disubstituted or trisubstituted. In a further embodiment aryl moieties are monosubstituted or disubstituted. Optional aryl substituents include d-4alkyl, C1-4alkoxy, halogen, nitro, trihalomethyl, trihalomethoxy, -N(C1-3alkyl)2 and -SO2-C1-4alkyl.
In another embodiment the term "aryl" refers to an optionally substituted aromatic group containing one, two or three conjugated or fused rings with at least one ring having a conjugated pi-electron system. In one embodiment aryl moieties are C6-14aryl. In another embodiment aryl moieties are unsubstituted, monosubstituted, disubstituted or trisubstituted phenyl. Optional aryl substituents include aryl, heteroaryl, heterocyclyl, cycloalkyl, halogen, alkoxy and alkyl.
As used herein, the term "aryloxy" as a group or a part of a group refers to an -O-aryl group wherein aryl is as herein defined.
As used herein, the term "biaryl" as a group or a part of a group refers to an aryl group which is directly substituted with a second aryl group, wherein aryl is as herein defined.
As used herein, the term "heteroaryl" as a group or a part of a group refers to an optionally substituted aromatic group comprising one to four heteroatoms selected from N,
O and S, the aromatic group containing one, two or three 5- or 6- membered conjugated
or fused rings with at least one ring having a conjugated pi-electron system. Examples of monocyclic heteroaryl groups (one ring) include optionally substituted thienyl, furyl, furazanyl, pyrrolyl, triazolyl, tetrazolyl, imidazolyl, oxazolyl, thiazolyl, oxadiazolyl, isothiazolyl, isoxazolyl, thiadiazolyl, pyranyl, pyrazolyl, pyrimidyl, pyridazinyl, pyrazinyl, pyridyl, triazinyl, tetrazinyl and the like. Examples of fused aromatic rings include quinolinyl, isoquinolinyl, quinazolinyl, quinoxalinyl, pteridinyl, cinnolinyl, phthalazinyl, naphthyridinyl, indolyl, isoindolyl, azaindolyl, indolizinyl, indazolyl, purinyl, pyrrolopyridinyl, furopyridinyl, benzofuranyl, isobenzofuranyl, benzothienyl, benzoimidazolyl, benzoxazolyl, benzoisoxazolyl, benzothiazolyl, benzoisothiazolyl, benzoxadiazolyl, benzothiadiazolyl, dibenzofuranyl and the like. In one embodiment heteroaryl moieties are pyridyl, imidazolyl, oxazolyl, benzofuranyl, dibenzofuranyl, benzothiazolyl, indolyl or indazolyl. In a further embodiment, heteroaryl moieties are pyridyl, imidazolyl, isoxazolyl, benzofuranyl, dibenzofuranyl, benzothiazolyl, indolyl or indazolyl. In a yet further embodiment optionally substituted heteroaryl moieties are benzofuranyl, pyridyl, dibenzofuranyl, imidazolyl and isoxazolyl. In one embodiment heteroaryl moieties are unsubstituted. In another embodiment heteroaryl moieties are monosubstituted, disubstituted or trisubstituted. In a further embodiment heteroaryl moieties are monosubstituted or disubstituted. Optional heteroaryl substituents include C^alkyl, C1-4alkoxy and halogen. In another embodiment optionally substituted heteroaryl moieties are benzofuranyl, pyridyl, dibenzofuranyl, imidazolyl and isoxazolyl. In one embodiment heteroaryl is unsubstituted, monosubstituted, disubstituted or trisubstituted; in one aspect heteroaryl is unsubstituted or monosubstituted. Optional heteroaryl substituents include aryl, heteroaryl, heterocyclyl, cycloalkyl, halogen, alkoxy and alkyl.
As used herein, the term "cycloalkyl" as a group or a part of a group refers to a saturated cyclic hydrocarbon group of 3 to 7 carbon atoms. Examples of such groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl or cycloheptyl.
In another embodiment cycloalkyl is unsubstituted or substituted at one, two or three positions. Optional cycloalkyl substituents include aryl, heteroaryl, heterocyclyl, cycloalkyl, halogen, alkoxy and alkyl.
As used herein, the term "cycloalkyloxy" as a group or a part of a group refers to an -O-cycloalkyl group wherein cycloalkyl is as herein defined.
As used herein, the term "heterocyclyl" as a group or a part of a group refers to an optionally substituted, 3- to 7-membered, saturated or partially saturated cyclic hydrocarbon group containing one to four heteroatoms selected from N, O and S. Examples of such groups include include pyrrolidinyl, azetidinyl, imidazolidinyl, oxoimidazolidinyl, pyrazolidinyl, oxazolidinyl, piperidinyl, piperazinyl, morpholinyl, thiomorpholinyl, thiazolidinyl, hydantoinyl, valerolactamyl, oxiranyl, oxetanyl, dioxolanyl, dioxanyl, oxathiolanyl, oxathianyl, dithianyl, dihydrofuranyl, tetrahydrofuranyl,
dihydropyranyl, tetrahydropyranyl, tetrahydropyridinyl, tetrahydropyrimidinyl, tetrahydrothiophenyl, tetrahydrothiopyranyl, diazepanyl, azepanyl and the like. In one embodiment heterocyclyl is an optionally substituted 5- or 6-membered group. In another embodiment heterocyclyl moieties are optionally substituted pyrrolidnyl, imidazolidinyl, piperidinyl, piperazinyl or morpholinyl. In a further embodiment heterocyclyl moieties are optionally substituted piperazinyl. In one embodiment heterocyclyl moieties are unsubstituted. In another embodiment heterocyclyl moieties are monosubstituted, disubstituted or trisubstituted. In a further embodiment heterocyclyl moieties are monosubstituted. Optional heterocyclyl substituents include C1-4alkyl, -C(O)C-MaIkVl, -C(O)OCi-4alkyl and oxo.
In another embodiment optional heterocyclyl substituents include aryl, heteroaryl, heterocyclyl, cycloalkyl, halogen, alkoxy and alkyl. Preferred heterocyclyl groups include piperidinyl, more preferably 1 -piperidinyl.
As used herein, the term "halogen" or "halo" refers to a fluorine (fluoro), chlorine (chloro), bromine (bromo) or iodine (iodo) atom. In one embodiment halogen substituents are fluorine or chlorine atoms.
As used herein, the term "pharmaceutically acceptable" used in relation to an ingredient (active ingredient such as an active ingredient, a salt thereof or an excipient) which may be included in a pharmaceutical formulation for administration to a patient, refers to that ingredient being acceptable in the sense of being compatible with any other ingredients present in the pharmaceutical formulation and not being deleterious to the recipient thereof.
As used herein, the term "N-phthalimido" refers to a phthalimide group which is bonded through the nitrogen atom.
As used herein, the term "proteases" are enzymes that catalyze the cleavage of amide bonds of peptides and proteins by nucleophilic substitution at the amide bond, ultimately resulting in hydrolysis. Proteases include: cysteine proteases, serine proteases, aspartic proteases, and metalloproteases. Protease "inhibitors" bind more strongly to the enzyme than the substrate and in general are not subject to cleavage after enzyme catalyzed attack by the nucleophile. They therefore competitively prevent proteases from recognizing and hydrolysing natural substrates and thereby act as inhibitors.
In one embodiment of the invention R1 represents d.8alkyl, -C1-8alkyleneNRERF,
-C1-8alkyleneNRGC(O)OCi-6alkyl or -C1-8alkyleneNRGC(O)C1-6alkyl. In another embodiment R1 represents C1-8alkyl, Ci-8alkyleneNRERF, or C1-8alkyleneNRGC(O)C1-6alkyl. In another embodiment R1 represents C1-8alkyleneNH2. In another embodiment R1
represents C1-8alkyleneNRGC(O)Oterf-butyl. In a further embodiment R1 represents C1-8alkyleneNRGC(O)methyl. In a yet further embodiment, R1 represents C1-8alkyl; In another embodiment R1 represents Ci-6alkyl; in a further embodiment R1 represents isobutyl or neopentyl; in one embodiment R1 represents isobutyl; in another embodiment R1 represents neopentyl.
In one embodiment R3 represents hydrogen or C1-3alkyl and R4 represents hydrogen or Ci-3alkyl; in another embodiment one of R3 and R4 represents or C1-3alkyl and the other represents hydrogen; in a further embodiment R3 and R4 both represent hydrogen.
In one embodiment of the invention X represents NR5 and A represents C(O) and R2 represents R2a or R2b wherein
R2a represents -NRH-aryl, -NRH-heteroaryl, -NRH-aryl-heteroaryl or -NRH-heteroaryl- aryl; and
R2b represents -C1-6alkyleneRA, aryl, biaryl, -aryl-heteroaryl, -heteroaryl-aryl, -aryl-heterocyclyl, -aryl-Ci-aalkylene-heterocyclyl, -aryl-O-C-i^alkylene-heterocyclyl, aryl-C^alkylene-heteroaryl, -aryl-heteraoaryl-C-i.aalkylene-heterocyclyl, -heteroaryl-aryl-C-i-aalkylene-heterocyclyl, aryloxy, heteroaryl, cycloalkyl, -cycloalkyl- aryl, cycloalkyloxy, heterocyclyl, -NRH-aryl-heterocyclyl, -NRH-cycloalkyl, -NR6C1- 6alkyleneRc, -OC1-6alkyleneRD or -OC1-6alkenyl; -aryl-d-salkylene-heterocyclyl-R11, - aryl-C1.3alkylene-heteroaryl-RK, C1-3alkyene(NH2)-aryl;
and
R5 represents hydrogen, C1-6alkyl, C1-6alkenyl, -C(O)R2a, -CLsalkylene-heterocyclyl, - C1-8alkyleneNRGC(O)C1-6alkyl, -C1-8alkyleneNRGC(O)OCi-6alkyl, -C1-8alkyleneNRERF, N-phthalidimido-Ci-8alkylene- or -C(O)Ci_6alkyl.
In another embodiment of the invention X represents NR5 and A represents C(O) and R2 represents -Ci-6alkyleneRA, aryl, -aryl-heteroaryl, -aryl-heterocyclyl,
-aryl-C^alkylene-heterocyclyl, -aryl-O-C-i-salkylene-heterocyclyl, aryloxy, heteroaryl, cycloalkyl, cycloalkyloxy, heterocyclyl, -NRH-aryl, -NRH-heteroaryl, -NRH-aryl-heteroaryl, -NRH-cycloalkyl, -NRBC1-6alkyleneRc, -OC1-6alkyleneRD or -OC1-6alkenyl. In another embodiment X represents NR5 and A represents C(O) and R2 represents aryl, -aryl-heteroaryl, -aryl-heterocyclyl, -aryl-C-i-aalkylene-heterocyclyl, heteroaryl, -NRH-aryl or -NRH-heteroaryl. In one embodiment wherein X represents NR5 and A represents C(O) and where R2 contains a heterocyclyl moiety, heterocyclyl is selected from optionally substituted pyrrolidnyl, imidazolidinyl, piperidinyl, piperazinyl or morpholinyl. In another
embodiment wherein X represents NR5 and A represents C(O) and where R2 contains a heterocyclyl moiety, heterocyclyl is selected from optionally substituted pyrrolidinyl, piperidinyl or piperazinyl.
In another embodiment of the invention X represents NR5 and A represents C(O) wherein R5 represents hydrogen, C1-6alkyl, -C(O)R2a, -C^alkylene-heterocyclyl, -C1-8alkyleneNRGC(O)OC1-6alkyl, -C1-8alkyleneNRERF, N-phthalidimido-C^salkylene- or -C(O)Ci-6alkyl. In another embodiment X represents NR5 and A represents C(O) wherein R5 represents hydrogen, C1-3alkyl, -C(O)R2a, -C1-6alkylene-piperidyl, -d-ealkyleneNC-i. 3alkylC(O)OC1-4alkyl, -C1-6alkyleneN(C1-3alkyl)2, N-phthalidimido-C^ealkylene- Or -C(O)C1- 3alkyl. In a further embodiment X represents NR5 and A represents C(O) wherein R5 represents hydrogen.
In another embodiment of the invention X represents NR5, A represents C(O) and R2 and R5 together with the carbon and nitrogen atoms to which they are respectively attached form a group selected from
In one embodiment of the invention X represents NR5, A represents -S02-, R2 represents C1-6alkyl or C1-6aralkyl, and R5 represents hydrogen, C1-6alkyl, C1-6alkenyl, -C1-8alkylene- heterocyclyl, -Ci-8alkyleneNRGC(O)Ci-6alkyl, -C1-8alkyleneNRGC(O)OC1.6alkyl,
-C1-8alkyleneNRERF, N-phthalidimido-C^salkylene- or -C(O)C1-6alkyl. In another embodiment invention X represents NR5, A represents -S02-, R2 represents Ci-6alkyl or C1-6aralkyl and R5 represents hydrogen.
In one embodiment of the invention X represents CH2, A represents C(O) and R2 represents -NRH-aryl, -NRH-heteroaryl, -NRHC1-6aralkyl, -NRHC1-6alkylene-heteroaryl, -NRH-aryl-heteroaryl, -NRH-aryl-heterocyclyl or -OCi-6aralkyl. In another embodiment X represents CH2, A represents C(O) and R2 represents -NH-aryl, -NH-heteroaryl, -NHC1. 6aralkyl, -NHC1-6alkylene-heteroaryl or -NH-aryl-heteroaryl. In one embodiment wherein X represents CH2, A represents C(O) and where R2 contains a heterocyclyl moiety, heterocyclyl is selected from optionally substituted pyrrolidinyl, imidazolidinyl, piperidinyl, piperazinyl or morpholinyl. In another embodiment wherein X represents CH2, A represents C(O) and where R2 contains a heterocyclyl moiety, heterocyclyl is selected from optionally substituted pyrrolidinyl, piperidinyl or piperazinyl.
In one embodiment RA represents hydrogen, halogen, -NRERF, C1-3alkyl, cycloalkyl, aryl, -aryl-d^alkylene-heterocyclyl or -aryl-O-C^salkylene-heterocyclyl. In another embodiment RA represents hydrogen. In one embodiment, where RA contains a heterocyclyl moiety, heterocyclyl is selected from optionally substituted pyrrolidinyl, imidazolidinyl, piperidinyl, piperazinyl or morpholinyl. In another embodiment, where RA contains a heterocyclyl moiety, heterocyclyl is selected from optionally substituted pyrrolidinyl, piperidinyl or piperazinyl.
In one embodiment RB represents hydrogen. In another embodiment RB represents C1-8alkyl. In another embodiment RB represents C1-6alkyl.
In one embodiment Rc represents hydrogen, halogen, -NRERF, cycloalkyl, aryl, -aryl-C-^alkylene-heterocyclyl, C1-3alkenylaryl, -NHC(O)C1-6alkyl, -NHC(O)OC1-6alkyl or -NHC(O)OC1-6aralkyl. In another embodiment Rc represents hydrogen. In one embodiment, where Rc contains a heterocyclyl moiety, heterocyclyl is selected from optionally substituted pyrrolidinyl, imidazolidinyl, piperidinyl, piperazinyl or morpholinyl. In another embodiment where Rc contains a heterocyclyl moiety, heterocyclyl is selected from optionally substituted pyrrolidinyl, piperidinyl or piperazinyl.
In one embodiment RD represents hydrogen, cyano, CCI3, C1-3alkyl, cycloalkyl, heterocyclyl, aryl, -NHC(O)C1-6alkyl, -NHC(O)OC1-6alkyl or -NRERF. In another embodiment RD represents hydrogen. In one embodiment where RD contains a heterocyclyl moiety, heterocyclyl is selected from optionally substituted pyrrolidinyl, imidazolidinyl, piperidinyl, piperazinyl or morpholinyl. In another embodiment where RD contains a heterocyclyl moiety, heterocyclyl is selected from optionally substituted pyrrolidinyl, piperidinyl or piperazinyl.
In one embodiment RE represents hydrogen or Ci-3alkyl. In another embodiment RE represents hydrogen. In a further embodiment RE represents C1-3alkyl.
In one embodiment RF represents hydrogen or Ci-3alkyl. In another embodiment RF represents hydrogen. In a further embodiment RF represents C1-3alkyl.
In one embodiment RG represents hydrogen or C1-3alkyl. In another embodiment RG represents hydrogen. In a further embodiment RG represents C1-3alkyl.
In one embodiment RH represents hydrogen, Ci-6alkyl, -C1-6alkyleneNHC(O)OC1-4alkyl or -C1-6alkyleneNRERF. In another embodiment RH represents hydrogen or C1-6alkyl. In a further embodiment RH represents C1-6alkyl. In another embodiment RH represents hydrogen.
The meaning of any functional group or substituent thereon at any one occurrence in Formula I, or any subformula thereof, is independent of its meaning, or any other functional group's or substituenfs meaning, at any other occurrence, unless stated otherwise. It is to be understood that the present invention covers all combinations of groups and embodiments described herein.
In an alternative embodiment of the invention there is provided at least one chemical entity selected from a compound of Formula A:
wherein:
X represents NRa or CHR 6b.;
R1 represents alkyl;
R and R independently represent hydrogen or alkyl;
R represents hydrogen, C1-3alkyl or -C(O)R 2. ;
R6 represents hydrogen;
when X represents NR5 then R2 represents aryl, heteroaryl, -NH-aryl, -NH-heteroaryl, alkoxy, -Oalkylaryl or -Oalkylheteroaryl;
when X represents CHR6 then R2 represents -NH-aryl or -NH heteroaryl;
and pharmaceutically acceptable salts and solvates thereof.
In one aspect of the invention, X represents NR5. In another aspect of the invention, X represents CHR6 wherein R6 represents hydrogen.
In one aspect of the invention R1 represents C1-4alkyl, for example isobutyl or neopentyl; in one embodiment R1 represents isobutyl.
When X represents NR5 then in one embodiment R2 represents aryl, heteroaryl, -NH-aryl, -NH-heteroaryl, alkoxy or -Oalkylaryl, in another aspect R2 represents aryl, heteroaryl, -NH-aryl or -NH-heteroaryl, in a further aspect R2 represents aryl, (substituted with alkoxy or halogen), heteroaryl (substituted with alkoxy), -NH-aryl (substituted with alkoxy, alkyl, halogen, imidazolyl or piperidinyl), or -NH-heteroaryl (substituted with halogen, alkoxy, alkyl).
In one embodiment R3 represents hydrogen;
In one embodiment R4 represents hydrogen;
In one embodiment R5 represents hydrogen or -C(O)R2, in another embodiment R5 represents hydrogen.
In one aspect, chemical entities useful in the present invention may be at least one chemical entity selected from:
Ex 1 : 2-(2-Cyano-4-pyhmidinyl)-2-(2,2-dimethylpropyl)-N-4-pyridinylhydrazine carboxamide;
Ex 2: 2-(2-Cyano-4-pyrimidinyl)-N-(4-fluorophenyl)-2-(2-methylpropyl)hydrazine carboxamidetrifluoroacetate;
Ex 3: 2-(2-cyano-4-pyrimidinyl)-N-[3-(methyloxy)phenyl]-2-(2-methylpropyl) hydrazinecarboxamide trifluoroacetate;
Ex 4: 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)-N-[3-(trifluoromethyl)phenyl] hydrazinecarboxamide trifluoroacetate; Ex 5: 2-(2-cyano-4-pyrimidinyl)-N-(2,6-dichloro-4-pyridinyl)-2-(2,2-dimethylpropyl) hydrazinecarboxamide trifluoroacetate;
Ex 6: 2-(2-cyano-4-pyrimidinyl)-N-(3-fluorophenyl)-2-(2-methylpropyl)hydrazine carboxamide trifluoroacetate;
Ex 7: 2-(2-cyano-4-pyrimidinyl)-N-[4-(methyloxy)phenyl]-2-(2-methylpropyl) hydrazinecarboxamide trifluoroacetate;
Ex 8: 2-(2-cyano-4-pyrimidinyl)-N-[2-(methyloxy)phenyl]-2-(2-methylpropyl) hydrazinecarboxamide trifluoroacetate;
Ex 9: 2-(2-cyano-4-pyrimidinyl)-N-(2-fluorophenyl)-2-(2-methylpropyl)hydrazine carboxamide trifluoroacetate; Ex 10: 2-(2-cyano-4-pyrimidinyl)-2-(2,2-dimethylpropyl)-N-[3-(methyloxy)phenyl] hydrazinecarboxamide trifluoroacetate;
Ex 11 : 2-(2-cyano-4-pyrimidinyl)-2-(2,2-dimethylpropyl)-N-[3-(trifluoromethyl)phenyl] hydrazinecarboxamide trifluoroacetate;
Ex 12: 2-(2-cyano-4-pyrimidinyl)-2-(2,2-dimethylpropyl)-N-(4-fluorophenyl)hydrazine carboxamide; Ex 13: 2-(2-cyano-4-pyrimidinyl)-2-(2,2-dimethylpropyl)-N-[4-(methyloxy)phenyl] hydrazinecarboxamide trifluoroacetate;
Ex 14: 2-(2-cyano-4-pyrimidinyl)-N-(3,5-dimethyl-4-isoxazolyl)-2-(2-methylpropyl) hydrazinecarboxamide trifluoroacetate;
Ex 15: 2-(2-cyano-4-pyrimidinyl)-2-(2,2-dimethylpropyl)-N-(2-fluorophenyl)hydrazine carboxamide trifluoroacetate;
Ex 16: 2-(2-cyano-4-pyrimidinyl)-2-(2,2-dimethylpropyl)-N-[2-(methyloxy)phenyl] hydrazinecarboxamide trifluoroacetate;
Ex 17: 2-(2-cyano-4-pyrimidinyl)-2-(2,2-dimethylpropyl)-N-(3-fluorophenyl)hydrazine carboxamide trifluoroacetate; Ex 18: 2-(2-cyano-4-pyrimidinyl)-N-(2,6-dichloro-4-pyridinyl)-2-(2-methylpropyl) hydrazinecarboxamide;
Ex 19: 2-(2-cyano-4-pyrimidinyl)-2-(2,2-dimethylpropyl)-N1 ,N1-bis[4-(methyloxy) phenyl]-
1 ,1-hydrazinedicarboxamide;
Ex 20: 2-(2-cyano-4-pyrimidinyl)-2-(2,2-dimethylpropyl)-N1 ,N1-bis(2-fluorophenyl)-1 ,1- hydrazinedicarboxamide;
Ex 21 : 2-(2-cyano-4-pyrimidinyl)-2-(2,2-dimethylpropyl)-N1 ,N1-bis[2-(methyloxy) phenyl]-
1 ,1-hydrazinedicarboxamide;
Ex 22: N'-(2-cyano-4-pyrimidinyl)-7-(methyloxy)-N'-(2-methylpropyl)-1 -benzofuran-2- carbohydrazide trifluoroacetate; Ex 23: N'-(2-cyano-4-pyrimidinyl)-3-(methyloxy)-N'-(2-methylpropyl)benzohydrazide trifluoroacetate;
Ex 24: N'-(2-cyano-4-pyrimidinyl)-3-fluoro-N'-(2-methylpropyl)benzohydrazide trifluoroacetate;
Ex 25: N'-(2-cyano-4-pyrimidinyl)-4-(methyloxy)-N'-(2-methylpropyl)benzohydrazide trifluoroacetate;
Ex 26: N'-(2-cyano-4-pyrimidinyl)-N'-(2,2-dimethylpropyl)-7-(methyloxy)-1 -benzofuran-2- carbohydrazide;
Ex 27: N'-(2-cyano-4-pyrimidinyl)-N'-(2-methylpropyl)-4-[(trifluoromethyl)oxy] benzohydrazide trifluoroacetate; Ex 28: N'-(2-cyano-4-pyrimidinyl)-4-fluoro-N'-(2-methylpropyl)benzohydrazide trifluoroacetate;
Ex 29: (2-chlorophenyl)methyl 2-(2-cyano-4-pyrimidinyl)-2-(2,2-dimethylpropyl) hydrazinecarboxylate;
Ex 30: 1 ,1-dimethylethyl 2-(2-cyano-4-pyrimidinyl)-2-(2,2-dimethylpropyl) hydrazinecarboxylate; Ex 31 : N2-(2-cyano-4-pyrimidinyl)-N2-(2-methylpropyl)-N1-4-pyridinylglycinamide;
Ex 32: N2-(2-cyano-4-pyrimidinyl)-N2-(2,2-dimethylpropyl)-N1-[4-(methyloxy)dibenzo
[b,d]furan-3-yl]glycinamide;
Ex 33: N2-(2-cyano-4-pyrimidinyl)-N1-(3-fluorophenyl)-N2-(2-methylpropyl) glycinamide;
Ex 34: N2-(2-cyano-4-pyrimidinyl)-N2-(2,2-dimethylpropyl)-N1-[3-(methyloxy)phenyl] glycinamide;
Ex 35: N2-(2-cyano-4-pyrimidinyl)-N2-(2,2-dimethylpropyl)-N1-[4-(1-piperidinyl) phenyl]glycinamide trifluoroacetate;
Ex 36: N2-(2-cyano-4-pyrimidinyl)-N2-(2,2-dimethylpropyl)-N1-[4-(1 H-imidazol-1- yl)phenyl]glycinamide; Ex 37: N2-(2-cyano-4-pyrimidinyl)-N2-(2-methylpropyl)-N1-{4-[(trifluoromethyl)oxy] phenyljglycinamide;
Ex 38: 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)-N-4-pyridinylhydrazine carboxamide;
Ex 39: 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)-N-[4-(trifluoromethyl)phenyl] hydrazϊnecarboxamide trifluoroacetate; Ex 40: N'-(2-cyano-4-pyrimidinyl)-Nl-(2,2-dimethylpropyl)-4-[(4-methyl-1-piperazinyl) methyl]benzohydrazide trifluoroacetate;
Ex 41 : N'-(2-cyano-4-pyrimidinyl)-N'-(2,2-dimethylpropyl)-4-{[2-(4-morpholinyl)ethyl] oxyjbenzohydrazide trifluoroacetate;
Ex 42: ethyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazinecarboxylate; Ex 43: propyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazinecarboxylate;
Ex 44: butyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazinecarboxylate;
Ex 45: 2-methylpropyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazine carboxylate;
Ex 46: 1 -methylethyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazine carboxylate;
Ex 47: 2,2,2-trichloro-1 ,1-dimethylethyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl) hydrazinecarboxylate trifluoroacetate;
Ex 48: 1 ,2,2-trimethylpropyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazine carboxylate;
Ex 49: 1-methylpropyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazine carboxylate trifluoroacetate; Ex 50: 1 -methyl-2-phenylethyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl) hydrazinecarboxylate trifluoroacetate;
Ex 51 : 1 ,3-dimethylbutyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazine carboxylate;
Ex 52: 2,2-dimethylpropyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazine carboxylate; Ex 53: 2-phenylbutyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazine carboxylate;
Ex 54: 3,3-dimethylbutyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazine carboxylate;
Ex 55: 3-methylbutyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazine carboxylate;
Ex 56: 1 ,1-dimethylethyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazine carboxylate;
Ex 57: 1 ,1-dimethylpropyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazine carboxylate;
Ex 58: 1 ,1-dimethylpentyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazine carboxylate; Ex 59: 1 ,1 ,2-trimethylpropyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazine carboxylate;
Ex 60: 1-methyl-1-phenylethyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl) hydrazinecarboxylate;
Ex 61 : 1 ,1-dimethyl-2-phenylethyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl) hydrazinecarboxylate;
Ex 62: 1 ,1-dimethyl-3-phenylpropyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl) hydrazinecarboxylate;
Ex 63: 1 ,1-dimethylethyl 2-(2-cyano-4-pyrimidinyl)-1-methyl-2-(2-methylpropyl) hydrazinecarboxylate; Ex 64: 1 ,1-dimethylethyl 2-(2-cyano-4-pyrimidinyl)-1-[3-(dimethylamino)propyl]-2-(2- methylpropyl)hydrazinecarboxylate;
Ex 65: butyl 2-(2-cyano-4-pyrimidinyl)-1 -[3-(dimethylamino)propyl]-2-(2-methyl propyl)hydrazinecarboxylate;
Ex 66: N'-(2-cyano-5-methyl-4-pyrimidinyl)-4-fluoro-N'-(2-methylpropyl)benzo hydrazide; Ex 67: N'-(2-cyano-5-methyl-4-pyrimidinyl)-Nl-(2,2-dimethylpropyl)-4-fluorobenzo hydrazide;
Ex 68: 1 ,1-dimethylethyl 2-(2-cyano-5-methyl-4-pyrimidinyl)-2-(2,2-dimethylpropyl) hydrazinecarboxylate;
Ex 69: 1,1-dimethylethyl 2-(2-cyano-6-methyl-4-pyrimidinyl)-2-(2-methylpropyl) hydrazinecarboxylate;
Ex 70: 1,1-dimethylethyl 2-(2-cyano-6-methyl-4-pyrimidinyl)-2-(2,2-dimethylpropyl) hydrazinecarboxylate;
Ex 71 : N'-(2-cyanopyrimidin-4-yl)-NI-(2,2-dimethylpropyl)-4-methoxybenzohydrazide;
Ex 72: NI-(2-cyanopyrimidin-4-yl)-N1-(2,2-dimethylpropyl)-3-
(trifluoromethoxy)benzohydrazide;
Ex 73: N'-(2-cyanopyrimidin-4-yl)-N1-(2,2-dimethylpropyl)-3-fluorobenzohydrazide; Ex 74: N'-(2-cyanopyrimidin-4-yl)-N1-(2,2-dimethylpropyl)-4-
(trifluoromethoxy)benzohydrazide;
Ex75: N'-(2-cyanopyrimidin-4-yl)-N1-(2,2-dimethylpropyl)-3-methoxybenzohydrazide;
Ex76: N'-(2-cyanopyrimidin-4-yl)-N'-(2,2-dimethylpropyl)-3,4-difluorobenzohydrazide;
Ex77: NI-(2-cyanopyrimidin-4-yl)-NI-(2,2-dimethylpropyl)-4-fluorobenzohydrazide; Ex78: N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-2-(3-methoxyphenyl)acetohydrazide;
Ex79: N'-(2-cyanopyrimidin-4-yl)-2-(4-fluorophenyl)-N'-isobutylacetohydrazide;
Ex80: N'-(2-cyanopyrimidin-4-yl)-N'-(2,2-dimethylpropyl)-4-fluoro-2-
(trifluoromethyl)benzohydrazide;
Ex 81 : N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-2-(4-methoxyphenyl)acetohydrazide; Ex 82: N'-(2-cyanopyrimidin-4-yl)-Nl-isobutylhexanohydrazide trifluoroacetate;
Ex 83: N'-(2-cyanopyrimidin-4-yl)-N'-isobutylpentanohydrazide trifluoroacetate;
Ex 84: N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-3,3-dimethylbutanohydrazide;
Ex 85: N'-(2-cyanopyrimidin-4-yl)-N'-isobutylpropanohydrazide;
Ex 86: N'-(2-cyanopyrimidin-4-yl)-N'-isobutylbutanohydrazide; Ex 87: N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-3-methylbutanohydrazide;
Ex 88: N1-(2-cyanopyrimidin-4-yl)-3-cyclopentyl-N'-isobutylpropanohydrazide;
Ex 89: N'-(2-cyanopyrimidin-4-yl)-N'-isobutylcyclopentanecarbohydrazide;
Ex 90: 4-chloro-N'-(2-cyanopyrimidin-4-yl)-N'-isobutylbutanohydrazide;
Ex 91 : 5-chloro-NI-(2-cyanopyrimidin-4-yl)-N'-isobutylpentanohydrazide; Ex 92: N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-2-methylpropanohydrazide;
Ex 93: N'-(2-cyanopyrimidin-4-yl)-N'-isobutylcyclohexanecarbohydrazide;
Ex 94: N'-(2-cyanopyrimidin-4-yl)-Nl-isobutyl-2,2-dimethylpropanohydrazide;
Ex 95: N'-(2-cyanopyrimidin-4-yl)-2-cyclopentyl-NI-isobutylacetohydrazide;
Ex 96: N'-(2-cyanopyrimidin-4-yl)-N'-isobutylbenzohydrazide; Ex 97: NI-(2-cyanopyrimidin-4-yl)-N'-isobutyl-2-phenylacetohydrazide trifluoroacetate;
Ex 98: N1-(2-cyanopyrimidin-4-yl)-N'-isobutyl-3-phenylpropanohydrazide;
Ex 99: N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-3,5,5-trinnethylhexanohydrazide;
Ex 100: N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-2(N,N-diπnethylamino)acetohydrazide;
Ex 101 : N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-2-methylbutanohydrazide; Ex 102: 1-acetyl-Nl-(2-cyanopyrimidin-4-yl)-Nl-isobutylpiperidine-4-carbohydrazide;
Ex 103: 2-(4-chlorophenyl)-N'-(2-cyanopyrimidin-4-yl)-N'-isobutylacetohydrazide;
Ex 104: N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-2-[4-(trifluoromethyl)phenyl]acetohydrazide;
Ex 105: N1-(2-cyanopyrimidin-4-yl)-2-[4-(dimethylamino)phenyl]-N'-isobutylacetohydrazide;
Ex 106: N'-(2-cyanopyrimidin-4-yl)-N1-isobutyl-2-[4-
(trifluoromethoxy)phenyl]acetohydrazide;
Ex 107: N1-(2-cyanopyrimidin-4-yl)-N'-isobutyl-2-(4-methylphenyl)acetohydrazide; Ex 108: N'-(2-cyanopyrimidin-4-yl)-N1-isobutyl-2-(4-nitrophenyl)acetohydrazide;
Ex 109: 2-biphenyl-4-yl-N'-(2-cyanopyrimidin-4-yl)-N'-isobutylacetohydrazide;
Ex 110: 2-(4-chlorophenyl)-N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-2- methylpropanohydrazide trifluoroacetate;
Ex 111 : N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-2-[4-(methylsulfonyl)phenyl]acetohydrazide trifluoroacetate;
Ex 112: N'-(2-cyanopyrinnidin-4-yl)-N'-isobutyl-2-[3-(trifluoronnethyl)phenyl]acetohydrazide trifluoroacetate;
Ex 113: 2-(2-chlorophenyl)-N'-(2-cyanopyrimidin-4-yl)-N1-isobutylacetohydrazide trifluoroacetate; Ex 114: N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-2-phenylpropanohydrazide trifluoroacetate;
Ex 115: N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-2-(3-nitrophenyl)acetohydrazide trifluoroacetate;
Ex 116: N1-(2-cyanopyrimidin-4-yl)-N'-isobutyl-2-(2-methylphenyl)acetohydrazide trifluoroacetate; Ex 117: NI-(2-cyanopyrimidin-4-yl)-N'-isobutyl-2-(2-methoxyphenyl)acetohydrazide trifluoroacetate;
Ex 118: N'-(2-cyanopyrimidin-4-yl)-2-(3-fluorophenyl)-N'-isobutylacetohydrazide trifluoroacetate;
Ex 119: 2-(4-bromophenyl)-N'-(2-cyanopyrimidin-4-yl)-N'-isobutylacetohydrazide trifluoroacetate;
Ex 120: N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-2-methyl-2-phenylpropanohydrazide;
Ex 121 : N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-4-[(4-methylpiperazin-1- yl)methyl]benzohydrazide trifluoroacetate;
Ex 122: N' N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-2-mesitylacetohydrazide; Ex 123: N'-(2-cyanopyrimidin-4-yl)-N'-(2,2-dimethylpropyl)-4-(4-methylpiperazin-1- yl)benzohydrazide;
Ex 124: N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-4-(2-morpholin-4-ylethoxy)benzohydrazide trifluoroacetate;
Ex 125: N'-(2-cyanopyrimidin-4-yl)-N'-(2,2-dimethylpropyl)-4-(morpholin-4- ylmethyl)benzohydrazide trifluoroacetate;
Ex 126: Nl-(2-cyano-4-pyrimidinyl)-NI-(2,2-dimethylpropyl)-4-[(4-propyl-1 - piperazinyl)methyl] benzohydrazide;
Ex 127: N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-4-(morpholin-4-ylnnethyl)ben2ohyclrazicle trifluoroacetate;
Ex 128: NI-(2-cyanopyrimidin-4-yl)-N'-(2,2-dimethylpropyl)-4-(4-propylpiperazin-1 - yl)benzohydrazide trifluoroacetate; Ex 129: N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-4-[(4-propylpiperazin-1- yl)methyl]benzohydrazide trifluoroacetate;
Ex 130: N'-(2-cyanopyrimidin-4-yl)-N'-(2,2-dimethylpropyl)-2-methyl-2-{4-[(4- methylpiperazin-1-yl)methyl]phenyl}propanohydrazide trifluoroacetate;
Ex 131 : N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-2-methyl-2-{4-[(4-methylpiperazin-1- yl)methyl]phenyl}propanohydrazide trifluoroacetate;
Ex 132: N'-(2-cyano-5-methylpyrimidin-4-yl)-N'-isobutyl-2-[4-(2-morpholin-4- ylethoxy)phenyl]acetohydrazide;
Ex 133: N1-(2-cyano-5-methylpyrimidin-4-yl)-N1-isobutyl-2-(4- methoxyphenyl)acetohydrazide trifluoroacetate; Ex 134: NI-(2-cyano-4-pyrimidinyl)-N'-(2,2-dimethylpropyl)-4-(1-piperazinyl methyl)benzohydrazide;
Ex 135: 2-(2-cyanopyrimidin-4-yl)-2-(2,2-dimethylpropyl)-N-[4-(6-methyl-1 ,3-benzothiazol-
2-yl)phenyl]hydrazinecarboxamide;
Ex 136: N-butyl-2-(2-cyanopyrimidin-4-yl)-2-isobutylhydrazinecarboxamide trifluoroacetate;
Ex 137: 2-(2-cyanopyrimidin-4-yl)-2-isobutyl-N-(2-phenylethyl)hydrazinecarboxamide trifluoroacetate;
Ex 138: 2-(2-cyanopyrimidin-4-yl)-N-ethyl-2-isobutylhydrazinecarboxamide trifluoroacetate; Ex 139: N-(2-chloroethyl)-2-(2-cyanopyrimidin-4-yl)-2-isobutylhydrazinecarboxamide trifluoroacetate;
Ex 140: N-benzyl-2-(2-cyanopyrimidin-4-yl)-2-isobutylhydrazinecarboxamide trifluoroacetate;
Ex 141 : 2-(2-cyanopyrimidin-4-yl)-N-cyclopentyl-2-isobutylhydrazinecarboxamide trifluoroacetate;
Ex 142: 2-(2-cyanopyrimidin-4-yl)-2-isobutyl-N-propylhydrazinecarboxamide;
Ex 143: 4-methoxyphenyl 2-(2-cyanopyrimidin-4-yl)-2-isobutylhydrazinecarboxylate;
Ex 144: 3-(trifluoromethyl)phenyl 2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazinecarboxylate; Ex 145: 2-methoxyphenyl 2-(2-cyanopyrimidin-4-yl)-2-isobutylhydrazinecarboxylate;
Ex 146: 2-chlorophenyl 2-(2-cyanopyrimidin-4-yl)-2-isobutylhydrazinecarboxylate;
Ex 147: 4-fluorophenyl 2-(2-cyanopyrimidin-4-yl)-2-isobutylhydrazinecarboxylate;
Ex 148: benzyl 2-(2-cyanopyrimidin-4-yl)-2-isobutylhydrazinecarboxylate;
Ex 149: allyl 2-(2-cyanopyrimidin-4-yl)-2-isobutylhydrazinecarboxylate;
Ex 150: Cyclohexyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazine carboxylateEx
151 : cyclopentylmethyl 2-(2-cyanopyrimidin-4-yl)-2-isobutylhydrazinecarboxylate trifluoroacetate;
Ex 152: ferf-butyl 4-[({[2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazino]carbonyl}oxy)methyl]piperidine-1-carboxylate;
Ex 153: 1 ,1-dimethylbutyl 2-(2-cyanopyrimidin-4-yl)-2-isobutylhydrazinecarboxylate;
Ex 154: 2-cyano-1 ,1-dimethylethyl 2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazinecarboxylate;
Ex 155: 2-fert-butoxy-1 ,1-dimethyl-2-oxoethyl 2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazinecarboxylate;
Ex 156: 3-[(te/t-butoxycarbonyl)amino]-1 ,1-dimethylpropyl 2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazinecarboxylate; Ex 157: te/t-butyl 4-[3-({[2-(2-cyanopyrimidin-4-yl)-2-isobutylhydrazino]carbonyl}oxy)-3- methylbutyl]piperazine-1-carboxylate;
Ex 158: 5-[(teAf-butoxycarbonyl)amino]-1 ,1-dimethylhexyl 2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazinecarboxylate;
Ex 159: A/2-(2-cyanopyrimidin-4-yl)-Λ/2-(2,2-dimethylpropyl)-Λ/1-(4- fluorophenyOglycinamide;
Ex 160: Λ/2-(2-cyanopyrimidin-4-yl)-Λ/2-(2,2-dimethylpropyl)-Λ/1-(pyridin-3- ylmethyl)glycinamide;
Ex 161 : N2-(2-cyanopyrimidin-4-yl)-N1-(4-fluorobenzyl)-N2-isobutylglycinamide;
Ex 162: N2-(2-cyanopyrimidin-4-yl)-N2-(2,2-dimethylpropyl)-N1-[4- (trifluoromethoxy)phenyl]glycinamide;
Ex 163: N2-(2-cyanopyrimidin-4-yl)-N2-(2,2-dimethylpropyl)-N1-[4-
(trifluoromethoxy)benzyl]glycinamide;
Ex 164: N2-(2-cyanopyrimidin-4-yl)-N2-(2,2-dimethylpropyl)-N1-[2-(4- fluorophenyl)ethyl]glycinamide; Ex 165: N2-(2-cyanopyrimidin-4-yl)-N2-(2,2-dimethylpropyl)-N1-[2-(6-fluoro-1H-indol-3- yl)ethyl]glycinamide;
Ex 166: N2-(2-cyanopyrimidin-4-yl)-N2-(2,2-dimethylpropyl)-N1-[2-
(trifluoromethoxy)benzyl]glycinamide;
Ex 167: N2-(2-cyanopyrimidin-4-yl)-N2-(2,2-dimethylpropyl)-N1-[3- (trifluoromethoxy)phenyl]glycinamide;
Ex 168: N2-(2-cyanopyrimidin-4-yl)-N2-(2,2-dimethylpropyl)-N1-(3- fluorobenzyl)glycinamide;
Ex 169: N2-(2-cyanopyrimidin-4-yl)-N2-(2,2-dimethylpropyl)-N1-(3- fluorophenyl)glycinamide;
Ex 170: N2-(2-cyanopyrimidin-4-yl)-N2-(2,2-dimethylpropyl)-N1-pyridin-3-ylglycinamide;
Ex 171 : N2-(2-cyanopyrimidin-4-yl)-N2-(2,2-dimethylpropyl)-N1-1 H-indazol-6-ylglycinamide; Ex 172: N2-(2-cyanopyrimidin-4-yl)-N2-(2,2-dimethylpropyl)-N1-(2- fluorobenzyl)glycinamide;
Ex 173: N2-(2-cyanopyrimidin-4-yl)-N2-(2,2-dimethylpropyl)-N1-pyridin-4-ylglycinamide;
Ex 174: N2-(2-cyanopyrimidin-4-yl)-N2-(2,2-dimethylpropyl)-N1-(4-morpholin-4- ylphenyl)glycinamide; Ex 175: N2-(2-cyanopyrimidin-4-yl)-N2-(2,2-dimethylpropyl)-N1-(5-methylisoxazol-3- yl)glycinamide;
Ex 176: fert-butyl 2-(2-cyanopyrimidin-4-yl)-2-isobutyl-1-(3-piperidin-1- ylpropyl)hydrazinecarboxylate;
Ex 177: ferf-butyl 2-(2-cyanopyrimidin-4-yl)-1-[3-(1 ,3-dioxo-1 ,3-dihydro-2H-isoindol-2- yl)propyl]-2-isobutylhydrazinecarboxylate;
Ex 178: te/t-butyl 2-(2-cyanopyrimidin-4-yl)-1-[4-(1 ,3-dioxo-1 ,3-dihydro-2H-isoindol-2- yl)butyl]-2-isobutylhydrazinecarboxylate;
Ex 179: te/t-butyl 2-(2-cyanopyrimidin-4-yl)-1-{4-[(ethoxycarbonyl)(methyl)amino]butyl}-2- isobutylhydrazinecarboxylate; Ex 180: terf-butyl 2-(2-cyanopyrimidin-4-yl)-1 -[5-(1 ,3-dioxo-1 ,3-dihydro-2H-isoindol-2- yl)pentyl]-2-isobutylhydrazinecarboxylate;
Ex 181 : tert-bu\y\ 2-(2-cyanopyrimidin-4-yl)-1-[4-(dimethylamino)butyl]-2- isobutylhydrazinecarboxylate;
Ex 182: terf-butyl 2-(2-cyanopyrimidin-4-yl)-1-[3-(dimethylamino)propyl]-2-(2,2- dimethylpropyl)hydrazinecarboxylate;
Ex 183: 1 ,1-dimethylethyl {3-[{[2-(2-cyano-4-pyrimidinyl)-2-(2- methylpropyl)hydrazino]carbonyl}(1 ,1-dimethylethyl)amino]propyl}carbamate;
Ex 184: N'-(2-cyanopyrimidin-4-yl)-Nl-isobutyl-4-methylpiperazine-1 -carbohydrazide;
Ex 185: N'-(2-cyanopyrimidin-4-yl)-N1-isobutylmorpholine-4-carbohydrazide; Ex 186: N'-(2-cyanopyrimidin-4-yl)-N'-isobutyl-2-oxoimidazolidine-1 -carbohydrazide;
Ex 187: benzyl [3-(terf-butyl{[2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazino]carbonyl}amino)propyl]carbamate;
Ex 188: benzyl [4-(terf-butyl{[2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazino]carbonyl}amino)butyl]carbamate; Ex 189: terf-butyl [4-(butyl{[2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazino]carbonyl}amino)butyl]carbamate;
Ex 190: tert-butyl [5-(butyl{[2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazino]carbonyl}amino)pentyl]carbamate;
Ex 191: tert-butyl [4-(tert-butyl{[2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazino]carbonyl}amino)butyl]carbamate; Ex 192: tert-butyl [5-(tert-butyl{[2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazino]carbonyl}amino)pentyl]carbamate;
Ex 193: benzyl [2-(fert-butyl{[2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazino]carbonyl}amino)ethyl]carbamate;
Ex 194: tert-butyl {3-[{[2-(2-cyanopyrimidin-4-yl)-2-isobutylhydrazino]carbonyl}(4- fluorophenyl)amino]propyl}carbamate;
Ex 195: Λ/-butyl-2-(2-cyanopyrimidin-4-yl)-2-isobutyl-W-pyridin-4-ylhydrazinecarboxannide trifluoroacetate;
Ex 196: tert-butyl [6-(butyl{[2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazino]carbonyl}amino)hexyl]carbamate; Ex 197: Λ/-(tert-butyl)-2-(2-cyanopyrimidin-4-yl)-Λ/-[4-(dimethylamino)benzyl]-2- isobutylhydrazinecarboxamide;
Ex 198: Λ/-(tert-butyl)-2-(2-cyanopyrimidin-4-yl)-A/-{(2E)-3-[4-(dimethylamino)phenyl]prop-
2-en-1-yl}-2-isobutylhydrazinecarboxamide;
Ex 199: 2-(2-cyanopyrimidin-4-yl)-Λ/-(4-fluorophenyl)-2-isobutyl-Λ/~ methylhydrazinecarboxamide;
Ex 200: Λ/-butyl-2-(2-cyanopyrimidin-4-yl)-Λ/-[4-(dimethylamino)benzyl]-2- isobutylhydrazinecarboxamide;
Ex 201 Λ/-butyl-2-(2-cyanopyrimidin-4-yl)-Λ/-{(2£)-3-[4-(dimethylamino)phenyl]prop-2-en-1- yl}-2-isobutylhydrazinecarboxamide; Ex 202: Λ/-butyl-2-(2-cyanopyrimidin-4-yl)-2-isobutyl-Λ/-{4-[(4-methylpiperazin-1- yl)methyl]benzyl}hydrazinecarboxamide trifluoroacetate;
Ex 203: 4-[[3-(4-fluorophenyl)-2-oxo-1-imidazolidinyl](2-methylpropyl) amino]-2- pyrimidinecarbonitrile;
Ex 204: 4-{(2,2-dimethylpropyl)[3-(4-fluorophenyl)-2-oxo-1-imidazolidinyl] amino}-2- pyrimidinecarbonitrile;
Ex 205: 4-[{3-[4-(methyloxy)phenyl]-2-oxo-1-imidazolidinyl}(2-methylpropyl) amino]-2- pyrimidinecarbonitrile;
Ex 206: : Λ/-(3-aminopropyl)-2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl) hydrazinecarboxamide trifluoroacetate; Ex 207: Λ/-(2-aminoethyl)-2-(2-cyanopyrimidin-4-yl)-2-isobutylhydrazinecarboxamide;
Ex 208: piperidin-4-ylmethyl 2-(2-cyanopyrimidin-4-yl)-2-isobutylhydrazinecarboxylate;
Ex 209: Λ/-(4-aminobutyl)-Λ/-butyl-2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazinecarboxamide trifluoroacetate;
Ex 210: A/-(5-aminopentyl)-Λ/-butyl-2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazinecarboxamide; Ex 211 : Λ/-(4-aminobutyl)-2-(2-cyanopyrimidin-4-yl)-2-isobutylhydrazinecarboxamide trifluoroacetate;
Ex 212: A/-(5-aminopentyl)-2-(2-cyanopyrimidin-4-yl)-2-isobutylhydrazinecarboxamide trifluoroacetate;
Ex 213: Λ/'-(5-amino-2,2-dimethylpentyl)-Λ/I-(2-cyanopyrimidin-4- yl)cyclohexanecarbohydrazide trifluoroacetate;
Ex 214: 3-amino-1 ,1-dimethylpropyl 2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazinecarboxylate trifluoroacetate;
Ex 215: Λ/-(6-aminohexyl)-Λ/-butyl-2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazinecarboxamide trifluoroacetate; Ex 216: /V-(3-aminopropyl)-2-(2-cyanopyrimidin-4-yl)-/V-(4-fluorophenyl)-2- isobutylhydrazinecarboxamide trifluoroacetate;
Ex 217: 1 ,1-dimethyl-3-piperazin-1-ylpropyl 2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazinecarboxylate;
Ex 218: Λ/-(4-aminobutyl)-2-(2-cyanopyrimidin-4-yl)-Λ/-(4-fluorophenyl)-2- isobutylhydrazinecarboxamide trifluoroacetate;
Ex 219: Λ/-(5-aminopentyl)-2-(2-cyanopyrimidin-4-yl)-Λ/-(4-fluorophenyl)-2- isobutylhydrazinecarboxamide trifluoroacetate;
Ex 220: /V-(6-aminohexyl)-2-(2-cyanopyrimidin-4-yl)-Λ/-(4-fluorophenyl)-2- isobutylhydrazinecarboxamide trifluoroacetate; Ex 221 : Methyl [5-({[2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazino] carbonyl}amino)pentyl]carbamate;
Ex 222: tert-butyl {5-[1-(2-cyanopyrimidin-4-yl)-2-(cyclohexylcarbonyl)hydrazino]-4,4- dimethylpentyl}carbamate;
Ex 223: te/ϊ-butyl {5-[1-(2-cyanopyrimidin-4-yl)-2-(4-fluorobenzoyl)hydrazino]-4,4- dimethylpentyljcarbamate;
Ex 224: Λ/-[4-(acetylamino)butyl]-Λ/-butyl-2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazinecarboxamide;
Ex 225: Λ/-[5-(acetylamino)pentyl]-/V-butyl-2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazinecarboxamide; Ex 226: Λ/-{5-[1-(2-cyanopyrimidin-4-yl)-2-(4-fluorobenzoyl)riydrazino]-4,4- dimethylpentyl}acetamide;
Ex 227: Λ/-{5-[1 -(2-cyanopyrimidin-4-yl)-2-(cyclohexylcarbonyl)hydrazino]-4,4- dimethylpentyl}acetamide;
Ex 228: methyl [4-({[2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazino]carbonyl}amino)butyl]carbamate; Ex 229: Λ/-[5-(acetylamino)pentyl]-2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazinecarboxamide;
Ex 230: Λ/-[6-(acetylamino)hexyl]-Λ/-butyl-2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazinecarboxamide;
Ex 231 : methyl [6-(butyl{[2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazino]carbonyl}amino)hexyl]carbamate;
Ex 232: 3-(acetylamino)-1 ,1-dimethylpropyl 2-(2-cyanopyrimidin-4-yl)-2- isobutyl hyd razi necarboxylate;
Ex 233: 3-(4-acetylpiperazin-1-yl)-1 ,1-dimethylpropyl 1-acetyl-2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazinecarboxylate; Ex 234: 3-(4-acetylpiperazin-1-yl)-1 ,1-dimethylpropyl 2-(2-cyanopyrimidin-4-yl)-2- isobutylhydrazinecarboxylate;
Ex 235: 1 ,1-dimethylethyl 2-(5-bromo-2-cyano-4-pyrimidinyl)-1-(3-bromo propyl)-2-(2- methylpropyl)hydrazinecarboxylate;
Ex 236: Λ/'-(2-cyano-4-pyrimidinyl)-A/I-(3,3-dimethylbutyl)-4-fluorobenzo hydrazide; Ex 237: 4-[(2,2-dimethylpropyl)(2,4-dioxo-1-imidazolidinyl)amino]-2-pyrimidinecarbonitrile;
Ex 238: 4-[(2,4-dioxo-1 -imidazolidinyl)(2-methylpropyl)amino]-2-pyrimidinecarbonitrile;
Ex 239: 4-[(5,5-dimethyl-2,4-dioxo-1 ,3-oxazolidin-3-yl)(2-methylpropyl) amino]-2- pyrimidinecarbonitrile;
Ex 240: /V-(2-cyano-4-pyrimidinyl)-4-fluoro-/\/'-(3-methylbutyl)benzo hydrazide; Ex 241 : 1 ,1-dimethylethyl 2-(2-cyano-5-methyl-4-pyrimidinyl)-2-(2-methyl propyl)hydrazinecarboxylate;
Ex 242: Λ/I-(2-cyanopyrimidin-4-yl)-Λ/I-isobutyl-1 -phenylmethanesulfonohydrazide;
Ex 243: Λ/I-(2-cyanopyrimidin-4-yl)-Λ/I-isobutylmethanesulfonohydrazide;
Ex 244: Λ/I-(2-cyano-4-pyrimidinyl)-Λ/-[3-(dimethylamino)propyl]-Λ/'-(2-methyl propyl)-2- phenylacetohydrazide trifluoroacetate; and free bases, solvates and alternative salts thereof.
The compounds of the present invention may be in the form of and/or may be administered as a pharmaceutically acceptable salt. Indeed, in certain embodiments of the invention, pharmaceutically acceptable salts of the compounds according to Formula I may be preferred over the respective free base or free acid because such salts impart greater stability or solubility to the molecule thereby facilitating formulation into a dosage
form. Accordingly, the invention is further directed to pharmaceutically acceptable salts of the compounds according to Formula I.
As used herein, the term "pharmaceutically acceptable salts" refers to salts that retain the desired biological activity of the subject compound and exhibit minimal undesired toxicological effects. For a review on suitable salts see Berge et al, J. Pharm. ScL, 1977, 66, 1-19. The term "pharmaceutically acceptable salts" includes both pharmaceutically acceptable acid addition salts and pharmaceutically acceptable base addition salts. These pharmaceutically acceptable salts may be prepared in situ during the final isolation and purification of the compound, or by separately reacting the purified compound in its free acid or free base form with a suitable base or acid, respectively. The salt may precipitate from solution and be collected by filtration or may be recovered by evaporation of the solvent.
A pharmaceutically acceptable acid addition salt can be formed by reaction of a compound of formula (I) with a suitable inorganic or organic acid (such as hydrobromic, hydrochloric, sulfuric, sulfamic, nitric, phosphoric, succinic, maleic, hydroxymaleic, acrylic, formic, acetic, hydroxyacetic, phenylacetic, butyric, isobutyric, propionic, fumaric, citric, tartaric, lactic, mandelic, benzoic, o-acetoxybenzoic, chlorobenzoic, methylbenzoic, dinitrobenzoic, hydroxybenzoic, methoxybenzoic salicylic, glutamaic, stearic, ascorbic, palmitic, oleic, pyruvic, pamoic, malonic, lauric, glutaric aspartic, p-toluenesulfonic, benzenesulfonic, methanesulfonic, ethanesulfonic, 2-hydroxyethanesulfonic, naphthalenesulfonic (e.g. 2-naphthalenesulfonic), p-aminobenzenesulfonic (i.e. sulfanilic), hexanoic, heptanoic, or phthalic acid), optionally in a suitable solvent such as an organic solvent, to give the salt which is usually isolated for example by crystallisation and filtration. A pharmaceutically acceptable acid addition salt of a compound of formula (I) can comprise or be for example a hydrobromide, hydrochloride, hydroiodide, sulfate, bisulfate, nitrate, phosphate, hydrogen phosphate, succinate, maleate, malate, formate, acetate, trifluoroacetate, saccharate, propionate, fumarate, citrate, tartrate, lactate, benzoate, salicylate, glutamate, aspartate, p-toluenesulfonate, benzenesulfonate, methanesulfonate, ethanesulfonate, naphthalenesulfonate (e.g. 2- naphthalenesulfonate), methanesulphonic, ethanesulphonic, p-toluenesulphonic, isethionate or hexanoate salt. In one embodiment there is provided the trifluoroacetic acid salts of the compounds of the invention.
A pharmaceutically acceptable base addition salt can be formed by reaction of a compound of formula (I) with a suitable inorganic or organic base (e.g. ammonia, triethylamine, ethanolamine, triethanolamine, choline, arginine, lysine or histidine), optionally in a suitable solvent such as an organic solvent, to give the base addition salt which is usually isolated for example by crystallisation and filtration. Pharmaceutically acceptable base salts include ammonium salts and salts with organic bases, including
salts of primary, secondary and tertiary amines, including aliphatic amines, aromatic amines, aliphatic diamines, and hydroxy alkylamines, such as methylamine, ethylamine, isopropylamine, diethylamine, ethylenediamine, ethanolamine, trimethylamine, dicyclohexyl amine, diethanolamine, cyclohexylamine and N-methyl-D-glucamine. Other suitable pharmaceutically acceptable base salts include pharmaceutically acceptable metal salts, for example pharmaceutically acceptable alkali-metal or alkaline-earth-metal salts such as hydroxides, carbonates and bicarbonates of sodium, potassium, lithium, calcium, magnesium, aluminium, and zinc; in particular pharmaceutically acceptable metal salts of one or more carboxylic acid moieties that may be present in the compound of Formula I.
Other non-pharmaceutically acceptable salts, for example oxalates may be used, for example in the isolation of compounds of the invention, and are included within the scope of this invention.
The invention includes within its scope all possible stoichiometric and non-stoichiometric forms of the salts of the compounds of Formula (I).
As used herein, the term "compounds of the invention" means both the compounds according to Formula I and salts and solvates thereof. The term "a compound of the invention" also appears herein and refers to both a compound according to Formula I and its salts and solvates.
The compounds of the invention may exist as solids or liquids, both of which are included in the invention. In the solid state, the compounds of the invention may exist as either amorphous material or in crystalline form, or as a mixture thereof. It will be appreciated that solvates of the compounds of the invention may be formed wherein solvent molecules are incorporated into the crystalline lattice during crystallisation. Solvates may involve non-aqueous solvents such as ethanol, isopropanol, DMSO, acetic acid, ethanolamine, and ethyl acetate, or they may involve water as the solvent that is incorporated into the crystalline lattice. Solvates wherein water is the solvent that is incorporated into the crystalline lattice are typically referred to as "hydrates." The invention includes all such solvates.
It will be further appreciated that all crystalline forms, polymorphs, geometric isomers, stereoisomers (including enantiomers and diastereomers) and tautomers of the compounds of the invention, or mixtures thereof, are contemplated to be within the scope of the present invention.
According to another aspect of the invention there is provided at least one chemical entity selected from a compound of Formula I or a pharmaceutically acceptable salt or solvate thereof for use in human or veterinary medical therapy.
The compounds of the invention are cysteine protease inhibitors, such as inhibitors of cysteine proteases of the papain superfamily, for example of the falcipain family, including falcipain-2 or falcipain-3. The compounds of the invention are also inhibitors of cysteine proteases of the papain superfamily, for example those of the cathepsin family such as cathepsins K, L, S and B, particularly cathepsin K.
The compounds of the invention may be useful for treating conditions in which cysteine proteases are implicated, including infections by Plasmodium falciparum which is the most virulent malaria-causing parasite, and by Plasmodium vivax, Pneumocystis carinii, Trypsanoma cruzi, Trypsanoma brucei, and Crithidia fusiculata; as well as in treating conditions such as schistosomiasis, tumor metastasis, metachromatic leukodystrophy, muscular dystrophy, amytrophy, chronic obstructive pulmonary disorder (COPD), atherosclerosis; and especially conditions in which cathepsin K is implicated, including diseases of excessive bone or cartilage loss and other bone and joint diseases such as osteoporosis, bone metastasis, gingival disease (including gingivitis and periodontitis), arthritis (including osteoarthritis and rheumatoid arthritis), Paget's disease; hypercalcemia of malignancy, and metabolic bone disease. In addition, metastatic neoplastic cells also typically express high levels of proteolytic enzymes that degrade the surrounding matrix, and certain tumors and metastatic neoplasias may be effectively treated with the compounds of the invention. Accordingly, the invention is directed to methods of treating such conditions.
In one aspect of the invention, there is provided at least one chemical entity selected from a compound of Formula I or a pharmaceutically acceptable salt or solvate thereof, for use in the treatment of a condition mediated by inhibition of a cysteine protease, particularly inhibition of a cysteine protease of the papain superfamily such as those of the falcipain family, including falcipain-2 or falcipain-3, for example malaria.
In another aspect of the invention, there is provided at least one chemical entity selected from a compound of Formula I or a pharmaceutically acceptable salt or solvate thereof for use in the treatment of a condition mediated by inhibition of a cysteine protease, particularly inhibition of a cysteine protease of the papain superfamily, such as those of the cathepsin family for example cathepsins K, L, S and B, particularly cathepsin K, for example conditions characterised by excessive bone loss such as osteoporosis and bone metastasis, and other bone and joint diseases such as osteoarthritis.
In another aspect of the invention there is provided the use of at least one chemical entity selected from a compound of Formula I or a pharmaceutically acceptable salt or solvate thereof in the manufacture of a medicament for the treatment of a condition mediated by inhibition of a cysteine protease, particularly inhibition of a cysteine protease of the papain superfamily such as those of the falcipain family, including falcipain-2 or falcipain-3, for example malaria.
In a further aspect of the invention there is provided the use of at least one chemical entity selected from a compound of Formula I or a pharmaceutically acceptable salt or solvate thereof in the manufacture of a medicament for the treatment of a condition mediated by inhibition of a cysteine protease, particularly inhibition of a cysteine protease of the papain superfamily, such as those of the cathepsin family for example cathepsins K, L, S and B, particularly cathepsin K1 for example conditions characterised by excessive bone loss such as osteoporosis and bone metastasis, and other bone and joint diseases such as osteoarthritis.
In another aspect of the invention there is provided a method for the treatment of a human or animal subject suffering from a condition mediated by inhibition of a cysteine protease, particularly inhibition of a cysteine protease of the papain superfamily such as those of the falcipain family, including falcipain-2 or falcipain-3, for example malaria, which method comprises administering an effective amount of at least one chemical entity selected from a compound of Formula I or a pharmaceutically acceptable salt or solvate thereof or a pharmaceutical composition comprising at least one chemical entity selected from a compound of Formula I or a pharmaceutically acceptable salt or solvate thereof.
In another aspect of the invention there is provided a method for the treatment of a human or animal subject suffering from a condition mediated by inhibition of a cysteine protease, particularly inhibition of a cysteine protease of the papain superfamily, such as those of the cathepsin family for example cathepsins K, L, S and B, particularly cathepsin K, for example conditions characterised by excessive bone loss such as osteoporosis and bone metastasis, and other bone and joint diseases such as osteoarthritis, which method comprises administering an effective amount of at least one chemical entity selected from a compound of Formula I or a pharmaceutically acceptable salt or solvate thereof or a pharmaceutical composition comprising at least one chemical entity selected from a compound of Formula I or a pharmaceutically acceptable salt or solvate thereof.
The compounds of the invention are cysteine protease inhibitors and can be useful in the treatment of a condition mediated by inhibition of a cysteine protease, particularly inhibition of a cysteine protease of the papain superfamily such as those of the falcipain family, including falcipain-2 or falcipain-3, for example in the treatment of malaria, or those of the cathepsin family for example cathepsins K, L, S and B, particularly cathepsin K, for
example in the treatment of conditions characterised by excessive bone loss such as osteoporosis and bone metastasis, and other bone and joint diseases such as osteoarthritis. Accordingly, the invention is further directed to pharmaceutical compositions comprising at least one chemical entity selected from a compound of Formula I or a pharmaceutically acceptable salt or solvate thereof.
As used herein "excessive bone loss" is a disease state in which the normal balance between bone resorption and formation is disrupted, and there is a net loss of bone at each cycle. Diseases which are characterised by excessive bone loss include, but are not limited to, osteoporosis and gingival diseases, excessive cartilage or matrix degradation including osteoarthritis and rheumatoid arthritis.
The methods of treatment of the invention comprise administering a safe and effective amount of at least one chemical entity selected from a compound of Formula I or a pharmaceutically acceptable salt or solvate thereof, or a pharmaceutical composition containing at least one chemical entity selected from a compound of Formula I or a pharmaceutically acceptable salt or solvate thereof, to a patient in need thereof.
As used herein, "treatment" means: (1) the amelioration or prevention of the condition being treated or one or more of the biological manifestations of the condition being treated, (2) the interference with (a) one or more points in the biological cascade that leads to or is responsible for the condition being treated or (b) one or more of the biological manifestations of the condition being treated, or (3) the alleviation of one or more of the symptoms or effects associated with the condition being treated. The skilled artisan will appreciate that "prevention" is not an absolute term. In medicine, "prevention" is understood to refer to the prophylactic administration of a drug to substantially diminish the likelihood or severity of a condition or biological manifestation thereof, or to delay the onset of such condition or biological manifestation thereof.
As used herein, "safe and effective amount" means an amount of the compound sufficient to significantly induce a positive modification in the condition to be treated but low enough to avoid serious side effects (at a reasonable benefit/risk ratio) within the scope of sound medical judgment. A safe and effective amount of a compound of the invention will vary with the particular compound chosen (e.g. depending on the potency, efficacy, and half- life of the compound); the route of administration chosen; the condition being treated; the severity of the condition being treated; the age, size, weight, and physical condition of the patient being treated; the medical history of the patient to be treated; the duration of the treatment; the nature of concurrent therapy; the desired therapeutic effect; and like factors, but can nevertheless be routinely determined by the skilled artisan.
As used herein, "patient" refers to a human or other animal.
The compounds of the invention may be administered by any suitable route of administration, including both systemic administration and topical administration. Systemic administration includes oral administration, parenteral administration, transdermal administration, rectal administration, and administration by inhalation. Parenteral administration refers to routes of administration other than enteral, transdermal, or by inhalation, and is typically by injection or infusion. Parenteral administration includes intravenous, intramuscular, and subcutaneous injection or infusion. Inhalation refers to administration into the patient's lungs whether inhaled through the mouth or through the nasal passages. Topical administration includes application to the skin as well as intraocular, optic, intravaginal, and intranasal administration.
The compounds of the invention may be administered once or according to a dosing regimen wherein a number of doses are administered at varying intervals of time for a given period of time. For example, doses may be administered one, two, three, or four times per day. Doses may be administered until the desired therapeutic effect is achieved or indefinitely to maintain the desired therapeutic effect. Suitable dosing regimens for a compound of the invention depend on the pharmacokinetic properties of that compound, such as absorption, distribution, and half-life, which can be determined by the skilled artisan. In addition, suitable dosing regimens, including the duration such regimens are administered, for a compound of the invention depend on the condition being treated, the severity of the condition being treated, the age and physical condition of the patient being treated, the medical history of the patient to be treated, the nature of concurrent therapy, the desired therapeutic effect, and like factors within the knowledge and expertise of the skilled artisan. It will be further understood by such skilled artisans that suitable dosing regimens may require adjustment given an individual patient's response to the dosing regimen or over time as individual patient needs change.
Typical daily dosages may vary depending upon the particular route of administration chosen. Typical daily dosages for oral administration range from about 0.01 to about 25 mg/kg, in one embodiment from about 0.1 to about 14 mg/kg. Typical daily dosages for parenteral administration range from about 0.001 to about 10 mg/kg; in one embodiment from about 0.01 to about 6 mg/kg The compounds of Formula I may also be used in combination with other therapeutic agents. The invention thus provides, in a further aspect, a combination comprising a compound of Formula I or a pharmaceutically acceptable derivative thereof together with a further therapeutic agent. When a compound of Formula I or a pharmaceutically acceptable derivative thereof is used in combination with a second therapeutic agent active against the same disease state the dose of each compound may differ from that when the compound is used alone. Appropriate doses will be readily appreciated by those skilled in the art. It will be
appreciated that the amount of a compound of the invention required for use in treatment will vary with the nature of the condition being treated and the age and the condition of the patient and will be ultimately at the discretion of the attendant physician or veterinarian.
The compounds of the present invention may be used alone or in combination with one or more additional active agents, such as other inhibitors of cysteine and serine proteases, antimalarial drugs or drugs to treat excessive bone loss.
Such other active agents include inhibitors of bone resorption or other bone diseases, for example bisphosphonates (i.e., alendronate, risedronate, etidronate, and ibandronate), hormone replacement therapy, anti-estrogens, calcitonin, and anabolic agents such as bone morphogenic protein, iproflavone, and PTH. In the alternative, such other active agents include antimalarial drugs, such as folates (e.g. chloroquine, mefloquine, primaquine pyrimethamine, quinine artemisinin, halofantrine, doxycycline, amodiquine, atovaquine [atovaquone], tafenoquine) and antifolates (e.g. dapsone, proguanil, sulfadoxine, pyrimethamine, chlorcycloguanil, cycloguanil) or antibacterial drugs such as azithromycin, doxycycline, ciprofloxacin and clindamycin.
The combinations referred to above may conveniently be presented for use in the form of a pharmaceutical formulation and thus pharmaceutical formulations comprising a combination as defined above together with a pharmaceutically acceptable carrier or excipient comprise a further aspect of the invention. The individual components of such combinations may be administered either sequentially or simultaneously in separate or combined pharmaceutical formulations by any convenient route.
When administration is sequential, either the compound of the present invention or the second therapeutic agent may be administered first. When administration is simultaneous, the combination may be administered either in the same or different pharmaceutical composition. When combined in the same formulation it will be appreciated that the two compounds must be stable and compatible with each other and the other components of the formulation. When formulated separately they may be provided in any convenient formulation, conveniently in such manner as are known for such compounds in the art.
Compositions The compounds of the invention will normally, but not necessarily, be formulated into pharmaceutical compositions prior to administration to a patient. Accordingly, in another aspect the invention is directed to pharmaceutical compositions comprising a compound of the invention and a pharmaceutically acceptable carrier and/or excipient. The carrier and/or excipient must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not deleterious to the recipient thereof.
The pharmaceutical compositions of the invention may be prepared and packaged in bulk form wherein a safe and effective amount of a compound of the invention can be extracted and then given to the patient such as with powders or syrups. Alternatively, the pharmaceutical compositions of the invention may be prepared and packaged in unit dosage form wherein each physically discrete unit contains a safe and effective amount of a compound of the invention. When prepared in unit dosage form, the pharmaceutical compositions of the invention typically contain from about 0.5 mg to about 1750 mg, e.g. from about 5 mg to about 1000 mg for oral dosage forms and from about 0.05 mg to about 700 mg, e.g. from about 0.5 mg to about 500 mg for parenteral dosage forms.
The pharmaceutical compositions of the invention typically contain one compound of the invention. However, in certain embodiments, the pharmaceutical compositions of the invention contain more than one compound of the invention. For example, in certain embodiments the pharmaceutical compositions of the invention contain two compounds of the invention. In addition, the pharmaceutical compositions of the invention may optionally further comprise one or more additional pharmaceutically active compounds. Conversely, the pharmaceutical compositions of the invention typically contain more than one pharmaceutically acceptable excipient. However, in certain embodiments, the pharmaceutical compositions of the invention contain one pharmaceutically acceptable excipient.
The compound of the invention and the pharmaceutically acceptable excipient or excipients will typically be formulated into a dosage form adapted for administration to the patient by the desired route of administration. For example, dosage forms include those adapted for (1) oral administration such as tablets, capsules, caplets, pills, troches, powders, syrups, elixers, suspensions, solutions, emulsions, sachets, and cachets; (2) parenteral administration such as sterile solutions, suspensions, and powders for reconstitution; (3) transdermal administration such as transdermal patches; (4) rectal administration such as suppositories; (5) inhalation such as aerosols and solutions; and (6) topical administration such as creams, ointments, lotions, solutions, pastes, sprays, foams, and gels.
Suitable pharmaceutically acceptable excipients will vary depending upon the particular dosage form chosen. In addition, suitable pharmaceutically acceptable excipients may be chosen for a particular function that they may serve in the composition. For example, certain pharmaceutically acceptable excipients may be chosen for their ability to facilitate the production of uniform dosage forms. Certain pharmaceutically acceptable excipients may be chosen for their ability to facilitate the production of stable dosage forms. Certain pharmaceutically acceptable excipients may be chosen for their ability to facilitate the carrying or transporting the compound or compounds of the invention once administered to the patient from one organ, or portion of the body, to another organ, or portion of the
body. Certain pharmaceutically acceptable excipients may be chosen for their ability to enhance patient compliance.
Suitable pharmaceutically acceptable excipients include the following types of excipients: Diluents, fillers, binders, disintegrants, lubricants, glidants, granulating agents, coating agents, wetting agents, solvents, co-solvents, suspending agents, emulsifiers, sweeteners, flavoring agents, flavor masking agents, coloring agents, anticaking agents, hemectants, chelating agents, plasticizers, viscosity increasing agents, antioxidants, preservatives, stabilizers, surfactants, and buffering agents. The skilled artisan will appreciate that certain pharmaceutically acceptable excipients may serve more than one function and may serve alternative functions depending on how much of the excipient is present in the formulation and what other ingredients are present in the formulation.
Skilled artisans possess the knowledge and skill in the art to enable them to select suitable pharmaceutically acceptable excipients in appropriate amounts for use in the invention. In addition, there are a number of resources that are available to the skilled artisan which describe pharmaceutically acceptable excipients and may be useful in selecting suitable pharmaceutically acceptable excipients. Examples include Remington's
Pharmaceutical Sciences (Mack Publishing Company), The Handbook of Pharmaceutical Additives (Gower Publishing Limited), and The Handbook of Pharmaceutical Excipients
(the American Pharmaceutical Association and the Pharmaceutical Press).
The pharmaceutical compositions of the invention are prepared using techniques and methods known to those skilled in the art. Some of the methods commonly used in the art are described in Remington's Pharmaceutical Sciences (Mack Publishing Company).
In one aspect, the invention is directed to a solid oral dosage form such as a tablet or capsule comprising a safe and effective amount of a compound of the invention and a diluent or filler. Suitable diluents and fillers include lactose, sucrose, dextrose, mannitol, sorbitol, starch (e.g. corn starch, potato starch, and pre-gelatinized starch), cellulose and its derivatives (e.g. microcrystalline cellulose), calcium sulfate, and dibasic calcium phosphate. The oral solid dosage form may further comprise a binder. Suitable binders include starch (e.g. corn starch, potato starch, and pre-gelatinized starch), gelatin, acacia, sodium alginate, alginic acid, tragacanth, guar gum, povidone, and cellulose and its derivatives (e.g. microcrystalline cellulose). The oral solid dosage form may further comprise a disintegrant. Suitable disintegrants include crospovidone, sodium starch glycolate, croscarmelose, alginic acid, and sodium carboxymethyl cellulose. The oral solid dosage form may further comprise a lubricant. Suitable lubricants include stearic acid, magnesium stearate, calcium stearate, and talc.
There is further provided by the present invention a process of preparing a pharmaceutical composition, which process comprises mixing at least one compound of formula (I) or a pharmaceutically acceptable derivative thereof, together with a pharmaceutically acceptable carrier and/or excipient.
All publications, including but not limited to patents and patent applications cited in this specification are herein incorporated by reference as if each individual publication were specifically and individually indicated to be incorporated by reference as though fully set forth.
Abbreviations
In describing the invention, chemical elements are identified in accordance with the Periodic Table of the Elements. Abbreviations and symbols utilized herein are in accordance with the common usage of such abbreviations and symbols by those skilled in the chemical arts. The following abbreviations are used herein:
ACN acetonitrile
AcOEt ethyl acetate
AFC 7-amido-4-trifluoromethylcoumarin
AMC 7-amido-4-methylcoumarin
CDCI3 deuterated chloroform
CHAPS 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate
CYS cysteine
DABCO 1 ,4-diazabicyclo[2.2.2]octane
DCM dichloromethane
DIPEA diisopropylamine
DMAP 4-dimethylamino pyridine
DMSO-d6 deuterated dimethylsulfoxide
DMSO dimethylsulfoxide
DTT dithiothreitol
E64 fraπs-epoxysuccinyl-L-leucylamido(4-guanidino)butane
EDCI N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide
EDTA (ethylenedinitrilo)tetraacetic acid
ES+ MS Positive Electrospray mass spectrometry
ES- MS • Negative Electrospray mass spectrometry
EtOH ethanol h hours
H-D-VLR-AFC HD-Valyl-Leucyl-Arginyl-7-Amido-4-trifluoromethylcoumarin
Hex hexane
HOBt 1 -hydroxybenzotriazole
HPLC high pressure liquid chromatography i-PrOH isopropanol kg kilogram(s)
KQKLR-AMC N-Acetyl-Lysyl-Glutaminyl-Lysyl-Leucyl-Arginyl-y-Amido^- methylcoumarin
MeOH methanol
MES 2-(N-morpholino)ethanesulfonic acid
MgSO4 magnesium sulfate min minutes mg miligram(s)
NaHCO3 sodium bicarbonate
Na2SO4 sodium sulfate
NMR Nuclear Magnetic Resonance spectroscopy
PtO2 platinum oxide
TEA triethylamine
TFA trifluoroacetic acid
THF tetrahydrofuran
Z-LR-AMC benzyloxycarbonyl-leucyl-arginyl-Z-amido^-methylcoumarin
Compound Preparation
The general procedures used to synthesise the compounds of Formula I are described in reaction Schemes 1-18 and are illustrated in the Examples.
I
Throughout the specification, general formulae are designated by Roman numerals I, II, III, IV etc. Subsets of compounds of Formula I are defined as Ia, Ib, lb(i), Ib(ii), Ic, lc(i), Id, Ie and If.
The semicarbazide compounds of Formula Ia, which are compounds of Formula I wherein R1 is C1-8alkyl, -C1-8alkyleneN(C1-3alkyl)2 -C1-8alkyleneNRGC(O)OCi-6alkyl or -C1.8alkyleneNRGC(O)C1-6alkyl, A is C(O), X is NR5 and R5 is hydrogen, C1-6alkyl, -C1-
8alkyleneN(C1-3alkyl)2, -Ci-8alkyleneNRGC(O)OC1-6alkyl or -C1-8alkyleneNRGC(O)C1-6alkyl, or -C(O)NRHR10 in which RH is C^alkyl, -C1-8alkyleneN(C1-3alkyl)2, -C1-8alkyleneNRGC(O)OC1-6alkyl or -C1-8alkyleneNRGC(O)C1-6alkyl and R10 is aryl, heteroaryl, aryl-heteroaryl, cycloalkyl or -C1-6alkyleneRc, wherein Rc is hydrogen, C1-3alkyl, aryl or halogen, may be prepared from the corresponding hydrazine compounds of Formula II, wherein R1 is C1-8alkyl, -C1-8alkyleneN(C1-3alkyl)2 -C1-8alkyleneNRGC(O)OC1-6alkyl or -C1-8alkyleneNRGC(O)C1-6alkyl and R5 is hydrogen, C1-
6alkyl, -C
1-8alkyleneN(C
1-3alkyl)
2, -C
1.
8alkyleneNR
GC(O)OC
1-6alkyl or
according to Scheme 1. This transformation may be carried out following one of two different procedures, procedure A or procedure B.
Procedure A: Compounds of Formula Ha, which are compounds of Formula II, wherein R5 is hydrogen, are reacted with one equivalent of the isocyanate, R10NCO, wherein R10 is as defined above for Formula Ia, in the presence of a suitable base such as triethylamine in a suitable solvent such as DCM to give compounds of Formula Ia wherein R5 is hydrogen and RH is hydrogen. Under these conditions, when an excess of the isocyanate R10NCO is added, compounds of Formula Ia wherein R5 is C(O)NRHR10 (N,N-bis aminocarbonyl compounds) are obtained.
Procedure B: A primary amine R10-NH2l or a secondary amine R10-NH-RH, wherein R10 and RH are as defined as above for Formula Ia, is dissolved in a suitable solvent such as dry THF and cooled to a suitable temperature, e.g. -100C to 100C, then reacted with triphosgene, and the resulting mixture is added to compounds of Formula Il in the presence of a suitable base such as triethylamine. This mixture may be stirred at a suitable temperature for a suitable length of time for complete reaction, for example at room temperature for 6 h, to give compounds of Formula Ia wherein R5, R10 and RH are as defined above for Formula Ia.
+triphosgene
Ia
Scheme 1
The acylhydrazide compounds of Formula Ib, which are compounds of Formula I wherein R1 is C1-8alkyl, -C1-8alkyleneN(C1-3alkyl)2 -C1-8alkyleneNRGC(O)OC1-6alkyl or -C1-8alkyleneNRGC(O)Ci-6alkyl, A is C(O), X is NR5 and R5 is hydrogen C1-8alkyl, -C1- 8alkyleneN(C1-3alkyl)2 -C1-8alkyleneNRGC(O)OCi-6alkyl or -C1-8alkyleneNRGC(O)Ci-6alkyl, and R2 is aryl, heteroaryl, cycloalkyl, heterocyclyl, -aryl-heterocyclyl, biaryl, aryl-heteroaryl, -aryl-Ci-salkylene-heterocyclyl, -aryl-O-d-salkylene-heterocyclyl or C1-6alkyleneRA, wherein RA is hydrogen, C1-3alkyl, halogen, -N(Ci-3alkyl)2, aryl, biaryl, cycloalkyl, -aryl-C-i^alkylene-heterocyclyl or -aryl-O-C^alkylene-heterocyclyl, may be prepared from the corresponding hydrazine compounds of Formula II, wherein R1 is C1-8alkyl, -C1- 8alkyleneN(C1-3alkyl)2 -C1-8alkyleneNRGC(O)OC1-6alkyl or -C1-8alkyleneNRGC(O)C1-6alkyl and R5 is hydrogen, C1-8alkyl, -C1-8alkyleneN(C1-3alkyl)2 -C1-8alkyleneNRGC(O)OC1-6alkyl or -C1-8alkyleneNRGC(O)C1-6alkyl, according to Scheme 2. Compounds of Formula Il are reacted with an acid chloride R2COCI, wherein R2 is as defined above for Formula Ib, in a suitable solvent such as pyridine to give compounds of Formula Ib.
Il Ib
Scheme 2
The alkoxycarbonyl hydrazine compounds of Formula Ic, which are compounds of Formula I wherein R1 is C1-8alkyl, -C1-8alkyleneN(C1-3alkyl)2 -C1-8alkyleneNRGC(O)OC1-6alkyl or -C1-8alkyleneNRGC(O)C1-6alkyl, A is C(O), X is NR5 and R5 is hydrogen, C1-8alkyl, -C1-8alkyleneN(C1-3alkyl)2 -C1-8alkyleneNRGC(O)OC1-6alkyl or -C1-8alkyleneNRGC(O)C1-6alkyl and R2 is OR11 in which R11 is aryl, cycloalkyl, C1-6alkenyl, biaryl, heteroaryl, -aryl-heteroaryl, or -C1-6alkyleneRD, wherein RD is hydrogen, C1-3alkyl, aryl, heteroaryl, heterocyclyl, cycloalkyl, heterocyclyl, CCI3, cyano, -NHC(O)C1-6alkyl, -NHC(O)OC1-6alkyl, or -C(O)C1-6alkyl, may be prepared from the corresponding hydrazine compounds of Formula II, wherein R5 is hydrogen, C1-8alkyl, -C1-8alkyleneN(Ci-3alkyl)2 - C1-8alkyleneNRGC(O)OC1-6alkyl or -C1-8alkyleneNRGC(O)C1-6alkyl and R1 is C1-8alkyl, -C1- 8alkyleneN(C1-3alkyl)2 -C1-8alkyleneNRGC(O)OC1-6alkyl or -C1-8alkyleneNRGC(O)C1-6alkyl, according to Scheme 3. Compounds of Formula Il are reacted with a chloroformate R11OCOCI, wherein R11 is as defined above for Formula Ic, in a suitable solvent such as DCM in the presence of a suitable base such as a mixture of diisopropylethylamine and DMAP to give compounds of Formula Ic. Chloroformates R11OCOCI are either
commercially available, or they may be obtained by reaction between the corresponding commercially available alcohol R11OH, wherein R11 is as defined above for Formula Ic, and triphosgene in a suitable solvent such as THF, which may be directly reacted with compounds of Formula II in the presence of a suitable base such as triethylamine in a suitable solvent, for example pyridine, to give compounds of Formula Ic.
Il Ic
Scheme 3
Compounds of Formula Ha which are compounds of Formula Il wherein R5 is hydrogen, may be prepared from compounds of Formula III, wherein R1 is wherein R1 is C1-8alkyl, -C1-8alkyleneN(Ci-3alkyl)2 -C1.8alkyleneNRGC(O)OCi-6alkyl or
-C1-8alkyleneNRGC(O)C1-6alkyl, according to Scheme 4 by deprotection in the presence of a suitable acid such as trifluoroacetic acid.
acidic deprotection
Scheme 4 Compounds of Formula III may be prepared from compounds of Formula IV, wherein R1 is
C1-8alkyl, -C1-8alkyleneN(C1-3alkyl)2 -C1-8alkyleneNRGC(O)OC1.6alkyl or
-C1-8alkyleneNRGC(O)C1-6alkyl, according to Scheme 5 by cyanation, by displacement of the chloro substituent of compounds of Formula IV using a variety of conditions, for example by treatment with potassium or sodium cyanide in the presence of a suitable base such as DABCO in a suitable solvent such as DMSO.
cyanation
Scheme 5 Compounds of Formula IV may be prepared from compounds of Formula V, wherein R
1 is
C1-8alkyl, -C1-8alkyleneN(C1-3alkyl)2 -C1-8alkyleneNRGC(O)OC1-6alkyl or
-C1-8alkyleneNRGC(O)Ci-6alkyl, according to Scheme 6 by reaction of compounds of Formula V with 2,4-dichloropyrimidine, a compound of Formula Vl1 (commercially available from FLUKA or SIGMA) in a suitable solvent such as EtOH, for example at room temperature for 3-4 days, for example according to the literature procedure given in Bagley J. R. et al., (1989) J. Med. Chem. 32, 663-671.
Scheme 6
Compounds of Formula V may be prepared from the compound of Formula VII by a reductive amination reaction with an aldehyde VIII, wherein R13 is one carbon shorter in chain length than R1, wherein R1 is C1-aalkyl, -Ci.8alkyleneN(C1-3alkyl)2 - C1-8alkyleneNRGC(O)OC1-6alkyl or -C1-8alkyleneNRGC(O)C1-6alkyl, according to Scheme 7. The compound of Formula VII, fe/f-butyl carbazate, is commercially available (ALDRICH). Reductive amination of the compound of Formula VII with aldehydes of Formula VIII is carried out in the presence of a suitable reducing agent such as hydrogen, and a suitable catalyst such as platinum or palladium or platinum oxide, in a suitable solvent such as i- PrOH, EtOH or a mixture thereof, for example according to the literature procedures given in Hilpert, H. (2001) Tetrahedron, 57, 7675-7683 or Dyker, H. et al, (2001) J. Org. Chem. 66, 3760-3766).
Scheme 7
Compounds of Formula Id, which are compounds of Formula I wherein R1 is C1-8alkyl, C1-
8alkyl, -C1-8alkyleneN(C1-3alkyl)2 -Ci-8alkyleneNRGC(O)OC1-6alkyl or
-C1-8alkyleneNRGC(O)C1-6alkyl, A is C(O), X is CH2 and R2 is -NRHR14, wherein R14 is aryl, heteroaryl, -C1-6aralkyl, -aryl-heterocyclyl, -aryl-heteroaryl, Ci-6alkylene-heteroaryl, and RH is hydrogen, C1-6alkyl, -C1-8alkyleneN(Ci-3alkyl)2 -C1-8alkyleneNRGC(O)OC1-6alkyl or -C1-8alkyleneNRGC(O)Ci-6alkyl, may be prepared from the corresponding compound of Formula IX, wherein R1 is as defined for Formula Id, according to Scheme 8. Compounds of Formula IX are treated with an amine R14-NH-RH, wherein R14 and RH are as defined above for Formula Id, in the presence of a suitable reagent such as HOBt and EDCI in a suitable solvent such as DCM to give compounds of Formula Id.
Scheme 8
Compounds of Formula IX may be prepared from compounds of Formula X, wherein R1 is as defined for Formula Id above, and R12 is C1-4alkyl, according to Scheme 9. The ester group of compounds of Formula X may be hydrolysed under basic conditions, for example by treatment with lithium hydroxide, in a suitable solvent such as THF, to give compounds of Formula IX.
Scheme 9
Compounds of Formula X may be prepared from compounds of Formula Xl, wherein R1 is as defined for Formula Id above and R12 is C1-4alkyl according to Scheme 10, by cyanation, by displacement of the chloro substituent of compounds of Formula Xl, for example by treatment with potassium or sodium cyanide in the presence of a suitable base such as DABCO in a suitable solvent such as DMSO.
cyanation
Xl X
Scheme 10
Compounds of Formula Xl may be prepared from compounds of Formula XII, wherein R1 is as defined for Formula Id above and R12 is C1-4alkyl, according to Scheme 11 , by reaction of compounds of Formula XII with 2,4-dichloropyrimidine XIII (commercially available from FLUKA or SIGMA) in a suitable solvent such as i-PrOH, for example under microwave conditions and heating to a suitable temperature, e.g. 110°C-150°C for 20 min, for example according to the literature procedure given in Guanglin L. et al., (2002) Tetrahedron Lett. 43, 5739-5742.
Scheme 11
Compounds of Formula XII may be prepared according to Scheme 12 by procedure A or B:
Procedure A: compounds of Formula XIV, wherein R12 is C1-4alkyl, may be reacted with a primary amine R1-NH2, wherein R1 is as defined for Formula I above, in the presence of a suitable base such as a mixture of isobutylamine and triethylamine, and in a suitable solvent such as THF, for example according to the literature procedure given in Guerrini R. et al., (2000) J. Med. Chem. 43, 2805-2813. Compounds of Formula XIV are commercially available.
Procedure B:
Compounds of Formula XII may be prepared from compounds of Formula XVIII wherein
R12 is C1-4alkyl, which are commercially available, by reductive amination with aldehydes
of Formula VIII, wherein R13 is one carbon shorter in chain length than R1. The reaction may be carried out using a suitable reducing agent such as hydrogen, in the presence of a suitable catalyst such as platinum or palladium or platinum oxide, in a suitable solvent such as i-PrOH, EtOH.
XIV XIl XVIII
Scheme 12
Aldehydes of Formula VIII are either commercially available, or they may be prepared according to Scheme 13 i) from the corresponding commercially available dimethyl or diethyl acetal compound of Formula XV wherein R13 is as defined above for compounds of
Formula VIII, by acid hydrolysis using a suitable acid such as hydrochloric acid, or ii) by oxidation of the commercially available alcohol compound of Formula XVI, wherein R13 is as defined above for compounds of Formula VIII, following standard procedures as the Swern oxidation or Dess-Martin oxidation.
C1 2alkyl — O acid 13
\ is hydrolysis /T~R Oxidation 13
/ *- O VIII /
C^alkyl — O H0 xv XV,
Scheme 13 Compounds of Formula lb(i), which are compounds of Formula I which are compounds of Formula I wherein R
1 is C
1-8alkyleneNH
2, A is C(O), X is NR
5 and R
5 is hydrogen or C
1- 6alkyl and R
2 is aryl, heteroaryl, cycloalkyl, heterocyclyl, -aryl-heterocyclyl, biaryl, aryl- heteroaryl, -aryl-C^alkylene-heterocyclyl, -aryl-O-C^alkylene-heterocyclyl or C
1- 6alkyleneR
A, wherein R
A is hydrogen, C
1-3alkyl, halogen, -N(C
1-3alkyl)
2, aryl, biaryl, cycloalkyl, -aryl-C-i-aalkylene-heterocyclyl or -aryl-O-C-i^alkylene-heterocyclyl, may be prepared from a compound of Formula Ib wherein R
5 and R
2 are as defined for Formula lb(i), by a deprotection reaction in the presence of a suitable acid such as trifluoroacetic acid, in a suitable solvent such as dichloromethane, or alternatively hydrobromic acid in a suitable solvent such as acetic acid according to Scheme 14.
Ib lb(i)
Scheme 14
Compounds of Formula Ib(ii) which are compounds of Formula I wherein R1 is -C1-8alkyleneNC(O)C1-6alkyl, A is C(O), X is NR5 and R5 is hydrogen or C1-6alkyl and R2 is aryl, heteroaryl, cycloalkyl, heterocyclyl, -aryl-heterocyclyl, biaryl, aryl-heteroaryl, -aryl-CLsalkylene-heterocyclyl, -aryl-O-C1-3alkylene-heterocyclyl or C1-6alkyleneRA, wherein RA is hydrogen, Ci-3alkyl, halogen, -N(C1-3alkyl)2, aryl, biaryl, cycloalkyl, -aryl-C^alkylene-heterocyclyl or -aryl-O-C-^alkylene-heterocyclyl, may be prepared from compounds of Formula lb(i) as defined above, according to Scheme 15, by treatment of lb(i) with an anhydride of Formula O[C(O)Ci-6alkyl]2 in a suitable solvent such as dichloromethane, at a suitable temperature, e.g. -100C to 1O0C.
Scheme 15
Compounds of Formula lc(i), which are compounds of Formula I wherein R
1 is C
1-8alkyl, A is C(O), X is NR
5 and R
5 is -C
1-8alkyleneN(C
1-3alkyl)2, -d-βalkylene-heterocyclyl, -C(O)C
1- 6alkyl, -Ci
-8alkyleneNR
GC(O)OC
1-6alkyl or -C
1-8alkyleneNR
GC(O)C
1-6alkyl wherein R
G is as defined for Formula I, or N-phthalidimido-CI-δalkylene- and R
2 is OR
11 in which R
11 is aryl, cycloalkyl, C
1-6alkenyl, biaryl, heteroaryl, -aryl-heteroaryl, or -C
1-6alkyleneR
D, wherein R
D is hydrogen, C
1-3alkyl, aryl, heteroaryl, heterocyclyl, cycloalkyl, heterocyclyl, CCI
3, cyano, -NHC(O)C
1-6alkyl, -NHC(O)OC
1-6alkyl, or -C(O)C
1-6alkyl, may be prepared from compounds of Formula Ic wherein R
1 is C
1-8alkyl, A is C(O), X is NR
5 and R
5 is hydrogen and R
2 is OR
11 in which R
11 is as defined for lc(i), according to Scheme 16 by treatment of Ic with an alkylating agent of Formula R
5-CI, R
5-Br or R
5-OSO
2Y, wherein R
5 is as defined for Formula lc(i) and Y is methyl or p-tolyl, in the presence of a suitable catalyst such as tetrabutylammonium hydrogensulfate and optionally sodium iodide in the presence of a base such as a mixture of potassium carbonate and sodium hydroxide, in a suitable solvent such as toluene, at elevated temperature, e.g. 90-170
0C.
Scheme 16
Compounds of Formula Ie, which are compounds of Formula I wherein R1 is C1-8alkyl, A is C(O), X is NR5 and R5 is hydrogen and R2 is a nitrogen-containing heterocyclyl group which is bonded through the nitrogen, or R2 is NRH-aryl or NRH-heteroaryl (wherein RH represents hydrogen, C1-6alkyl, -C1-6alkyleneNHC(O)C1-4alkyl or
-C1-6alkyleneNHC(O)OC1-4alkyl), or R2 is -NRBC1.6alkyleneRc (wherein RB is C1-8alkyl and Rc is aryl, -aryl-d-salkylene-heterocyclyl, Ci-3alkenylaryl, -NHC(O)OC1-6alkyl, -NHC(O)C1-6alkyl, -NHC(O)OCi.6alkyl, -NHC(O)C1-6aralkyl), may be prepared from compounds of Formula Il wherein R1 is C1-8alkyl and R5 is hydrogen according to Scheme 17, by treatment with triphosgene followed by treatment with an amine R2H, wherein R2 is as defined for Formula Ie, in the presence of a suitable base such as caesium carbonate or triethylamine in a suitable solvent such as THF at an appropriate temperature, e.g. -10°C to 10°C.
triphosgene
I" Ie
Scheme 17
Compounds of Formula If, which are compounds of Formula I wherein R1 is C1-8alkyl, -Ci- 8alkyleneN(C1-3alkyl)2 -C1-8alkyleneNRGC(O)OC1-6alkyl or -C1-8alkyleneNRGC(O)C1-6alkyl, A is -SO2-, R2 is C1-6alkyl or C1-6arakyl, X is NR5 and R5 is hydrogen or C1-6alkyl, C1-6alkenyl, -C(O)R2a, -C1-8alkylene-heterocyclyl, -C1-8alkyleneNRGC(O)C1-6alkyl,
-C
1-8alkyleneNR
GC(O)OC
1-6alkyl, -Ci
-8alkyleneN(C
1-3alkylene), N-phthalidimido-C.,.
8alkylene- and R
2 is Ci
-6alkyl or C
1-6aralkyl, may be prepared from compounds of Formula Il wherein R
1 is C
1-8alkyl, -C
1-8alkyleneN(C
1-3alkyl)
2 -C
1-8alkyleneNR
GC(O)OC
1-6alkyl or -Ci.
8alkyleneNR
GC(O)C
1-6alkyl, and R
5 is hydrogen or C-,.
6alkyl, C
1-6alkenyl, -C(O)R
2a, -Ci- aalkylene-heterocyclyl, -Ci
-8alkyleneNR
GC(O)Ci
-6alkyl, -Ci
-8alkyleneNR
GC(O)OCi
-6alkyl, - Ci
-8alkyleneN(Ci
-3alkylene) or N-phthalidimido-C
Lsalkylene-, according to Scheme 18, by treatment of compounds Il with a sulfonyl chloride R
2SO
2CI, wherein R
2 represents Ci-
6alkyl or Ci
-6aralkyl; in a suitable solvent such as pyridine. Sulfonyl chlorides R
2SO
2CI may be commercially available or they may be prepared from the corresponding sulfonic acids R
2SO
2OH by treatment of the sulfonic acids with thionyl chloride in a suitable solvent such as toluene at elevated temperatures such as 90-170°C.
If
Scheme 18
It will be readily apparent to those skilled in the art that other compounds of Formula I may be prepared using methods analogous to those outlined above, or by reference to the experimental procedures detailed in the Examples provided herein.
Those skilled in the art will also appreciate that in the preparation of the compound of Formula I or a solvate thereof, it may be necessary and/or desirable to protect one or more sensitive groups in the molecule or the appropriate intermediate to prevent undesirable side reactions. Suitable protecting groups for use according to the present invention are well known to those skilled in the art and may be used in a conventional manner. See, for example, "Protective groups in organic synthesis" by T.W. Greene and P.G.M. Wuts (John Wiley & sons 1991) or "Protecting Groups" by PJ. Kocienski (Georg Thieme Verlag 1994). Examples of suitable amino protecting groups include acyl type protecting groups (e.g. formyl, trifluoroacetyl, acetyl), aromatic urethane type protecting groups (e.g. benzyloxycarbonyl (Cbz) and substituted Cbz), aliphatic urethane protecting groups (e.g. 9-fluorenylmethoxycarbonyl (Fmoc), t-butyloxycarbonyl (Boc), isopropyloxycarbonyl, cyclohexyloxycarbonyl) and alkyl or aralkyl type protecting groups (e.g. benzyl, trityl, chlorotrityl). Examples of suitable oxygen protecting groups may include for example alky silyl groups, such as trimethylsilyl or fe/t-butyldimethylsilyl; alkyl ethers such as tetrahydropyranyl or terf-butyl; or esters such as acetate.
Examples
The following examples illustrate the invention. These examples are not intended to limit the scope of the invention, but rather to provide guidance to the skilled artisan to prepare and use the compounds, compositions, and methods of the invention. While particular embodiments of the invention are described, the skilled artisan will appreciate that various changes and modifications can be made without departing from the spirit and scope of the invention
Intermediates Intermediate 1 : 1 ,1 -Dimethylethyl-2-(2-methylpropyl)hydrazinecarboxylate
A solution of 1 ,1-dimethylethyl hydrazinecarboxylate (ALDRICH, 9.2 g, 70 mmol) in i- PrOH (50 ml) was treated at O0C with /-butylaldehyde (6.4 ml, 70 mmol) over 15 min and stirring at O0C for 2 h, then the mixture was stirred 5 h at room temperature. To this solution containing the intermediate hydrazone was added PtO2 and the suspension was hydrogenated at room temperature and 2.6 bar for 48 h. The suspension was filtered and the solvent was removed under reduced pressure to give the title compound. 1H NMR (300 MHz, CDCI3) δ ppm: 6.02 (br.s, 1 H), 3.92 (br.s, 1 H), 2.66 (d, 2H), 1.73 (m, 1H), 1.46 (s , 9H), 0.93 (d, 6H) [ES+ MS] m/z 189 (MH+).
Intermediate 2: 1,1-Dimethylethyl 2-(2,2-dimethylpropyl)hydrazinecarboxylate
The title compound was prepared by a method analogous to that described above for Intermediate 1 , replacing /-butylaldehyde with trimethylacetaldehyde. 1H NMR (300 MHz, CDCI3) δ ppm: 8.19 (s, 1 H), 3.34 (br.s, 1 H), 2.46 (d, 2H), 1.37 (s, 9H), 0.85 (s, 9H) [ES+ MS] m/z 203 (MH+).
Intermediate 3: 1 ,1 -Dimethylethyl-2-(2-chloro-4-pyrimidinyl)-2-(2-methylpropyl)- hydrazinecarboxylate
A mixture of Intermediate 1 (12.07 g, 64 mmol), 2,4-dichloropyrimidine (19.07 g, 128 mmol), triethylamine (10.7 ml, 145 mmol) and dry EtOH (100 ml) was stirred at room temperature for 3 days. The mixture was concentrated under reduced pressure and the residue partitioned between DCM and 1M ammonium chloride. The organic layer was treated with brine and dried over MgSO4. The residue was purified by flash chromatography (eluent: Hex/AcOEt mixtures 97:3 to 50:50) to give the title compound. 1H
NMR (300 MHz, DMSO-d6) δ ppm: 9.77 (br.s, 1H), 8.15 (d, 1 H), 6.52 (br.s, 1 H), 3.76
(br.s, 1 H), 3.17 (br.s, 1H), 1.92 (m, 1 H), 1.43 (s, 9H), 0.87 (m, 6H). [ES- MS] m/z 299 (MH').
Intermediate 4: 1,1-Dimethylethyl-2-(2-chloro-4-pyrimidinyl)-2-(2,2-dimethylpropyI)- hydrazinecarboxylate
The title compound was prepared by a method analogous to that described above for Intermediate 3, replacing Intermediate 1 with Intermediate 2. 1H NMR (300 MHz, CDCI3) δ ppm: 8.12 (d, 1 H), 6.64 (br.s, 1 H), 6.58 (d, 1H), 1.50 (s, 9H)1 1.00 (s, 9H). [ES- MS] m/z 313 (MH').
Intermediate 5: 1 ,1 -Dimethylethyl-2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)- hydrazinecarboxylate
Potassium cyanide (3.5 g, 54.6 mmol) was added to a suspension of Intermediate 3 (13.69 g, 45.51 mmol) and DABCO (5.10 g, 45.51 mmol) in a mixture of DMSO/H
2O 85/15 (195 ml) at room temperature. The reaction mixture was stirred at 100
0C for 1.7 h, poured into ice water (250 ml). The white solid was filtered off and dried. The compound was purified by flash chromatography (eluent: Hex/AcOEt mixtures 4:1 to 3:1) to give the title compound.
1H NMR (300 MHz, DMSO-d6) δ ppm: 9.88 (br.s, 1 H), 8.37 (d, 1 H), 6.80 (br.s, 1 H), 3.60 (br.s, 1 H), 3.25 (br.s, 1 H), 1.95 (m, 1H), 1.43 (s, 9H), 0.88 (m, 6H). [ES- MS] m/z 290 (MH-).
Intermediate 6: 1,1-Dimethylethyl-2-(2-cyano-4-pyrimidinyl)-2-(2,2-dimethylpropyl)- hydrazinecarboxylate .
The title compound was prepared prepared by a method analogous to that described above for Intermediate 5, replacing Intermediate 3 with Intermediate 4. 1H NMR (300 MHz, DMSO-d6): 9.89 (br.s, 1H), 8.39 (d, 1H), 6.79 (br.s, 1h), 3.96 (d, 1H), 3.10 (d, 1H), 1.43 (s, 9H), 0.93 (s, 9H). [ES+ MS] m/z 306 (MH+).
Intermediate 7: 4-[1 -(2-Methylpropyl)hydrazino]-2-pyrimidinecarbonitrile
Intermediate 5 (8.2 g, 28.15 mmol) was dissolved in a mixture of TFA (12 ml) in DCM (120 ml). The reaction mixture was stirred at room temperature overnight. More TFA (5 ml) was added and after 2 h the reaction was finished. The mixture was treated with water (2 x 100 ml), 10% NaHCO3 (100 ml), dried over Na2SO4 and evaporated under reduced pressure to give the title compound. 1H NMR (300 MHz, DMSO-d6): 8.17 (d, 1 H), 7.23 (d, 1 H), 4.93 (s, 2H), 3.52 (d, 1 H), 2.17 (m, 1 H), 0.84 (d, 6H). [ES+ MS] m/z 292 (MH+).
Intermediate 8: 4-[1 -(2,2-Dimethylpropyl)hydrazino]-2-pyrimidinecarbonitrile
The title compound was prepared by a method analogous to that described above for Intermediate 7, replacing Intermediate 5 with Intermediate 6. 1H NMR (300 MHz, DMSO- d6): 8.19 (d, 1 H), 7.22 (d, 1 H), 4.99 (s, 2H), 3.55 (s, 2H), 0.94 (s, 9H). [ES+ MS] m/z 206 (MH)+
Intermediate 9: Ethyl W-(2-methylpropyl)glycinate
Ethyl bromoacetate (1.2 ml, 11.07 mmol) was added dropwise to a solution of isobutylamine (1.1 ml, 11.07 mmol) and triethylamine (1.9 ml, 13.28 mmol) in THF (30 ml) at O0C. The reaction mixture was warmed to room temperature and stirred overnight. The precipitate was filtered and the filtrate was concentrated under reduce pressure. The residue was purified by flash chromatography (eluent: Hex/AcOEt mixtures 4:1 to 3:1) to give the title compound. 1H NMR (300 MHz, DMSO-d6) δ ppm: 3.60 (s, 3H), 3.29 (s, 3H), 2.28 (d, 2H), 1.60 (m, 1 H), 0.84 (d, 6H). Reference: J. Med. Chem 2000, 43, 2805-2813
Intermediate 10: Methyl
The title compound was prepared by a method analogous to that described above for Intermediate 9, replacing ethyl bromoacetate with methyl bromoacetate and isobutylamine with neopentylamine. 1H NMR (300 MHz, DMSO-d6) δ ppm: 3.60 (s, 3H), 3.31 (br.s, 2H), 2.24 (s, 2H), 0.84 (s, 9H).
Intermediate 11: Ethyl W-(2-chloro-4-pyrimidinyl)-W-(2-methylpropyl)glycinate
To a solution of Intermediate 9 (0.89 g, 5.59 mmol) and triethylamine (1 ml, 7.27 mmol) in i-PrOH (50 ml) was added 2,4-dichloropyrimidine (0.83 g, 5.59 mmol) and the mixture was heated at reflux for 24 h. The reaction mixture was concentrated under reduce pressure and the residue was purified by flash chromatography (eluent: Hex/AcOEt mixtures 4:1 to
3:1) to give the title compound. 1H NMR (300 MHz, DMSO-d6) δ ppm: 8.07 (d, 1 H), 6.87
(d, 1 H), 4.25 (br.s, 2H), 4.11 (q, 2H), 3.28 (d, 2H), 1.93 (m, 1 H), 1.18 (dt, 3H), 0.87 (d, 6H).
Intermediate 12: Methyl /V-(2-chloro-4-pyrimidinyl)-/V-(2,2-dimethylpropyl)gIycinate
The title compound was prepared by a method analogous to that described above for Intermediate 11 , replacing Intermediate 9 with Intermediate 10. 1H NMR (300 MHz, DMSO-d6) δ ppm: 8.07 (d, 1H), 6.99 (d, 1H), 4.26 (br.s, 2H), 3.63 (s, 3H), 3.33 (br.s, 2H), 0.92 (s, 9H).
Intermediate 13: Ethyl W-(2-cyano-4-pyrimidinyl)-Λ/-(2-methyIpropyl)glycinate
Potassium cyanide (0.29 g, 4.46 mmol) was added to a suspension of Intermediate 11 (0.61 g, 2.23 mmol) and DABCO (0.25 g, 2.23 mmol) in a mixture of DMSO/H2O 85/15 (24 ml) at room temperature. The reaction mixture was stirred for 4 h at 60 0C, poured into ice water (50 ml) extracted with DCM (3x100 ml). The combined extracts were dried over Na2SO4 and concentrated under high vacuum. The residue was purified by flash chromatography (eluent: Hex/AcOEt mixtures 3:1 to 1 :1). 1H NMR (300 MHz, DMSO-d6) δ ppm: 8.27 (d, 1 H), 7.17 (d, 1 H), 4.29 (s, 2H), 4.12 (q, 2H), 3.33 (br.s, 2H), 1.94 (m, 1 H), 1.18 (t, 3H), 0.88 (d, 6H).
Intermediate 14: Methyl Λ/-(2-cyano-4-pyrimidinyl)-Λ/-(2,2-dimethylpropyl)glycinate
The title compound was prepared by a method analogous to that described above for Intermediate 13, replacing Intermediate 11 with Intermediate 12. 1H NMR (300 MHz, DMSO-d6) δ ppm: 8.28 (d, 1 H), 7.28 (d, 1 H), 4.32 (br.s, 2H), 3.64 (s, 3H), 3.37 (br.s, 2H), 0.93 (S, 9H).
Intermediate 15: W-(2-Cyano-4-pyrimidinyl)-Λ/-(2-methylpropyl)gIycine
A mixture of Intermediate 13 (115 mg, 0.44 mmol) and lithium hydroxide (22 mg, 0.53 mmol) in THF/H2O 2/1 (3 ml) was stirred at room temperature overnight. The reaction mixture was concentrated and taken up in 2N hydrochloric acid (10 ml), extracted with AcOEt (3x10 ml). The organic extracts were combined, washed with brine and dried over Na2SO4, filtered and evaporated to give the title compound as a solid. 1H NMR (300 MHz, DMSO-d6) δ ppm: 12.80 (br.s, 1H), 8.26 (d, 1 H), 7.13 (d, 1 H), 4.23 (s, 2H), 3.30 (d, 2H), 1.94 (m, 1H), 0.88 (d, 6H).
Intermediate 16: Λ/-(2-Cyano-4-pyrimidinyl)-W-(2,2-dimethyIpropyl)glycine
The title compound was prepared by a method analogous to that described above for Intermediate 15, replacing Intermediate 13 with Intermediate 14. 1H NMR (300 MHz, DMSO-d6) δ ppm: 12.80 (br.s, 1H), 8.26 (d, 1 H), 7.25 (d, 1 H), 4.22 (s, 2H), 3.34 (br.s, 2H), 0.93 (s, 9H).
Intermediate 17: 4-[(4-methyl-1-piperazinyl)methyl]ben2oic acid
A solution of N-methylpiperazine (ALDRICH, 1.46 ml, 13.1 mmol) in dimethylformamide (5 ml) was cooled to 0° C and, then, potassium carbonate (K2CO3, 1.81 g, 13.1 mmol) was added. This mixture was stirred at 0° C for 30 min. Then, methyl 4-(bromomethyl) benzoate (ALDRICH, 3 g, 13.1 mmol) was added. The reaction mixture was allowed to warm up to room temperature and stirred for 17 h. The mixture was concentrated under reduce pressure. The residue was dissolved in DCM and washed with water, the aqueous
layer was extracted with DCM. The organic layers were combined, washed with water, dried over MgSO4 filtered and the solvent removed under reduce pressure to give the corresponding methyl 4-[(4-methyl-1-piperazinyl)methyl]benzoate. 1H RMN (300 MHz, CDCI3-d6): 7.97 (d, 2H), 7 40 (d, 2H), 3.90 (s, 3H), 3.55 (s, 2H), 2.47 (br. m, 8H), 2.28 (s, 3H).
A solution of lithium hydroxide (ALDRICH, 337 mg, 14.1 mmol) in H2O (10 ml) was added to a solution of the methyl ester (1.4 g, 5.63 mmol) in MeOH (20 ml) and the mixture was heated at reflux for 2h. The mixture was concentrated under reduced pressure. The residue was dissolved in DCM and 2N hydrochloric acid was added to give pH 5. The aqueous layer was partitioned with n-Butanol (5 times) and the fractions were combined, dried over MgSO4, filtered and evaporated under reduce pressure to give the title compound as a white solid. 1H RMN (300 MHz, DMSO-d6): 12.83 (br. m, 1 H), 11.05 (br. m, 1 H), 7.90 (d, 2H), 7.43 (d, 2H), 4.36 (m, 1 H), 3.60 (s, 2H), 3.38-2.80 (br. m, 8H), 2.68 (s, 3H). [ES+ MS] m/z 235 (MH)+.
Intermediate 18: 4-{[2-(4-morpholinyl)ethyl]oxy}benzoic acid
Triphenylphosphine (ALDRICH, 26 g, 98.6 mmol) and 4-(2-hydroxyethyl) morpholine (ALDRICH, 8.6g, 66 mmol) were added to a solution of methyl 4-hydroxybenzoate (ALDRICH, 10 g, 66 mmol) in dry THF (200 ml) under nitrogen atmosphere. The mixture was cooled to O0C and a solution of diisopropyl azodicarboxylate (ALDRICH, 17.3 g, 85.8 mmol) in dry THF (80 ml) was added drop wise. The reaction mixture was stirred a room temperature overnight. The solvent was removed under reduce pressure and the residue was treated with ethyl ether (50 ml). The triphenylphosphane oxide was precipitated and filtered. The solvent was evaporated and treated again with ethyl ether (50 ml). This procedure was repited twice to eliminate the triphenylphosphane oxide. To end, the last residue was purified by flash chromatography (eluent: DCM/MeOH mixtures 40:1 to 20:1 ) to give methyl 4-{[2-(4-morpholinyl)ethyl]oxy}benzoate. Sodium hidroxyde 2N (50 ml) was added to solution of the methyl ester above obtained (16 g, 60 mmol) in dioxane (60 ml). The reaction mixture was stirred overnight. Hydrochloric acid 2N was added up to pH 7 and the mixture evaporated under reduce pressure. The residue was purified by flash chromatography (eluent: DCM/MeOH mixtures 9:1 to 5:1 ) to give the title compound. 1H
NMR (300 MHz, DMSO-d6) δ: 2.45 (m, 4H), 2.69 (t, 2H), 3.56 (t, 4H), 4.14 (t, 2H)1 7.00 (d, 2H), 7.87 (d, 2H). [ES+ MS] m/z 252 (MH+).
Intermediate 19: 1,1-dimethylethyl 2-(2-chloro-5-methyl-4-pyrimidinyl)-2-(2- methylpropyl)hydrazinecarboxylate
To a solution of 2,4-dichloro-5-methylpyrimidine (1.5 g, 9 mmol) and Intermediate 1 (0.8 g, 4 mmol) in EtOH (8 mL), TEA (0.7 ml_, 5 mmol) was added and the resulting reaction mixture was stirred at room temperature for 72 h, then refluxed for 40 h to reach completion. The mixture was concentrated under reduced pressure and the residue partitioned between DCM and 1 M ammonium chloride. The organic layer was washed with water and brine and dried over anhydrous MgSO4. The residue was purified by flash chromatography (elute: Hex/EtOAc mixtures 100:0 to 1 :1) to give the title compound. 1H NMR (300 MHz, CDCI3) δ ppm: 7.98 (br.s, 1 H), 6.64 (s, 1 H), 4.10 (m, 1H), 3.54 (m, 1 H), 2.22 (s, 3H), 2.03 (m, 1 H), 1.48 (br.s, 9H), 0.95 (d, 6H) [ES+ MS] m/z 315 (MH+), [ES- MS] m/z 313 (MH").
Intermediate 20: 1,1-dimethylethyl 2-(2-cyano-5-methyl-4-pyrϊmidinyl)-2-(2- methylpropyl)hydrazinecarboxylate
The title compound was prepared by a method analogous to that described above for
Intermediate 5, replacing Intermediate 3 with Intermediate 19. 1H NMR (300 MHz, DMSO- d6) δ ppm: 9.93 (br.s, 1 H), 8.19 (s, 1 H), 3.98 (br.s, 1 H), 3.05 (br.s, 1 H), 2.23 (s, 3H), 1.93 (m, 1 H), 1.41 (br.s, 9H), 0.86 (br.s, 6H); [ES+ MS] m/z 306 (MH+).
Intermediate 21 : 5-methyl-4-[1-(2-methylpropyl)hydrazino]-2-pyrimidinecarbonJtrile
The title compound was prepared by a method analogous to that described for Intermediate 7, replacing Intermediate 5 with Intermediate 20. 1H NMR (300 MHz, DMSO- d6) δ ppm: 7.97 (s, 1 H), 4.84 (br.s, 2H), 3.52 (d, 2H), 2.44 (s, 3H), 2.18 (m, 1 H), 0.87 (d, 6H) [ES+ MS] m/z 206 (MH+).
Intermediate 22: 1,1-dimethylethyl 2-(2-chIoro-5-methyI-4-pyrimidinyI)-2-(2,2- dimethylpropyl)hydrazinecarboxylate
The title compound was prepared by a method analogous to that described for Intermediate 19, replacing Intermediate 1 with Intermediate 2.
1H NMR (300 MHz, CDCI
3) δ ppm: 7.90 (br.s, 1 H), 6.67 (s, 1 H), 4.72 (m, 1 H), 2.73 (m, 1 H), 2.23 (s, 3H), 1.47 (br.s, 9H), 0.97 (s, 9H) [ES+ MS] m/z 329 (MH
+), [ES- MS] m/z 327 (MH
").
Intermediate 23: 4-[1 -(2,2-dimethylpropyl)hydrazino]-5-methyl-2- pyrimidinecarbonitrile
The title compound was prepared by a method analogous to that described for Intermediate 7, replacing Intermediate 5 with Example 68. 1H NMR (300 MHz, DMSO-d6)
δ ppm: 7.98 (s, 1H), 4.91 (br.s, 2H), 3.64 (s, 2H), 2.46 (s, 3H), 0.94 (d, 9H) [ES+ MS] m/z 220 (MH+).
Intermediate 24: 1,1-dimethylethyl 2-(2-chloro-6-methyI-4-pyrimidinyl)-2-(2- methylpropyl)hydrazinecarboxylate
The title compound was prepared by a method analogous to that described for Intermediate 19, replacing 2,4-dichloro-5-methylpyrimidine with 2,4-dichloro-6- methylpyrimidine. 1H NMR (300 MHz, CDCI3) δ ppm: 6.50 (br.s, 1H), 6.37 (s, 1 H), 4.15 (m, 1H), 3.55 (m, 1H), 2.35 (s, 3H), 2.03 (m, 1H), 1.48 (br.s, 9H), 0.94 (d, 6H) [ES+ MS] m/z 315 (MH+), [ES- MS] m/z 313 (MH").
Intermediate 25: 1,1-dimethylethyl 2-(2-chloro-6-methyl-4-pyrimidinyl)-2-(2,2- dimethylpropyl)hydrazinecarboxylate
The title compound was prepared by a method analogous to that described for Intermediate 24, replacing Intermediate 1 with Intermediate 2. 1H NMR (300 MHz, CDCI3) δ ppm: 6.56 (br.s, 1H), 6.40 (s, 1 H), 4.15 (m, 1 H), 3.50 (m, 1H), 2.36 (s, 3H), 1.49 (br.s, 9H), 0.98 (s, 9H) [ES+ MS] m/z 329 (MH+), [ES- MS] m/z 327 (MH").
Intermediate 26: 4-[(4-propyl-1-piperazinyl)methyl]benzoic acid.
A mixture of 4-(bromomethyl)benzoic acid (ALDRICH, 3.0Og., 13.9 mmol), 1-N- propylpiperazine dihydrobromide (ALDRICH, 4.05 g., 13.9 mmol) and anhydrous potassium carbonate (ALDRICH, 3.86 g., 27.9 mmol) in dry acetonitrile 120ml was stirred at ambient temperature for 18 hours; the solvent was removed by evaporation. Then, 2N HCI was added until pH value approximately 2 and the product was extracted with n- butanol. The combined organic layers were washed with brine, dried over MgSO4 and concentrated under reduced pressure. The crude product was used without any further purification.
Intermediate 27: 4-(4-propyM-piperazinyl)benzoic acid.
A mixture of ethyl 4-(1-piperazinyl)benzoate (CHESS, 937mg, 4mmol), triethylamine (FLUKA, 1.12mL, 8mmol) and 1 -bromopropane (ALDRICH, 0.4mL, 4.4mmol) in dry dimethylformamide (6mL) was stirred 18 hours at room temperature. The solvent was removed under reduce pressure and the residue was dissolved in ethanol/NaOH 2N 1:1 (25mL) and refluxed for 4 hours. The solvent was concentrated under reduced pressure to 5mL; the solid obtained was filtered to obtain 1.1g of the title compound (sodium salt). 1H NMR (300 MHz1 D2O) δ ppm: 7.65 (d, 2H), 6.93 (d, 2H), 3.08 (m, 4H), 2.49 (m, 4H), 2.20 (m , 2H), 1.34 (d, 2H) and 0.70 (t, 3H). [ES+ MS] m/z 249 (MH)+; [ES- MS] m/z 247 (M-H)"
Intermediate 28: 2-methyl-2-(4-methylphenyl)propanoic acid
To a solution of diisopropylamine (ALDRICH, 15.2 mL, 109 mmol) in 108 mL of anhydrous THF at -2O
0C under N
2 was added n-BuLi (55.7 mL, 106 mmol, 1.9M solution in hexane) dropwise via cannula. After stirring for 2 hours, p-tolylacetic acid (ALDRICH, 3.99 g, 26.5
mmol) was added in 26 mL of THF. The mixture was stirred at -5
0C for 2 hours and, then, treated with iodomethane (9.9 mL, 159 mmol). The reaction mixture was allowed to gradually warm up to room temperature, stirred at that temperature for 15 hours, and then, quenched by pouring it into 20 mL of HCI (1 N). The aqueous layer was extracted with Et
2O (3x100 mL). The organic layer was treated with brine and dried over MgSO
4 and concentrated in vacuo to give 4.86 g of the title compound as a brown oil.
1H NMR (300 MHz
1 DMSO-d6) δ ppm: 12.22 (br.s, 1 H), 7.21 (m, 2H), 7.12 (m, 2H), 2.25 (s, 3H), 1.43 (s, 6H). No molecular peak is detected by MS.
Intermediate 29: methyl 2-methyl-2-(4-methylphenyI)propanoate
To a stirred mixture of Intermediate 28 (4.86 g, 26.5 mmol) and K2CO3 (14.6 g, 106 mmol) in acetone (150 mL), methyl iodine (ALDRICH, 16 mL, 265 mmol) was added at room temperature, and, then, the reaction mixture was stirred at 5O0C for 3 hours. The mixture was filtered, the filtrate was evaporated to dryness, and the residue was chromatographed (silica gel, 1:1 hexane/ethyl acetate) to obtain 4.37 g of the desired product. 1H NMR (300 MHz, DMSO-d6) δ ppm: 7.15 (m, 4H), 3.56 (s, 3H), 2.25 (s, 3H), 1.46 (s, 6H).
Intermediate 30: methyl 2-methyl-2-{4-[(4-methyl-1-piperazinyl)methyl]phenyl} propanoate
To a stirred solution of Intermediate 29 (3.4g, 17.7 mmol) in CCI4 (55mL) in a 250 mL round-bottom flask at room temperature and under Argon, NBS (3.46g, 19.5 mmol) and azoisobutyronitrile (FLUKA, 290 mg, 1.77 mmol) were added. The mixture was stirred at 8O0C for 18 hours. The mixture was cooled down to room temperature, filtered and the filtrate was concentrated under vacuum. The resulting orange residue was used without further purification.
To the crude bromide in ACN (150 mL), K2CO3 (4.89g, 35.4 mmol) and N- methylpiperazine (ALDRICH, 1.96 mL, 17.7 mmol) were added at room temperature. After 15 h, the reaction solution was concentrated to dryness and the residue was diluted with CH2CI2 (10OmL) and washed with H2O (3x50 mL). The organic layer was dried with Na2SO4 and purified by flash chromatography on silica gel (hexane/ethyl acetate) to give 2.82 g of the title compound. 1H NMR (300 MHz, DMSO-d6) δ ppm: 7.23 (s, 4H), 3.57 (s, 3H), 3.40 (s, 2H), 2.33 (m, 8H), 2.15 (s, 3H), 1.47 (s, 6H). [ES+ MS] m/z 291 (MH)+.
Intermediate 31 : 2-methyl-2-{4-[(4-methyl-1-piperazinyl)methyl]phenyI}propanoic acid
Intermediate 30 (2.82 g, 9.71 mmol) was dissolved in a mixture of 9.3:0.7 of THF:H2O. To the stirred solution was added lithium hydroxide monohydrate (ALDRICH, 1.63 g, 38.84 mmol) and the reaction mixture was refluxed for 48 h. THF was removed under vacuum and the remaining residue was purified by preparative HPLC (ACN/H2O 0.1 % of TFA, from 0% to 30% of ACN) to give 1.37 g of the acid. 1H NMR (300 MHz, DMSO-d6: 7.35 (s, 4H), 3.86 (br.s, 2H), 3.12-3.42 (m, 8H), 2.78 (s, 3H), 1.46 (s, 6H). [ES+ MS] m/z 277 (MH)+.
Intermediate 32: 4-(chloromethyl)-ΛT-(2-cyano-4-pyrimidinyl)-Λf-(2,2-dimethyl propyl) benzohydrazidβ.
To a stirred solution of Intermediate 8 (200 mg, 0.97 mmol) in dry THF (3 mL), 4- (chloromethyl)benzoyl chloride (ALDRICH, 184 mg, 0.97 mmol) and potassium carbonate (268 mg, 1.94 mmol) were added and the resulting reaction mixture was stirred at room temperature for 18 hours. The mixture was filtered and the solvent was evaporated in vacuo. The crude product was used without any further purification.
Intermediate 33: 4-Amino-2-methyl-2-butanol.
A mixture of 3-hydroxy-3-methylbutanenitrile (FLUKA, 0.7 ml, 6.8 mmol), ethanol (50 ml) and concentrated hydrochloric acid (1.1 ml) was added PtO2 (159 mg) and the suspension was hydrogenated at room temperature and 35 psi for 20 hours. The reaction was filtered and the solvent was removed under reduced pressure to give the title compound. 1H NMR (300 MHz, DMSO-d6) δ ppm: 7.96 (br.s, 2H), 3.82 (br.s, 1 H), 2.81 (m, 2H), 1.64 (m, 2H), 1.10 (s, 6H). [ES+ MS] m/z 104 (MH)+.
Intermediate 34: 1,1-dimethylethyl (3-hydroxy-3-methyIbutyl)carbamate.
A mixture of Intermediate 33 (701.5 mg, 6.8 mmol), 1 N NaOH (11.2 ml, 11.2 mmol) and bis(1 ,1-dimethylethyl) dicarbonate (ALDRICH, 2.226 g, 10.2 mmol) in terf-butyl alcohol (127 ml) was stirred at room temperature for 72 hours. The solvent was removed under reduced pressure and the crude was suspended in 1N ammonium chloride (40 ml), 1N HCI was added until pH = 6-7, the aqueous layer was extracted with ethyl acetate (3x40 ml). The organic extracts were dried over MgSO4 and concentrated under vacuum to give the title compound. 1H NMR (300 MHz, DMSO-d6) δ ppm: 6.64 (m, 1H), 4.22 (s, 1H), 2.98 (m, 2H), 1.47 (t, 2H), 1.35 (s, 9H), 1.06 (s, 6H). [ES+ MS] m/z 204 (MH)+.
Intermediate 35: 1,1-dimethylethyl 4-(3-hydroxy-3-methylbutyl)-1-piperazine carboxylate.
The title compound was prepared by a method analogous to that described above for Intermediate 34, using 2-methyl-4-(1-piperazinyl)-2-butanol (UBICHEM) as starting
material.
1H NMR (300 MHz, DMSO-d6) δ ppm: 3.27 (m, 4H), 2.45-2.25 (m, 6H), 1.51 (t, 2H), 1.38 (s, 9H), 1.07 (s, 6H). [ES+ MS] m/z 273 (MH)
+.
Intermediate 36: 1,1-dimethylethyl (5-hydroxy-1,5-dimethylhexyl)carbamate
The title compound was prepared by a method analogous to that described above for intermediate 34, using 6-amino-2-methyl-2-heptanol (AVOCADO) as starting material. 1H NMR (300 MHz, DMSO-d6) δ ppm: 6.58 (d, 1 H), 4.03 (s, 1 H), 3.39 (m, 1H), 1.35 (s, 9H), 1.26 (m, 6H), 1.02 (s, 6H), 0.97 (d, 3H). [ES+ MS] m/z 246 (MH)+.
Intermediate 37: ethyl (4-chlorobutyl)methylcarbamate
Ethyl chloroformate (ALDRICH, 6.88 mL, 72 mmol) in dry toluene (10 ml.) was slowly added onto a solution of 1-methyl-pyrrolidine (ALDRICH, 5 mL, 48 mmol) in dry toluene (20 mL) at 850C (exothermic reaction !!) and stirring was continued at that temperature for 2 hours. The reaction mixture was cooled down to room temperature, diluted with EtOAc (150 mL) and extracted with 1 N HCI (3 x 180 mL). The organic layer was washed with sat NaHCO3, then, with brine, it was dried over NaSO4 and concentrated to dryness. Intermediate 37 was obtained as a yellow oil which was used without further purification. 1H NMR (300 MHz, CDCI3) δ ppm: 4.11 (q, 2H, J = 7.1 Hz), 3.56 (t, 2H, J = 6.3 Hz), 3.29 (m, 2H), 2.88 (bs, 3H), 2.0-1.5 (m, 4H), 1.25 (t, 3H, J = 7.1 Hz).
Intermediate 38: 5-(1,3-dioxo-1,3-dihydro-2H-isoindol-2-yl)pentylmethane sulfonate. .
A mixture of 5-amino-1-pentanol (ALDRICH, 0.333 mL, 3.2mmol), phthalic anhydride (PROBUS, 533mg, 3.6mmol) in dry toluene (Panreac, 10 mL) was heated at 1000C for 4
hours. The reaction mixture was diluted with 7OmL of dichloromethane, washed twice with 5OmL of hydrochloric acid 1N and with 5OmL of aqueous sodium bicarbonate saturated solution, dried over sodium sulfate and evaporated to dryness. The residue was purified by flash cromatography using dichloromethane/methanol 20:1 as eluant to give 2-(5- hydroxypentyl)-1H-isoindole-1 ,3(2H)-dione which was dissolved in 3OmL of dichloromethane (PANREAC) and treated with triethylamine (SIGMA, 0.443 mL, 3.18mmol) and methanesulfonyl chloride (ALDRICH1 0,217 mL, 2.8mmol). The reaction mixture was stirred overnight at room temperature, then, diluted with 5OmL of dichloromethane, washed with 5OmL of hydrochloric acid 1 N, 5OmL of aqueous sodium bicarbonate saturated solution and 5OmL of brine, dried over sodium sulfate and evaporated to dryness to give Intermediate 39. [ES+ MS] m/z: 312 (MH)+.
Intermediate 39: 4-(dimethylamino)butyl methanesulfonate
N' // \\
/ O ' " O 4-(Dimethylamino)-1-butanol (MERCK, 328 mg, 2.8 mmol) was treated with triethylamine (SIGMA, 0.443 mL, 3.18mmol) and methanesulfonyl chloride (ALDRICH, 0.217 mL, 2.8mmol) at O0C. The reaction mixture was stirred overnight at room temperature, then, diluted with 5OmL of dichloromethane, washed with 5OmL of hydrochloric acid 1 N, 5OmL of aqueous sodium bicarbonate saturated solution and 5OmL of brine, dried over sodium sulfate and evaporated to dryness give Intermediate 39. [ES+ MS] m/z: 196 (MH)+.
Intermediate 40: 1,1-dimethylethyl {3-[(1,1-dimethylethyl)amino]propyl} carbamate
Cesium hydroxide monohydrate (ALDRICH, 1.1 g, 6.8 mmol) was added to a suspension of molecular sieves 4A in anhydrous DMF under nitrogen atmosphere. The suspension was stirred 10 minutes, terf-butylamine (ALDRICH, 500 mg, 6.8 mmol) was added and the reaction was stirred 30 minutes at room temperature. Then, 2-(Boc-amino)propyl bromide (ALDRICH, 1.8 g, 8.6 mmol) was added and the suspension was stirred under nitrogen at room temperature overnight. The reaction crude was filtered, DMF was evaporated under vacuum and the residue was partitioned between AcOEt and NaOH 1 N. The organic phase was dried with MgSO
4, evaporated to dryness and the crude was purified in silica (DCM/MeOH) to give the desired compound.
1H NMR (300 MHz, CDCI
3) δ ppm: 6.86 (m,
1 H), 4.2 (br s, 1 H), 2.96 (q, 2H, J = 6 Hz), 2.54 (m, 2H), 1.50 (m, 2H), 1.36 (s, 9H), 1.06 (s, 9H). [ES+ MS] m/z 231 (MH)
+.
Intermediate 41: phenylmethyl {3-[(1,1-dimethylethyl)amino]propyl}carbamate
Benzyl N-(3-hydroxypropyl)carbamate (ALDRICH, 500 mg, 2.39 mmol) was dissolved in 12 imL of dry DCM under nitrogen atmosphere and cooled at O0C in an ice bath. Triethylamine (FLUKA, 0.5 mL, 3.58 mmol) and toluene-4-sulfonyl chloride (ALDRICH, 528 mg, 2.77 mmol) were added and the resultant solution was stirred at room temperature overnight. Reaction was diluted with more DCM and washed with water. Organic phase was dried with MgSO4, filtered and evaporated. Residue was immediately dissolved in 10 mL of anhydrous acetonitrile and sodium bicarbonate (PANREAC, 507 mg, 6.04 mmol) and terf-butylamine (ALDRICH, 367 mg, 5.02 mmol) were added. The suspension was heated at 50-550C for 3 hours and more sodium bicarbonate (PANREAC, 507 mg, 6.04 mmol) and terf-butylamine (ALDRICH, 367 mg, 5.02 mmol) were added. Reaction was heated at 50-550C overnight, then cooled down to room temperature, filtered over Celite and evaporated to dryness. The residue was purified in silica using mixtures of DCM/Methanol. 1H NMR (300 MHz, DMSO) δ ppm: 8.19 (br s, 2H), 7.30-7.43 (m, 5H), 5.02 (s, 2H), 3.09 (q, 2H, J = 6.1 Hz), 2.85 (m, 2H), 1.70 (m, 2H), 1.23 (s, 9H). [ES+ MS] m/z 265 (MH)+.
Intermediate 42: phenylmethyl {4-[(1,1-dimethylethyl)amino]butyl}carbamate
The title compound was prepared by a method analogous to that procedure described above for intermediate 41 using 4-(Z-amino)-1-butanol (FLUKA) as starting reagent. 1H
NMR (300 MHz, DMSO) δ ppm: 7.26-7.38 (m, 5H), 4.99 (s, 2H), 2.94-3.02 (m, 2H), 2.43- 2.49 (m, 2H), 1.30-1.51 (m, 4H), 1.00 (s, 9H). [ES+ MS] m/z 279 (MH)+.
Intermediate 43: 1,1-dimethylethyl [4-(butylamino)butyl]carbamate (M356/132/1)
The title compound was prepared by a method analogous to that described above for intermediate 41 , using 4-(Boc-amino)-1-butanol (FLUKA) as starting reagent. [ES+ MS] m/z 245 (MH)
+.
Intermediate 44: 1,1-dimethylethyl [5-(butyIamino)pentyl]carbamate
The title compound was prepared by a method analogous to that described above for Intermediate 41, using 5-(Boc-amino)-1-pentanol (FLUKA) as starting reagent. [ES+ MS] m/z 259 (MH)+.
Intermediate 45: 1,1-dimethylethyl {4-[(1,1-dimethylethyl)amino]butyl}carbamate
The title compound was prepared by a method analogous to that described above for intermediate 41 using 4-(Boc-amino)-1-butanol (FLUKA) as starting reagent. 1H NMR (300 MHz, DMSO) δ ppm: 6.76 (m, 1 H), 4.35 (t, 1 H, J = 5.1 Hz), 2.85-2.91 (m, 2H), 2.42 (t, 2H, J = 6.5 Hz), 1.28-1.44 (m, 13H), 0.99 (s, 9H). [ES+ MS] m/z 245 (MH)+.
Intermediate 46: 1,1-dimethylethyl {5-[(1,1-dimethylethyl)amino]pentyl}carbamate
The title compound was prepared by a method analogous to that described above for intermediate 41 using 5-(Boc-amino)-1-pentanol (FLUKA) as starting reagent.
1H NMR (300 MHz, DMSO) δ ppm: 6.75 (m, 1 H), 4.32 (t, 1 H, J = 5.1 Hz), 2.84-2.90 (m, 2H), 2.42 (t, 2H, J = 6.9 Hz), 1.21-1.43 (m, 15H), 0.99 (s, 9H). [ES+ MS] m/z 259 (MH)
+.
Intermediate 47: phenylmethyl {2-[(1,1-dimethylethyl)amino]ethyl}carbamate
The title compound was prepared by a method analogous to that described above for Intermediate 41 , using benzyl N-(2-hydroxyethyl)carbamate (ALDRICH) as starting reagent. 1H NMR (300 MHz, DMSO) δ ppm: 7.27-7.38 (m, 5H), 7.17 (m, 1H), 4.99 (s, 2H), 2.98-3.05 (m, 2H), 2.48-2.52 (m, 2H), 0.98 (s, 9H). [ES+ MS] m/z 251 (MH)+.
Intermediate 48:
A mixture of 1 ,1-dimethylethyl (3-bromopropyl)carbamate (FLUKA, 0.50 g, 2.1 mmol), 4- fluoraniline (ALDRICH, 0.2 ml, 2.1 mmol), sodium hydroxide (168 mg, 4.2 mmol) and dry DMF (5 ml) was stirred in Microwave oven at 8O0C, 900 s. The reaction mixture was concentrated under vacuum and the residue was purified by flash chromatography (eluant: Hex/AcOEt mixtures 100:0 to 40:60) to give the title compound. 1H NMR (300 MHz, CDCI3-d6): 6.81-6.91 (m, 2H), 6.59-6.64 (m, 2H), 4.63 (br., 1H), 3.12-3.27 (m, 4H), 1.73-1.82 (m, 2H), 1.44 (s, 9H).
Intermediate 49: Λ/-butyl-4-pyridinamine
A mixture of 4-chloropyridine hydrochloride (ALDRICH, 0.2 g, 1.33 mmol) and butylamine (ALDRICH, 0.263 imL, 2.67 mmol) in 4 mL of ethanol was heated in a microwave oven at
175 0C for 1 hour. Solvent was evaporated and the residue was purified using preparative HPLC (LUNA 50x250 mm, gradient 10% ACN, 0.1%TFA/water, 0.1%TFA to 100% ACN, 0.1%TFA). 1H NMR (300 MHz, DMSO) δ ppm: 13.12 (br s, 1H), 8.59 (m, 1H), 8.25 (d, 1H1 J = 7.0 Hz), 8.09 (d, 1 H, J = 6.5 Hz), 6.86-6.93 (m, 2H), 3.29 (q, 2H, J = 6.7 Hz), 1.59 (qt, 2H1 J = 6.9 Hz)1 1.34-1.47 (m, 2H)1 0.95 (t, 2H1 J = 7.3 Hz). [ES+ MS] m/z 151 (MH)+.
Intermediate 50: 1,1-dimethylethyl [6-(butylamino)hexyl]carbamate
H CL .N
N H O The title compound was prepared by a method analogous to that described above for intermediate 41 , using 6-(Boc-amino)-1-hexanol (FLUKA) as starting reagent. 1H NMR (300 MHz1 DMSO) δ ppm: 6.80 (m, 1 H)1 4.36 (m, 1 H)1 2.87-2.96 (m, 2H)1 2.69-2.75 (m, 2H), 2.46-2.54 (m, 2H)1 1.17-1.46 (m, 21 H)1 0.88-0.93 (m, 3H). [ES+ MS] m/z 273 (MH)+.
Intermediate 51 : {[4-(dimethylamino)phenyl]methyl}(1 ,1 -dimethylethyl)amine
A suspension of 4-dimethylaminobenzaldehyde (ALDRICH, 300 mg, 2 mmol), tert- butylamine (ALDRlCH1 0.525 mL, 5 mmol) and MgSO4 (200 mg) in 10 mL of chloroform was refluxed overnight. Reaction was evaporated and the residue dissolved in 10 mL of methanol. Solution was cooled in an ice bath and sodium borohydride (ALDRICH1 76 mg, 2 mmol) was added. Reaction was stirred at room temperature for 4h, was poured into water and was extracted with DCM. Organic phase was dried with MgSO4, filtered and evaporated and the residue was purified in silica with mixtures DCM/Methanol. 1H NMR (300 MHz1 CDCI3) δ ppm: 7.21 (d, 2H1 J = 7.0 Hz), 6.70 (d, 2H1 J = 7.0 Hz), 3.62 (s, 2H), 2.90 (s, 6H)1 1.16 (s, 9H). [ES+ MS] m/z 207 (MH)+.
Intermediate 52: {(2E)-3-[4-(dimethylamino)phenyl]-2-propen-1 -yl}(1 ,1 -dimethyl ethyl)amine
The title compound was prepared by a method analogous to that described above for Intermediate 51 , using 4-dimethylaminocinnamaldehyde (ALDRICH) as starting reagent. 1H NMR (300 MHz, CDCI3) δ ppm: 7.23-7.27 (m, 2H), 6.64-6.67 (m, 2H), 6.43 (d, 1 H, J = 15.0 Hz), 6.11 (dt, 1 H, J = 15.0, 6.0 Hz), 3.34 (d, 2H, J = 6.0 Hz), 2.93 (s, 6H), 1.15 (s, 9H). [ES+ MS] m/z 233 (MH)+.
Intermediate 53: 4-[(butylamino)methyl]-N,N-dimethylaniline
The title compound was prepared by a method analogous to that described above for Intermediate 51 , using 4-dimethylaminobenzaldehyde (ALDRICH) and butylamine (ALDRICH) as starting reagent. 1H NMR (300 MHz, CDCI3) δ ppm: 7.17 (d, 2H, J = 7.0 Hz), 6.71 (d, 2H, J = 7.0 Hz), 3.68 (s, 2H), 2.92 (s, 6H), 2.58-2.60 (m, 2H), 1.27-1.51 (m, 4H), 0.89 (t, 3H, J = 6.5 Hz). [ES+ MS] m/z 207 (MH)+.
Intermediate 54: 4-[(1 E)-3-(butylamino)-1 -propen-1 -yl]-N,N-dimethylaniline
The title compound was prepared by a method analogous to that described above for intermediate 51 , using 4-dimethylaminocinnamaldehyde (ALDRICH) and butylamine (ALDRICH) as starting reagents. 1H NMR (300 MHz, CDCI3) δ ppm: 7.23-7.27 (m, 2H), 6.64-6.68 (m, 2H), 6.41 (d, 1 H, J = 15.0 Hz), 6.10 (dt, 1 H, J = 15.0, 6.0 Hz), 3.37 (d, 2H, J = 6.0 Hz), 2.94 (s, 6H), 2.64 (t, 2H, J = 6.5 Hz), 1.27-1.51 (m, 4H), 0.91 (t, 3H, J = 6.5 Hz). [ES+ MS] m/z 233 (MH)+.
Intermediate 55: 4-[(4-methyl-1-piperazinyl)methyl]benzaldehyde trifluoroacetate
A solution of 4-(hydroxymethyl)benzaldehyde dimethyl acetal (FLUKA, 1 g, 5.49 mmol) in
27 mL of anhydrous DCM was cooled in an ice bath at O0C. Triethylamine (FLUKA, 1.15 mL, 8.23 mmol) and toluene-4-sulfonyl chloride (ALDRICH, 1.21 g, 6.37 mmol) were added and the reaction was stirred at room temperature for 5 hours. Water was added and the organic layer was washed, dried over MgSO4 and evaporated to dryness. The tosilate was immediately dissolved in 24 mL of dry acetonitrile, sodium bicarbonate (PANREAC, 1.17 g, 13.88 mmol) and N-methylpiperazine (ALDRICH, 1.27 mL, 11.52 mmol) were added and the resultant suspension was stirred at room temperature overnight. Reaction was filtered over Celite and the solvent was evaporated to dryness. The residue was purified using preparative HPLC (LUNA column 50x250 mm, gradient: 0% ACN-water, 0.1%TFA to 60% ACN-water, 0.1%TFA). 1H NMR (300 MHz DMSOd6) δ ppm: 10.05 (s, 1 H), 7.95 (d, 2H, J = 8.3 Hz), 7.60 (d, 2H, J = 8.1 Hz), 3.75 (s, 2H), 2.92- 3.22 (m, 6H), 2.83 (s, 3H), 2.37-2.49 (m, 2H). [ES+ MS] m/z 219 (MH)+.
Intermediate 56: W-({4-[(4-methyl-1 -piperazinyl)methyl]phenyl}methyl)-1 -butanamine
(M409/101/2)
The title compound was prepared by a method analogous to that described above for Intermediate 51 , using n-butylamine (ALDRICH) and Intermediate 55 as starting reagents.
1H NMR (300 MHz, DMSO-d
6) δ ppm: 8.82 (br s, 2H), 7.37-7.47 (m, 4H), 2.83-4.20 (m, 12H), 2.76 (s, 3H), 2.30-2.45 (m, 2H), 1.57 (qt, 2H, J = 6.0 Hz), 1.30 (sex, 2H, J = 7.3 Hz), 0.86 (t, 3H, J = 7.5 Hz). [ES+ MS] m/z 276 (MH)
+.
Intermediate 57: /V-(2-chloroethyl)-/V-(4-fluorophenyl)urea
A mixture of 2-chlordethylamine hydrochloride (ALDRICH, 2g, 17.2 mmol) and di-/so- propylethyl-amine (ALDRICH, 3.6 mL, 20.7 mmol) in THF (80 mL) was cooled down to O0C in an ice bath and 4-fluorophenyl isocyanate (ALDRICH, 2.35 mL, 20.7 mmol) was added dropwise. The solution was allowed to reach room temperature and was stirred for 5h. AcOEt was added and the organic phase was washed three times with 1 N NH4CI, brine, dried over MgSO4 and the solvent was evaporated. The residue was purified in silica with mixtures of hexane/AcOEt. 1H NMR (300 MHz DMSO-Of6) δ ppm: 8.70 (br s, 1 H), 7.41-7.45 (m, 2H), 7.07-7.13 (m, 2H), 6.41 (t, 1 H, J = 5.6 Hz), 3.69 (t, 2H, J = 6.2 Hz), 3.45 (q, 2H, J = 6.0 Hz). [ES+ MS] m/z 217 (MH)+.
Intermediate 58: 1-(4-fluorophenyl)-2-imidazolidinone
Intermediate 57 (2.71 g, 12.5 mmol) was dissolved in 35 mL of anhydrous THF and was added dropwise to a suspension of sodium hydride, 60% dispersion in mineral oil
(ALDRICH, 1.05 g, 26.3 mmol) in 50 mL of anhydrous THF. The resultant suspension was stirred at room temperature for 2h and at reflux for 40 minutes. The reaction was cooled, the solvent was evaporated and the residue was partitioned between DCM and water. Aqueous layer was extracted with more DCM and the combined organic phases were washed with brine, dried over MgSO
4 and evaporated. The residue was purified in silica with mixtures of hexane/AcOEt.
1H NMR (300 MHz DMSO-Qf
6) δ ppm: 7.56-7.63 (m, 2H), 7.15-7.22 (m, 2H), 6.99 (br s, 1 H), 3.84-3.89 (m, 2H), 3.40-3.45 (m, 2H). [ES+ MS] m/z 181 (MH)
+.
Intermediate 59: 1-amino-3-(4-fluorophenyl)-2-imidazolidinone
Intermediate 58 (1.84 g, 10.21 mmol) was dissolved in 37.5 mL of acetic acid and 3.5 mL of water. The solution was cooled to O0C in an ice bath and a solution of sodium nitrite (PANREAC, 940 mg, 13.62 mmol) in 2.7 mL of water was added dropwise. The reaction was stirred at O0C for 5 minutes and was diluted with 21 mL of water. The resultant precipitate was filtered, washed with water and dried. This product was dissolved in 15 mL of dioxane and 15 mL of 4N sulfuric acid. The solution was cooled to 1O0C in an ice-water bath and zinc powder (ALDRICH, 982 mg, 15 mmol) was added in portions. The suspension was stirred at 1O0C for 4h. 400 mg of zinc powder (ALDRICH, 6.1 mmol) were added and the reaction was stirred at room temperature overnight. The formed precipitate was filtered and was partitioned between DCM and brine. The organic layer was dried over MgSO4, filtered and evaporated to give the desired compound. 1H NMR (300 MHz DMSO-CZ6) δ ppm: 7.59-7.66 (m, 2H), 7.17-7.25 (m, 2H), 4.50 (br s, 2H), 3.71-3.76 (m, 2H), 3.45-3.50 (m, 2H). [ES+ MS] m/z 196 (MH)+.
Intermediate 60: 1 -(4-f luorophenyl)-3-[(2-methylpropyl)amino]-2-imidazolidinone
Intermediate 59 (0.475 g, 2.43 mmol) was dissolved in 2-propanol (20 mL) and iso- butylaldehyde (ALDRICH, 0.18 mL, 1.98 mmol) was added. The solution was refluxed for 2h and, then, cooled down to room temperature. Acetic acid (PANREAC, 0.534 mL) and platinum (IV) oxide (ALDRICH, 53 mg) were added and the mixture was hydrogenated at
30 psi overnight. Two more additions of platinum (IV) oxide were needed and after 72 h reaction was completed. Catalyst was filtered over celite, filtrate was evaporated and the residue was purified using preparative HPLC (LUNA column 50x250 mm, ACN:H2O, 0.1 %TFA, gradient 30-100%) to give the title compound. 1H NMR (300 MHz DMSO-Cf6) δ ppm: 7.58-7.65 (m, 2H), 7.19-7.26 (m, 2H), 3.74-3.79 (m, 2H), 3.50-3.55 (m, 2H), 2.70 (d, 2H, J = 6.9 Hz), 1.75 (qt, 1 H, J = 6.7 Hz), 0.95 (d, 6H, J = 6.6 Hz). [ES+ MS] m/z 252 (MH)+.
Intermediate 61 : 1-[(2,2-dimethylpropyl)amino]-3-(4-fluorophenyl)-2-imidazolidinone
The title compound was prepared by a method analogous to that described above for intermediate 60, using trimethylacetaldehyde (ALDRICH) as starting reagent. 1H NMR (300 MHz DMSO-CZ6) δ ppm: 7.58-7.65 (m, 2H), 7.16-7.25 (m, 2H) ), 4.82 (br s, 1 H), 3.72- 3.77 (m, 2H)1 3.48-3.53 (m, 2H), 2.66 (s, 2H), 0.94 (s, 9H). [ES+ MS] m/z 266 (MH)+.
Intermediate 62: 1-[(2-chloro-4-pyrimidinyl)(2-methylpropyl)amino]-3-(4-fluoro phenyl)-2-imidazolidinone
Intermediate 60 (53 mg, 0.21 mmol) was dissolved in 2 mL of 2-propanol. 2,4- dichloropyrimidine (ALDRICH, 63 mg, 0.42 mmol) and DIEA (ALDRICH, 73 μl_, 0.42 mmol) were added and the solution was refluxed for 48h. DIEA (ALDRICH, 73 μL, 0.42 mmol) was added and the reaction was refluxed for 24h. Solvent was evaporated, residue was dissolved in 2 mL of anhydrous dioxane and DIEA (ALDRICH, 73 μL, 0.42 mmol) was added. Reaction was refluxed for 72h. 2,4-dichloropyrimidine (ALDRICH, 63 mg, 0.42 mmol) and DIEA (ALDRICH, 73 μL, 0.42 mmol) were added and the solution was refluxed for 24h more. Solvent was evaporated and the residue was dissolved in DCM and washed with NH4CI 1 N (3x), and brine. The organic phase was dried over MgSO4 filtered and evaporated. Crude product was purified in preparative HPLC (SunFire column 19x150 mm, gradient 30%ACN; 0.1 %TFA to 100%ACN, 0.1%TFA) to yield 9 mg (7%) of the title
compound as a yellow oil. 1H NMR (300 MHz CDCI3) δ ppm: 8.16 (d, 1H, J = 5.5 Hz), 7.46-7.53 (m, 2H), 7.02-7.10 (m, 2H), 6.50 (d, 1H, J = 6.0 Hz), 3.51-3.99 (m, 6H), 2.13 (qt, 1 H, J = 7.0 Hz), 0.98-1.04 (m, 6H). [ES+ MS] m/z 364 (MH+).
Intermediate 63: 1 -[(2-chloro-4-pyrimidinyl)(2,2-dimethylpropyl)amino]-3-(4-f luoro phenyl)-2-imidazolidinone
The title compound was prepared by a method analogous to that described for Intermediate 62 using Intermediate 61 as starting material. 1H NMR (300 MHz CDCI3) δ ppm: 8.19 (d, 1H, J = 6.1 Hz), 7.46-7.53 (m, 2H), 7.02-7.10 (m, 2H), 6.48 (d, 1H, J = 6.2 Hz), 3.46-4.01 (m, 6H), 1.06 (m, 9H). [ES+ MS] m/z 378 (MH)+.
Intermediate 64: 1,1-dimethylethyl {2-[(1,1-dimethylethyl)amino]ethyl}carbamate
Caesium hydroxide monohydrate (ALDRICH, 1.1 g, 6.8 mmol) was added to a suspension of molecular sieves 4A in anhydrous DMF under nitrogen atmosphere. The suspension was stirred 10 minutes, tert-butylamine (ALDRICH, 500 mg, 6.8 mmol) was added and the reaction was stirred 30 minutes at room temperature. Then, 2-(Boc-amino)ethyl bromide (FLUKA, 1.8 g, 8.6 mmol) was added and the suspension was stirred under nitrogen at room temperature overnight. Reaction was filtered, DMF was evaporated under vacuum and the residue was partitioned between AcOEt and NaOH 1N. The organic phase was dried with MgSO
4, and evaporated to dryness. The crude was purified in silica (DCM/MeOH):
1H NMR (300 MHz, DMSO) δ ppm: 6.82 (m, 1H), 3.06 (m, 2H), 2.69 (m, 2H), 1.37 (s, 9H), 1.12 (s, 9H). [ES+ MS] m/z 217 (MH)
+.
Intermediate 65: 1,1-dimethylethyl {2-[{[2-(2-cyano-4-pyrimidinyl)-2-(2-methyl propyl)hydrazino]carbonyl}(1,1-dimethylethyl)amino]ethyl}carbamate
To a solution of Intermediate 7 (30 mg, 0.16 mmol) in anhydrous THF (4 mL) under nitrogen atmosphere at O0C was added triphosgene (22 mg, 0.075 mmol). The resultant solution was stirred at O0C for 2 hours. Triethylamine (0.59 mL, 0.42 mmol) was added, the mixture was stirred 10 minutes and intermediate 64 (74 mg, 0.32 mmol) dissolved in THF (2 mL) was added dropwise. Reaction was allowed to reach room temperature and was stirred overnight. Solvent was evaporated and the residue was partitioned between DCM and 1 N NH4CI. The organic phase was washed with sat. NaHCO3, water and brine. The solvent was evaporated and the residue was purified in silica (DCM/MeOH). [ES+ MS] m/z 434 (MH)+.
Intermediate 66: 1,1-dimethylethyl {5-[1-(2-cyano-4-pyrimidinyl)-2-(cyclohexyl carbonyl)hydrazino]-4,4-dimethylpentyl}carbamate
Side-product isolated in the purification step of Example 223.
The compound was purified by preparative HPLC (ACN/H2O, from 50% to 100% of ACN) to give the title compound. 1H NMR (300 MHz, DMSO-d6) δ ppm: 10.48 (br.s, 1 H), 8.34 (d, 1 H), 6.74 (m, 1 H) 4.05 (m, 1 H), 2.99 (m, 1 H), 2.84 (m, 2H), 1.64-1.83 (m, 6H), 1.35 (s, 9H), 1.16-1.40 (m, 8H), 0.87 (s, 6H). [ES- MS] m/z 457 (M-H)".
Intermediate 67: 1,1-dimethylethyl {4-[(4-fluorophenyl)amino]butyl}carbamate
A mixture of 1-fluoro-4-iodobenzene (ALDRICH, 0.5 g, 1.17 mmol), N-(Boc)-1 ,4- butanediamine (FLUKA, 0.33 mL, 1.76 mmol), copper(l) iodide (ALDRICH, 22 mg, 0.117 mmol) and L-proline (ALDRICH, 14 mg, 0.117 mmol) in DMSO (3 mL) was heated at 6O
0C under nitrogen atmosphere overnight. Solvent was evaporated and the residue was dissolved with AcOEt and washed with 1 N NH
4CI and brine. The organic layer was dried with MgSO
4 and the solvent evaporated to afford crude product which was purified in silica (hexane/AcOEt).
1H NMR (300 MHz, CDCI
3) δ ppm: 6.84-6.92 (m, 2H), 6.49-6.56 (m, 2H), 4.55 (br s, 1 H), 3.07-3.19 (m, 4H), 1.53-1.68 (m, 4H), 1.44 (s, 9H). [ES+ MS] m/z 283 (MH
Intermediate 68: 1,1-dimethylethyl {4-[{[2-(2-cyano-4-pyrimidinyl)-2-(2-methyl propyl)hydrazino]carbonyl}(4-fluorophenyl)amino]butyl}carbamate
To a solution of Intermediate 7 (30 mg, 0.16 mmol) in anhydrous THF (4 mL) under nitrogen atmosphere at O0C was added triphosgene (22 mg, 0.075 mmol). The resultant solution was stirred at O0C for 2 hours. Triethylamine (0.059 mL, 0.42 mmol) was added, reaction was stirred 10 minutes and Intermediate 67 (90 mg, 0.32 mmol) in THF (2 mL) was added dropwise. Reaction was allowed to reach room temperature and was stirred overnight. Solvent was evaporated and the residue was partitioned between DCM and NH4CI 1 N. The organic layer was washed with sat. NaHCO3, water and brine. The solvent was evaporated and the residue was used in the following step without further purification). 1H NMR (300 MHz CDCI3) δ ppm: 8.25 (d, 1 H, J = 6.1 Hz), 7.32-7.45 (m, 2H), 7.19-7.26 (m, 2H), 6.68 (d, 1H, J = 6.3 Hz), 6.24 (br s, 1H), 4.61 (br s, 1H), 3.65-3.70 (m, 2H), 3.36 (br s, 1 H), 3.07-3.15 (m, 2H), 1.93 (m, 1 H), 1.49-1.56 (m, 4H), 1.41 (s, 9H), 0.90 (d, 6H, J = 6.7 Hz). [ES+ MS] m/z 500 (MH+).
Intermediate 69: 1,1-dimethylethyl {5-[(4-fluorophenyl)amino]pentyI}carbamate
The title compound was prepared by a method analogous to that described above for Intermediate 67, using N-Boc-1 ,5-diaminopentane (FLUKA) as starting reagent. 1H NMR (300 MHz1 CDCI3) δ ppm: 6.84-6.92 (m, 2H), 6.49-6.56 (m, 2H), 4.52 (br s, 1H), 3.04-3.16 (m, 4H), 1.47-1.68 (m, 6H), 1.44 (s, 9H). [ES+ MS] m/z 297 (MH)+.
Intermediate 70: 1,1-dimethylethyl {5-[{[2-(2-cyano-4-pyrimidinyl)-2-(2-methyl propyl)hydrazino]carbonyl}(4-fluorophenyl)amino]pentyl}carbamate
The title compound was prepared by a method analogous to that described above for intermediate 68, using intermediate 69 as starting reagent. 1H NMR (300 MHz CDCI3) δ ppm: 8.23 (d, 1 H, J = 6.1 Hz), 7.33-7.44 (m, 2H), 7.18-7.24 (m, 2H), 6.68 (d, 1 H, J = 6.1 Hz), 6.24 (br s, 1 H), 4.55 (br s, 1 H), 3.62-3.66 (m, 2H), 3.34 (br s, 1 H), 3.04-3.10 (m, 2H), 1.93 (m, 1 H), 1.23-1.61 (m, 15H), 0.89 (d, 6H, J = 6.4 Hz). [ES+ MS] m/z 514 (MH+).
Intermediate 71 : 1,1-dimethylethyl {6-[(4-fluorophenyl)amlno]hexyl}carbamate
The title compound was prepared by a method analogous to that described above for intermediate 67 using ferf-butyl N-(6-aminohexyl)carbamate hydrochloride (ALDRICH) as starting reagent. 1H NMR (300 MHz, CDCI3) δ ppm: 6.84-6.92 (m, 2H)1 6.49-6.56 (m, 2H), 4.49 (br s, 1 H), 3.03-3.15 (m, 4H), 1.32-1.66 (m, 17H). [ES+ MS] m/z 311 (MH)+. 1H NMR
(300 MHz, CDCI3-d6): 6.84-6,90 (m, 2H), 6.50-6.54 (m, 2H), 4.50 (br., 1 H), 3.02-3.15 (m, 4H), 1.35-1.67 (m, 17H).
Intermediate 72: 1,1-dimethyIethyl {6-[{[2-(2-cyano-4-pyrimidinyl)-2-(2-methyI propyl)hydrazino]carbonyl}(4-fluorophenyl)amino]hexyl}carbamate
The title compound was prepared by a method analogous to that described above for intermediate 68, using intermediate 71 as starting reagent. 1H NMR (300 MHz CDCI3) δ ppm: 8.24 (d, 1 H, J = 6.1 Hz), 7.38 (m, 2H), 7.19-7.25 (m, 2H), 6.69 (d, 1 H, J = 6.1 Hz), 6.17 (br s, 1H), 4.53 (br s, 1H), 3.62-3.67 (m, 2H), 3.34 (br s, 1H), 3.03-3.10 (m, 2H), 1.92 (m, 1 H), 1.23-1.64 (m, 17H), 0.89 (d, 6H, J = 6.6 Hz). [ES+ MS] m/z 528 (M
Intermediate 73: 1,1-dimethyIethyl (4-hydroxy-4-methylpentyl)carbamate
To a solution of 5-amino-2,2-dimethyl-pentanol (APOLLO, 2.39 g, 18.2 mmol) in DCM (40 mL) was treated at O0C with di-terf-butyl dicarbonate (ALDRICH, 5.02 ml, 21.84 mmol). The reaction mixture was stirred at room-temperature for 2 hours, and them washed with water and brine, dried over MgSO4 and the residue was purified (silica gel 1 :1 hexane/ethyl acetate) to obtain the desired product. 1H NMR (300 MHz, DMSO-d6) δ ppm: 6.74 (m, 1 H), 4.40 (t, 1 H), 3.04 (d, 2H), 2.83 (m, 2H), 1.36 (s, 9H), 1.29 (m, 2H), 1.09 (m, 2H), 0.75 (s, 6H).
Intermediate 74: 1,1-dimethylethyl (5-{2-[(4-fluorophenyl)carbonyl]hydrazino}-4,4- dimethylpentyl)carbamate
A suspension of Intermediate 73 (3.86 g, 16.68 mmol), tetrabutylammonium chloride (FLUKA, 464 mg. 1.67 mmol), tetramethyl-1-piperidinyl-oxy (ALDRICH, 261 mg, 1.67 mmol), in a mixture 0.5N NaHCO3/0.05N K2CO3 (56 mL) and DCM (56 mL) was vigorously stirred at room temperature for 2 h. The organic layer was recovered, dried over MgSO4, and concentrated under vacuum to yield quantitatively the expected aldehyde that was used without further purification. A solution of the aldehyde (3.2 g, 14.2 mmol) in MeOH (48 mL) was treated with 4-fluorobenzhydrazide (ALDRICH, 2.2 g, 14.2 mmol) and stirring at room-temperature overnight. To this solution containing the intermediate hydrazone was added PtO2 and the suspension was hydrogenated at room temperature and 2.6 bar for 18 hours. The suspension was filtered and the solvent was removed under reduced pressure to give the title compound. 1H NMR (300 MHz, DMSO-d6) δ ppm: 9.17 (s, 1 H), 7.86 (dd, 2H), 7.28 (dd, 2H), 6.75 (m, 1 H), 4.60 (m, 1 H), 4.10 (m, 1 H), 2.86 (m, 2H), 1.36 (s, 9H), 1.11-1.23 (m, 4H), 0.82 (s, 6H).
Intermediate 75: 1,1-dimethylethyl (5-{1-(2-chloro-4-pyrimidinyl)-2-[(4-fluoro phenyl)carbonyl]hydrazino}-4,4-dimethylpentyl)carbamate
A mixture of Intermediate 74 (580 mg, 1.58 mmol), 2,4-dichloropyrimidine (ALDRICH, 353 mg, 2.37 mmol), N,N-diisopropylethylamine (0.5 mL, 3.16 mmol) and dry EtOH (9 ml) was stirred at room temperature for 5 days. The mixture was concentrated under reduced pressure and the residue partitioned between DCM and 1 M ammonium chloride. The organic layer was treated with brine and dried over MgSO4. The residue was purified by flash chromatography (eluant: Hex/AcOEt mixtures 97:3 to 50:50) to give the title compound. 1H NMR (300 MHz, DMSO-d6) δ ppm: 10.40 (m, 1H), 8.13 (d, 1H), 7.95 (m, 2H), 7.38 (m, 2H), 6.73 (m, 1 H), 6.53 (m, 1 H), 4.57 (m, 1 H), 4.05 (m, 1 H), 3.11 (m, 1 H), 2.84 (m, 3H), 1.63 (m, 2H), 1.35 (s, 9H), 0.86 (s, 6H).
Intermediate 76: 1,1-dimethylethyl (5-{1-(2-cyano-4-pyrimidinyl)-2-[(4-fluoro phenyl)carbonyl]hydrazino}-4,4-dimethylpentyl)carbamate
Potassium cyanide (128 mg, 1.96 mmol) was added to a suspension of Intermediate 75 (473 mg, 0.98 mmol) and DABCO (110 mg, 0.98 mmol) in a mixture of DMSO/H2O 85/15 (5 ml) at room temperature. The reaction mixture was stirred at 80 0C for 2 h, poured into ice water (15 ml). The white solid was filtered off and dried. The compound was purificated by preparative HPLC (ACN/H2O, from 50% to 100% of ACN) to give the title compound. 1H NMR (300 MHz, DMSO-d6) δ ppm: 11.19 (br.s, 1H), 8.36 (d, 1H), 7.97 (m, 2H), 7.39 (m, 2H), 6.94 (m, 1 H) 6.72 (br.s, 1 H), 4.13 (m, 1 H)1 3.66 (m, 1 H), 3.19 (m, 2H), 2.85 (m, 3H), 1.69 (m, 2H), 1.34 (s, 9H), 0.93 (s, 6H). [ES- MS] m/z 469 (M-H)".
Intermediate 77: Λ/1-(5-amino-2,2-dimethylpentyl)-ΛT-(2-cyano-4-pyrimidinyl)-4- fluorobenzohydrazide trifluoroacetate.
Intermediate 76 (47 mg, 0.1 mmol) was dissolved in a mixture of TFA (0.5 ml) in DCM (5 ml). The reaction mixture was stirred at room temperature for 2 hours. The mixture was treated with water (2 x 100 ml), 10% NaHCO3 (100 ml), dried over Na2SO4 and evaporated under reduced pressure. The compound was purificated by preparative HPLC (ACN/H2O 0.1% of TFA, from 10% to 100% of ACN) to give the title compound. 1H NMR (300 MHz, DMSO-d6: 11.23 (br.s, 1H), 8.35 (d, 1H), 7.98 (dd, 2H), 7.63 (m, 3H), 7.41 (dd, 2H), 6.96 (m, 1 H), 4.14 (m, 1 H), 3.24 (m, 2H), 2.73 (m, 2H), 1.34 (m, 4H), 0.98 (m, 6H). [ES+ MS] m/z 371 (MH)+.
Intermediate 78: 4-[1-(2)2-dimethylpropyl)hydrazino]-6-methyl-2-pyrimidine carbonitrile.
The title compound was prepared by a method analogous to that described for Intermediate 7, replacing Intermediate 5 with Example 70 and used in the next step without purification.
Intermediate 79: W-(3,3-dimethylbutyl)-4-fluorobenzohydrazide.
A solution of 3,3-dimethylbutyraldehyde (ALDRICH, 0.78 ml_, 6.22 mmol) in MeOH (22 ml) was treated with 4-fluorobenzoic hydrazide (ALDRICH, 959 mg, 6.22 mmol) and was stirred at room temperature overnight. To this solution containing the intermediate hydrazone was added PtO
2 and the suspension was hydrogenated at room temperature and 35 pSi for 12 h. The suspension was filtered and the solvent was removed under reduced pressure to give the title compound.
1H NMR (300 MHz, DMSO-d6) δ ppm: 10.04 (br.s, 1 H), 9.48 (s, 1 H), 7.85 (m, 2H), 7.30 (m, 2H), 2.93 (m, 1 H), 2.77 (m, 1 H), 1.36 (dd, 2H), 1.74 (m, 4H), 1.38 (s, 9H), 0.99 (m, 1 H). [ES+ MS] m/z 239 (MH)
+.
Intermediate 80: Λr-(2-chIoro-4-pyrimidinyl)-ΛT-(3,3-dimethylbutyl)-4-fluorobenzo hydrazide.
A mixture of Intermediate 79 (800 mg, 4.33 mmol), 2,4-dichloropyrimidine (ALDRICH, 1 g, 6.71 mmol), N,N-diisopropylethylamine (FLUKA, 1.2 ml, 6.71 mmol) and dry EtOH (12 ml) was stirred at room temperature for 5 days. The mixture was concentrated under reduced pressure and the residue partitioned between DCM and 1 M ammonium chloride. The organic layer was treated with brine and dried over MgSO
4. The residue was purified by flash chromatography (eluant: Hex/AcOEt mixtures 100:1 to 50:50) to give the title compound.
1H NMR (300 MHz, DMSO-d6) δ ppm: 11.10 (s, 1 H), 8.11 (d, 1 H), 7.99 (dd, 2H), 7.39 (dd, 2H), 6.66 (br.s, 1 H), 3.56 (m, 1 H), 3.32 (m, 1 H)
1 1.54 (t, 2H), 0.91 (s, 9H). [ES+ MS] m/z 351 (MH)
+.
Intermediate 81 : 1-[(2,2-dimethylpropyl)amino]-2,4-imidazolidinedione.
To a solution of 1-aminohydantoin hydrochloride (ALDRICH, 3 g, 20 mmol) in /-PrOH (80 mL), trimethylacetaldehyde (ALDRICH, 1.74 ml, 20 mmol) and 3A molecular sieves (2 g) were added and the resulting reaction mixture was then heated to reflux. After 24 hours, it seems the reaction has almost reached completion, and hence the reaction mixture was filtered. The filtrate was then added, under an inert atmosphere, to a suspension of platinum (IV) oxide (ALDRICH, 0.4 g) in /-PrOH (10 mL) to which glacial acetic acid (2 mL) had been previously added. The resulting reaction mixture was then hydrogenated at room temperature and 2.5 bar for 24 hours. The suspension was filtered and more catalyst (PtO2, 0.3 g) was added to the filtrate. The mixture was then hydrogenated at room temperature and 2.5 bar for further 4 h, before the reaction reached completion. The suspension was then filtered and the solvent was removed under reduced pressure. The crude reaction mixture was purified by flash chromatography (hex/EtOAc 2:1) to give the title compound. 1H NMR (300 MHz, DMSO-d6) δ ppm: 10.71 (br.s, 1H), 4.91 (m, 1H), 3.96 (S, 2H), 2.58 (s, 2H), 0.87 (s, 9H). [ES+ MS] m/z 186 (MH)+.
Intermediate 82: 1 -[(2-chloro-4-pyrimidinyl)(2,2-dimethylpropyl)amino]-2,4- imidazolidinedione.
To a solution of intermediate 81 (2.2 g, 12 mmol) in EtOH (40 ml_), 2,6-dichloropyrimidine
(ALDRICH, 2.2 g, 15 mmol) and N.N-diisopropylethylamine (FLL)KA, 2.5 mL, 15 mmol) were added and the resulting reaction mixture was refluxed for 5 days. Then, solvent was evaporated under reduced pressure despite the reaction not reaching completion. The crude mixture was then purified by flash chromatography (eluent DCM/MeOH mixtures
25:1 to 15:1), and then HPLC to give the title compound. 1H NMR (300 MHz, DMSO-d6) δ ppm: 11.30 (br.s, 1 H), 8.26 (d, 1 H), 7.00 (br.s, 1 H), 4.39- 4.24 (AB system, 1 H), 4.14- 4.03
(AB system, 1 H), 3.70- 3.49 (AB system, 1 H), 3.43- 3.34 (AB system, 1 H), 0.97 (s, 9H). [ES+ MS] m/z 298 (MH)+.
Intermediate 83: 1 -[(2-methylpropyl)amino]-2,4-imidazolidinedione.
To a solution of 1-aminohydantoin hydrochloride (ALDRICH, 3 g, 20 mmol) in /-PrOH (80 mL), /so-butyraldehyde (ALDRlCH, 1.46 ml, 20 mmol) and 3A molecular sieves (2 g) were added and the resulting reaction mixture was then heated to reflux. After 16 hours, it seems the reaction has almost reached completion, and hence the reaction mixture was filtered. The filtrate was then added, under an inert atmosphere, to a suspension of platinum (IV) oxide (ALDRICH, 0.4 g) in /-PrOH (10 mL) to which glacial acetic acid (2 mL) had been previously added. The resulting reaction mixture was then hydrogenated at room temperature and 35 p.s.i for 5 hours. The suspension was filtered and the solvent was removed under reduced pressure. The crude reaction mixture was purified by flash chromatography (hex/EtOAc 1:1) to give the title compound.
1H NMR (300 MHz, DMSO- d6) δ ppm: 10.70 (br.s, 1 H), 5.01 (m, 1 H), 3.95 (s, 2H), 2.58 (m, 2H), 1.62 (m, 1 H), 0.86 (d, 6H), [ES- MS] m/z 170 (M-H)-.
Intermediate 84: 1-[(2-chloro-4-pyrimidinyl)(2-methylpropyl)amino]-2,4- imidazolidinedione.
To a solution of intermediate 83 (0.5 g, 3 mmol) in /-PrOH (3 ml_), N1N- diisopropylethylamine (FLUKA, 0.9 ml_, 5 mmol) and 2,6-dichloropyrimidine (ALDRICH, 0.5 g, 3.3 mmol) were added. After stirring well, the resulting reaction mixture was irradiated with microwaves to a temperature of 150 0C. After 6 hours, solvent was evaporated under reduced pressure despite the reaction not reaching completion. The crude mixture was then purified by flash chromatography (eluant DClWMeOH mixtures 100:1 to 20:1), and then HPLC to give the title compound. 1H NMR (300 MHz, DMSO-d6) δ ppm: 11.25 (br.s, 1 H), 8.23 (d, 1 H), 7.01 (d, 1H), 4.25- 4.12 (AB system, 2H), 3.51 (d, 2H), 2.01 (m, 1 H), 0.92 (m, 6H). [ES- MS] m/z 282 (M-H)".
Intermediate 85: 4-fluoro-W-[(1 £)-3-methyIbutylidene]benzohydrazide
A solution of 4-fluorobenzhydrazide (ALDRICH, 1.00g, 6.5 mmol) in dry methanol (PANREAC, 23 mL) was added isovaleraldehyde (ALDRICH, 0.835 mL, 7.8 mmol). The mixture was stirred at room temperature overnight. The solvent was removed under reduced pressure and the oil crude was resuspended in DCM and extracted with brine (3x25 mL). The organic layer was dried over MgSO4, concentrated under vacuum to give the title compound. 1H NMR (300 MHz, DMSO-d6) δ ppm: 11.42 (s, 1 H), 7.91 (m, 2H), 7.72 (t, 1H), 7.32 (m, 2H), 2.15 (m, 2H), 1.84 (m, 1 H), 0.93 (d, 6H).
Intermediate 86: 4-fluoro-ΛT-(3-methylbutyl)benzohydrazide
A solution of intermediate 85 (1.31 g, 5.9 mmol) in dry methanol (PANREAC, 40 mL) was added PtO2 (134 mg), the mixture was hydrogenated at room temperature and 39 psi overnight. The reaction was filtered and the solvent was removed under reduced pressure to give the title compound. 1H NMR (300 MHz, DMSO-d6) δ ppm: 11.42 (br.s, 1 H)1 10.02 (br.s, 1 H), 7.87 (m, 2H), 7.28 (m, 2H), 2.78 (m, 2H), 1.65 (m, 1 H), 1.33 (m, 2H), 0.86 (d, 6H). [ES+ MS] m/z 225 (MH)+.
Intermediate 87: W-(2-chIoro-4-pyrimidinyl)-4-fluoro-W-(3-methylbutyl)benzo hydrazide.
A solution of intermediate 86 (441 mg, 1.97 mmol) in dry ethanol (PANREAC, 7 mL) was added 2,4-dichloropyrimidine (ALDRICH, 587 mg, 3.94 mmol) and N,N- diisopropylethylamine (LANCASTER, 0.858 mL, 4.92 mmol). The mixture was stirred at room temperature for 5 days, and concentrated under vacuum. The oil crude was resuspended in DCM1 extracted with ammonium chloride 1N (2x10 mL), H2O (10 mL), and brine (10 mL). The organic layer was dried over Na2SO4, concentrated under vacuum and the crude product is purified by flash chromatography (eluant: HexaneVλcOEt mixtures 99:1 to 50:50) to give the title compound. 1H NMR (300 MHz, DMSO-d6) δ ppm: 11.09 (br.s, 1H), 8.11 (d, 1H), 8.00 (m, 2H), 7.39 (m, 2H), 6.66 (m, 1H), 3.80 (m, 2H), 1.63 (m, 1H), 1.49 (m, 2H), 0.90 (d, 6H). [ES+ MS] m/z 337 (MH)+.
Examples
Example 1: 2-(2-Cyano-4-pyrimidinyl)-2-(2,2-dimethylpropyl)-Λ/-4-pyridinylhydrazine - carboxamide
A solution of 4-aminopyridine (ALDRICH, 85 mg, 0.9 mmol) in dry THF (ALDRICH, 10 ml) under nitrogen atmosphere was cooled to O0C. Triphosgene (93 mg, 0.31 mmol) was added and the reaction mixture was stirred for 5 min. Then, triethylamine (0.204 ml, 1.46 mmol) was added, the reaction mixture was stirred again for 5 min and, finally, Intermediate 8 (75 mg, 0.36 mmol) was added. The resultant suspension was stirred at room temperature for 6 h and concentrated under reduced pressure. The crude product was purified by flash chromatography (eluent: DCM:MeOH 90:1 to 50:1) to give the title compound. 1H NMR (300 MHz, DMSO-d6): 9.91 (br., 2H), 9.60 (br., 1H), 8.36-8.39 (m, 3H), 7.47 (m, 2H), 6.99 (br.s, 1 H), 4.11 (m, 1 H), 0.97 (s, 9H). [ES+ MS] m/z 326 (MH+).
Example 2: 2-(2-Cyano-4-pyrimidinyl)-W-(4-fluorophenyl)-2-(2-methylpropyl) hydrazinecarboxamide trifluoroacetate
Intermediate 7 (20 mg, 0.105 mmol) and 4-fluorophenyl isocyanate (ALDRICH, 0.014 ml, 0.126 mmol) were dissolved in DCM (1 ml). The reaction mixture was stirred at room temperature overnight. The reaction was diluted with DCM (3 ml) and 1 N hydrochloric acid was added (2 ml), the aqueous layer was extracted with DCM (2x2 ml). The combined extracts were washed with saturated NaHCO3, dried over MgSO4 and concentrated under vacuum. The crude product was purified by preparative HPLC (XTERRA 19x150 mm, ACN:H2O, 0.1 %TFA, gradient 30-100%) to give the title compound. 1H NMR (300 MHz, DMSO-d6): 9.05 (s, 1H), 8.99 (s, 1 H), 8.35 (d, 1 H), 7.45 (m, 2H), 7.09 (m, 2H), 6.98 (m, 1H), 3.96 (br.s, 1 H), 3.25 (br.s, 1 H), 2.04 (m, 1H), 0.91 (br.s, 6H). [ES+ MS] m/z 329 (MH+).
Examples 3-18 were prepared by methods analogous to that described for Example 2 replacing 4-fluorophenyl isocyanate with the isocyanates indicated in Table 1.
Table 1
Example 19: 2-(2-Cyano-4-pyrimidinyl)-2-(2,2-dimethylpropyl)-N
1,Λ/
1-bis[4-
(methyloxy)phenyl]-1 ,1 -hydrazinedicarboxamide
To a solution of Intermediate 8 (20 mg, 0.097 mmol) and triethylamine (0.054 ml, 0.388 mmol) in DCM (1 ml), 4-methoxyphenyl isocyanate (ALDRICH, 0.019 ml, 0.1455 mmol) was added at O0C. The reaction mixture was stirred at room temperature overnight. The reaction was diluted with DCM (3 ml) and 1 N hydrochloric acid (2 ml) was added, the aqueous layer was extracted with DCM (2x2 ml). The combined extracts were washed with saturated NaHCO3, dried over MgSO4 and concentrated under vacuum. The crude product was purified by preparative HPLC (XTERRA 19x150 mm, ACN:H2O, 0.1%TFA, gradient 40-100%) to give the title compound. 1H NMR (300 MHz, DMSO-d6): 10.08 (br.s, 1H), 8.49 (m, 1H), 7.34-7.37 (m, 5H), 6.88-6.93 (m, 4H), 3.80 (m, 1H), 3.72 (s, 3H), 3.68 (m, 1H), 1.04 (s, 9H). [ES+ MS] m/z 504 (MH+).
Examples 20 and 21 were prepared by methods analogous to that described for Example 18, replacing 4-methoxyphenyl isocyanate with the isocyanates indicated in Table 2.
Example 22: ΛP-(2-cyano-4-pyrimidinyl)-7-(methyloxy)-ΛP-(2-methylpropyI)-1 ■ benzofuran-2-carbohydrazide trifluoroacetate
Preparation of 7-methoxy-benzofuran-2-carbonyl chloride:
To a solution of 7-methoxy-benzofuran-2-carboxylic acid (AVOCADO, 75 mg, 0.375 mmol) in toluene at O0C was added thionyl chloride (0.12 ml, 0.75 mmol). The mixture was stirred under reflux for 4 hours. Solvent was removed under reduce pressure and the residue was used without any purification at the following step.
Intermediate 7 (36 mg, 0.188 mmol) and 7-methoxy-benzofuran-2-carbonyl chloride (prepared as above) were dissolved in pyridine (1 ml). The reaction mixture was stirred at room temperature overnight. 1N hydrochloric acid (3 ml) was added and the aqueous layer was extracted with DCM (2x2ml). The combined extracts were washed with saturated NaHCO3, dried over MgSO4 and concentrated under vacuum. The crude product was purified by preparative HPLC (XTERRA 19x150 mm, ACN:H2O, 0.1%TFA, gradient 30-100%) to give title compound. 1H NMR (300 MHz, DMSO-d6): 11.44 (s, 1H),
8.34 (d, 1H), 7.73 (s, 1H), 7.26-7.38 (m, 2H), 7.13 (m, 1H), 6.99 (br.s, 1H), 3.97 (s, 3H),
3.46 (br.s, 1 H), 1.97-2.06 (m, 1 H), 0.94 (d, 6H). [ES+ MS] m/z 366 (MH+).
Examples 23-28 were prepared by methods analogous to that described for Example 22 using Intermediate 7 or 8 and replacing 7-methoxy-benzofuran-2-carbonyl chloride with the acid chlorides indicated in Table 3.
Example 29: (2-ChIorophenyl)methyl 2-(2-cyano-4-pyrimidinyl)-2-(2,2- dimethylpropyl)hydrazinecarboxylate
To a solution of Intermediate 8 (40 mg, 0.195 mmol) and diisopropylethylamine (0.041 ml, 0.234 mmol) in DCM (1 ml) at O0C, DMAP (5 mg, 0.04 mmol) and 2-chlorobenzyl chloroformate (0.033 ml, 0.214 mmol) were added. The reaction mixture was stirred at room temperature for 48 h. The reaction was diluted with DCM (3 ml) and 1 N hydrochloric acid (2 ml) was added, the aqueous layer was extracted with DCM (2x2 ml). The combined extracts were washed with saturated NaHCO3, dried over MgSO4 and concentrated under vacuum. The crude product was purified by preparative HPLC (LUNA 19x250 mm, ACN:H2O, 0.1 %TFA, gradient 40-100%) to give the title compound. 1H NMR (300 MHz, DMSO-d6): 10.37 (br.s, 1 H), 8.41 (d, 1 H), 7.56-7.50 (m, 2H), 7.44-7.38 (m, 3H), 6.90 (br.s, 1 H), 5.23 (s, 2H), 4.04 (br.s, 1 H), 3.13 (br.s, 1 H), 0.94 (s, 9H). [ES+ MS] m/z 374 (MH+).
Example 30: 1,1-Dimethylethyl 2-(2-cyano-4-pyrimidinyl)-2-(2,2-dimethylpropyl)- hydrazinecarboxylate
The preparation of this compound is described above as Intermediate 6.
Example 31 : Λf-^-Cyano^-pyrimidinyO-Λ^-^-methylpropylJ-Λ/M- pyridinylglycinamide
To a mixture of Intermediate 15 (42 mg, 0.18 mmol), EDCI (FLUKA, 38 mg, 0.20 mmol) and HOBt (ALDRICH, 27 mg, 0.20 mmol) in DCM (1 ml) was added 4-aminopyridine (ALDRICH, 21 mg, 0.20mmol). The mixture was stirred under nitrogen atmosphere at room temperature overnight. The reaction was purified directly by flash chromatography (eluent: Hex/AcOEt mixtures 7:3 to 5:5) to give the title compound. 1H NMR (300 MHz, DMSO-d6) δ ppm: 10.55 (br.s, 1 H), 8.42 (d, 2H), 8.27 (br.d, 1 H), 7.53 (dd, 2H), 7.17 (d, 1H), 4.42 (S1 2H), 3.33 (d, 2H), 2.07-1.89 (m, 1H), 0.90 (d, 6H). [ES+ MS] m/z 311 (MH+).
Examples 32-37 were prepared by methods analogous to that described for Example 31 using Intermediate 15 or 16 and replacing 4-aminopyridine with the amines indicated in Table 4.
Table 4
Example 38: 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)-Λ/-[4- (trifluoromethyl)phenyl]hydrazinecarboxamide trifluoroacetate
The title compound was prepared by a method analogous to that described above for Example 1 replacing Intermediate 8 with Intermediate 7. 1H NMR (300 MHz, DMSO-d6): 9.29-9.42 (br.m, 2H), 8.33-8.38 (m, 3H), 7.43 (m, 2H), 7.00 (br.m, 1H), 3.90 (br.m, 1H), 3.65 (br.m, 1H), 1.98-2.07 (m, 1H), 0.90 (m, 6H). [ES+ MS] m/z 312 (MH+).
Example 39 was prepared by a method analogous to that described above for Example 1 replacing Intermediate 8 with Intermediate 7.
Table 5
Example 40: Λr-(2-cyano-4-pyrimidinyl)-ΛT-(2,2-dimethylpropyl)-4-[(4-methyl-1 - piperazinyl)methyl]benzohydrazide trifluoroacetate
To a solution of Intermediate 17 (50mg, 0.20 mmol) in DCM (10 ml) was added DCC (ALDRICH, 163 mg, 0.8 mmol) and stirred at room temperature for 1 hour. To this solution was added Intermediate 8 (45 mg, 0.22 mmol) and stirring at room temperature overnight. Then DCC (ALDRICH, 41 mg, 0.20 mmol) and triethylamine (ALDRICH1 0.31 μl, 0.22 mmol) were added and stirred at room 7 hours. After that, triethylamine (ALDRICH, 0.31 μl, 0.22 mmol) was added to give basic pH and the reaction was stirred at room temperature overnight. The reaction was evaporated and the product purified by column chromatography DCM/MeOH mixtures 100:0 to 0:100. The product was purified by preparative HPLC (Luna 21x250 mm, ACN:H2O, 0.1% TFA, gradient 25-100%) to give the title compound. 1H NMR (DMSO-d6, 300 MHz) δ ppm: 11.16 (m, 1 H), 9.45 (m, 1 H), 8.36 (m, 2H), 7.90 (d, 2H), 7.50 (d, 2H), 6.88 (br., 1 H), 4.09-4.16 (m, 1 H), 2.90-3.35 (br.m, 7H),
2.77 (s, 3H), 2.25-2.39 (m, 2H), 0.99 (s, 9H). [ES+ MS] m/z 422 (MH)+.
Example 41 : ΛP-(2-cyano-4-pyrimidinyl)-ΛT-(2,2-dimethylpropyl)-4-{[2-(4- morpholinyl)ethyl]oxy}benzohydrazide trifluoroacetate
The title compound was prepared by a method analogous to that described above for Example 40, replacing Intermediate 17 with Intermediate 18. 1H NMR (300 MHz, DMSO- d6): 11.08 (m, 1 H), 10.10 (m, 1 H), 8.36 (m, 1 H), 7.92 (d, 2H), 7.15 (d, 2H), 6.85 (br.m, 1H), 4.43-4.46 (br.m, 4H), 3.94-4.02 (br.m, 2H), 3.48-3.76 (br.m, 6H), 3.17-3.26 (br.m, 2H), 0.99 (s, 9H). [ES+ MS] m/z 439 (MH+).
Examples 42-44 were prepared by methods analogous to that described for Example 29 using Intermediate 7 and replacing 2-chlorobenzyl chloroformate with the chloroformate indicated in Table 6.
Table 6
Example 45: 2-methylpropyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazine- carboxylate
To a solution of Intermediate 7 (65 mg, 0.34 mmol) in pyiridine (5 ml) at O0C, isobutyl chloroformate (56 mg, 0.41 mmol) in THF (2 ml) was added dropwise. The reaction mixture was stirred at room temperature for 4 h. The reaction was concentrated under vacuum, diluted with DCM (5 ml), washed with citric acid (10%) and brine. The organic layer was dried over Na2SO4 and concentrated under vacuum. The crude product was purified by preparative HPLC (XTERRA 19x150 mm, ACNiH2O, 0.1%TFA, gradient 10- 100%) to give. the title compound. 1H NMR (300 MHz, DMSO-d6): 8.31 (d, 1 H), 6.81 (d, 1H), 6.73 (br.s,1 H), 3.95-3.98 (d, 2H), 3.60 (br.m, 2H)1 1.90-2.13 (m, 2H), 0.85-1.03 (m, 12H). [ES+ MS] m/z 292 (MH+).
Examples 46 and 47 were prepared by methods analogous to that described for Example 45 using Intermediate 7 and replacing isobutyl chloroformate with the chloroformate indicated in Table 6a.
Table 6a
Example 48: 1 ,2,2-Trimethylpropyl 2-(2-cyano-4-pyrimidinyl)-2-(2- methylpropyl)hydrazinecarboxylate
Triphosgene (ALDRICH, 202 mg, 0.68 mmol) was added to a stirred solution of 3,3- dimethyl-2-butanol (0.51 mmol) in dry THF at -5° C under nitrogen for 8 hours. Then TEA (0.128 mL, 0.92mmol) was added and stirred at 0° C. After 10 minutes, a solution of intermediate 7 (65 mg, 0.34 mmol) in pyridine was added and the mixture was stirred at room temperature for 4 hours. Solvent was removed under reduced pressure and the residue was dissolved in DCM and washed with citric acid 10 % and brine. The crude was purified by preparative HPLC (XTERRA 19x150 mm, ACN:H
2O, gradient 0-100%) to give
the title compound.
1H NMR (300 MHz, DMSO-d6): 8.30 (d, 1H), 6.79 (br.m, 1H), 4.67 (br.m, 1H), 2.06 (m, 1 H), 1.19 (br.m, 3H), 0.93-1.00 (m, 15H). [ES+ MS] m/z 320 (MH
+).
Examples 49-55 were prepared by methods analogous to that described for Example 48 using Intermediate 7 and replacing 3,3-dimethyl-2-butanol with the alcohol indicated in Table 7.
Table 7
Example 56: 1 ,1 -Dimethylethyl-2-(2-cyano-4-pyrimidinyI)-2-(2-methylpropyl)- hydrazinecarboxylate
The preparation of this compound is described above as Intermediate 5.
Example 57: 1,1-dimethylpropyl 2-(2-cyano-4-pyrimidinyl)-2-(2- methylpropyl)hydrazinecarboxylate.
A solution of Intermediate 7 (100 mg, 0.52 mmol) and carbonyldiimidazole (ALDRICH, 255 mg, 1.57 mmol) in dry THF (ALDRICH, 3 ml) under nitrogen atmosphere was warmed to reflux overnight. Then the mixture of reaction was cooled to room temperature
and a solution of 2-methyl-2-butanol (ALDRICH, 0.174 ml, 1.6 mmol) and diisopropylethylamine (0.273 ml, 1.5 mmol) in dry THF (ALDRICH, 3 ml) under nitrogen atmosphere was added. The new reaction mixture was warmed to reflux for 20 hours. The reaction was diluted with AcOEt (10 ml) and treated with ammonium chloride 1 M (3 x 15 ml) and brine (15 ml), dried over MgSO4 and evaporated under reduced pressure. The crude product was purified by preparative HPLC (LUNA 20x250 mm, ACN:H2O, gradient 40-100%) to give the title compound. 1H NMR (300 MHz, DMSO-d6): 9.88, 9.43 (br.m, 1 H, rotamers), 8.37 (d, 1H), 6.79 (m, 1 H), 3.82 (br.m, 1 H), 3.22 (br.m, 1 H), 1.94 (m, 1 H), 1.76 (m, 2H), 1.39, 1.30 (m, 6H, rotamers), 0.89, 0.60 (m, 9H, rotamers). [ES+ MS] m/z 306 (MH+).
Examples 57-61 were prepared by methods analogous to that described for Example 57 using Intermediate 7 and replacing 2-methyl-2-butanol with the alcohol indicated in Table 8.
Table 8
Example 63: 1,1-Dimethylethyl 2-(2-cyano-4-pyrimidinyl)-1-tnethyl-2-(2- methylpropyl)-hydrazinecarboxylate.
A mixture of Intermediate 5 (200 mg, 0.68 mmmol), tetrabutylammonium hydrogen sulfate (ALDRICH, 46 mg, 0.136 mmol), potassium carbonate (ALDRICH, 282 mg, 2.04 mmol), sodium hydroxide (PANREAC, 82 mg, 2.04 mmol) and iodomethane (ALDRICH, 171 μL, 2.04 mmol) in dry toluene (PANREAC, 5 ml) was stirred at room temperature for 24 hours. The reaction mixture was diluted with 20 ml of toluene, washed with 10 ml of water and 10 ml of brine, dried over sodium sulfate and evaporated to dryness. The residue was purified by PLC plates 20x20 (silica gel 60 F254, 2mm Merck 1.05717.0001 ) using dichloromethane/methanol 20:1 as eluent to give the title compound. 1H NMR (300 MHz, CDCI3) : 8.30 (d, 1 H), 6.50 (d, 1 H), 3.70 (br.s, 1 H), 3.43 (br.s, 1 H), 3.15 (br.s, 3H), 2.06 (m, 1H), 1.51-1.36 (m, 9H), 0.99 (m, 6H). [ES MS] m/z: 306 (MH+).
Example 64: 1,1-Dimethylethyl 2-(2-cyano-4-pyrimidinyl)-1-[3-
(dimethylamino)propyl]-2-(2-methylpropyl)hydrazinecarboxylate.
A mixture of Intermediate 5 (200 mg, 0.68 mmmol), tetrabutylammonium hydrogen sulfate (ALDRICH, 46 mg, 0.136 mmol), potassium carbonate (ALDRICH, 282 mg, 2.04 mmol), sodium hydroxide (PANREAC, 82 mg, 2.04 mmol), sodium iodide (FLUKA, 40 mg, 0.068 mmol) and 3-dimethylaminopropyl chloride hydrochloride (ALDRICH, 322 mg, 2.04 mmol) in dry toluene (PANREAC, 5 ml) was heated at 14O0C for 3 hours using a microwave oven SmithCreator Personal Chemistry. The reaction mixture was diluted with 20 ml of toluene, washed with 10 ml of water and 10 ml of brine, dried over sodium sulfate and evaporated to dryness. The residue was purified by PLC plates 20x20 (silica gel 60 F254, 2mm Merck 1.05717.0001) using dichloromethane/methanol 20:1 as eluent to give the title compound. 1H NMR (300 MHz, DMSOd6): 8.40 (d,1H), 6.81 (br.s, 1H), 3.53 (br.m, 4H), 2.20 (br.m, 2H), 2.08 (s, 6H)1 1.99 (br.m, 1 H), 1.67 (br.m, 2H), 1.43 (br.m, 4H), 1.26 (br.m, 5H), 0.93 (d, 6H). [ES MS] m/z: 377 (MH+).
Example 65: Butyl 2-(2-cyano-4-pyrimidinyl)-1-[3-(dimethylamino)propyl]-2-(2- methylpropyl)hydrazinecarboxylate
The title compound was prepared by a method analogous to that described above for Example 64, replacing Intermediate 5 with Example 44. 1H NMR (300 MHz, CDCI3): 8.27 (d, 1H), 6.59 (d, 1H), 4.09 (br.s, 2H), 3.57 (br.m, 4H), 2.31 (br.m, 2H), 2.21 (br.m, 6H), 2.08 (m, 1 H), 1.88-1.59 (m, 6H), 098 (m, 6H), 0.85 (br.m, 3H). [ES MS] m/z: 377 (MH+).
Examples 66-67 were prepared by methods analogous to that described for Example 22 using Intermediate 21 or 23 and replacing 7-methoxy-benzofuran-2-carbonyl chloride with the acid chlorides indicated in Table 3a.
Table 3a
Example 68: 1,1-dimethylethyl 2-(2-cyano-5-methyl-4-pyrimidinyl)-2-(2,2- dimethylpropyl)hydrazinecarboxylate
Potassium cyanide (0.415 g, 6 mmol) was added to a suspension of Intermediate 22 (1.05 g, 3 mmol) and DABCO (0.358 g, 3 mmol) in a mixture of DMSO/H
2O 9:1 (14 mL) at room temperature. The reaction mixture was stirred at 80
0C overnight, then it was poured into iced water. The light pink solid that precipitated was filtered off, washed abundantly with water and dried under air. The compound was purified by flash chromatography (eluent: Hex/EtOAC mixtures 30:1 to 1:1) to give the title compound.
1H NMR (300 MHz, d
6- DMSO) δ ppm: 9.94 (br.s, 1H), 8.20 (s, 1H), 4.36 (d, 1H), 2.81 (d, 1H), 2.27 (s, 3H), 1.41 (br.s, 9H), 0.93 (s, 9H); [ES - MS] m/z 318 (M - H)\
Example 69: 1,1-dimethylethyl 2-(2-cyano-6-methyl-4-pyrimidinyl)-2-(2- methylpropyl)hydrazinecarboxylate
The title compound was prepared by a method analogous to that described above for Example 68, replacing Intermediate 22 with Intermediate 24. 1H NMR (300 MHz, d6- DMSO) δ ppm: 9.80 (br.s, 1 H), 6.63 (s, 1 H), 3.78 (br.s, 1 H), 3.28 (br.s, 1 H), 2.34 (s, 3H), 1.93 (m, 1 H), 1.44 (br.s, 9H), 0.87 (br.s, 6H); [ES+ MS] m/z 306 (MH+).
Example 70: 1,1-dimethylethyl 2-(2-cyano-6-methyl-4-pyrimidinyl)-2-(2,2- dimethylpropyl)hydrazinecarboxylate
The title compound was prepared by a method analogous to that described above for Example 68, replacing Intermediate 22 with Intermediate 25. 1H NMR (300 MHz, d6- DMSO) δ ppm: 9.82 (br.s, 1 H), 6.61 (s, 1 H), 3.97 (m, 1 H), 3.10 (m, 1 H)1 2.35 (s, 3H), 1.44 (br.s, 9H), 0.92 (s, 9H); [ES+ MS] m/z 320 (MH+).
Examples 71-133 were prepared by methods analogous to that described for Example 22 using Intermediate 7, 8 or 21 as indicated in Table 3b and the corresponding acid or acid chloride replacing 7-methoxy-benzofuran-2-carboxylic acid or 7-methoxy-benzofuran-2- carbonyl chloride with the acids or the acid chlorides indicated in Table 3b.
Table 3b
Example 126: Λ/I-(2-cyano-4-pyrimidinyl)-Λ/'-(2,2-dimethylpropyl)-4-[(4-propyl-1 ■ piperazinyl)methyl] benzohydrazide.
Intermediate 26 (1 g, 3.8 mmol) was dissolved in thionyl chloride (5 ml). The reaction mixture was stirred at room temperature for 18 hours. The solvent was evaporated in vacuo and the crude product was used without any further purification. To a stirred solution of the crude (817 mg, 2.91 mmoles) in dry THF (15 ml_), DIPEA (0.68 mL, 3.88 mmol), Intermediate 8 (200 mg, 0.97 mmol) and potassium tert-butoxide (152 mg, 1.36 mmol) were added and the resulting reaction mixture was stirred at room temperature for 18 hours. The mixture was filtered and the solvent was evaporated in vacuo. The crude reaction mixture was purified by HPLC (H
2O containing 0.1% TFA : ACN, 10:90 to 0:100) to give the title compound.
1H NMR (300 MHz, DMSO) δ ppm: 11.26 (s, 1 H), 8.35 (d, 1 H), 7.93 (d, 2H), 7.55 (d, 2H), 6.89 (m, 1 H), 4.11 (m, 1H), 3.91 (s, 2H), 3.49-2.52 (br, 11 H), 1.62 (m, 2H), 0.99 (s, 9H), 0.89 (t, 3H). [ES+ MS] m/z 450 (MH)
+.
Example 134: W-(2-cyano-4-pyrimidinyl)-W-(2,2-dimethylpropyl)-4-(1 -piperazinyl methyl) benzohydrazide.
To a stirred solution of Intermediate 32 (360 mg, 1.0 mmol) in ACN (5 mL), piperazine (ALDRICH, 104 mg, 1.2 mmol), potassium carbonate (276 mg, 2.0 mmol) and sodium iodide (10 mg, 0.07 mmol) were added and the resulting reaction mixture was stirred at room temperature for 48 hours. The mixture was filtered and the solvent was evaporated in vacuo. The crude reaction mixture was purified by HPLC (H2O containing 0.1% TFA : ACN, 10:90 to 0:100) to give the title compound. 1H NMR (300 MHz, DMSO) δ ppm: 11.21 (s, IH), 8.78 (br, IH), 8.36 (d, IH), 7.92 (d, 2H), 7.57 (d, 2H), 6.88 (br, IH), 3.86 (br, 2H), 3.19 (br, 5H), 2.79 (m, 4H), 0.99 (s, 9H)1 [ES+ MS] m/z 408 (MH)+.
Examples 135-142 were prepared by methods analogous to that described for Example 2 using Intermediates 7 or 8 and replacing 4-fluorophenyl isocyanate with the isocyanates indicated in Table 1a.
Table 1a
Examples 143-149 were prepared by methods analogous to that described for Example 29 using intermediate 7 and replacing 2-chlorobenzyl chloroformate with the chloroformates indicated in Table 6b.
Table 6b
Example 150: Cyclohexyl 2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazine carboxylate.
A solution of cyclohexanol (ALDRlCH, 0.063 g, 0.63 mmol) in dry THF (5 ml_) under nitrogen atmosphere was cooled at -5° C. Triphosgene (ALDRICH, 0.088 g, 0.47 mmol) was added and mixture was stirred for 1,5 hours at -5° C. Triethylamine (ALDRICH, 0.237 mL, 1.7 mmol) was added and the mixture was stirred for 10 minutes more. Finally, Intermediate 7 (0.065 g, 0.63 mmol) in pyridine (1mL) was added. The resultant mixture was warmed up to room temperature, diluted with DCM (5mL) and concentrated under vacuum. The resultant oil was again diluted in DCM and washed with citric acid 10% (10 ml) and brine. The organic layer was dried over Na2SO4 and concentrated under vacuum. The crude product was purified by preparative HPLC (XTERRA 19x150 mm, ACNiH2O, 0.1%TFA, gradient 10-100%) to give the title compound. 1H NMR (300 MHz, CDCI3) δ ppm: 8.30 (d, 1H), 6.80 (d, 1H) 6.65 (s, 1H) 4.74 (m, 1H) 3.99-3.30 (br, 2H) 2.06 (m, 1H) 2 -1.18 (m, 10H) 0.97 (d, 6H); [ES- MS] m/z 316 (M-H)".
Examples 151-158 were prepared by methods analogous to that described for Example 150 using Intermediate 7 and replacing cyclohexanol with the alcohols indicated in Table 7a.
Table 7a
Examples 159-175 were prepared by methods analogous to that described for Example 31 using Intermediate 15 or 16 and replacing 4-aminopyridine with the amines indicated in Table 4a.
Table 4a
Examples 176-182 were prepared by methods analogous to that described for Example 64 using Intermediates 5 or 6 and replacing 3-dimethylaminopropyl chloride with the alkylating reagents indicated in Table 8.
Table 8
Example 183: 1,1-dimethylethyl {3-[{[2-(2-cyano-4-pyrimidinyl)-2-(2- methylpropyl)hydrazino]carbonyl}(1,1-dimethylethyl)amino]propyl}carbamate
To a solution of Intermediate 7 (30 mg, 0.16 mmol) in anhydrous THF (4 ml_) under nitrogen at O0C was added triphosgene (22 mg, 0.075 mmol). The resultant solution was stirred at O0C for 2 hours. Triethylamine (59 μl, 0.42 mmol) was added, the mixture was stirred 10 minutes and Intermediate 40 (74 mg, 0.32 mmol) dissolved in THF (2 mL) was added dropwise. The mixture was allowed to reach room temperature and stirred overnight. The solvent was evaporated and the residue was partitioned between DCM and 1N NH4CI. The organic layer was washed with sat. NaHCO3, water and brine. The solvent was evaporated and the residue was purified in silica (DCM/MeOH) to obtain the desired compound. 1H NMR (300 MHz CDCI3) δ ppm: 8.97 (br s, 1H), 8.30 (d, 1H, J = 5.1 Hz),
6.87 (m, 1 H), 6.70 (d, 1 H, J = 6.1 Hz), 3.86 (m, 1H), 2.91-3.30 (m, 6H), 1.96 (m, 1 H), 1.69 (m, 2H), 1.37 (s, 9H), 1.34 (s, 9H), 0.91 (m, 6H). [ES+ MS] m/z 448 (MH)+.
Examples 184-202 were prepared by methods analogous to that described for Example 183 using Intermediate 7 and replacing intermediate 40 with the amines indicated in Table
9.
Table 9
Example 203: 4-[[3-(4-fluorophenyl)-2-oxo-1-imidazolidinyl](2-methylpropyl) amino]- 2-pyrimidinecarbonitrile.
Intermediate 62 (5 mg, 0.0137 mmol) was dissolved in 7 mL of a mixture DMSO/water 9:1. 1,4-Diazabicyclo[2.2.2]octane (ALDRICH, 4 mg, 0.037 mmol) and potassium cyanide (ALDRICH, 2 mg, 0.034 mmol) were added and the suspension was stirred for 24h. Ice- water was added over the reaction and the precipitate was filtered, washed with water and dried to give the title compound as a white solid. 1H NMR (300 MHz CD3OD) δ ppm: 8.38 (d, 1H, J = 6.3 Hz), 7.58-7.63 (m, 2H), 7.09-7.18 (m, 3H), 3.99-4.15 (m, 2H), 3.56-3.92 (m, 4H), 2.21 (m, 1H), 1.04-1.09 (m, 6H). [ES+ MS] m/z 355 (MH+).
Example 204: 4-{(2,2-dimethylpropyl)[3-(4-f luorophenyl)-2-oxo-1 -imidazolidinyl] amino}-2-pyrimidinecarbonitrile.
The title compound was prepared by a method analogous to that described for Example 203 using Intermediate 63 as starting material. [ES+ MS] m/z 369 (MH)+.
Example 205:. 4-[{3-[4-(methyloxy)phenyl]-2-oxo-1-imidazolidinyl}(2-methylpropyl) amino]-2-pyrimidinecarbonitrile.
The title compound was prepared by a method analogous to that described for Example 203 replacing 4-fluorophenyl isocyanate with 4-methoxyphenyl isocyanate (ALDRICH). [ES+ MS] m/z 367 (MH)+.
Example 206: W-(3-aminopropyl)-2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyI) hydrazinecarboxamide trifluoroacetate
Trifluoroacetic acid (ALDRICH) was dropped onto a solution of example 183 (45 mg, 0,1 mmol) in dichloromethane (5 mL). After 1 hour at room temperature, the solvent was evaporated and the residue was partitioned between dichloromethane and sat. NaHCO3.
The organic layer was dried over MgSO4, filtered and evaporated. 1H NMR (300 MHz
DMSO-CZ6) δ ppm: 8.92 (br s, 1H), 8.34 (d, 1H, J = 6.3 Hz), 7.51 (br s, 3H), 6.84 (m, 1H),
3.94 (m, 1 H), 3.05-3.11 (m, 3H), 2.73 (t, 2H, J = 7.6 Hz), 1.99 (qt, 1 H, J = 6.7 Hz), 1.64 (qt, 2H1 J = 6.9 Hz), 0.88 (m, 6H). [ES+ MS] m/z 292 (MH)+.
Examples 207-220 were prepared by methods analogous to that described for Example 206 replacing Example 183 with the Examples or Intermediates indicated in Table 10.
Table 10
Example 221: Methyl [5-({[2-(2-cyano-4-pyrimidinyl)-2-(2-methylpropyl)hydrazino] carbonyl}amino)pentyl]carbamate.
Example 212 (126 mg, 0.29 mmol) was dissolved in DCM (7 mL) and DIEA (ALDRICH, 0,152 mL, 0.87 mmol) was added. The reaction was cooled to O0C in an ice bath and methyl chloroformate (ALDRICH, 0,027 mL, 0.35 mmol) was added dropwise. Reaction was stirred while was allowed to reach room temperature. After 5h reaction was diluted with DCM and washed with sat. NaHCO3 and brine. Organic layer was dried with MgSO4, solvent was evaporated and desired product was obtained. 1H NMR (300 MHz DMSO-Cf6) δ ppm: 8.66 (br. s, 1H), 8.33 (d, 1 H, J = 6.3 Hz), 7.06 (m, 1H), 6.84 (m, 1H), 6.54 (m, 1H), 3.91 (m, 1 H), 3.49 (s, 3H), 2.88-3.10 (m, 5H), 1.98 (qt, 1 H, J = 6.6 Hz), 1.81 (s, 3H), 1.14- 1.41 (m, 6H), 0.82-0.91 (m, 6H). [ES+ MS] m/z 378 (MH)+.
Example 222: 1,1-dimethylethyl {5-[1-(2-cyano-4-pyrimidinyl)-2-(cyclohexyl carbonyl)hydrazino]-4,4-dimethylpentyl}carbamate.
The preparation of this compound is described above as Intermediate 66.
Example 223: 1,1-dimethylethyl (5-{1-(2-cyano-4-pyrimidinyl)-2-[(4-fluoro phenyl)carbonyl]hydrazino}-4,4-dimethylpentyl)carbamate.
The preparation of this compound is described above as Intermediate 76.
Examples 224-234 were prepared by methods analogous to that described for Example 221 replacing Example 212 and methyl chloroformate with the Examples or Intermediates and acylating reagents indicated in Table 11.
Table 11
Example 235: 1,1-dimethylethyl 2-(5-bromo-2-cyano-4-pyrimidinyl)-1-(3-bromo propyl)-2-(2-methylpropyI)hydrazinecarboxylate.
The title compound was prepared by a method analogous to that described above for Example 22, using Intermediate 78 and 4-fluorobenzoyl chloride (ALDRICH). [ES+ MS] m/z 342 (MH)+.
Example 236: ΛT-(2-cyano-4-pyrimiclinyl)-Λ/I-(3,3-dimethylbutyl)-4-fluorobenzo hydrazide.
Potassium cyanide (ALDRICH, 291 mg, 4.48 mmol) was added to a suspension of Intermediate 83 (786 mg, 2.24 mmol) and DABCO (ALDRICH, 252 mg, 2.24 mmol) in a mixture of DMSO/H2O 9:1 (10 ml) at room temperature. The reaction mixture was stirred at 80 0C for 2 h, poured into ice water (15 ml). The white solid was filtered off partitioned between water and DCM, collected by evaporation of the DCM layers and dried. The compound was purified by flash chromatography (eluant: Hex/AcOEt mixtures 100:0 to 0:100) to give the title compound. 1H NMR (300 MHz, DMSO-d6) δ ppm: 11.17 (br.s, 1 H),
8.31 (d, 1H), 8.00 (dd, 2H), 7.39 (dd, 2H), 6.93 (m, 1H), 4.03 (m, 1H), 3.58 (m, 1 H), 1.55
(t, 2H), 0.92 (s, 9H). [ES- MS] m/z 340 (M-H)".
Example 237: 4-[(2,2-dimethylpropyl)(2,4-dioxo-1 -imidazolidinyl)amino]-2- pyrimidinecarbonitrile.
To a solution of Intermediate 85 (16 mg, 0.5 mmol) in a 9:1 DMSO-water mixture (2 ml), potassium cyanide (90 mg, 1.4 mmol) and 1 ,4-diazabicyclo[2.2.2]octane (ALDRICH, 165 mg, 1.5 mmol) were added and the resulting reaction mixture was stirred at room temperature for 16 hours. Product was extracted from the reaction mixture with n-butanol several times and the combined organic layers were washed with brine and dried over anhydrous sodium sulphate. The crude product was then purified by preparative HPLC to yield the title compound as the corresponding trifluoroacetate salt. 1H NMR (300 MHz,
DMSO-d6): 11.35 (br.s, 1 H), 8.47 (d, 1 H), 7.31 (br.s, 1 H), 4.42- 4.28 (br., 1 H), 4.17- 4.08 (br., 2H), 3.46- 3.32 (br., 2H), 0.98 (s, 9H). [ES+ MS] m/z 289 (MH)+.
Example 238: 4-[(2,4-dioxo-1 -imidazoIidinyl)(2-methylpropyl)amino]-2- pyrimidinecarbonitrile.
To a solution of Intermediate 87 (54 mg, 0.19 mmol) in a 9:1 DMSO-water mixture (2 ml), potassium cyanide (35 mg, 0.5 mmol) and 1 ,4-diazabicyclo[2.2.2]octane (ALDRICH, 101 mg, 0.9 mmol) were added and the resulting reaction mixture was heated at 90
0C for 16 hours. Solvent was evaporated under vacuum and the crude mixture was then purified by preparative HPLC to yield the title compound as the corresponding trifluoroacetate salt.
1H NMR (300 MHz, CDCI3): 8.59 (br., 1H), 7.56 (br.s, 1 H), 7.03 (d, 1H), 4.86- 4.06 (AB system, 2H), 3.92- 3.81 (AB system, 1H), 3.66- 3.55 (AB system, 1H), 2.08 (m, 1 H), 1.00 (dd, 6H). [ES- MS] m/z 273 (M-H)
".
Example 239: 4-[(5,5-dimethyl-2,4-dioxo-1,3-oxazolidin-3-yl)(2-methylpropyl) amino]- 2-pyrimidinecarbonitrile.
A solution of Example 155 (148 mg, 0.39 mmol) in dry DCM (SCHARLAU, 15 ml.) at O0C was added trifluoroacetic acid (ALDRICH, 5 mL) dropwise. The reaction mixture was stirred at room temperature for 7 hours. Then 1-hydroxy-1 H-benzotriazol hydrate (ALDRICH, 58 mg, 0.43 mmol) and Λ/-(3-dimethylaminopropyl)-Λ/'-ethylcarbodiimide hydrochloride (EDCI) (ALDRICH, 83 mg, 0.43 mmol) were added. The new mixture was stirred at room temperature for 7 hours and the crude was extracted with ammonium chloride 1 N (15 ml), H2O, and brine. The organic extracts were dried over MgSO4, concentrated under vacuum and the crude product is purified by flash chromatography (eluant: DCM) to give the title compound. 1H NMR (300 MHz, DMSO-d6): 8.61 (d, 1 H), 7.54 (m, 1H), 3.65 (m, 2H), 1.95 (m, 1 H), 1.68 (s, 6H), 0.95 (d, 6H). [ES+ MS] m/z 304 (MH)+.
Example 240: W-(2-cyano-4-pyrimidinyl)-4-fluoro-W-(3-methylbutyl)benzo hydrazide.
A solution of Intermediate 93 (480 mg, 1.42 mmol) in DMSO (PANREAC, 4.5 mL) was added H2O (0.5 mL), 1 ,4-diazabicyclo[2.2.2]octane (ALDRICH, 160 mg, 1.42 mmol) and potassium cyanide (ALDRICH, 111 mg, 1.71 mmol). The mixture was stirred at room temperature for 2 hours, poured into ice water (40 mL). The solid was filtered off, dried, and resuspended in DCM (15 mL), extracted with H2O (15 mL), and brine (15 mL). The organic layer was dried over Na2SO4, concentrated under vacuum and the crude product is purified by flash chromatography (eluant: Hexane\AcOEt mixtures 99:1 to 1:99) to give
the title compound. 1H NMR (300 MHz, DMSO-d6) δ ppm: 11.16 (br.s, 1H), 8.32 (d, 1H), 8.01 (m, 2H), 7.40 (m, 2H), 6.94 (m, 1 H), 3.80 (m, 2H), 1.63 (m, 1 H), 1.52 (m, 2H), 0.91 (d, 6H). [ES+ MS] m/z 328 (MH)+.
Example 241 : 1,1-dimethylethyl 2-(2-cyano-5-methyl-4-pyrimidinyl)-2-(2-methyl propyl)hydrazinecarboxylate.
The preparation of this compound is described above as Intermediate 20.
Examples 242 and 243 were prepared by methods analogous to that described for Example 22 using Intermediate 7 and replacing 7-methoxy-benzofuran-2-carbonyl chloride with the sulfonic acid chlorides indicated in Table 3c.
Table 3c
Example 244: A/'-(2-cyano-4-pyrimidinyl)-W-[3-(dimethylamino)propyl]-W-(2-methyl propyl)-2-phenylacetohydrazide trifluoroacetate.
A mixture of Example 97 (150 mg, 0.485 mmol), tetrabutylammonium hydrogen sulfate (ALDRICH, 33 mg, 0.097 mmol), potassium carbonate (ALDRICH, 201 mg, 1.455 mmol), sodium hydroxide (58 mg, 1.455 mmol), sodium iodide (7.3 mg, 0.048 mmol) and dimethylaminopropyl chloride hydrochloride (230 mg, 1.455 mmol) in dry toluene (4 mL) was heated at 14O0C for 3.5 hours using a microwave oven SmithCreator Personal Chemistry. The reaction mixture was diluted with dichloromethane (20 mL), washed with water (10 mL), then, with brine (10 mL), dried over sodium sulfate and evaporated to dryness. The residue was purified by preparative HPLC to obtain the desired compound. 1H NMR (300 MHz, DMSOd6, 8O0C) δ ppm: 9.35 (bs, H); 8.41 (d, 2H), 7.46-6.86 (m, 5H); 4.07-3.35 (m, 6H); 3.10 (m, 2H); 2.76 (s, 6H); 2.28-1.81 (m, 3H); 0.99 (m, 6H).
Biological Assays
The compounds of this invention may be tested in one of several biological assays to determine the concentration of compound which is required to have a given pharmacological effect.
Determination of Falcipain-2, Falcipain-3, Vivapain-2, Cathepsin K Cathepsin S, Cathepsin L, and Cathepsin B proteolytic catalytic activity Assays for Falcipain-2, Falcipain-3, and Vivapain-2 are carried out with parasitic recombinant enzymes. Cathepsins K, S, L, and B are carried out with human recombinant enzymes. Standard assay conditions for the determination of kinetic constants used a fluorogenic peptide substrate, typically H-D-VLR-AFC (Falcipain-2, Falcipain-3, Vivapain-2), Z-FR-AFC (Cathepsin K, L, B), or KQKLR-AMC (Cathepsin S) and are determined in 100 mM sodium acetate, pH 5.5, containing 10 mM DTT and 0.5 mM CHAPS (Falcipain-2, Falcipain-3, Vivapain-2), and 100 mM sodium acetate, pH 5.5, containing 5 mM L-cysteine, 1mM CHAPS and 5mM EDTA (Cathepsin K, L, B), or 5OmM MES, pH 6.5, containing 0.5mM CHAPS, 1OmM L-CYS, 5mM EDTA (Cathepsin S). Stock substrate solutions are prepared at 20 mM in DMSO. The activity assays contained 30 uM substrate (Falcipain-2, Falcipain-3, Vivapain-2), 20 uM substrate (Cathepsin K), 25uM substrate (Cathepsin B), 5uM substrate (Cathepsin L), and 3OuM substrate (Cathepsin S).
All assays contained 1% DMSO. Independent experiments found that this level of DMSO had no effect on enzyme activity or kinetic constants. All assays are conducted at ambient temperature as end point assays being quenched after 60 minutes with the exception of Cathepsin S at 90 minutes, with 16.6 uM E-64 in 1% DMSO. Product formation (AFC or AMC) is determined from fluorescence (excitation at 405nM; emission at 53OnM, AFC, or excitation at 360 nM; emission at 460 nM, AMC) monitored with a LJL Aquest (Molecular Devices) fluorescent plate reader. In the case of kinetic reads (used in mechanism of action studies), the reaction is not quenched but is read in the plate reader every 3 minutes for approximately 90 minutes. In addition, the mechanism of action studies for Falcipain-2 utilize Z-LR-AMC as the substrate. Product formation is determined from the fluorescence of AMC, measured with a LJL Acquest (Molecular Devices) fluorescent plate reader (excitation at 36OnM; emission at 46OnM).
Inhibition studies Potential inhibitors are evaluated using the quenched read (endpoint) method.
Assays are carried out in the presence of variable concentrations of test compound. Reactions are initiated by addition of enzyme and substrate to wells containing inhibitor stamped in 100% DMSO. For endpoint assays, the reaction is quenched with the addition of E64. Dose response data is fit to an IC50 curve with preset fitting tools according to equation 1 :
y = a + (b-a)/(1+(1O*/10c)d) (1 )
where y is the response at a particular inhibitor concentration x, a is the minimum response value, b is the maximum response value, c is the IC50, and d is the slope of the IC50 curve. Assuming the compound is a competitive inhibitor, the apparent Ki can be calculated from IC50, as shown in equation 2:
IC50 = appK, (1 +[S]/ KM) (2)
where appK, is the apparent K1, S is the concentration of substrate, KM is the Michaelis binding constant for substrate, and K/ is the binding constant of a competitive inhibitor for free enzyme. For a more direct measurement of the Ki and the binding mechanism, we performed mechanism of action studies that included a titration of substrate and inhibitor with a kinetic read. If the progress curves for each of these kinetic assays are linear, the measured rates (v) were fit to equation 3:
v = VmS/ [(KM (1 + [I] /K1) + [S] ( 1+ [I] Z aK1)] (3)
where Vm is the maximum velocity, S is the concentration of substrate with Michaelis constant of KMl [I] is the concentration of inhibitor, K, is the binding constant of inhibitor for
free enzyme, and aKs is the binding constant of inhibitor for a potential enzyme-substrate complex.
For those compounds whose progress curves were nonlinear, with a decrease in enzyme activity over time characteristic of time-dependent inhibition, the progress curves were fit to equation 4 to yield the kobs-
[AMC] = vs t + (v0 - vss) [1 - exp (-kObst)] / kobs (4)
where [AMC] is the concentration of product formed over time t, vø is the initial reaction velocity and Vs is the final steady state rate. The kobs values were fit to equations 5 and 6, describing a one-step and two-step time dependent binding mechansim respectively:
.■ kobs = koff( 1 + [l]/appK,) (5)
kobs = koff + kon ([l]/( appK, + [l]) (6)
appK, = K1 (1+[S]/ KM) [I)
Equation 7 describes the apparent K| for competitive compounds and was substituted into equations 5 and 6 to generate the relevant binding constants from the fitting routine. In addition, the initial and final velocities were fit to equation 3 to further define the binding mechanism and potency. A complete discussion of this kinetic treatment has been fully described (Morrison et al., Adv. Enzymol. Relat. Areas MoI. Biol., 1988, 61, 201).
In vitro models to evaluate activity against bone metastasis
Compounds can be evaluated for their actvity against bone metastasis using published in vitro models as follows: prostate cancer bone metastases model. in rat (Liepe K. et al., (2005) Anticancer Research 25(2A), 1067-1073 and Neudert M. et al., (2003) International Journal of Cancer 107(3), 468-477); models of prostate and breast cancer metastases to bone in mice (Angelucci A. et al., (2004) International Journal of Oncology 25(6), 1713- 1720 and Sasaki A. et al., (1995) Cancer Research 55(16), 3551-3557); and other models in various species for evaluating bone metastasis (Rosol T. J. et al, (2003) Cancer. 97, 748-757).
Activity in cathepsin K assay
Examples 14, 32 and 46 of the present invention were found to have an IC50 of between 200nm and 100nm in the assay for cathepsin K.
All other exemplified compounds were found to have IC50 of 100nm or lower in the assay for cathepsin K.
Activity in falcipain-2 assay
All exemplified compounds were tested in the assay for falcipain-2 at two separate locations. Examples of compounds of the invention which were found to have an IC50 of 100nm or lower in at least one of the two assays for falcipain-2 include Examples 18, 22, 66, 94, 97, 103, 104, 106, 109, 110, 112, 116, 119, 122, 129, 130, 131, 132, 133, 143, 144, 146, 153, 157, 182, 214 and 234.