1216
P. D. KLIMSTRA AND R. E. COUNSELL
l-nl. d
Hypocholesterolemic Agents. 11.' 20-Halo-5a-androstane Derivatives P. D. I(LIMSTR.4
hh'D
R. E. COUNSELL
Division of Chemical Research of G. D. Sea& & Co., Chicago 80, Illinois Received M a y 18, 19692 The synthesis of a number of 2~-halo-5a-androstan-3~-ol-l$-onesand their derivatives is described. Physical data have been obtained to confirm the chemical evidence for the axial orientation of the fluorine in the 2&fluoro-3-keto-L~androstanes. Preliminary biological studies have shown some of these compounds t o possess interesting physiological properties.
A recent report by Hellman and associates* describing the hypocholesterolemic effect produced by parenterally administered androsterone (5a-androstan-3a-ol-17-0ne) stimulated our interest in 2bhalo-5a-androstan-3a-ol-lT-ones and their derivatives. I t was hoped that the introduction of a 28-halogen into the androsterone moleculc would not only enhance the cholesterol-lowering effect, but also produce compounds possessing oral artivit y. A search of the literature rrvealed that the synthesis of 28-bromo5a-androstan-3a-ol-IT-onc had been described by Fajkos, ct aL3 By using a slight modification of this procedure, the 2p-chlor0, bromo, and iodo-5a-androstan-3a-ol1 i-onr analogs iIIb-d) n ere prepared by diaxial cleavage of 2a,:~a-cpo~y-3a-androstan-li-one(I). -111 at tempt to prepare the cori r-ponding L'p-fluoro-:,a-androststrt-:3cu-ol17-one (Ira) from I with boroii trifluoride rthcrate in a l)cn7erie-rtlic1r d u t i o n ' led mainly to a I11gh mrlting product nith analysis for C38H571' O i j -1fair yield of IIa I\ as obtained, hon (11, I\ hcn a ~ h l t ) i ( ~ (
1
~
(1) Paper I R I Counsell, P D Iilliii7tiri,R I: Ranney and D I ( uoh. J . l l r d I i i i C h c m , 6 , 720 ( 1 4 G 3 i Z ) I, l i ~ l l i i i n n I I I, Bradlon, B Ziiiiioff I> I< I'ukushima and T r I . a l l ~ ~ 1 1I~ 1C i i ? Lridocrinoi a u d l i e t a b 19, 9311 i 1 9 i 9 1 3) J i ,*ihod , i d I' 'jorm cdi C Z ( , Chem L'umm 24, 3117 (1'1X I T H lTcnlrv,t 11id ?' I I t t ~ g l i d s r r n t t t i r e l > tssignril t l i t b t r r i r t i i r i I )
?
0
November, 1962
HYPOCHOLESTEROLEMIC AGENTS. I1
1217
form solution of I was added slowly to a large excess (20-30-fold) of anhydrous hydrogen fluoride in chloroform-tetrahydrofuran. The 17-carbonyl group in the above halohydrins was reduced smoothly and in good yield with lithium-tri-tert-butoxyaluminohydride to give the corresponding 2p-halo-5a-androstane-3a,l7pdiols (IIIa-d). The reduction of IIc with sodium borohydride, on the due to the basicity other hand, gave 2a,3a-epoxy-5a-androstan-l7p-ol of the reducing media. Similar treatment of IIb, however, gave the desired 2p-chloro-3a,l7p-diol (IIIb). The acetates (IIe-g) of the 2p-halohydrins were prepared by conventional methods. The observa tion6 that 2 a-fluoro-5 a-androstan- 17p-01-3-one acetate (X) caused marked inhibition of methylcholanthrene-induced rat mammary cancer as well as a strong anabolic response in the young female rat prompted us t3 prepare the corresponding 2p-fluor0 epimer. Diaxial fission of 2a,3a-epoxy-5a-androstan-17@-01acetate (VII) with anhydrous hydrogen fluoride as described above gave the desired fluorohydrin (VIII). Chromic acid oxidation of VI11 furnished 2~-fluor0-5a-androstan-17~-01-3-one acetate (IX). A similar oxidation of IIa gave 2p-fluoro-5a-androstane-3,17-dione (IV). Treatment of IV and IX with hydrobromic acid in acetic acid led to the previously described 2a-fluor0 epimers, V6 and X.6 An interest in the possible myotrophic activity of the 2-fluor0 steroids prompted us to prepare some 2-fluoro-5a-androst-1-ene derivatives.* Bromination and dehydrohalogenation of V gave 2fluoro-~~a-androst-l-ene-3,17-dione (VI). The preparation of 2fluoro-5a-androst-1-en-17~-ol-3-one acetate (XII) involved treating X I in glacial acetic acid with anhydrous hydrogen fluoride. Except for a recently issued patentj9this is the first description of 2fi-fluoro-3-ketosteroids. Consequently, spectroscopic and other physical data were obtained to confirm the chemical evidence for the axial orientation of the fluorine group in compounds IV and IX. In contrast with 2-bromo- and 2-chloro-3-ketosteroids,10 the infrared spectrum was of no value for differentiating the equatorial and axial 2-fluoro-3-keto-5a-androstane epimers. For example, the C-3 carbonyl stretching region of both IV and IX \vas displaced by about 24 cm.-l to a higher frequency. I n the ultraviolet, however, the presenw of the axial fluorine in compound IX produced a bathochromica
’
( 8 ) S Nakanishi K Morita, a n d E V Jensen, J . A m . Chem. Soc , 81,5259 (1959) (7) J Edrrards a n d H J. Ringold, J . A m Chem. SOC.,81,5262 (1959). (8) The preparation a n d myotrophic activity of 2-bromo a n d 2-chloro-5cr-androstI-Pnr derivatives was included in a previous publication, R. E. Counsell a n d P D Klimstra, J . M c d Pharm Chem , 6,477(1962) (9) R E Counsell a n d P. D. Klimstra, U S Patent 2,980,710(lQ6l) ( I O ) E G. Cummins and J E Page, J. Chem SOC, 3847 (1957)
1’. D. KLIMSTRA AND It. E. COUNHELT.
1218
I, ltlt’ = 0 VII, R = OSC, 1t’ = 11
Vol. 5
1121, S = F, RR’ = 0, R ” = kI 11, X = C1, RR’ = 0, R ” = H r. X = Hr. RR’ = 0. R ” = H
g; X = I, ‘ RR’ = 0, R ” = BC 111%.X = F.R = OH. R’ = R ” = II t>: X = C1, R = OH; R’ = R ” = €1 r, S = Br, It = OH, R’= R ” = €1
(1, X VI11 Y
V, IZR‘=O X, R=OAc, li’ - H
= =
I, R = OH, R’= H ” = H F, It = OAc, It’ = It’’ = If
lV, RR‘=O IX. R=OAc, R ’ z H
V
VI, KR‘=O X I I , R=OAc, R ’ = H
OAc
O
H XI
shift ill the lo\\ iiiteiisiimycurboiiyl ubsorption I)aiid of alwui, +!)nip. l’his shift is slight,ly less than t’hat reported ii.c., 15) for ot’lier axial a-fluoroketosteroids.ll -4s expected, the introduction of the equatorial 2a-fluor0 group left the ultraviolet absorption maximum unaffected. Additional evidcncc. for the axial configuration of t,he fluorine group in 2P-fluoro-~Scu-an(~rostaiie-3,17-dione(IT:) \vas ohtitined hy nurlear magnetic resonance measurements. ‘l‘lie C-1‘3 methyl prot’ons of 11- displayed a split, peak ((3and 65 c.p.s.) possihly dur t o a 5 lmnd
+
(
11)
(’. l)jf.rnssi, “Optical Kotatorp t)iayersil,rl.” .\lv(;raW-f~iil Hook ( ~ ~ J l l l ) , a l l yI. n r . , W. Klyne. .I. A m . Chim. Sor.., 79, 1.501;
X P W York, N . Y . , 19tj0, 1~1,. 11.5-131: C . Iljerassi and
f 19.77).
( 1 2 ) Tlii: n.iii.r, spectrii w e w furnisl~edby U r . It. ‘r. 1)illon ani1 Air. .J. 1)anias:oiis uf uiii‘ .inalytical Division. Measurements were obtained on t h e Varian model . I 4 0 at (XI 3 1 c . lit ~leuteriochlorofi~srii using terramethylsilane as a n internal atnndard.
20
1
400
300
500
A, mu.
Fig. 1.-Rotatory dispersion curves (methanol) of 2P-fluoro-5~androstan-l7~-ol3-one acetate ( I X ) and 2~-fluoro-5~-androstan-l7~-ol-3-one acetate (X).
spin-spin coupling between the C-19 methyl protons and t'he C-2 axial fluorine.'" I n contrast', the 2a-fluoro isomer (V) gave the usual C-19 methyl group singlet (68 c.P.s.). The possibility that the split peak is due to a mixture of A-ring conformers, however, cannot' be rompletely eliminated. The fact' that t'he fluorohydrin (IIa) also gave a split' peak (56 and 58 c.P.s.) for the C-19 methyl protons would tend t o discount' this latter p0ssibi1ity.l~ Recent'ly, optical rotatory dispersion measurements have been very useful in det'ermining the configuration of a-haloketones. l1 As shown in 1;ig. 1 , the ORD curvel5 of X is not sufficient,lydifferent from that of (13) D. R. Davis, R . P. Lutz, and J. D . Roberts, J . Am. Chem. Soc., 83,246 (1961). (14) We are indebted t o Dr. R. H. Bible of our Chemical Research Division for his assistance in the interpretation of the n.m.r. data. KlynE, Westfield College, London, England, for (16) We are greatly indebted t o Prof. 1%'. determining the O R D curves of these compoundsfor us.
.jw:mdro\f 2111-1 7p-ol-:3-onc acc.tatci.*' thiiy vorifirming t h i t csclriat ori:il vonfiguration of t hc fluoriiie atom. h i (LoIitra\t itti :Axial cdroino and a-rhlorokctonch. honevcr. T S g:ir :I ncgati\c t ~ u r w 'l'liib anomalous behavior of axial fluorine* has 1)ec.n noted p r iously. ~ * A recent report 1)y ,Illinger and aisotktes'F described the iiw of dipole moment data to prove the equatorial orientation oi 2a-fiuorocholestan-3-one I)>- direct cori-c>latioii\\-it11 c~ilrulatc d valuw. 1'1~liminary measurements galre dipole momcnt values 11 hich primaril! prove the a d configuratioii of thc fluoriiie in 2~-fliioi,o-.ioc-:~~itlrostane-3,17-dione (IV) when c3ompare.d with its cialculatcd x uluc~.IT Biological Activity.'*--The assay iised t o e ~ a l u n l ethe h?-pocholesterolemic activity of the 2fl-halo-5a-aiidr an-3oc-ol-l7-ollr~sn a h 011 adaptation of that used by (kirattiiii i i n ( onorlier. l'hc compounds were administered int raperit oiieally t o Trit oil-st imulat cd adult male rats. At a dose of 100 ing. 'kg.,2p-ioclo-Sa-androst~i~3a-01-17-one (IId) and 2~-chloro-5a-aiidr.ostai~-3a-~~l-l7-oiie (TI]) whibited significant inhibition of the Triton-induc~dliypercliolt+t crcilemia These compounds po of thc standard diphen\-leth~-lat.c.tic. :ic~itI (1,io~ol~w.pcc.ti\ ely By comparison, androsterone TI as innctil e 111 this a w i y iintlcr sirnihr vonditions. Sone of the 3fl-halo-Scr-androstane derivativc.: \m.: effectiw in 1 ~ ' ducing the plasma cholesterol level when administered orally t o rats made hypercholesterolemic with 0.2c7, propylthiouracil. Compounds IId and IIe, howes.er, did show some activity in thik ashay ~r-hcnatlministered subcutaneously I n comparing compounds IX and X, interesting biological results were noted. While 3cr-fluoro-.;a-androstan-liU-ol-8-oiieacetate (Xi exhibited anabolic activity in the immature rat as measured by the increase in weight of the levator ani muscle, the 2p-fluoro isomer (IX) was inactive at the same dose level. Both compounds, however, exhibited antiestrogenic activity. Studies are now in progresb to erald a t e 2fl-fluoro-5a-androstan-1 i@-ol-S-one acctate as an inhibitor of mammary fibroadenoma. ( 8
(1G)
N. L. Allinger, H. M. Blatter, >I. .I. DaRooge, and L. .i, I'rpiheig, .7. O w . Chenr., 26,
25.50 (1961).
(17) We are extremely grateful t o Dr. Norman 1,. Ailinger of JI-aynf. State I'nivemity for (leternlining and interpreting the dipole moment d a t a for t h i s compound. Tti~st'data will hr reported by Dr. -4llinger in J. Ore. Chem.. in press. (18) We are grateful t o Drs. Francis J. Saiinderu, Robert E. Ranney, Donald L. C u o k , n n ~ 1 M r . E. F. Xutting of our Biological Research Division for furnishing 11swith this information. (19) S. Garsttini. C. Morpurgo. B. Miirelli, R. Paolrtti, and S . Ikssprini, Arch. In!. Phnrrniirodyn. Therap.. 109, 400 (1957).
November, 1962
HYPOCHOLESTEROLENIC AGENTS. I1
1221
Experimentalz0 2p-Fluoro-5a-androstan-3a-oI-17-one(IIa).-To a solution of anhydrous hydrogen fluoride (20.8 g.) in purified tetrahydrofuran (38 ml.) and chloroform (14 ml.) cooled in a Dry Ice-isopropyl alcohol bath was added dropwise and with rapid stirring 2a,3a-epoxy-5a-androstan-17-one4(I, 12.0 g.) in chloroform (64 ml.). The addition required 2.5 hr. The reaction mixture remained 1.5 hr. in the cold and was then allowed to come to room temperature. The contents were poured slowly into 15% aqueous potassium carbonate solution (800 ml.). Chloroform (100 ml.) waa added and the solution stirred rapidly. Additional 15% aqueous potassium carbonate solution (250 ml.) was added and the organic layer separated. The aqueous layer was extracted with chloroform and the combined organic layers were washed successively with water, 5% sodium bicarbonate solution and water. The extract was dried over anhydrous potassium carbonate containing Darco and the solvent removed in vacuo. The resulting glass waa taken up in benzene and adsorbed onto silica gel (1 kg.). Elution with benzene-ethyl acetate (7: 1) gave IIa (6.6 g.), map.184-185', [ a I z a D f97'. Anal. Calcd. for C19H29FOz:C, 73.99; H, 9.48; F, 6.16. Found: C, 74.10; H, 9.53; F, 5.90. 2~-Chloro-5a-androstan-3a-ol-17-one (IIb). General Method.-A hetero(I, 4.0 g.), concd. hydrogeneous mixture of 2a,3a-epoxy-5a-androstan-17-one4 chloric acid*' (110 ml,), and chloroform (250 ml.) was placed on a mechanical shaker for 20 min. The chloroform layer waa separated and washed successively with 5% sodium bicarbonate solution and water. The extract waa dried over anhydrous sodium sulfate containing Darco. Removal of the solvent in vacw left a white solid which was recrystallized from ethanol-water to give I I b (2.7 g.), ~ m.p. 197-199'dec., [ a I N$92". Anal. Calcd. for C10H20C102:C, 70.24; H, 9.00; C1, 10.92. Found: C, 70.13; H, 8.89; C1, 10.81. 2p-Fluoro-5a-androstan-3~-ol-17-one Acetate (IIe). General Method.-A solution of 2p-fluoro-5a-androstan-3a-ol-17-one (IIa, 0.4 g.) in acetic anhydride (2.0 ml.) and dry pyridine (4.0 ml.) was stirred at room temperature for 3.5 hr. The solution was poured into ice and water (30 ml.). An oil separated which mas extracted with ether. The extract was washed successively with 501, hydrochloric acid, 5% sodium bicarbonate solution, and water. The organic phase waa dried over anhydrous potassium carbonate containing Darco. Removal of the solvent in vacw left an oil which crystallized from ethanol-water to give IIe (0.32g.), m.p. 123-125', [a]zsD 113.5'. Anal. Calcd. for CzlHalFOa: C, 71.97; H, 8.92. Found: C, 72.18; H, 8.65. 2p-Fluoro-5a-androstane-3a,l7j3-diol (IIIa). General Method.-A solution of lithium-tri. tert-butoxyaluminohydride2a (2.5 g.) in tetrahydrofuran (15 ml.) was added to 2~-fluoro-5a-androstan-3a-ol-17-one (IIa, 1.0 g.) in tetrahydrofuran (15 ml.) cooled in an ice bath. After stirring for 3 hr. the colution was poured into a mixture of ice and water (150 ml.) containing acetic acid (15 ml.). The
+
(20) Optical rotations, spectra, and analytical data were furnished by our Analytical Department under the supervision of Dr. R. T. Dillon. Optical rotations and infrared spectra were obtained in chloroform and ultraviolet epectra in methanol unless otherwise indicated. Velting points are uncorrected. (21) Substitution of 48% aqueous hydrobromic acid (80 ml.) and 47% aqueous hydriodic e u d (50 ml.) in this procedure afforded compounds IIc and IId, respectively. (22) Obtained commercially from Metal Hydrides, Inc.
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Novpmbpr, 1962
HYPOCHOLESTEROLEMIC AGENTS. TI
1223
precipitate was collected, washed with water, and air dried. Recrystallization from ethanol-water gave IIIa (0.8g.), m.p. 242-244'. Anal. Calcd. for ClsHs1F02: C, 73.51; H, 10.07. Found: C, 73.68; H, 10.07. 2p-Iod0-5~androstane-3~,17p-diol(IIId).-A heterogeneous mixture of 2~,3~-epoxy-5a-androstan-17p-o14 (2.0 g.) and 47% aqueous hydriodic acid (23 ml.) in chloroform (75 ml.) was agitated mechanically for 20 min. at room temperature. A small amount of dark brown precipitate which formed was collected and discarded. The tan-colored chloroform layer was separated and washed successively with dil. sodium thiosulfate solution (20 ml., 0.004 N ) , 5y0aqueous sodium bicarbonate solution and water. The extract was dried over anhydrous sodium sulfate containing Darco. Removal of the solvent i n vucw left a solid which was recrystallized from acetone to give IIId (1.2 g.), m.p. 141-142', dec., [O,Iz6D+54' (dioxane). I O54.54; ~ : H, 7.47. Found: C, 54.63; H, 7.54. Anal. Calcd. for C ~ ~ H ~ I C, 2p-Fluoro-5a-androstane-3,17-dione (IV).-To a solution of 2p-fluoro-5arandrostan-3~-01-17-one (IIa, 0.5 g.) in acetone (20 ml.) was added dropwise and with stirring a standard chromic acid solution28until the color of the reagent persisted. The excess of chromic acid was decomposed by adding a drop of iscpropyl alcohol. The organic layer was decanted from its inorganic salts and poured into ice and water. The precipitate was collected, washed with water, and air dried. Recrystallization from methanol-water gave IV (320 mg.), m.p. 142-143", €% 42, Amax 3.4, 5.75 p , [o(]2iD +123". Anal. Calcd. for ClgHz,F02: C, 74.47; H, 8.88. Found: C, 74.16; HI 8.93. 2a-Fluoro-5a-androstane-3,17-dione (V).-A solution of 2p-fluoro-5a-androstane-3,17-dione (IV, 100 mg.) in glacial acetic acid (3.5 ml.) and 48% aqueous hydrobromic acid (0.5 ml.) was allowed to stand for 20 hr. at room temperature. The reaction mixture was poured into ice and water and the precipitate collected, washed with water, and air dried to give V, m.p. 203-205" dec. (reported6m.p. 204-205'), 8%60, Amaz 3.4, 5.75p, [a]"D +138'. 2-Fluoro-5~-androst-l-en-3,17-dione (VI).-To 2~-fluoro-5~-androstane-3,17dione (VI 3.0 g.) in glacial acetic acid (25 ml.) and 48y0 aqueous hydrobromic acid (0.1 ml.) cooled to 10-15' was added dropwise with stirring over 0.5 hr. a solution of bromine (1.6 9.) and sodium acetate (0.82 g.) in glacial acetic acid (15 ml.). The reaction was stirred for 2 hr. and poured into ice and water (250 ml.) containing sodium acetate (0.8 g.). The precipitate was collected and washed successively with water and 5% sodium bicarbonate solution. The crude 2-bromo derivative (1.3 g.) in dimethylformamide (10 ml.) containing lithium chloride (0.45 g.) and lithium carbonate (0.27 g.) was refluxed for 2.5 hr. in a nitrogen atmosphere. Ether (20 ml.) was added and the total solution poured into cold water. The aqueous layer was extracted with ether and the combined extract washed successively with 5% hydrochloric acid, 5% sodium bicarbonate solution, and water. The extract was dried over anhydrous potassium carbonate containing Darco and the solvent removed in vacuo. The oily residue was dissolved in benzene and adsorbed onto silica gel (50 g.). Crystallization of the benzeneethyl acetate (19: 1) eluate from acetone-hexane gave VI (0.3 g.), m.p. 188-190", 236-237 5 emnl 8600, $122'. Anal. Cdcd. for C1,H~jFO5:C, 74.97; H, 8.23. Found: C, 74.74; H, 8.55. (23) (1946).
K. Bowden, I. hl. Heilbron, E. R . H. .Jones, and B. C. L.Weedon, J. Chem. Soc., 39
1224
R. E. COUNSELL, P. D. KLIMSTU,A K D It. E. 1Zasxti.l-
L'ol. 5
2B-Fluoro-5a-androstne-3a,l7@-diol17-Monoacetate (VIII).---Treatment ol 2a,3a-epoxy-5a-androstan-17fl-o1 acetate4 (VII, 5.6 g.) with hydrogen fluoride as described above gave pure VI11 (2.8 g.), m.p. 204.5-205.5', [a]% 12". Anal. Calcd. for CzlHaFO~:C, 71.56; H, 9.41. Found: C, 71.67; H, 9.47. 2p-Fluoro-5a-androstan-l7@-ol-3-one 17-Acetate (IX).-Chromic acid oxidation of 2p-fluoro-5a-androstane-3a,17@-diol17-monoacetate (VIII, 1.0 g.) as described above gave I X (0.4 g.), m.p. 153-154', e% 24.6, A,, 3.38, 5.75, 7.91 p, [ a ] D 2 s +38'. Anal. Calcd. for C21HS1F08: C, 71.97; H, 8.92. Found: C, 71.91; H, 9.20. 2a-Fluoro-5a-androstan-l7~-ol-3-one 17-Acetate (X).-A sample of 26fiuoro-5a-androstan-17p-ol-3-one 17-acetate (IX, 100 mg.) was epimerized as described above to give X as white needles (70 mg.), m.p. 192-194' (reported6 m.p. 190-193'), e% 18.5, A,,,3.38,5.75, 7.92p, [a]% +46.5". 2-Fluoro-5a-androst-l-en-l7@-ol-3-one 17-Acetate (XII).-To a solution of anhydrous hydrogen fluoride (7.0 g.) in glacial acetic acid (60 ml.) was added with rapid stirring during 20 min. a solution of la,2cu-epoxy-5a-androstan-l7j3ol-&one 17-acetatez4(XI, 4.0 g.) in glacial acetic acid (140 ml.). The solution warmed to 40" and was controlled by means of a water bath. After allowing the solution to stand at room temperature for 17 hr., it waa poured into ice and water (650 ml.). The resulting semi-solid was extracted with ether. The extracts were washed with 5y0 sodium bicarbonate solution and dried over anhydrous potassium carbonate containing Darco. Solvent removal in vacuo left a solid which waa recrystallized twice from acetone-hexane t o give XI1 (2.05 g.), m.p. 172-175', 238-237.6 Emax 8200, [ C Y ] % +59". Anal. Calcd. for C21HzsFOs: C, 72.38; H, 8.39. Found: C, 72.84; H, 8.45.
+
(24)
W.M. Hoehn, J . Ore. Chem., 98,929 (1958).
.
Hypocholesterolemic Agents 111. N-Methyl-N-(dialkylamino)alkyl-17~aminoandrost-5-en-3~-01Derivatives R. E. COUNSELL, P. D. KLIMSTRA AND R. E. RANNEY Division of Chemical Research of G. D. Searle & Go., Chicago 80, Illinois Received June 16, 1968
A series of N-methyl-N-( dialky1amino)alkyl derivatives of 17j3-aminoandrost5-en-3p-01 sterically similar to cholesterol was synthesized by the Leuckart reductive amination of readily available 3,!3-hydroxyandrost-5-en-17-one followed by reduction with lithium aluminum hydride. The ability of 17-ketosteroids t o form stable Schiff bases when condensed with primary amines afforded an alternate path to these compounds. Several compounds in this series were found to exhibit pronounced oral hypocholesterolemic activity when evaluated in rats.
One approach to the development of hypocholesterolemic agents has involved the synthesis and biological evaluation of compounds
