PH26228A - An intermediate for the preparation of phosphorous containing CoA reductase inhibitors - Google Patents

Description

. a bri _1- HX8b Ki : ud

PHOSPHORUS-CON1AINING HMG-CoA REDUCTASE 5

INHIBITORS, NEW INTERMEDIATES AND METHOD BB . J . “ } The present invention relates to new phosphorus-containing compounds which inhibit the activity of 3-hydroxy-3-methylglutaryl-coenzyme A | R reductase and thus is useful in inhibiting } 15 . cholesterol biosynthesis, to hypocholesterolemic : compositions containing such compounds, to new B intermediates formed in the preparation of such 4 compounds and to a method of using such compounds : for such purposes. :

BR

§ : F. M. Singer et al., Proc. Soc. Exper. :

Biol. Med., 102, 370 (1959) and F. H. Hulcher,

Arch. Biochem. Biophys., 146, 422 (1971) disclose 26 that certain mevalonate derivatives inhibit the : biosynthesis of cholesterol.

Endo et al in U. S. Patents Nos. 4,049,495, 4,137,322 and 3,983,140 disclose a fermentation ” oo . }

HX8b “2 product which is active in the inhibition of cholesterol biosynthesis. This product is called P compactin and was reported by Brown et al., : (J. Chem. Soc. Perkin I. 1165 (1976)) to have a OF complex mevalonolactone structure. be

GB 1,586,152 discloses a group of synthetic compounds of the formula :

CH

0 :

R,0 : +0 L Tr

Ry E : rs 26% : 15

Ry : in which E represents a direct bond, a Cy 4 alkylene bridge or a vinylene bridge and the various R's represent a variety of substituents.

The activity reported in the U.K. patent : is less than 1% that of compactin.

U. S. Patent Mo. 4,375,475 to willard et al discloses hypocholesterolemic and hypolipemic compounds having the structure

Ho & 0

H O vd 2 ; ¢ 5

R R

1 5 3 2 4

Rj { 1

BAD ORIGINAL 9

26228 2

HX8b ih -3= = wherein A is H or methyl; E is a direct bond," -CH,-, -CH,-CH,-, -CH,-CH,-CH,~ or -CH=CH-; Ry. R, ] : and R, are each selected from H, halogen, C,_, © alkyl, Cig haloalkyl, phenyl, phenyl substituted by halogen, C,_, alkoxy, C,_g alkanoyloxy, } © Cy_4 alkyl, or C,_, haloalkyl, and OR, in which R, i is H, C,_g alkanoyl, benzoyl, phenyl, halophenyl, 5 phenyl C,_, alkyl, C,_g alkyl, cinnamyl, C;_, uk : haloalkyl, allyl, cycloalkyl-C, ,-alkyl, adamantyl-C, ,-alkyl, or substituted phenyl :

C,.3~alkyl in each of which the substituents are selected from halogen, C,_, alkoxy, C,_, alkyl, or

Ci-4 haloalkyl; and the corresponding dihydroxy acids resulting from the hydrolytic opening of the lactone ring, and the pharmaceutically acceptable : salts of said acids, and the C,_, alkyl and ’ phenyl, dimethylamino or acetylamino substituted

C,.3-alkyl esters of the dihydroxy acids; all of the compounds being the enantiomers having a 4 R : 4 configuration in the tetrahydropyran moiety of the te, B trans racemate shown in the As fF, 198 7% J : j WO 84/02131 (PCT/EPS8 /00308), (bas d on tA / U. S. application Serial No. 443,668, filed hy ;

November 22, 1982, and U. S. application Serial y { No. 548,850, filed November 4, 1983), filed in the i : ~ name of Sandoz AG discloses heterocyclic analogs of mevalono lactone and derivatives thereof having the structure : i soe I

R, R : 4 . : 6 N 3

Ry

Co

HX8b ; yp. : ‘ L ! s } 4 i wherein one of R and Rj is and the . i

Rg

R5a l other is primary or secondary Ci-6 alkyl, Cag f cycloalkyl or phenyl=-(CH,) -. wherein R, is hydrogen, Ci-a alkyl, Ci-4 alkoxy, (except t-butoxy), trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy, v

Rg is hydrogen, Ci-3 alkyl, Cy-3 alkoxy. : trifluoromethyl, fluoro, chloro, phenoxy or - benzyloxy,

Re, is hydrogen, Ci-2 alkyl, Ci-2 alkoxy, fluoro or chloro, and mis 1, 2 or 3, with the provisos that both Rg and Rg, must be hydrogen when R, is hydrogen, Rg, must be hydrogen when Rg is hydrogen, not more than one of

R, and Rg is trifluoromethyl, not more than one of

Ry and Rg is phenoxy and not more than one of R, and R. is benzyloxy,

R, 1s hydrogen, C;_, alkyl, C5 _¢ cycloalkyl, Cia alkoxy (except t-butoxy). trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy,

R, is hydrogen, Cy_j alkyl, C,_, alkoxy, trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy, with the provisos that Rg must be hydrogen when R, is hydrogen, not more than one of

R, and Ry is trifluoromethyl, not more than one of

R, and Ry is phenoxy, and not more than one of R, and Ry is benzyloxy.

’ . : p : ro . ER © - ak 262 2R i vo HX8b pi -5- iad

X. is =-(CH,) - or -CH=CH- (n=0, 1, 2 or 3), 3

Re i =n 4 3 2 1 "

Z is -CH-CH,—C—CH,~-COOH IT ¥ 2 2 ol 5 OH du £ wherein R, is hydrogen or C,_ 5 alkyl in > . free acid form or in the form of a physiologically- Ke hydrolysable and -acceptable ester or a § lactone #1 thereof or in salt form. h

GB 2162-179-A discloses naphthyl analogues ) of mevalolactone useful as cholesterol . h biosynthesis inhibitors having the structure

Me Me $ ;

XN 2 B e a! ; wherein R, = 1-3C alkyl; : ' 2 is a gp. of formula 2, or Z,: :

H H i - (HCH, CHCH,, COOR,, = *+H

OH OH 0 : 0 : . ¢ : Ek (2;) (2,) :

R, = H, a hvdrolysable ester gp. or a : cation. : - vr wey I ro. Sti AEA

HX8b i fy i

European Patent No. 164-698-A discloses } i, preparation of lactones useful as anti-hyper- i cholesterolemic agents by treating an amide with ie an organic sulphonyl halide R>S0,X, then removing fe! the protecting group Pr. of

B

Rt Ry ; ‘ 0 5 | p

Pro 2.4 FR 50.X HOw! 0 Es

H NR-R —_— B : { OH 1 H 13 0 5 x: : wherein X = halo; :

Pr = a carbinol-protecting group; : rl = H or CHy; 4 r3, RY = H, 1-3C alkyl or phenyl-(1-3C j alkyl), the phenyl being optionally substituted by ) 1-3C alkyl, 1-3C alkoxy or halo; :

RZ = a group of formula (A) or (B): 0

NE rR ? 7c

CH, H ; 7 g

Hy R R 0d. .B :£ a (A) (B)

9 RO OR HX8b

I un LL

Q = R®-c- or R®-cH ; 2 ! bt

R™ = H or OH; eo

R = H or CHy; : a, b, c and d = optional double bonds; rR’ = phenyl or benzyloxy, the ring in each . case being optionally substituted by 1-3C alkyl or ly halo; r8, rR? = 1-3cC alkyl or halo; i

R® = 1-3C alkyl, phenyl or mono- or ) : di-(1-3C alkyl)phenyl.

Anderson, Paul Leroy, Ger. Offen. b

DE 3,525,256 discloses naphthyl analogs of i mevalonolactones of the structure i

Me Me ; 3 2) H OH i ]

H 2 - 7 s° !

Semc” GHCH CHCH COR TJ 3 ~g :

OH OH i ° C 1 °

R Q

Q’

I

,oo1 _ 1, 7 _ wherein R™ is alkyl, 2 =Q, Q ; R =H, or a : hydrolyzable ester group useful as inhibitors of . cholesterol biosynthesis and in treatment of . atherosclerosis. ,, ([,shcd a 2, 1989 (Xs

WO 8202-943, (based on U.S. application wf

Serial No. 460,600, filed January 24, 1983) filed in the name of Sandoz AG discloses mevalono-lactone analogues useful as hypolipoproteinaemic agents having the structure

HX8b . -8- he . $e :

X-2 R HL

RO 1 >a R ne 1 Ay 3 A

RO ' 4 x 1 Ry :

He wherein the two groups Ro together form a radical of formula i i 8 7 6 5 c=C-~-C=C or -(CH,) 4" d i

R, Ry i wherein R, is hydrogen, C,_, alkyl, g

Ci_g alkoxy, (except t-butoxy), trifluoromethyl, i fluoro, chloro, phenoxy or benzyloxy, bv

Ry is hydrogen, C;_; alkyl, Cy 4 alkoxry, : trifluoromethyl, fluoro, chloro, phenoxy or § benzyloxy, with the proviscs that not more than 2 one of R, and Ry is trifluoromethyl, not more than Cb one of R, and Rg is phenoxy, and not more than one of R, and Ry is benzyloxy,

Ry is hydrogen, C; alkyl, fluoro, chloro or benzyloxy, i R, is hydrogen, Ci-a alkyl, Ci-4 alkoxy, (except t-butoxy), trifluoromethyl, fluoro, chloro, phenoxy or benzyloxy,

Rg is hydrogen, C, 4 alkyl, Cy _5 alkoxy, trifluoromethyl, fluoro, chloro, phenoxy or : benzyloxy,

Re, is hydrogen, Ci.2 alkyl, Ci.2 alkoxy, fluoro or chloro, and with the provisos that not a0 omy

CL NAL

26 HX8b i -) ! 3 more than one of R, and Rg is trifluoromethyl, not more than one of R, and Rg is phenoxy and not more 5 than one of R, and Rg is benzyloxy,

ACH2)q ¢ 5 X is -(CH,) -, c=c I: -(CHy) 4 H wherein n is 0, 1, 2 or 3 and both g's are 0 or ; one is 0 and the other is 1,

Z is :

Re 5 4 3] 2 1 ~CH-CH,—(-CH, ~COOH 11

OH bu wherzin Rg is hydrogen or C, 4 alkyl, with ! the general proviso that -X-Z and the R, bearing phenyl group are ortho to each other; in free acid form or in the form of a physiologically-hydrolysable and acceptable ester or a § lactone thereof or in salt form.

Published Pee. 148d

European patent application 127,848-A (Merck & Co, Inc.) discloses derivatives of 3-hydroxy-5- \ : thia-w-aryl-alkanoic acids having the structural x ’ formula: fo

HO 0 :

Cs $0), :

E

HX8b . } -10- Ce wherein Z is: A ai rY rR? z Y 3 > | Ng3 TC -

R® ! nis 0, 1 or 2; \

E is -CH,-, ~CH,-CH, =, ~CH,~CH,~CH,~, i, -CH=CH-CH, =; or -CH,-CH=CH-; .

R;, R, and R, are, e.g., hydrogen, chloro, : bromo, fluoro, c,-alkyl, phenyl, substituted phenyl 8 or OR, in which R, is, e.g., hydrogen, !

C,_galkanoyl, benzoyl, phenyl, substituted : phenyl, C,_g¢lkyl, cinnamyl, C,_ghaloalkyl, allyl, " cycloalkyl-C,_jalkyl, adamantyl-C, _,-alkyl, or : phenyl €1-3 alkyl; x

RY, R® and r® are hydrogen, chloro, bromo, t fluoro or Ci-3 alkyl; and

X is, e.g., hydrogen, C; 4 alkyl, a cation derived from an alkali metal or is ammonium. :

Those compounds have antihypercholesterolemic activity by virtue of their ability to inhibit ! 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) ; reductase and antifungal activity. Ho

French, PRION BRRKEITIAN 00% A filed Veg on April 1, 1986,\(Sano i sA) discloses 3-carboxy-2-hydroxy-propane-phosphonic acid derivatives including salts thereof which are useful as hypolipaemic agents and have the formula:

26228 ua } -11- ;

R 0 OR

R,00C~-CH 12 id 3 i 1 2 2° J

OH OR, I o wherein Ry and R, = H, lower alkyl or optionally substituted aralkyl; . Ry and R, = H, lawer alkyl or optionally substituted aryl or aralkyl. kb

These comounds are disclosed as giving \ greater reductions in cholesterol, triglyceride and phospholipid levels than meglutol. published May 22 1959 !

European patent application 142,146-A, (Merck Bb : & Co., Inc) discloses mevinolin-like compounds of Lr the structural formula: 2] - . HO 0 i £0

E .

Z wherein: ~ rR! is, e.g., hydrogen or C, _4alkyl; : E is -CH,CH,, -CH=CH-~, or -(CH,) -i and

Z is .

HX8b . ~12-

LC

. .

PS . rR’ X bg =, ’

Rg ‘ 3 wherein X is -O- or -NR? wherein rR? is hydrogen or i

C1-3alkyls V

R" is C,_galkyl; and 3

R® is hydrogen or CH; . oo 2) x 3 rR} wherein r10, gl! and RL? are independently, e.d., hydrogen, halogen or C,_,alkyl; 3) 14 . 'n !

In accordance with the present invention, a there is provided phosphorus-containing compounds

262 HX8b cs 13 Gi

Fi which inhibit the enzyme 3-hydroxy-3-methylglutaryl- on coenzyme A reductase (HMG-CoA Reductase) and thus i are useful as hypocholesterolemic agents and ot " include the following moiety 2 hoi 9 I -P-CH, -CH~CH, -CO- L

X OH ” ( CH, ) n Pr : B ; wherein X is -0- or -NH-, n is 1 or 2 and 2 is a 4 "hydrophobic anchor". a bl

The term hydrophobic anchor as employed + herein refers to a lipophilic group which when . dh linked to the HMG-like upper side chain of the iN molecule by the appropriate linker ("X"), binds to 3 a hydrophobic pocket of the enzyme not utilized in binding the substrate HMG CoA, resulting in 8 enhanced potency relative to compounds where Z=H. >

In preferred embodiments, the compounds of the invention have the formula I ] 0 H :

I R-P-CH., -C~CH.,, ~CO,R® ~P-CH,-C-CH,-CO;

X OH i (CHy)p 2 including salts thereof, wherein R is OH, lower ) alkoxy or lower alkyl; ;

R® is H or lower alkyl;

HX8b ; ~-14-

X is -0- or -NH-; 3 n is 1 or 2; cy

Z is a hydrophobic anchor; | bo and including pharmaceutically acceptable v salts thereof. i

The terms "salt" and "salts" refer to basic - salts formed with inorganic and organic bases.

Such salts include ammonium salts, alkali metal Lo salts like, lithium, sodium and potassium salts iv (which are preferred), alkaline earth metal salts 5 like the calcium and magnesium salts, salts with : organic bases, such as amine like salts, e.g., a dicyclohexylamine salt, benzathine, N-methyl-D~ x glucamine, hydrabamine salts, salts with amino acids like arginine, lysine and the like. The 3 nontoxic, pharmaceutically acceptable salts are 5 preferred, although other salts are also useful, : e.g., in isolating or purifying the product. E

Examples of hydrophobic anchors which may be : included in accordance with the present invention include, but are not limited to p22 rt . gr! r% 10] (o (3 p22 Zs R2

262 _~ -15- EL r® I br 0 | Sb alkyl

R ’ ox i

R 6a 3 . Rr i ( Yq : : 10 wherein the dotted ,ines represent optional double : bonds, for example, i iN 0 Co 0 ’ g alkyl alkyl ' ' rR R2 ' i

R (R%%) R 6 (R°%) I

RS R i

Fo) alkyl alkyl

R° "LL ! f ' 6a 5 r62 25 R™%) R (R" gq -_er oT poem. J

HX8b » -16- Li jo i bh

Le

Fu hes ny

R Es o 0 — _alkyl ! alkyl . 1 or E

JD “X 4

R R (R™7) 3 v 0 y

R> (R62) ; q

L i . i wherein rt, RZ, r%2 and r2P may be the same or different and are each independently selected from 3 : H, halogen, lower alkyl, haloalkyl, phenyl, substituted phenyl or orY wherein RY is H, alkanoyl, benzoyl, phenyl, halophenyl, phenyl-lower alkyl, lower alkyl, cinnamyl, haloalkyl, allyl, cycloalkyl-lower alkyl, adamantyl-lower alkyl or substituted phenyl-lower . alkyl. : where Z is

: HX8b -17- a.

Ake g® bes . 5.18 0 | i be > alkyl 3 :

R 6a N

R of ( )q J 9 1 . ™ : R® and R°' are the same or different and ; are H, lower alkyl or OH; py 0 0 hi 6 . i" u a

R® is lower alkyl-C such as CH,-CH,-C-C-, ) CH, R 2 or arylCH,-; : r%2 is lower alkyl, hydroxy, oxo or halogen; L ~~ qis 0, 1, 2 or 3, and §

R’ is H or lower alkyl; E

Thus, the compounds of formula I encompass L 0 x

IA R-P-CH,-CH-CH,-CO,R 0 CH and C ] (CH) pn z ! . 0 ! 1 Xx

IB R-P-CH,-CH-CH,=CO,R

NH OH

(CHy)y,

Z .

The term "lower alkyl" or "alkyl" as employed herein alone or as part of another group includes

HX8Db i -18~ . ¥ both straight and branched chain hydrocarbons, containing 1 to 12 carbons in the normal chain, [ preferably 1 to 7 carbons, such as methyl, ethyl, ba propyl, isopropyl, butyl, t-butyl, isobutyl, I pentyl, hexyl, isohexyl, heptyl, 4,4~dimethyl- ef " pentyl, octyl, 2,2,4-trimethylpentyl, nonyl, decyl, undecyl, dodecyl, the various branched chain isomers thereof, and the like as well as such » groups including a halo-substituent, such as F, Br, Ls

Cl or I or CFy, an alkoxy substituent, an aryl # substituent, an alkyl-aryl substituent, a haloaryl i substituent, a cycloalkyl substituent, an alkyl- i cycloalkyl substituent, hydroxy, and alkylamino h substituent, an alkanoylamino substituent, an aryl- : carbonylamino substituent, a nitro substituent, a cyano substituent, a thiol substituent or an . alkylthio substituent. %

The term "cycloalkyl" as employed herein if alone or as part of another group includes \ saturated cyclic hydrocarbon groups containing 3 to 12 carbons, preferably 3 to 8 carbons, which : include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclodecyl and cyclododecyl, any of which groups may be substi- tuted with 1 or 2 halogens, 1 or 2 lower alkyl ; groups, 1 or 2 lower alkoxy groups, 1 or 2 hydroxy groups, 1 or 2 alkylamino groups, 1 or 2 alkanoyl- amino groups, 1 or 2 arylcarbonylamino groups, 1 or 2 amino groups, 1 or 2 nitro groups, 1 or 2 cyano groups, 1 or 2 thiol groups, and/or 1 or 2 alkyl- thio groups.

The term "aryl" or "Ar" as employed herein refers to monocyclic or bicyclic aromatic groups

HX8b + =19~ . containing from 6 to 10 carbons in the ring portion, such as phenyl, naphthyl, substituted l phenyl or substituted naphthyl wherein the Ie substituent on either the phenyl or naphthyl may ph be 1, 2 or 3 lower alkyl groups, halogens (Cl, Br I or F), 1, 2 or 3 lower alkoxy groups, 1, 2 or 3 hydroxy groups, 1, 2 or 3 phenyl groups, 1, 2 or 3 alkanoyloxy group, 1, 2 or 3 benzoyloxy groups, 1, § 2 or 3 haloalkyl groups, 1, 2 or 3 halophenyl - groups, 1, 2 or 3 allyl groups, 1, 2 or 3 cyclo- : alkylalkyl groups, 1, 2 or 3 adamantylalkyl groups, : i 1, 2 or 3 alkylamino groups, 1, 2 or 3 alkanoyl- : amino groups, 1, 2 or 3 arylcarbonylamino groups, : 1, 2 or 3 amino groups, 1, 2 or 3 nitro groups, 1, 2 or 3 cyana groups, 1, 2 or 3 thiol groups, and/or oo 1, 2 or 3 alkylthio groups with the aryl group A preferably containing 3 substituents.

The term "aralkyl", "“aryl-alkyl" or "aryl-lower alkyl" as used herein alone or as part of another group refers to lower alkyl groups as discussed above having an aryl substituent, such as benzyl.

The term "lower alkoxy", "alkoxy", or "aryloxy" or "aralkoxy" as employed herein alone or as part of another group includes any of the above lower alkyl, alkyl, aralkyl or aryl groups linked ] to an oxygen atom.

The term "lower alkylthio", "alkylthio", "arylthio" or "aralkylthio" as employed herein alone or as part of another group includes any of the above lower alkyl, alkyl, aralkyl or aryl groups

N linked to a sulfur atom.

HX8b » ~-20- . The term "lower alkylamino", talkylamino", "arylamino", "arylalkylamino" as employed herein a alone or as part of another group includes any of 5 the above lower alkyl, alkyl, aryl or arylalkyl po groups linked to a nitrogen atom.

The term "alkanoyl" as used herein as part ! of another group refers to lower alkyl linked to a carbonyl group. 3

The term "halogen" or "halo" as used herein > refers to chlorine, bromine, fluorine, iodine and

CFy, with chlorine or fluorine being preferred. : preferred are those compounds of formula 1 a which have the following structure

II R O CH H CH i

Ne Ne “Neo p . a :

CH, OH z i wherein R is OH, OLi; R® is Li or H;

X is 0 or NH; and

Z is rt RZ wherein rl is phenyl which includes an alkyl and/or halo substitutent g2a ! or rt is benzyloxy which includes a halo substituent; rR? and p42 are the same and are halogen or lower alkyl;

HX8b to -21- o rR! ly

Z may also preferably be i

Lo

OO i rR? a wherein rR! and R? are as defined immediately above with respect to the compound of formula II, or

RS. - 0 : Av % =z . : / 2

R> o wherein R° is H, CH, or OH and r® is - 0 #

ZN ~ ;

CH C, ¢ or (substituted)phenylmethyl \ 3 7 “hy 7 :

Cy R wherein R' is H or CH, . t

The compounds of formula I of the invention i may be prepared according to the following : reaction sequence and description thereof. t ae HX8 b -

Cl

I

I

Fe ais _ _ vt > > 2 = - —

Mm ™ ©

ON ~~ o PON = o 2 H “ iO nn zo i

NZ = ° 5 3

EQ pr 28 i n N nO a w > & 7 = H © ~N 0 a, 9 i

NG ez K © x i 0 pe: i > 0 ~ BE o=a-8 ~ ° A 1 ~ y 0 ’ o ma £2 © 5 [-4 oN [ag] z 3 i 8 Ten 1 : i x > i

Nu ~ 8] nN A 4 i 0 ~ x ~ —- 0 br > Q Q ww MN x e >0 = - Hog 0 Lx O uo—n 0 3 L — —- 1 wn 2 : ~ $m ~ = oz — 0 om 0 rd Q 3 TS MM Q won { > 0 UO | = ur he) £ 23 o=a-0 ee © = 50 Ve m= 9 > ~~ -

Z alamo . a <5 . \ 1 an gg 0 gL ~~ ~ I

J

7 ~ : . = ~— oO H > o

Z . © ~ = 4 a —_ © @ v ! ™ ™~ or n= n QO wu 0

Tr LU = O | 3 b= wi - Q p=U=U WN 2 ex = m © & © ved Q <l — oN g. > 0 [ 8g =1 22% 0) ee O18 = ca

Coe E- ~ red oN oN Q 7 v4 > peed

-23- HX8 b } - e fin, = - i > 3

A 4 — }

LJ] © SU oN —_~ ys o ™ ix

Q = hi = < = : v T° i = od bd

Oned-—0n / > . 5 wn hi ~N AN feo ’ ’ = © .

U V ,

I or ozZao-0 : % 3 a i © 3 — i >

A - i ~+ j= i a 0 b apf }

H +

Q Lo] 2 0

OQ O°] O-~

QO ~| cw :

Qo cod orl ; 1 [] TM \ xr ond Q . 1

QO a oad £2 ol > 0 ! ox ~ a. om i - —_ i 3 fl ©

Mm ox > ~~

- 24- HX8b pi a = iat oe oR 0 i

Le = of : L — 2 z 5 3 —- ¥ = : . 5 o 2 x

O o = ~ ~ 5 = +t Sy O : 2 8 x EEF 5 no a 1 5 - :

Q~= NN = . 7

P 7 ™~ U 3 = = . on c 2 Oo ‘

O “~ vet 1 iv ; 5 ~ c i

Oo=om X~0 — i = 3 3: g 0 ~ : 0 0 \ x re & OQ =n

A : 3,

A 3 wy e a 0 2 = — > f a > i © —e 8 § ot oH no o + ~ m 0 Hoa 3 fe O 0 —~ 0. rood 0 MH Be 40 @ > oo , - rd ‘

A 8 RY

QO Q 5 < — 8 32 a m = ~ 0 =

Sox Soe ~ $ - & : = = | 38 fa 3

Oo x Ze 5

Fl NE - cl = - oN ~ HN oo. —- = ia a = 0 0 | = T & = ~ | fa =] 0 1 © s Oo hed =~ it oN . bt

Mo 3

HX8b j -25- ‘ 2 ’

As seen in the above reaction sequence, 3 compounds of Formula I may be rrepared by : subjecting iodide A to an Arbuzov reaction by ik heating iodide A g | h

Gets 0-$1-C(CHj) 4

Cells .

A 1-CH,-C-CH,-CO, alkyl )

H i and phosphonite/phosphite III :

I11 R®-P(Oalkyl), 4 ’ ; wherein R® is lower alkyl or lower alkoxy, employing standard Arbuzov conditions and ’ procedures to form phosphinate/phosphonate IV I :

Iv R®-P—CH,—CH-CH,~C0,alkyl

Oalkyl SRC)

Cells Cells oo 25 Phosphinate/phosphonate IV is a novel compound and as such is a part of the present invention.

Phosphinate/phosphonate IV is then subjected to a phosphorus ester cleavage by treating a solution of compound IV in an inert organic solvent, such as methylene chloride, sequentially with bis(trimethylsilyl)trifluoroacetamide (BSTFA) and trimethylsilyl bromide, under an inert atmosphere

HX8b -26- such as argon to form the phosphinic acid VA where :

R® in IV is lower alkyl, that is, 8 o i 1 i

VA lower alkyl-P-CH,-CH-CH,-CO alkyl Fe

OH 0 __-Si-C(CHy);

CeHg CeHg i

L or phosphonic acid VB (wherein R? in IV is lower alkoxy), that is 2 :

VB HO-P-CH, -CH-CH, -C0 alkyl

OH ? _- Si-C(CHy )4

CegHg CeHg compounds VA and VB are novel intermediates ! and as such as part of the present invention. where phosphonic acid VB is obtained, it is esterified by treating VB in dry pyridine with : alcohol vC rPon (where RP is lower alkyl) and dicycl >hexyl carbodiimide and the resulting : reaction mixture is stirred under an inert atmosphere, such as argon, to form phosphonic meno alkyl ester VI

HX8b = -27- » 0 i

VI RPo-F-ca -CH~-CH.,CO, alkyl i 27% 2-72 0 _ SLCC) i

CgHg CeHg "

Ester VI or phosphinic acid VA is then dissolved in an inert organic solvent, such as, 3 methylene chloride, benzene or tetrahydrofuran (THF) and treated with trimethylsilyldiethylamine Ss and stirred under an inert atmosphere such as 4 argon; the mixture is evaporated and then dissolved = in methylene chloride (or other appropriate inert organic solvent). The resulting solution is cooled T to a temperature within the range of from about 0°C x to about 25°C, treated with oxalyl chloride and 5 then evaporated to give crude phosphonochloridate. ¥

The phosphonochloridate is dissolved in inert 3 organic solvent such as methylene chloride, q benzene, pyridine or THF; the solution is cooled to i a temperature within the range of from about -20°C ! to about 0°C and treated with \

B z- (CH, ) -¥H

Co employing a molar ratio of VI or VA:B of within the range of from about 0.5:1 to about 3:1 and ] preferably from about 1:1 to about 2:1, followed by triethylamine and catalytic 4-dimethylaminopyridine (DMAP) to form adduct VII

HX8b » -28~ I

L

0 i

VII rS A H bey -p—©C p—CH-CH,-CO,alkyl 2

X ? he > (CH), ~Si-C(CH3), H

Z CeHg CeHg wherein RC is lower alkyl or lower alkoxy. =

Compound VII is subjected to silyl ether cleavage 5 by treating a VII in an inert organic solvent such as tetrahydrofuran, with glacial acetic acid and 3 tetrabutylammonium fluoride to form ester VIII 3 : 3» 0 CA

VIII RC-P-CH, -CH-CH,~CO,R alkyl 1

B-CH,-CH-CH,=t92 y i

X OH RE

(CHy py ] 2 ) (R® = alkyl)

The ester VIII may then be hydrolyzed to the corresponding alkali metal salt or acid, that is, where R® is alkali metal or H by treatment with strong base such as lithium hydroxide in the \ presence of dioxane, tetrahydrofuran or other inert ‘ organic solvent, under an inert atmosphere such as argon, at 25°C, employing a molar ratio of base:ester VIII of within the range of from about : 1:1 to about 1.1:1 to form the corresponding alkali metal salt

HX8b -29- . 0 i

VIIIA R-P-CH,~CH-CH,~CO,alkali metal Es

X on £ ’ (CHp py fr . z ; wherein R is lower alkyl or lower alkoxy. b : #

Compound VIIIA may then be treated with strong 1 acid such as HCl to form the corresponding acid ’ ‘

VIIIB ” . 15 0 ;

VIIIB R-P-CH,~CH-CH,~CO,H :

X OH 5 (CH) ; 2

The ester VIII wherein R is lower alkoxy may be converted to the corresponding di-alkali metal salt by treating ester VIII with strong base at 50-60°C employing a molar ratio of base:ester

VIII of within the range of from about 2:1 to : : about 4:1 to form VIIIC 2 : ;

VIIIC alkali metalO-P-CH,-CH-CH,-CO,alkali metal

X OH

(CHy)p

HX8Db Pe -30- 5

The di-alkali metal salt VIIIC may be I converted to the corresponding acid wherein R is §

OH by treatment with strong acid such as HCl to 3 form VIIID i 4 0 ! i

VIIID HO-P- CH, -CH~CH, ~COoH -

X OH g i (CHy)p . z : i

The iodide starting material A may be : prepared starting with the bromide C on ;

C Br-CH,-CH-CH,C0,alkyl (prepared employing procedures as described ! in Tetrahedron Lett. 26, 2951 (1985)) which is dissolved in solution in dimethyl formamide i (DMF) with imidazole and 4-dimethylamino pyridine and the resulting solution is treated with t-butyldiphenyl silyl chloride under an inert atmosphere such as argon to form the silyl ether D

Cells Cols

N/ 051-C(CH; )g

D Br-CH,-CH-CH,CO,alkyl

HX8b Lp -31- H

Lf . 5

A solution of silyl ether D in an inert organic i solvent such as methyl ethyl ketone or DMF Be is treated with sodium iodide under an inert : atmosphere such as argon, to form iodide A. Ri

The starting compound B

B 2- (CH, ) XH . ) 5 may be prepared as described below depending upon o the definition of Z and X. 5

Thus, compounds of formula B wherein 2 is 5 rR + £3 : ie 3 p22 _ b rR? A 1h. and X is 0, that is, compounds of the structure rl B on 2a -

R ( CH, 'n OH i r2 may be prepared by treating aldehyde E

HX8b -32- Lo

Is

Er gl :

E a 2a . ir

R O »— (CH,), _,~CHO £ / bP,

N\

Rr? with a reducing agent such as lithium aluminum 8 hydride or sodium borohydride. re

Compounds of formula B where 2 is : : gt 3

RZ (5% Cf rR” » and X is N, that is compounds of the structure rR: p2 2a

R CH) "NH, ny: may be prepared by oxidizing the aldehyde E by treating E in solution with acetone with, for example, Jones reagent to form the acid F

Ltn

HX8b ath -33- i rl rs

F | E 2a

R (CH, ),,_,~COOH g wi 2 J 0 which in suspension with methylene chloride is : Bi treated with oxalyl chloride to form the KE corresponding acid chloride which is dissolved in B an inert organic solvent such as tetrahydrofuran, B and treated with a mixture of concentrated ammonium pe hydroxide in tetrahydrofuran to form an amide of i the structure J i

Bt rt Be

G | 0 - 2a 1] he

R (CHy Jpg ~C-NH, % }

Rr?

Amide G is then reduced to the corresponding amide

B2 by treating G with a reducing agent such as lithium aluminum hydride. starting compounds of formula B wherein 2 is

HX8b | Pe ~34- ” 3 = . 1 I.

R> :

R> Fe

H | i (R a)q alkyl b i and X is O or -NH-, that is, compounds of the L structure i tl ; 15H : i : -(CH, ),~¥H i (rq alkyl where X is O are disclosed by C. H. Heathcock et al, Cr

J. Org. Chem. 50, 1190 (1985). Compounds of formula H' were X is NH may be prepared by the : { reductive amination of

RS, 0 CHO

J : alkyl

Jl

R ;

R (82) q : (prepared as disclosed by C. H. Heathcock et al, supra)

. s } gad # te : da

HX8b oe -35- a a - - ’ . ff by treating J with ammonium acetate and sodium i cyanoborohydride in the presence of an alcohol - i solvent such as methanol. a starting compound of formula B wherein 2 is p rr . 5 Si kid 9 ; 5 oY: A } KE r2P RZ and X is O, that is, compounds of the structure

R22 rl L : 3 M rk LL bS (0703 (cH, ) ,-OH wv Ig ® . . go rR? BZ . o , { 2 1984 of pri published Ju. # } 25 are disclosed in WO 8402-903-A and GB 2,162,179A , y both filed in the name of sandoz. 7

Starting compounds of formula B wherein 2 R is } i - - t

:

HX8b vo ~36- " bed

He f g%2 rl ¥ (513; :

B

R2P RZ ; + it and X is NH, that is, compounds of the structure :

R22 rt

N 3 st (Fe r2P r? may be prepared by the reductive amination of the aldehyde O p22 rt 0 (0103 (CH,) CHO r2P RZ by treating O with ammonium acetate and sodium cyanoborohydride in the presence of an alcohol solvent such as methanol.

pts

HX8b ry : -37- : in

The compounds of the invention may be a prepared as racemic mixtures and may later be. a resolved to obtain the S-isomer which is preferred. However, the compounds of the invention may be prepared directly in the form of f their S-isomers as described herein and in the ’ es : working examples set out hereinafter. EB

The compounds of the invention are ¥ - inhibitors of 3-hydroxy-3-methyl-glutaryl coenzyme : 10 A (HMG-CoA) reductase and thus are useful in ’ inhibiting cholesterol biosynthesis as demonstrated by the following tests. 7 1) Rat Hepatic HMG-CoA Reductase . B

Rat hepatic HMG-CoA reductase activity is measured using a modification of the méthod described by Edwards (Edwards, P.A., et al., kb

J. Lipid Res. 20:40, 1979). Rat hepatic p microsomes are used as a source of enzyme, and the B enzyme activity is determined by measuring the E conversion of the 14~_pgmG-CoA substrate to : 14c_pevalonic acid. iE . a. Preparation of Microsomes L : 25 Livers are removed from 2-4 ) cholestyramine-fed, decapitated, Sprague Dawley : rats, and hcmogenized in phosphate buffer A ! (potassium phosphate, 0.04 M, pH 7.2; KCl, 0.05 M; : sucrose, 0.1 M; EDTA, 0.03 M; aprotinin, 500 KI units/ml). The homogenate is spun at 16,000 x g for 15 minutes at 4°C. The supernatant is removed and recentrifuged under the same conditions a second time. The second 16,000 x g supernatant is

HX8b = -38~ be spun at 100,000 x g for 70 minutes at 4°C. pelleted microsomes are resuspended in a minimum s volume of buffer A (3-5 ml per liver), and homogenized in a jlass/glass homogenizer. : c Dpithiothreitol is added (10 mM), and the x preparation is aliquoted, quick frozen in acetone/dry ice, and stored at -80°C. The specific activity of the first microsomal preparation was 0.68 nmole mevalonic acid/mg protein/minute. : b. Enzyme Assay

The reductase is assayed in 0.25 ml which y contains the following components at the indicated " final concentrations: E 0.04 M Potassium phosphate, pH 7.0 : 0.05 M KCl : ; 0.10 M sucrose 0.03 M EDTA - 0.01 M pithiothreitol 3.5 mM NaCl 1% Dimethylsulfoxide : 50-200 pg Microsomal protein : 100 ym 1%C-[DL]HMG-CoA (0.05 uCi, 30-60 mCi/mmole) . 2.7 mM NADPH (nicotinamide adenine dinucleotide phosphate)

Reaction mixtures are incubated at 37°C. Under conditions described, enzyme activity increases linearly up to 300 pg microsomal protein per reaction mixture, and is linear with respect to a hi a aa ORIGINAL J

Naame -

Le LF oo ~ HX8b ts -39- Eh i) incubation time up to 30 minutes. The standard incubation time chosen for drug studies is 20 : minutes, which results in 12-15% conversion of N

HMG-CoA substrate to the mevalonic acid product. = ‘1 [DL-]HMG-CoA substrate is used at 100 pM, twice Bi ) the concentration needed to saturate the’ enzyme ir under the conditions described. NADPH is used in iE excess at a level 2.7 times the concentration 8 required to achieve maximum enzyme velocity.

To " B

Standardized assays for the evaluation of inhibitors are conducted according to the following procedure. Microsomal enzyme is incubated in the presence of NADPH at 37°C for 15 . minutes. DMSO vehicle with or without test compound is added, and the mixture further } incubated for 15 minutes at 37°c. The enzyme . assay is initiated by adding 140 _pmG-CoA substrate. After 20 minutes incubation at 37°C x the reaction is stopped by the addition of 25 pl : of 33% KOH. 3H-mevalonic acid (0.05 uci) is eC added, and the reaction mixture allowed to stand 4 "at room temperature for 30 minutes. Fifty pl SN ii

HCl is added to lactonize the mevalonic acid. } > : 25 Bromophenol blue is added as a pH indicator to 7 monitor an adequate drop in pH. Lactonization is % allowed to proceed for 30 minutes at room OF temperature. Reaction mixtures are centrifuged for 15 minutes at 2800 rpm. The supernatants are : layered onto 2 grams AG 1-X8 anion exchange resin : 4 (Biorad, formate form) poured in 0.7 cm (id) glass 1 columns, and eluted with 2.0 ml H,0. The first 0.5 ml is discarded, and the next 1.5 ml is : .

HX8b ver 40 . collected and counted for both tritium and carbon - 14 in 10.0 ml opti-fluor scintillation fluid. I”

Results are calculated as nmoles mevalonic acid Le produced per 20 minutes, and are corrected to 100% 4 recovery of tritium. Drug effects are expressed. - as Igo values (concentration of drug producing 50% inhibition of enzyme activity) derived from composite dose response data with the 95% L confidence interval indicated. I conversion of drugs in lactone form to i their sodium salts is accomplished by solubilizing the lactone in DMSO, adding a 10-fold molar excess E of NaOH, and allowing the mixture to stand at room temperature for 15 minutes. The mixture is then i 35 partially neutralized (pH 7.5-8.0) using 1N HCl, : 3 and diluted into the enzyme reaction mixture. ] 2) cholesterol synthesis in Freshly Isolated Rat

Hepatocytes ee : Compounds which demonstrate activity as inhibitors of HMG-CoA reductase are evaluated for their ability to inhibit 14. _ acetate incorporation into cholesterol in freshly isolated rat hepatocyte suspensions using methods originally described by capuzzi et al. (Capuzzi, D.M. and Margolis, S..

Lipids, 6:602, 1971). a. Isolation of Rat Hepatocytes sprague Dawley rats (180-220 grams) are anesthetized with Nembutol (50 mg/kg). The abdomen is opened and the first branch of the portal vein is tied closed. Heparin (100-200 units) is injected directly into the abdominal x

HX8b Pr -41- Co vena cava. A single closing suture is placed on the distal section of the portal vein, and the portal vein is canulated between the suture and the first branching vein. The liver is perfused : at a rate of 20 ml/minute with prewarmed (37°C), Bh oxygenated buffer A (HBSS without calcium or ! magnesium containing 0.5 mM EDTA) after severing Lf the vena cava to allow drainage of the effluent. i

The liver is additionally perfused with 200 ml of prewarmed buffer B (HBSS containing 0.05% : : bacterial collagenase). Following perfusion with ’ buffer B, the liver is excised and decapsulated in 60 ml Waymouth's medium allowing free cells to : : disperse into the medium. Hepatocytes are isolated by low speed centrifugation for 3 minutes at 50xg at room temperature. Pelleted hepatocytes are washed once in Waymouth's medium, counted and assayed for viability by trypan blue exclusion. y

These hepatocyte enriched cell suspensions . routinely show 70-90% viability. SE b. }¥c-Acetate Incorporation into 3 . Cholesterol gE

Hepatocytes are resuspended at 5x10° cells ot i 25 per 2.0 ml in incubation medium (IM) [0.02 M ;

Tris-HCl (pH 7.4), 0.1 M KCl, 3.3 mM sodium FE citrate, 6.7 mM nicotinamide, 0.23 mM NADP, 1.7 mM : glucose-6-phosphate]. 5

Test compounds are routinely dissolved in 4 . 30 DMSO or DMSO:H,0 (1:3) and added to ‘the IM. Final A

DMSO concentration in the IM is < 1.0%, and has no & significant effect on cholesterol synthesis. .

HX8b -42-

Incubation is initiated by adding 14. _acetate (58 mCi/mmol, 2 uCi/ml), and placing the cell suspensions (2.0 ml) in 35 mm tissue es culture dishes, at 37°C for 2.0 hours. Following re incubation, cell suspensions are transferred to : - glass centrifuge tubes and spu. at 50xg for 3 : minutes at room temperature. Cell pellets are resuspended and lysed in 1.0 ml H,O0, and placed in n an ice bath. ¥

Lipids are extracted essentially as . described by Bligh, E. G. and W. J. Dyer, Can. J. >

Biochem. and Physiol., 37:911, 1959. The lower ¢ organic phase is removed and dried under a stream : of nitrogen, and the residue resuspended in (100 pl) chloroform:methanol (2:1). The total sample : is spotted on gilica gel (LK6D) thin-layer plates : and developed in hexane:ethyl ether:acetic acid A (75:25:1). Plates are scanned and counted using a 1

BioScan automated scanning system. Radiolabel in I the cholesterol peak (RF 0.28) is determined and expressed at total counts per peak and as a : percent of the label in the total lipid extract.

Cholesterol peaks in control cultures routinely contain 800-1000 cpm, and are 9-20% of the label : 25 present in the total lipid extract; results compatable with capuzzi, et al., indicating 9% of extracted label in cholesterol.

Drug effects (% inhibition of cholesterol synthesis) are determined by comparing % of label in cholesterol for control and drug treated cultures. Dose response curves are constructed from composite data from two or more studies, and

HX8b -43- | . "results are expressed as Ic, values with a 95% BE . confidence interval. -. 3) Cholesterol Synthesis in Human skin Fibroblasts 3

Compound selectivity favoring greater - ik inhibitory activity in hepatic tissue would be an attribute for a cholesterol synthesis inhibitor.

Therefore, in addition to evaluating cholesterol 5 synthesis inhibitors in hepatocytes, these Lh compounds are also tested for their activity as | “ inhibitors of cholesterol synthesis in cultured y fibroblasts. : a. Human Skin Fibroblast Cultures 3

Human skin fibroblasts (passage 7-27) are B grown in Eagles' minimal essential medium (EM) # containing 10% fetal calf serum. For each b experiment, stock cultures are trypsonized to E disperse the cell monolayer, counted, and plated in 35 mm tissue culture wells (5x10° cells/2.0 : 1 : ml). Cultures are incubated for 18 hours at 37°C in 5% €O,/95% humidified room air. Cholesterol i biosynthetic enzymes are induced by removing the serum containing medium, washing the cell monolayers, and adding 1.0 ml of EM containing ] ’ 1.0% fatty acid free bovine serum albumin, and I incubating the cultures an additional 24 hours. b. 14-_Acetate Incorporation into

Cholesterol : Induced fibroblast cultures are washed with

EMEM, 4, (Earle's minimal essential medium). Test compounds are dissolved in DMSO or DMSO:EM (1:3)

HX8b ~44~ (final DMSO concentration in cell cultures < 1.0%), added to the cultures, and the cultures preincubated for 30 minutes at 37°C in 5% €0,/95% humidified room air. Following preincubation with drugs, [1-1%ciNa acetate (2.0 pCi/ml, 58 mCi/mmole) is added, and the cultures reincubated for 4 hours.

After incubation, the culture medium is removed, and the cell monolayer (200 pg cell protein per culture) is scraped into 1.0 ml of H,O. Lipids in the lysed cell suspension are extracted into chloroform:methanol as described for hepatocyte suspensions. The organic phase is dried under nitrogen, and the residue resuspended in ) chloroform:methanol (2:1) (100 pl), and the total sample spotted on silica gel (LK6D) thin-layer plates, and analyzed as described for hepatocytes.

Inhibition of cholesterol synthesis is determined by comparing the percent of label in the cholesterol peak from control and drug-treated cultures. Results are expressed as Ico values, and are derived from composite dose response curves from two or more experiments. A 95% confidence interval for the Ig value is also calculated from the composite dose response curves.

A further aspect of the present invention is a pharmaceutical composition consisting of at least one .f the compounds of formula I in association with a pharmaceutical vehicle or diluent. The pharmaceutical composition can be formulated employing conventional solid or liquid vehicles of diluents and pharmaceutical additives of a type appropriate to the mode of desired administration. The compounds can be administered

HX8b . -45- cet by an oral route, for example, in the form of u tablets, capsules, granules or powders, or they oo can be administered by a parenteral route in the rh form of injectable preparations, such dosage forms gd containing from 1 to 2000 mg of active compound i : per dosage, for use in the treatment. The dose to be administered depends on the unitary dose, the symptoms, and the age and the body weight of the patient. : =

The compounds of formula I may be | Lo administered in a similar manner as known ; compounds suggested for use in inhibiting : hy cholesterol biosynthesis, such as lovastatin, in ph: mammalian species such as humans, dogs, cats and i the like. Thus, the compounds of the invention may : be administered in an amount from about 4 to 2000 A mg in a single dose or in the form of individual . doses from 1 to 4 times per day, preferably 4 to 5 200 mg in divided dosages of 1 to 100 mg, suitably } B 0.5 to 50 mg 2 to 4 times daily or in sustained release form. : 2 }

HX8b ~46—

The following working Examples represent preferred embodiments of the present invention.

Unless otherwise indicated, all temperatures are expressed in degrees Centigrade. Flash : chromatography was performed on either Merck 60 or whatmann LPS-1 silica gel. Reverse phase chromatography was performed on CHP-20 MCI gel resin supplied by Mitsubishi, Ltd.

Example 1 (S)-4-[[[4'-Fluoro-3,3',5-trimethyl(1,1'-biphenyl]- 2-y1 methoxy Jmethoxyphosphinyl]-3-hydroxy-butanoic acid, monolithium salt

A. N-(2,4-Dimethylbenzylidene)benzeneamine

Ref. Merck U. S. Patent No. 4,375,475, pg. 39.

A solution of freshly distilled 2,4-dimethylbenzaldehyde (Aldrich, 6.97 ml, 50 mmole) and distilled aniline (Aldrich, 4.56 ml, 50 mmole) in dry toluene (80.0 ml) was refluxed for 3.0 hours under argon in a flask equipped with a

Dean-Stark apparatus. The mixture was cooled, then evaporated in vacuo to a yellow oil. The crude oil was purified by Kugelrohr distillation’ (0.5 mm Hg, 160-180°C) to give 8.172 ¢ (78.1%) of desired title benzeneimine as a light yellow oil which crystallized on standing to a low melting solid. TLC (4:1) Hex-acetone, RF=0.67 and 0.77 (geometric isomers), U.V. and I.

HX8b -47- i

B. N a zy #2

H,C Pd ) oy

CH, 2

Ref. Merck U.S. Patent No. 4,375,475, pg. 39. BE

A mixture of Part A benzeneimine (6.0 g, | Ie 28.7 mmol) in glacial HOAc (144 ml) was treated L with palladium (II) acetate (6.44 g, 28.7 mmole) : and the clear, red homogeneous solution refluxed - under argon for one hour. The resulting turbid E mixture was filtered warm through a packed %" bed ! of Celite into 900 ml of H,O. Precipitated orange - solid was collected by filtration and dried 5 in vacuo at 65°C over P,Og for 16.0 hours to give I 10.627 g (85.5%) of desired title palladium br complex as an orange solid with m.p. = : Co 194°-196°C. (Literature m.p. of a recrystallized analytical sample = 203°-205°C). .

C. 4'-Fluoro-3,3',5-trimethyl(1,1'~ biphenyl ]-2-carboxaldehyde (1) Bromo [4-fluoro-3-methylphenyl]- ' magnesium

Ref. Merck U. S. Patent No. 4,375,475, PP- 37 and 38. ,

The title Part C(1l) Grignard reagent was prepared by adding 5-bromo-2-fluorotoluene (22.5 g, 60.9 mmole, Fairfield Chemical Co.) dropwise

HX8b -48-~ at a rate sufficient to maintain the reaction at reflux to stirred magnesium turnings (1.35 g, 55.4 mmole, 8.0 eq.) in dry Et,0 (70.0 ml). The reaction was initiated in an ultrasound device.

After bromide addition was complete, the mixture was stirred for one hour under argon at room temperature, refluxed for 15 minutes and then allowed to cool to room temperature. (2) 4'-Fluoro-3,3',5-trimethyl[1,1'~- biphenyl]-2-carboxaldehyde

In a second flask, a mixture of the Part B dipalladium complex (3.0 g, 6.92 mmole) and triphenylphosphine (14.52 g, 55.4 mmole, 8.0 eq.) in dry benzene (100 ml) was stirred at room temperature under argon for 30 minutes. Freshly prepared and filtered (glass wool plug) part C (1)

Grignard reagent was then added in one portion by means of a cannula to this solution and the mixture was stirred for 1.5 hours at room temperature under argon. 6.0 N HCl (35 ml) was added, the mixture stirred an additional hour at room temperature, then filtered through packed celite (%" bed). The filtrate was extracted with } 25 Et,0 (250 ml), the extract washed with brine (2 X 100 ml), dried over anhydrous MgSO, and evaporated in vacuo to give 13.35 g of a viscous orange oil which crystallized on standing. The crude orange solid was purified by flash chromatography on silica gel (700 9) eluting with hexane, followed by (95:5) hexane-Et,0. Product fractions were evaporated to give 1.507 g (89.9%) of desired title aldehyde as a light yellow solid with m.p. = he v (BAD ORIGINAL 9

‘ i

HX8b - ~49- = ! 72°-75°C) (Literature reports m.p. = 73°-74°C). pi

TLC: (95:5) Hex-Et,0, Rf = 0.40, U.V. and PMA. | =

D. 4'-Fluoro-3,3',5-trimethyl[(1,1'~ Bh biphenyl ]-2-methanol

A cooled (0°C, ice bath) solution of dry

Et,0 (15.0 ml) was treated with LiAlH, (259 mg, 6 6.82 mmole, 0.55 eq.) and the gray suspension ig k treater dropwise over 15 minutes with a solution B 10 . of the Part C aldehyde (3.0 g, 12.4 mmole) in dry ih

Et,0 (15 ml). The mixture was stirred at room ; temperature under argon for 30 minutes, then i cooled back to 0°C and quenched by sequential : dropwise addition of 260 ul HO, 260 pl of 15% . ;

NaOH and 780 pl H,O. The suspension was diluted with EtOAc, filtered through anhydrous Na,SO0, over " packed Celite (%" bed) and the colorless filtrate evaporated in vacuo to give 2.99 g (98.8%) of a { white solid. Trituration of the crude solid with Pe cold hexane and drying in vacuo afforded 2.467 g (81.6%) of desired title alcohol as a white solid with m.p. 102-103°C. TLC: (9:1) Hex-EtOAc, Rf = 0.24, U.V. and PMA. : 25, E. (S)-3-[[(1,1-Dimethylethyl)diphenyl- silyl]oxy]-4-(hydroxymethoxyphosphinyl)- : putanoic acid, methyl ester (1) (S)-4-Bromo-3-hydroxybutanoic acid, : methyl ester “(1)(a) [R-(R%,R*)]-2,3,4-trihydroxy- butanoic acid, calcium salt, hydrate

HX8b -50~-

Ref. Carbohydrate Research 72, pp. 301-304 (1979).

Calcium carbonate (50 g) was added to a solution of D-isoascorbic acid (44.0 g, 250 mmol) in H,0 (625 ml), the suspension cooled to 0°C (ice bath) and treated portionwise with 30% H,0, (100 ml). The mixture was stirred at 30°-40°C (oil . bath) for 30 minutes. Darco (10 g) was added and the black suspension heated on a steam bath until evolution of O, ceased. The suspension was filtered through celite, evaporated jh vacuo (bath temperature 40°C). The residue was taken up in

H,0 (50 ml), warmed on a steam bath and CH,OH was added until the solution was turbid. The gummy precipitated solid was collected by filtration and air dried to give 30.836 g (75.2%) of desired calcium salt as a powdery white solid.

TLC (7:2:1) iPrOH-NH,OH-H,0, Rf = 0.19, PMA. (1)(b) [S-(R*,S*)]~2,4~Dibromo-3- hydroxybutancic acid, methyl estore

Ref. Bock, K. et al., Acta scondinavica (B) 37, pp. 341-344 (1983) part (l)(a) calcium salt (30 g) was : 25 dissolved in 30-32% HBr in acetic acid (210 ml) and stirred at room temperature for 24 hours.

Methanol (990 ml) was then added to the brown solution and it was stirred overnight. The mixture was evaporated to an orange oil, taken up in CH,OH (75 ml), refluxed for 2.0 hours and evaporated.

The residue was partitioned petween EtOAc (100 ml) and H,0, the organic phase washed with H,0 (2x) and prine then dried over anhydrous Na, SO, and [Raion ma

HX8b oe -51- B "evaporated to give 22.83 g (90.5%) of crude 2 dibromide as a light orange oil. TLC (1:1) 4

EtOAc-Hex, Rf = 0.69, UV & PMA. 5 (1)(c) (S)-4-Bromo-3-hydroxybutanoic acid, methyl ester oo

Ref. the same as for preparation of (1)(b). 3

An argon purged solution of the dibromide aj (20.80 g, 75.4 mmol) and anhydrous NaOAc (21.0 g) in LL

EtOAc (370 ml) and glacial HOAc (37 ml) was & treated with 5% Pd/C (1.30 g) and the black p : suspension stirred under of H, (1 atm) B while monitoring H, uptake. After 2.0 gq hours H, uptake was complete, the mixture was 5 filtered through Celite, the filtrate washed with 5 saturated NaHCO, and brine then dried over : pr anhydrous MgSO, and evaporated to give crude : 2 dibromoester as a brown oil. The crude oil vas Bi "combined with another batch (starting from 36.77 g is of the dibromide) and vacuum distilled to give A be 25.77 g (61.3%) of desired title bromoester Co as.a clear oil with b.p. = 79°-80°C (1.0 mm Hg). . TLC (1:1) EtOAc-Hex, Rf = 0.44, PMA. :

Anal Calcd for CHgO,Br: C, 30.48; H, 4.60; :

Br, 40.56 | :

Found: C, 29.76; H, 4.50; Br, 39.86 (2) (s)-4-Bromo-3-[{(1,1-dimethylethyl)- diphenylsilyl]oxy]butanoic acid, methyl ester : " A solution of part E(1) bromohydrin (4.0 g, 20.4 mmol), imidazole (6.94 4, 5.0 eq.), and ‘

HX8b “52 4-dimethylamino pyridine (4-DMAP) (12 mg, 0.005 eq.) in dry DMF (40 ml) was treated with t-butyl- diphenylsilyl chloride (5.84 ml, 1.1 eq.) and the homogeneous mixture stirred overnight under argon at room temperature. The mixture was partitioned between 5% KHSO, and EtOAc, the organic phase washed with H,0 and brine, dried over anhydrous

Na,SO, and evaporated to give 9.32 ¢ (100%) of crude silyl ether as a clear, viscous oil. TLC (3:1) Hex-EtOAC, Rf silyl ether = 0.75, y.v. and

PMA. ’ (3) (S)-4-Todo-3-[[(1,1-dimethylethyl)- diphenylsilyl]oxy]butanoic acid, 5 methyl ester

A solution of the crude Fart E(2) bromide (9.32 g, 201 mmole) in methyl ethyl ketone (60 ml, dried over 48 sieves) was treated with sodium iodide (15.06 g, 100.5 mmole, 5.0 eq.) and the yellow suspension refluxed for 5.0 hours under argon. The mixture was cooled, diluted with EtOAc, filtered, the filtrate washed with dilute NaHSO4 (until colorless) and brine then dried over anhydrous Na,SO, and evaporated in vacuo to give 10.17 g of a yellow oil. The crude oil was purified by flash chromatography on silica gel (600 g) eluting with (3:1) Hexane-CH,Cl,. product fractions were combined and evaporated to give 7.691 gq (74.2%, overall yield for poth steps) of desired title iodide as a clear, colorless, viscous. 0il. TLC (3:1) Hex-ETOAcC, product. Rf = 0.75, U.V. and PMA. (Note: product iodide co-spots with starting bromide).

BAD ORIGINAL 9

HX8b -53- 4 (4) (S)-4-(Diethoxyphosphinyl)-3-{[(1,1- dimethyléthyl)diphenylsilyl]oxy]- * butanoic acid, methyl ester i

A solution of the iodide (7.691 g) in oo i . 5 triethyl phosphite (20 ml) was heated at 155°C " (oil bath) fc.- 3.5 hours under argon. The mixture was cooled and excess phosphite distilled off - in vacuo (0.5 mm Hg, 75°C) to leave a yellow oil , (~8.0 g). The crude oil was purified by flash nx chromatography on silica gel (400. g) eluting with (4:1) Hexane-acetone. Product fractions were B . evaporated to give 3.222 g (41.1%) of desired title " phosphonate as a clear, colorless, viscous oil. b

TLC (1:1) Hex-acetone, Rf = 0.51, U.V. and PMA. -

Additionally 2.519 g (61.1% corrected yield) of ) starting Part (3) iodide was recovered. 4 3 (5) (5)-3-[[(1,1-Dimethylethyl)diphenyl- b : . silyl]oxy]-4-phosphonobutanoic acid, methyl ester : A solution of the Part (4) phosphonate (9.85 g, 20.0 mmole) in dry CH,Cl, (60 ml) was treated sequentially with bistrimethylsilyltri- fluoroacetamide (BSTFA) (5.31 ml, 32.0 mmole, 1.6 oo 25 eq.) and trimethylsilyl bromide (TMSBr) (6.60 ml 50.0 mmole, 2.5 eq.) and the clear mixture stirred overnight under argon at room temperature. 5%

KHSO, (80 ml) was added and the mixture was extracted with EtOAc. The aqueous phase was saturated with NaCl and re-extracted with EtOAc.

The combined organic layers were washed with brine, dried over anhydrous Na,SO, and evaporated in vacuo to give crude title phosphonic acid as a

HX8b -54- © yiscous oil. TLC (7:2:1) iPrOH-NH,OH-H,0, Rf = 0.30, U.V. and PMA. (6) (s)-3-[[(1,1-Dimethylethyl)diphenyl- silyl]oxy]-4-(hydroxymethoxyphosphinyl)- putanoic acid, methyl ester _

Part (5) crude phosphonic acid (~20.0 mmole) in dry pyridine (25 ml) was treated with dried

CH,OH (over 3A sieves, 1.62 ml, 40.0 mmole, 2.0 eq.) and dicyclohexyl carbodiimide (DCC) (4.54 gm, 22.0 mmole, 1.10 eq.) and the resulting white suspension stirred under argon at room temperature overnight. Pyridine was removed in vacuo, then azeotroped with benzene (2 x 15 ml). The residual oil was dissolved in EtOAC, filtered and washed with 1.0 N HCl and brine, dried over anhydrous

Na, SO, and evaporated in vacuo to give 8.272 g of crude title ester as an oil containing a small amount of precipitated dicyclohexyl urea (pcu). TLC (7:2:1) iPrOH-NH,-OH H,0, Rf = 0.60,

U.v. and PMA.

F. (S)-4-([[4'-Fluoro-3,3',5-trimethyl- } (1,1'-biphenyl]-2-yl]methoxyphosphinyl]- 3-t-butyldiphenylsilyloxybutanoic acid, methyl ester

Part i crude phosphonic acid mono methyl ester (6.595 gm, ~14.7 mmole) was dissolved in dry

CH,Cl, (30 ml), treated with distilled trimethyl- silyldiethylamine (5.60 ml, 29.4 mmole, 2.0 ed.) and stirred under argon at room temperature for 1 hour. The mixture was evaporated in vacuo, chased with benzene (1 x 30 ml) and dried in vacuo.

HX8Db -55- 5

The light yellow viscous oil was dissolved in dry y

CH,Cl, (30 ml) and DMF (dried over 4R sieves, 2 5 drops), the clear solution cooled to -10°C i (salt/ice bath) and treated dropwise via syringe wh with distilled oxalyl chloride (1.41 ml, 16.2 A mmole, 1.1 eq.). Vigorous gas evolution was evident and the solution became deeper yellow in color. The mixture was stirred under argon at -10°C for 15 minutes then allowed to stir at room 3 temperature for 1. hour. The mixture was evaporated - in vacuo, chased with benzene (1 x 30 ml) and dried . in vacuo to give crude phosphonochloridate as a 3 yellow oil. ’

To a solution of the crude phosphono- 4 chloridate (~ 14.7 mmole) in dry CH,Cl, (10 ml) was added dropwise a solution of the Part D biphenyl alcohol (2.06 g, 8.43 mmole) in dry i pyridine (15 ml) and the resulting mixture stirred ; at room temperature under argon for 16 hours.

The mixture was evaporated to dryness and the residue partitioned between 5% KHSO, and EtOAc.

The organic phase washed with saturated NaHCO, and . brine then dried over anhydrous Na,SO, and evaporated in vacuo to give 8.290 ¢g of a brown } 25 oil. The crude product was purified by flash : chromatography on silica gel (370 g) eluting with (70:30) Hexane-acetone. Product fractions were combined and evaporated to give 3.681 g (66%) of the desired title phosphonate as a pale yellow oil. TLC (3:2) Hexane-acetone, Rf = 0.59, U.V. and

PMA.

HX8b -56-

G. (s)-4-[[[4'-Fluoro-3,3',5-trimethyl- : (1,1'-biphenyl]-2-yl])methoxyphosphinyl]- 3-hydroxybutanoic acid, methyl ester

A mixture of the Part F silyl ether (1.103

S g, 1.66 mmole) in dry THF (20.0 ml) was treated with glacial acetic acid (380 pl, 6.64 mmole, 4.0 eq.) and a 1.0 M tetrabutylammonium fluoride solution (4.98 ml, 4.98 mmole, 3.0 eq.) and the clear yellow solution stirred overnight at room temperature under argon. The mixture was ’ partitioned between cold H,0 and EtOAc, the organic phase washed with saturated NaHCO, and brine, dried over anhydrous Na,SO, and evaporated to a viscous yellow oil (1.174 g). The crude oil was purified by flash chromatography on silica gel (47 9) eluting with (85:15)

CH,Cl,-Acetone. Product fractions were evaporated to give 679 mg (93.1%) of desired title alcohol as a clear viscous oil. TLC (1:1) Hexane-~acetone, Rf 20 .= 0.41, U.V. and PMA.

H. (S)-4-[[[4'-Fluoro-3,3', 5-trimethyl- (1,1'-biphenyl]-2-yl]methoxy]methoxy= phosphinyl]-3-hydroxy-butanoic acid, monolithium salt } A solution of the Part G methyl ester (184 mg, 0.420 mmole) in dioxane (5.0 ml) was treated with 1.0 N LiOH (0.50 ml, 1.2 eq.) and the mixture stirred at room temperature under argon for 3 hours. The mixture was diluted with H,0, filtered through a 0.4 um polycarbonate membrane and evaporated in vacuo. The residue dissolved in H,0 (75 ml), frozen and lyophilized. The crude acid

HX8b ! -57 = . "was dissolved in a minimum amount of H,0 and wo chromatographed on a 100 ml bed of CHP-20 resin a eluting with a H,0/CH,CN linear gradient system. fe

Product fractions were evaporated, dissolved in o

H,0 (50 ml), filtered through 0.4 um polycarbonate membrane. and lyophilized to give 174 mg (89.1% based on weight of hydrate) of desired title . mono-lithium salt as a white solid. TLC (7:2:1) ra i PrOH-NH,OH-H,0, Rf = 0.58 , U.V. and PMA. i | 5

Anal Calcd for C, H,cOGPFLi +:1.95 moles H,0 (MW CB 465.46): C, 54.19; H, 6.26; F, 4.08; P. 6.65 bi . Found: C, 54.19; H, 6.21; F, 4.29; P, 6.43

H' NMR (400 HMz): i 6 1.74-2.08 ppm (2H, m, -PO(OCH,)CH,-) 2.30 (34, s, aromatic methyl) i 2.32 (3H, d, aromatic methyl a to fk fluorine, Jur? -2 Hz) 2.35-2.62 (2H, m, -CH,CO,Li) 2 2.46 (3H, s, aromatic methyl) 3.57 & 3.63 (3H, 2 doublets, -OP (OCH, )~, 2 diastereomers, Jy_.,= 10.3 } 25 Hz) 4.28 (1H, m, -CH,CH(OH)CH,CO,Li)

S

4.97 (2H, m, PhCH, OP (OCH, )R) 6.87-7.25 (5H, m, aromatic H's)

HX8b -58-

Example 2 (S)-4-[[[4'-Fluoro-3,3"', 5-trimethyl{l,1"- biphenyl ]-2-yl}methoxy]hydroxyphosphinyl]=3- hydroxybutanoic acid, dilithium salt

A solution of the Example 1 diester (374 mg, 0.853 mmole) in dioxane (8.0 ml) was treated with 1.0 N LiOH (2.6 ml, 3.0 eq.) and heated at 50°C (oil bath) for 5.0 hours under argon. A white precipitate was evident. The mixture was diluted with H,0 and filtered. The aqueous solution was extracted once with Et,O0, filtered through a 0.4 pm polycarbonate membrane and concentrated in vacuo. The crude product was chromatographed on CHP-20 resin (100 ml bed) eluting with a H,0/CH5CN linear gradient system.

Product fractions were evaporated in vacuo, taken up in H,0 (50 ml), filtered through a 0.4 pm polycarbonate membrane and lyophilized to give 260 mg (67.1% based on hydrate weight) of desired title : 20 di-lithium salt as a white solid. TLC (7:2:1).

PrOH-NH,OH-H,0, Rf=0.47, U.V. and FMA.

Anal Calcd for C,oH206PELL, + 1.77 moles H,0: c, 52.88, H, 5.67; F, 4.18; P, 6.82 ,5 Found: C, 52.88; H, 5.26; F, 4.24; P, 6.43 a.

H! NMR (400 MHz, CD,OD):

J

6 1.69 ppm (2H, m, -OPCH, CH(OH)-) 2.26-2.42 (2H, m, CH,CO,L1) 2.30 (34H, s, aromatic methyl) 2.31 (3H, 4, aromatic methyl «a to F, Jgp=1-9 Hz)

HX8b PE -59- Noi 2.38 (3H, s, aromatic methyl) i 4.22 (1H, m, -CH(OH)CH,-) w g 2 4.75 (2H, m, PhCH, OP-) of © 6.86-7.23 (5H, m, aromatic protons)

Example 3 by (35)-4-[[[4'-Fluoro-3,3',5-trimethyl[1,1'-biphenyl]- EB . 10 2-y1 methoxy ]methylphosphinyl]-3-hydroxybutanoic 5 i acid, monolithium salt _ .

A. (S)-4-{(Chloro)methylphosphinyl]-3- ry : [[(1,1-dimethylethyl)diphenylsilyl]oxy]- io butanoic acid, methyl ester %

The title phosphinochloridate compound is 4 prepared as described in Example 6 Part B first gs three paragraphs. :

B. (35)-4-[[[4'-Fluoro-3,3', 5-trimethyl- ; [1,1'-biphenyl]-2-yl]methoxy]methyl- ’ phosphinyl}-3-t-butyldiphenylsilyloxy- y . butanoic acid, methyl ester

A cooled (0°C, ice bath) solution of Part A 3 phosphinochloridate (~2.2 mmole) and Example 1 : part C(2) biphenyl alcohol (429 mg, 2.2 mmole, 1.0 ; eq) in dry CH,Cl, (10 ml) was treated with Et, N (425 pl, 3.04 mmole, 1.4 eq) and 4-DMAP (27 mg, : 0.22 mmole) and the orange solution stirred at room temperature overnight under argon. The mixture was partitioned between 5% KHSO, and EtOAc, the organic layer washed with brine, dried over anhydrous

Na,SO, and evaporated to give 1.1 g of an orange oil. The crude oil was purified by flash

HX8b -60- chromatography on LPS-1 silica gel (44 9g) eluting with (1:1) EtOAc:Hexane. Product fractions were combined and evaporated to give 298 mg (21%) of desired coupled title product as a pale yellow 0il. Also 460 mg (67% corrected yield) of starting

Example 1 Part Cc(2) biphenyl alcohol was recovered.

TLC (1:1) EtOAc:Hex, Rf=0.18 UV and PMA.

C. (35)-4-[[[4'-Fluoro-3,3',5-trimethyl~ [1,1'-biphenyl]-2-yl]methoxy]methyl- - phosphinyl]-3-hydroxybutanoic acid, methyl ester

A solution of the Part B silyl ether (298 mg, 0.46 mmole) in dry THF (6.0 ml) was treated with glacial HOAc (110 ul, 1.84 mmole, 4.0 eq) and a 1.0 M in THF solution of tetrabutylammonium fluoride (1.43 ml, 3.1 eq) and the resulting solution stirred overnight under argon at room temperature. The mixture was partitioned between cold H,0 and EtOAc, the organic phase washed with saturated NaHCO, and brine, dried over anhydrous Na,SO, and evaporated to a yellow oil (273 mg): The crude oil was purified by flash chromatography on LPS-1 silica gel (11 g) eluting with (3:2) Hex-acetone. product fractions were combined and evaporated to give 150 mg (80%) of desired title alcohol as a viscous oil.

TLC (1:1) Hex :acetone, Rf=0.23, UV and PMA. : 30 D. (35)-4-[[[4'-Fluoro-3,3', 5-trimethyl- (1,1'-biphenyl]-2-yl])methoxy]methyl- phosphinyl]-3-hydroxybutanoic acid, monolithium salt

HX8b -61~

A solution of the Part C methyl ester (150 mg, 0.367 mmole) in dioxane (3.0 ml) was treated with 1.0 N LiOH (0.44 ml, 1.2 eq.) and the resulting white suspension was stirred at room temperature under argon for 2 hours. The mixture was diluted with H,0, filtered through a 0.4 pm polycarbonate membrane and evaporated in vacuo to a ‘ colorless glass.

The crude product was taken up in a minimum amount of H,0 and chromatographed on HP-20 (100 ml . bed) eluting with a H,0/CH,CN linear gradient.

Product fractions were evaporated, taken up in H,0 (50 ml), filtered through a 0.4 ym polycarbonate membrane and lyophilized to give 130 mg (79% based on hydrate weight) of desired title lithium salt as a white solid. TLC (8:1:1) CH,Cl,-CH,O0H-HOAC,

Rf=0.52, UV and PMA. ~ Anal Calcd for C, HygOgFL1P + 1.73 moles H,0 (MW 445.49): C, 56.61; H, 6.44: F. 4.26; P, 6.95

Found: C, 56.67; H, 6.36; F, 4.31; Pp, 7.43 ul NMR (400 MHz):

Oo 5 1.49 ppm (3H, 4, ~OP (CH, )- Ig_p=14.7 Hz) 0 1.83-2.0 (2H, m, 2- (cH, )CH,-) 2.27-2.40 (2H, m, CH,CO,Li) 2.30 (6H, s, 2 aromatic methyl's) 2.44 (3H, s, aromatic methyl) 4.26 (1H, m, -CH,CH(OH)CH,CO,L1)

HX8b -62- i i 4.87 (2H, m, ArCH,OP(CH;)-) ; 6.90-7.20 (SH, m, aromatic H's) . a

Fe

Example 4 = (8)-4-[[(2,4-Dichloro-6-[ (4-fluorophenyl)methoxy]- phenyl methoxy Jmethoxyphosphinyl]-3-hydroxy- butanoic acid, monolithium salt

A. 2, 4-Dichloro-6=(4-fluorophenylmethoxy)- i ’ benzaldehyde oy (Ref.: J. Med Chem., 1986, 29, 167) J

A solution of 13.77 g (72.5 mmol) of oo 4, 6-dichloro-2-hydroxybenzaldehyde in 100 ml of -

DMF was stirred and 12.02 g (87 mmol) of K,CO4 was added. This mixture was heated to ~70°C for 60 ‘ minutes, then 11.7 ml of 4-fluorobenzyl bromide ~ was added. The resulting solution was stirred at ir 70°C for 3.5 hours, then this was poured onto ice .

H,0 (1.5 1), filtered and washed with H,O0, and recrystallized from Et,0/petroleum ether.

Yield: 17.88 g (83%) of off-white crystals, m.p. 107-108°C.

B. 2,4-Dichloro-6-[(4-fluorophenyl)- methoxy ]benzenemethanol cold (0°C, ice bath), dry Et,0 (10.0 ml) was treated with LiAlH, (158 mg, 4.16 mmole, 0.6 eq.) and the grey suspension treated dropwise’with a solution of Part A aldehyde (2.06 g, 6.93 mmole) in 10 ml dry THF. The mixture was warmed to room temperature and stirred for an hour under argon. The mixture was cooled back to 0°C (ice bath) and quenched by sequential dropwise addition

HX8b -63- of H,0 (160 pl), 15% NaOH (160 pl) and H,0 (475 pl).

Precipitated salts were removed by filtration through anhydrous Na,SO, over packed Celite (%" bed). The clear filtrate was evaporated to give 2.052 g (98.9%) of crude alcohol as white crystals. One trituration with cold hexane afforded 1.892 g (91.2%) of pure title alcohol as a white crystalline solid with m.p. = 72-73°C.

TLC (4:1) Hex-acetone, Rf=0.31, UV and

PMA. :

Anal Calcd for Cy4H110,C1,F (MW 301.142):

Cc, 55.84; H, 3.68; Cl, 23.55; F, 6.31

Found: C, 55.97; H, 3.71; Cl, 23.42; F, 6.30

C. (s)-4-[[(2,4~Dichloro=-6-[(4-fluoro- phenyl )methoxy }phenyl Jmethoxy]methoxy- phosphinyl]-3-t-butyldiphenylsilyloxy- butanoic acid, methyl ester : A solution of the Example 1 Part E(6) methyl ester (~3.84 mmole) in dry CH,Cl, (10 ml) was treated with distilled trimethylsilyl diethylamine (1.46 ml, 7.68 mmole, 2.0 eq.) and the resulting solution stirred at room temperature under argon for 1.0 hour. The mixture was evaporated in vacuo, chased with benzene (1 x 20 ml) and dried in vacuo to give crude silylated phosphonic acid mono methyl ester as a colorless oil.

A solution of the crude ester (~3.84 mmole) in dry CH,C1, (10 ml) and dry DMF (1 drop) was cooled to ~-10°C (salt, ice path) and treated

: HX8b -64~ © dropwise with distilled oxalyl chloride (368 pul, 4.22 mmole, 1:1 eq.). Gas evolution was evident , from the clear yellow mixture. The mixture was " . gtirred at room temperature under argon for one i hour, evaporated in vacuo, chased with benzene (2 x : 20 ml) to give crude phosphonochloridate as a viscous yellow oil.

The crude phosphonochloridate (~3.84 mmole) in dry CH,C1, (10 ml) at 0°C (ice bath) was treated e : 10 with Part B alcohol (1.15 g, 3.84 mmole, 1.0 eq.) x followed by Et,N (805 pl, 5.76 mmole, 1.5 eq.) and i 4-DMAP (47 mg, 0.384 mmole, 0.1 eq.) and the brown mixture stirred overnight at room temperature ‘ under argon. The mixture was partitioned between 5% KHSO, and EtOAc, the organic phase washed with ’ g brine, dried over anhydrous Na,SO, and evaporated by to give 3.197 g of a dark brown oil. The crude product was purified by flash chromatography on silica gel (160 g) eluting with (7:3) Hex-EtOAC. . k

Product fractions were combined and evaporated to . give 594 mg (21.1%) of desired title phosphonate as oo a yellow oil. Additionally, 688 mg (52.4% corrected yield ) of starting Part B alcohol was recovered. :

TLC (1:1) Hex-acetone, Rf=0.29, UV and PMA. : 25

D. (s)-4-[[[2,4-Dichloro-6-[(4-fluoro- phenyl )methoxy ]phenyl Jmethoxy]methoxy- phosphinyl]-3-hydroxy butanoic acid, methyl ester

A solution of the Part C silyl ester (578 mg, 0.788 mmole) in dry THF (8 ml) was treated with glacial HOAc (180 pl, 3.2 mmole, 4.0 eq.) followed by 1.0 M solution of n-Bu,NF in THF (2.36

HX8b -65- ml, 2.36 mmole, 3.0 eq.) and the resulting pale yellow solution stirred overnight under argon a. room temperature. The mixture was poured into : cold H,0 and extracted with EtOAc (2X). The organic phase was washed with saturated NaHCO, and brine, dried over anhydrous Na,SO, and evaporated to give 625 mg of a yellow oil. The crude product was purified by flash chromatography on silica gel (31 g) eluting with (7:3) Hexane-acetone.

Product fractions were combined and evaporated to give 339 mg (86.9%) of desired title alcohol as a clear, colorless, viscous oil.

TLC (1:1) Hex-acetone, Rf = 0.25, UV and PMA.

E. (s)-4-[{[[2,4-Dichloro-6-[(4-fluoro- phenyl )methoxy ]phenyl Jmethoxy]methoxy- phosphinyl]-3-hydroxybutanoic acid, monolithium salt

A solution of the Part D phosphonate (132 mg, 0.267 mmole) in dioxane (2.5 ml) was treated with 1.0 N LiOH (0.32 ml, 1.2 eq.) and the mixture stirred under argon at room temperature for 4.0 hours. A white precipitate was noted. The mixture was diluted with H,O0, filtered and the : 25 filtrate evapora ed to dryness in vacuo. The : residue was chromatographed on HP-20 resin (100 ml bed) eluting with a H,0/CH,CN linear gradient system. Product fractions were combined and evaporated, taken up in H,0, filtered through a 0.4 pum polycarbonate membrane and lyophilized to give 108 mg (79% based on hydrate weight) of desired title lithium salt as a white solid."

HX8b } -66- 1

TLC (20:1:1) CH,Cl,- CH,OH-HOAC, Rf = 0.41, Con

Uv and PMA. } -

Anal Calcd for Cj gH, g0,Cl,FliP + 1.42 moles H,0 (MW 511.72): C, 44.59; H, 4.10; CL, 13.86; .

F, 3.71; P, 6.05

Found: C, 44.22; H, 4.09; Cl, 13.91; F, 3.72;

P, 6.11 Cw

H NMR (400 MHz): CE : bi 5 1.98-2.11 ppm (2H, m, OP (OCH, )CH,CH(OH)- i 2.26-2.45 ppm (2H, m, -CH(OH)CH, CO, Li) fe 3.63 & | 5 3.62 (3H, 2 doublets, 2 diastereomers,

OP (OCH, )CH, =, Jgp~11 Hz) T 4.23 (1H, m, (~CH,CH(OH)CH,CO,L1) i 5.16 (2H, s, F-PhCH,0) . ) 5.24 (2H, 4, ArCH,OF, Jgp=6-2 Hz) 3 7.13-7.53 (6H, m, aromatic H's) y

Example 5 . (35)-4-[[ [2,4-Dichloro-6-[ (4-fluorophenyl)- : } 25 methoxy phenyl Imethoxyhydroxyphosphinyl]=3- : hydroxybutanoic acid, dilithium salt !

A mixture of the Example 4 Part D diester (210 mg, 0.424 mmole) in dioxane (4.0 ml) was : treated with 1.0 N LioH (1.30 ml, 3.0 eq.) and the colorless solution heated at 50°C (oil bath) under argon for 3.5 hours. A white precipitate was evident after 15 minutes. The mixture was diluted with H,0, filtered and the filtrate evaporated

HX8b -67~ in vacuo. The residue was dissolved in a minimum amount of H,0 and chromatographed on HP-20 resin (100 ml bed) eluting with a H,0/Cil3CN linear gradient. Product fractions were combined and evaporated. The residue was taken up in H,0 (50 ml), filtered through a 0.4 um polycarbonate membrane and lyophilized to give 175 mg (81% based on hydrate weight) of desired title dilithium salt as a white solid.

TLC (8:1:1) CH,Cl,-CH;0H-HOAC, Rf = 0.07, UV and PMA.

Anal Cacld for C,gHy g07CL,FLL,P + 1.70 moles H,O0 (MW 509.62): C, 42.42; H, 3.84; F, 3.73; cl, 13.91; P, 6.08

Found: C, 42.46; H, 3.90; F, 3.93; Cl, 13.42;

P, 5.66

HINMR (400 MHZ) : 0 6 1.73-1.92 opm (2H, m, ~OB (OLi )~CH,,CH(OH)- 2.27 (1H, dd, -CH(OH)CH,CO,Li, Jy,=0.8

Hz) 2.39 (1H, dd, ~CH(OH)CH,CO, Li, J=4 4

Hz) 4.26 (1H, m, CH,CH(OH)CH,CO,L1) 5.00 (2H, s, F-Ph-CH,OAr) 7.03-7.53 (6H, m, aromatic H's).

HX8b -68-

Example 6 (35)-4-[[2,4-Dichloro-6-[(4-fluorophenyl)methoxy]- phenyl Jmethoxy Jmethylphosphinyl]-3-hydroxybutanoic acid, methyl ester

A. (s)3-[[(1,1-Dimethylethyl)diphenylsilyl]- oxy] -4-(ethoxymethylphosphinyl)butanoic acid, methyl ester = oo

A mixture of the Example 1 Part E(3) iodide (4.68 g, 9.18 mmole) in methyl diethoxy- phosphine (Strem Chemicals, 5.0 g, 36.7 mmole) was heated at 100°C (oil bath) for 2.5 hours, then at 150°C for three additional hours under argon. A white precipitate slowly formed in the yellow solution. Excess phosphine was distilled off in vacuo (0.5 mm Hg) and the crude product purified by flash chromatography on silica gel eluting with (65:35) Hexane-acetone. Product fractions were combined and evaporated to give 1.590 g (38%) of desired title phosphinic ester (mixture of diastereomers) as a clear viscous oil.

TLC (3:2) Hex-acetone, Rf (2 diasterecmers) = 0.19 and 0.22, UV and PMA.

B. (3s)-4-[[2,4-Dichloro-6-[(4-fluoro- : 25 phenyl )methoxy ]phenyl ]methoxy Jmethyl- phosphinyl]-3-t-butyldiphenylsilyl- butanoic acid, methyl ester

A solution of Part A phosphinic ester (605 mg, 1.3 mmole) in dry CH,Cl, (6.0 ml) was treated with bis(trimethylsilyl)trifluoroacetamide (BSTFA) (280 pl, 1.05 mmole, 0.8 eq.) and trimethylsilyl bromide (TMSBr) (210 pl, 1.57 mmole, 1.2 eq.) and the resulting solution stirred at room temperature

HX8b -69- under argon overnight. 5% KHSO, (15 ml) was added and the mixture extracted with EtOAc. The organic phase was washed with brine, dried over anhydrous

Na,SO, and evaporated in vacuo to give crude phosphinic acid as a colorless oil.

A solution of the crude phosphinic acid (~1.3 mmole) in dry CH,Cl, (6.0 ml) was treated with distilled trimethylsilyl diethylamine (270 pl, 1.44 mmole, 1.1 eq.) and the clear mixture was stirred at room temperature under argon for 1.0 hour. The mixture was evaporated in vacuo, chased with benzene (1 x 15 ml), and dried in vacuo.

A cooled (0°C, ice bath) solution of the crude silylated phosphinic acid (~1.3 mmole) in dry

CH,Cl, (6.0 ml) and DMF (1 drop) was treated dropwise via syringe with distilled oxalyl chloride . (130 pl, 1.44 mmole, 1.1 eq.). Gas evolution was evident. The mixture was stirred at room temperature under argon for one hour then evaporated in vacuo, chased with benzene (2 x 15 ml) and dried in vacuo to give crude phosphino- - chloridate as a yellow oil.

A cooled (0°, ice bath) solution of phosphinochoridate (~1.3 mmole) and Example 1 Part

E(6) alcohol (392 mg, 1.3 mmole) in dry CH,Cl, (6.0 ml) was treated with Et3N (275 pl, 1.97 mmole, 1.5 eq.) and 4-DMAP (16 mg, 0.13 mmole, 0.1 eq) and the resulting yellow mixture stirred under argon at room temperature overnight. The mixture . was partitioned between 57 KHSO, and EtOAc, the organic phase washed with brine, dried over anhydrous Na,SO, and evaporated to give 908 mg of

HX8b -70- crude product as a dark yellow oil. The crude product was purified by flash chromatography on silica gel (45 g) eluting with (3:2) Hex-EtOAc.

Product fractions were combined and evaporated to give 266 mg (28.3%) of desired title product as a clear, colorless oil. Also 197 mg (57%, corrected yield) of the starting alcohol was recovered.

Cc. (3S)-4-[[2,4-Dichloro-6-[(4-fluoro- phenyl )methoxy]phenyl methoxy methyl- phosphinyl]-3~hydroxybutanoic acid, methyl ester

A solution of the Part B silyl ester (275 mg, 0.38 mmole) in dry THF (6.0 ml) was treated with glacial HOAc (90 pl, 1.53 mmole, 4.0 eq.) and a 1.0 M solution in THF of tetrabutylammonium fluoride (1.2 ml, 3.1 eq.). The resulting solution was stirred overnight under argon at room temperature. The mixture was partitioned between cold H,0 and EtOAc, the organic phase washed with saturated NaHCO, and brine, dried over anhydrous

Na,SO, and evaporated to give 258 mg of a yellow oil. The crude product was purified by flash chromatography on LPS-1 silica gel (8 g) eluting with (1:1) Hexane-acetone. Product fractions were combined and evaporated to give 142 mg (77%) of desired title alcohol as a clear, colorless, oil.

TLC (1:1) Hexane-acetone, Rf = 0.20, UV and PMA. go ORIGINAL J 5 d

HX8b -71=-

D. (35)-4-[[2,4-Dichloro-6-[(4-fluoro- phenyl )methoxy phenyl Jmethoxy]methyl- : phosphinyl]-3-hydroxybutanoic acid, monolithium salt

A solution of the Part C methyl ester (142 mg, 0.296 mmole) in dioxane (3.0 ml) was treated with 1.0 N LiOH (0.36 ml, 1.2 eq.) and the resulting white suspension stirred under argon at room temperature for 2.0 hours. The mixture was ~ 10 diluted with H,0, filtered through a 0.4 pm polycarbonate membrane and the filtrate evaporated in vacuo.

The crude product was dissolved in a minimum amount of H,0 and chromatographed on a 100 ml bed of HP-20 resin eluting with a H,0/CH;CN linear gradient. Product fractions were combined and evaporated. The residue was taken up in H,0, filtered through a polycarbonate membrane and lyophilized to give 93 mq (63% based on hydrate weight) of desired title lithium salt as a white solid. -

TLC (8:1:1) CH,Cl,- CH,OH-HOAC, Rf = 0.51, UV and

PMA.

Anal Calcd for C, gH; g07CL, FLIP + 1.38 moles H,0 (MW 495.94): C, 46.01; H, 4.42; F, 3.83; Cl, 14.30;

P, 6.24

Found: C, 46.10; H, 4.49; F, 3.82; Cl, 14.32;

P, 6.43 ul NMR (400 MHz):

HX8b 72 9 8 1.53 ppm (3H, d, -OP (CH, )CH,-, Iy-p = 14.6 Hz) ? 1.87-2.10 (2H, m, ~OP (OCH; )CH,-) 2.27 (1H, dd, -CH(OH)CH,CO,Li,

Ig-g=8-4 Hz, Jy-p=1-1 Hz) 2.38 (1H, dd, ~CH(OH)CH, CO, Li,

Ig-u=4-7 Hz, Jy.p=1-1 Hz) 4.29 (1H, m, -CH,CH(OH)CH,CO, Li) 5.16+5.18 (4H, m, ArCH,OP and F-PhCH,0)-) 7.11-7.52 (6H, m, aromatic)

Example 7 (S)-4-[[[4'-Fluoro-3,3"',5-trimethyl(1,1'-biphenyl- 2-y1]methyl]amino |methoxyphosphinyl]-3-hydroxy- butanoic acid, monolithium salt

A. 4'-Fluoro-3,3',5-trimethyl(l,1'- } biphenyl }-2-carboxylic acid

A solution of the Example 1 Part C(2) aldehyde (1.0 g, 4.13 mmole) in acetone (10.0 ml) at 0°C (ice bath) was treated dropwise with 8.0 N Jones reagent (4.1 ml, excess) and the resulting brown-green suspension stirred overnight under argon at room temperature. Excess oxidant was destroyed by adding isopropanol (10.0 ml) and precipitated chromium salts removed by filtration through a %" pad of Celite. The filtrate was evaporated, taken up in EtOAc, washed with 1.0 N

HCl (2X), saturated NH, Cl (2X) and brine, then dried over anhydrous Na,SO, and evapo.ated to give 1.011 g of a green solid with m.p. 153-154°C.

HX8b ~-73~ : The crude acid was purified via the dicyclohexylamine salt. To a solution of the . crude acid in EtOAc (5.0 ml) was added dicyclohexyl amine (DCHA) (823 pl, 1.0 eq.). The solution diluted with hexane and precipitated crystalline salt was collected to give 997 mg (55% from aldehyde, m.p. 181-183°C) of desired product as an . off-white crystalline DCHA salt.

The title free acid was regenerated from the DCHA salt by partitioning the salt between 57

KHSO, and EtOAc. The organic phase was washed with brine, dried over anhydrous Na,SO, and evaporated in vacuo to give 554 mg (52% from aldehyde) of desired title acid.

TLC (9:1) CH,Cl,-CH;O0H, Rf=0.37, UV and PMA.

B. 4'-Fluoro-3,3',5-trimethyl(1,1'- biphenyl]-2-carboxamide

A suspension of the Part A acid (554 mg, 2.14 mmole) in dry CH, Cl, (6.0 ml) and dry DMF (1 drop) at 0°C (ice bath) was treated dropwise via syringe with distilled oxalyl chloride (205 pl, 2.35 mmole, 1.1 eq.) and the clear yellow solution stirred under argon at room temperature for one hour. The mixture was evaporated in vacuo, chased with benzene (2X) and dried in vacuo to give crude acid chloride as a yellow oil.

A cooled (0°C, ice bath) mixture of THF (3.0 ml) and concentrated NH, OH (2.0 ml, excess) was treated dropwise with a THF solution (3.0 ml) of the crude acid chloride and the bright orange solution stirred at room temperature under argon for 1.0 hour. The mixture was partitioned

HX8b -74~ between H,0 and EtOAc, the organic phase washed with saturated NaHCOg5, H,0 and brine, then dried over anhydrous Na,sSO, and evaporated to give 528 mg (96.1%) of crude amide as a light orange solid. One recrystallization from EtOAc~-hexane afforded 435 mg (79.1%) of purified title amide as pale yellow needles with m.p. 197-198°C. TLC (1:1)

Et,0-Acetone Rf=0.83, UV and PMA.

Cc. 4'-Fluoro-3,3',5-trimethyl(1l,1'- biphenyl]=~2-methanamine

A cooled (0°C, ice bath) solution of dry

THF (5.0 ml) was treated with solid LiAlH, (125 mg, 3.3 mmole) and the gray suspension treated dropwise over five minutes with a solution of the

Part B amide (424 mg, 1.65 mmole) in THF (5.0 ml).

The resulting suspension was stirred at room temperature under argon for 2.5 hours, then refluxed for 45 minutes. The mixture was cooled to 0°C (ice bath) and quenched by sequential dropwise addition of 125 pl H,0. 125 ul of 15% NaOH and 375 wl H,0. Precipitated aluminum salts were removed . by filtration through anhydrous Na,SO, over packed

Celite. The clear filtrate was evaporated in vacuo to give the crude amine as a clear oil.

TLC (7:3) Et,0-acetone, Rf=0.60, UV and PMA. The amine was purified as the HCl salt.

A solution of the crude amine (~1.65 mmole) in absolute EtOH (8.0 ml) was treated with concentrated HCl (152 pl, 1.82 mmole) and the mixture stirred for 15 minutes at room temperature under argon. The mixture was evaporated in vacuo to a white crystalline solid. laao opvoma- !

HX8b -75m-

The solid was triturated with cold Et,0, collected by filtration and dried in vacuo to give 426 mg (92.4%) of title amine-HC1l as fine white crystals.

D. (S)-4-[[[[4'-Fluoro-3,3',5-trimethyl- [1,1'-biphenyl]-2~-yl]methyl]amino]- methoxyphosphinyl]-3-t-butyldiphenyl- silyloxy butanoic acid, methyl ester

A solution of Example 1 Part E(6) methyl ester (~2.0 mmole) in dry CH,C1, (5.0 ml) was treated with distilled trimethylsilyl diethylamine (758 pl, 4.0 mmole, 2.0 eq.) and the clear mixture stirred at room temperature under argon for one hour. The mixture was evaporated in vacuo, chased with benzene (1 -x 15 ml) and dried in vacuo.

A cooled (0°C) solution of the crude silyl phosphonate in dry CH,Cl, (7.0 ml) and DMF (1 drop) was treated dropwise with distilled oxalyl chloride (192 pl, 2.2 mmole, 1.1 eq.). Gas evolution was evident from the clear yellow mixture. The solution was stirred at room temperature for one hour, evaporated in vacuo, chased with benzene (2 x 15 ml), and dried in vacuo to give the crude phosphonochloridate as a yellow, viscous oil.

A cooled (0°C) solution of the phosphonochloridate and Part C biphenyl amine-HCl (416 mg, 1.49 mmole) in dry CH,Cl, (10 ml) was treated with Et,N (641 pl, 4.6 mmole, 2.3 eq.) and 4-DMAP (24 mg, 0.2 mmole, 0.1 eq.) and the clear yellow mixture stirred overnight at room temperature under argon. The mixture was partitioned between 5% KHSO, and EtOAc, the

HX8b -76- organic phase washed with brine, dried over anhydrous Na,SO, and evaporated in vacuo to give 1.19 g of a yellow oil. The crude product was purified by flash chromatography on silica gel (60 g) eluting with (7:3) hexane-acetone. ’ Product fractions were evaporated to give 588 mg (59.5%) of desired title phosphonamide as a pale yellow, viscous oil.

TLC (7:3) Hexane-acetone, Rf=0.20, UV and PMA.

E. (s)-4-[[[[4'-Fluoro-3,3',5-trimethyl- [1,1'-biphenyl)-2-yl]methyl]amino]- me thoxyphosphinyl]-3-hydroxybutanoic acid, methyl ester ~ A solution of Part D silyl ether (588 mg, 0.888 mmole) in dry THF (10.0 ml) was treated with glacial HOAc (203 pl, 3.55 mmole, 4.0 eq.) and a 1.0 M solution in THF of tetrabutylammonium fluoride (2.66 ml, 2.66 mmole, 3.0 eq.) and the resulting solution stirred overnight under argon at room temperature. The mixture was poured into cold H,0 and extracted with EtOAc. The organic phase was washed with saturated NaHCO, and brine, then dried over anhydrous Na,SO, and evaporated in vacuo to give 605 mg of an orange oil. The crude product was purified by flash chromatography : on silica gel (36 g) eluting with (1:1) Hexane- acetone. Product fractions were combined ard evaporated to give 196 mg (50.4%) of desired title alcohol as a light orange oil.

TLC (1:1) Hexane-acetone, Rf=0.16, UV and PMA. laa ORIGINAL 2) 'BAD ORIGINAL oI : \

HX8b -77

F. (S)-4-[[[[4'-Fluoro-3,3',5-trimethyl- [1,1'-biphenyl]-2-yl]methyl]amino}- me thoxyphosphinyl }-3-hydroxybutanoic acid, monolithium salt

A solution of the Part E diester (105 mg, 0.240 mmole) in dioxane (2.0 ml) was treated with 1.0 N LiOH (288 pl, 1.2 eq.) and the white suspension stirred under argon at room temperature for 4.0 hours. The mixture was diluted with H,0, filtered, the filtrate evaporated in vacuo. The residue was chromatographed on HP-20 (100 ml bed) resin eluting with a H,0/CH,CN linear gradient.

Product fractions were combined and evaporated.

The residue was taken up in H,0 (50 ml), filtered through a 0.4 pm polycarbonate membrane and lyophilized to give 70 mg (62.7% based on weight of hydrate) of desired title lithium salt as a white solid.

TLC (20:1:1) CH,Cl,-CH;OH-HOAC, Rf=0.19, UV and

PMA.

Anal Calcd for C,H, NOPFL1 + 2.41 moles H,O0 (MW 472.75): C, 53.35; H, 6.57; N, 2.96; F, 4.02;

P, 6.55

Found: C, 53.35; H, 6.52; N, 2.98; F, 4.05;

P, 6.59

HX8b =T78 =~ ul NMR (400 MHz): 0 1.79-1.97 ppm (2H, m, -B(ocH, )CH,-) 2.26-2.44 ppm (2H, m, -CH,,CO, Li 5 2.29 (3H, s, aromatic methyl) 2.31 (3H, d, aromatic methyl «o to fluorine, Jgp=1-4 Hz) 2.47 (3H, aromatic methyl) 3.46 & : 10 3.50 (3H, 2 doublets, 2 : diastereomers, Jyp=10-5 Hz) 0 : 3.96 (2H, m, PhCH, NHP (OCH, ) - 4.17 (1H, m, (-CH,CH(OH)CH,CO,L1) 6.84-7.21 (5H, m, aromatic protons)

Examples 8 to 20

Following the procedures as outlined heretofore and as described in the previous working Examples, the following additional compounds may be prepared. ° .

R-P-CH,-C-CH,=CO -R

X OH

(Cty) 2 . . a So. . *

BAD ORIGINAL oh [7 WE

HX8 b -79-

Ex.

No. Roz oon EA RY 8. OH (O)y—cn, 1 0 H 0

Ef;

H

3 . H - 3 C,H: CH3 © 2 “Hs

CH, cl 10. cH, 0) 1 © Li 0G

H

Cog 11. CH,0 (0)-c#y0 2 NH H § iy 12. OH © 2 oH > © 13. CHO - 1 © Li 49 (O)—cu -0 or ~ NT h,

. HX8b -80-

EX.

No. rR Zz 0 00n Xx _R* 14. CoH, | 1 NH CH, (©) en, 2 o, 15. OK C.H_-CH 2 0 OK 2757,

C=0 0

CH, 16. ONa 2 0 H _CH_-0 —@

Se CH

CL :

Pe : } CH,O 1 NH H 17 3 r~(O)-cx -0 2

CH of LY

H i HXS8b ’ -81l-

Ex. xX

No. R Ze nn x —R_ ——

C,H =C—C—0 1 0 H 18 Hs 4 YO 3 3 CH 3 7

OH

19. HO (o)- 2° =

HX8b -82=-

Example 21 (S)-4-Diisopropyloxyphosphinyl)-3- [[(1,1-dimethyl- ethyl )diphenylsilyl] oxy]-butanoic acid, methyl ester

The Example 1 Part E(3) iodide (45.1 mmol., 21.70 g) was stirred under high vacuum for 30 minutes. Freshly distilled triisopropyl phosphite (0.451 mol., 93.92 gq, 113.37 ml.) was added in one portion and the reaction mixuture was stirred under argon and heated in a 155°C oil bath for 16.5 hours. The mixture was then cooled to room temperature. Excess triisopropyl phosphite and volatile reaction products were removed by short path distillation (10 mm Hg) followed by Kugelrohr distillation (0.50 mm Hg, 100°c, 8 hours). The product was further purified via flash chroma- tography (95 mm diam. column, 6"/Merck silica gel, 6/3/1 Hexane/acetone/toluene eluent, 2"/min flow rate, 50 ml fractions) to afford 17.68 g (33.96 mmol, 75% yield) of the title isopropylphosphona’e as a clear viscous oil.

TLC: Silica gel Rg=0.32 (6:3:1 Hexane/acetone toluene) :

LgNMR: (270 MH, CDClj) 5 7.70-7.65 (m,4H) 7.45-7.35 (m, 6H) 4.57-4.44 (m,3H) 3.59 (s,3H) 2.94 and 2.88 (2xd, 1H J=3.7 Hz) 2.65 and 2.60 (2xd, 1H J=7.4 Hz) 2.24-1.87 (Series of m, 2H) 1.19 and 1.12 (2xd, 12H J=6.3 Hz) 1.01 (s, SH) ano ORIGINAL 9

HX8b -83~-

Example 22

S)-4-(Hydroxymethoxyphosphinyl)-3- {{(1,1-dimethyl~ ethyl )diphenylsilyl]oxy]butanoic acid, methyl ester, dicyclohexylamine (1:1) salt

The Example 21 isopropyl phosphonate (30.5 mmol, 10.66 g) was stirred under argon, at room temperature, in 80 ml of dry CH,Cl,. This solution was treated dropwise (5 min) with bistrimethylsilyltrifluoroacetamide (BSTFA) (32.8 mmol, 8.44 g, 8.71 ml), followed by dropwise : addition (10 min) of trimethylsilylbromide (TMSBr) (51.3 mmol, 7.84 g, 6.75 ml). After stirring at room temperature for 20 hours, the reaction mixture was quenched with 200 ml of 5% aqueous

KHSO, and stirred vigorously for 15 minutes. The aqueous layer was extracted 3 times with ethylacetate. The organic extracts were combined, washed once with brine, dried over Na,S0, and concentrated in vacuo. The residue was azeotroped 2 times with 50 ml of toluene. The precipitate which formed was suspended in toluene and filtered. The filtrate was concentrated and the azeotrope/filter process repeated. The resulting filtrate was evaporated in vacuo and then pumped under high vacuum for 5 hours. The resulting viscous clear oil was stirred under argon, at room temperature, in 50 ml of dry pyridine. This solution was treated in one portion with dicyclohexylcarbodiimide (DCC) (22.6 mmol, 4.65 g), followed by addition of methanol (41.0 mmol, 1.31 g, 1.67 ml). After stirring at room temperatu.e for 20 hours, the reaction mixture was filtered through a celite pad in a sintered glass funnel. ) Ce Lo

BAD Oniun Zl

HX8Db -84-

The celite was washed with ethyl acetate and the combined filtrates were evaporated in vacuo. The residue was redissolved in ethyl acetate and : washed 2 times with 5% aqueous KHSO, and once with brine. The organic extract was dried over Na,SO,, filtered, the filtrate concentrated and azeotroped . 2 times with toluene, suspended in toluene and filtered. The resulting filtrate was again concentrated, azeotroped, filtered and the filtrate evaporated in vacuo and placed under high vacuum for 6 hours to afford the phosphonate monoester as a clear viscous oil (10.2 g, >100% yield). TLC: silica gel R=0.50 (7:2:1 nPrOH/NH,OH/H,0) . The phosphonate monoester (1.21 g was pumped under high vacuum for 4 hours, affording 1.16 g (2.57 mmol)] was dissolved in 10 ml of dry ethyl ether and treated dropwise with dicyclohexylamine (2.65 mmol, 0.481 4g. 0.528 ml). ~The resulting homogeneous solution sat at room temperature for 7 hours resulting in significant crystal formation. The mixture was stored at -20°C for 16 hours and then warmed to room temperature and filtered. The crystals were washed with cold, dry ethyl ether and then pumped under high vacuum over P,0¢ for 18 hours. The crystals were subsequently pumped under high vacuum at 45°C for 4 hours, affording 1.25g (1.98 mmol, 77% yield) of the title dicyclohexylamine salt as a white powdery solid, m.p. 155-156°C.

TLC: Silica gel R=0.57 (20% MeOH/CH,Cl,) 'H NMR: (270 MH, , CDC1,) [BAD ORIGINAL 9

HX8b -85- 7.71-7.65 (m, 4H) 7.40-7.32 (m, 6H) 4.02 (m, 1H) 3.52 (s, 3H) 5 3.28 and 3.22 (m, 1H) 3.11 (4, 3H J=11 Hz) 2.77-2.64 (m, 2H) 2.62-2.56 (m, 1H) 1.92-1.08 (Series of m, 22H) 1.00 (S, 9H)

Mass Spec: (FAB) 632 (Msi)

IR: (KBr) 3466-3457 (broad) 3046, 3016, 2997, 2937, 2858, 2836, 2798, 2721, 2704, 2633, 2533, 2447, 1736, 1449, 1435, 1426, 1379, 1243, 1231, 1191, 1107, 1074, 1061, 1051. 820 CM-1

Anal Calcd for CoH 0(PSi-C) Hysol:

C,64.63; H,8.61; N,2.22

Found: C, 64.51; H, 8.49; N, 2.18

Claims (1)

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