J. Med. Chem. 1993,36, 243-248
`
`243
`
`Phenyl-Substituted Prostaglandins: Potent and Selective Antiglaucoma Agents
`
`Bahram Resul:pt
`Johan Stjernschantz,t Kiyo No,? Charlotta Liljebris,? Giiran SelBn,? Maria Astin,t
`Maritha Karlsson,t and Laszlo Z. Bib*
`Kabi Pharmacia AB Ophthalmics, Uppsala, Sweden, and Columbia University, College of Physicians and Surgeons,
`Department of Ophthalmology, New York, New York 10032
`Received September 10, 1992
`
`A series of phenyl-substituted analogues of prostaglandin Fa, (PGF2,) were prepared and evaluated
`for ocular hypotensive effect and side effects in different animal models. In addition, the activity
`of the analogues on FP receptors was studied in vitro. The results were compared with those of
`PGF2, and ita isopropyl ester. The phenyl-substituted PGF2, analogues exhibited good intraocular
`pressure reducing effect, were more selective, and exhibited a much higher therapeutic index in
`the eye than PGFZ, or ita isopropyl ester. The analogues exhibited high activity on FP receptors
`in a stereoselective manner for the 15,-hydroxyl group.
`
`on
`
`4 a,b
`
`S c b
`
`Introduction
`Glaucoma, a potentially blinding eye disorder, is char-
`acterized by increased intraocular pressure (IOP), exca-
`vation of the optic nerve head, and gradual loss of visual
`field. Initial treatment usually involves topical application
`of muscarinic agonists, particularly pilocarpine, or adr-
`energic agonists or antagonists, e.g. epinephrine and
`timolol, respectively. If treatment with such topically
`applied drugs is not effective, systemic administration of
`carbonic anhydrase inhibitors or surgical intervention may
`be employed.
`Recently, attention has been focused on prostaglandins
`(PGs), primarily prostaglandin Fz, esters as IOP-lowering
`
`sub~tances.~~~J Several studies indicate that PG:s of the
`F2, type reduce IOP by increasing uveoscleral outflow of
`aqueous hum0r.~9~ This is a new principle of reducing
`IOP for therapeutic purpose.
`PGF2, isopropyl ester 1 b (Figure 1) significantly reduces
`IOP.lp2J However, this substance is not suitable for
`therapeutic use due to side effects such as superficial
`irritation and vasodilation in the conjunctiva. Conse-
`quently, the therapeutic selectivity of PGF2, in the eye is
`low. In an attempt to increase ocular selectivity, we have
`prepared a number of phenyl-substituted PGFk analogues
`and found that such analogues with modified omega chains
`containing an aromatic ring possess high selectivity and
`display good IOP-lowering activity. In this paper we report
`the synthesis and biological activity of two 17-phenyl-
`PGF2, analogues, 3 and 5, and their 15-epimers,4 and 6
`(Figure 1). The biologic activity of these analogues has
`been compared with that of PGF2, and its 15-epimer.
`Chemistry
`7
`a Reagents. (a) DBU, i-Pr Uacetone; (b) DDQ (3.0equiv)/dioxane;
`Synthesis. 17-Pheny1-18,19,20-trinor-PGF~,
`isopropyl
`(c) CeC1~7Hz0, NaBHJmethanol.
`ester (3b) and (15R)-17-phenyl-18,19,20-trinor-PGF~.
`isopropyl ester (4b) are prepared as shown in Scheme I.
`silica gel, using ethyl acetate, to achieve separation of the
`The commercially available compound 3 is esterified in
`isomers 3b and 4b. Compound 2b is prepared in a similar
`acetone with isopropyl iodide in the presence of DBU6 to
`way using PGF2,.
`yield 3b which is oxidized in dioxane with DDQ7y8 to give
`13,14-Dihydro-l7-phenyl-l8,19,20-trinor-PGF~,
`isopro-
`the enone 7. The unsaturated ketone 7 is reduced in
`pyl ester (Sb) and (15S)-13,14-dihydro-17-phenyl-18,19,-
`methanol with sodium borohydride in the presence of
`20-trinor-PGF% isopropyl ester (6b) are prepared as shown
`cerium chloride heptahydrates-ll to give the epimeric
`in Scheme 11. The primary alcohol of the bicyclic lactone
`mixture of 3b and 4b which is then chromatographed on
`W-14 is oxidized using the Pfitzner-Moffat method1"17
`(DCC, DMSO, and &Pod) in dimethoxyethane (DME)
`to afford the aldehyde 9. Without isolation the aldehyde
`0 1993 American Chemical Society
`
`6 a,b
`
`a R=H
`b R i - P I
`Figure 1. Structures of compounds referred to in the text by
`numbers.
`Scheme IJ
`Y
`
`on
`
`3s
`
`h
`
`3b
`
`+ Kabi Pharmacia AB Ophthalmics.
`t Columbia University.
`
`0022-26231931 1836-Q243$Q4.00lQ
`
`Micro Labs Exhibit 1047
`Micro Labs v. Santen Pharm. and Asahi Glass
`IPR2017-01434
`
`

`

`244 Journal of Medicinal Chemistry, 1993, Vol. 36, No. 2
`Scheme 118
`
`Resul et al.
`
`OPPB
`
`0
`
`8
`
`9
`
`0
`
`IO
`
`i‘
`i
`
`d”
`
`5a + 6a - 5b + 6b
`
`h
`a Reagents: (a) DCC, DMSO, H3POdDME; (b) (CH30)zPO-
`CH~CO(CHZ)ZC&, NaH/DME; (c) CeC1~7Hz0, NaBHdMeOH; (d)
`Pd/C, 1 M NaOH/EtOH; (e) KzC03/MeOH; (0 DIBAL/THF, -78
`OC; (g) Ph3P(CHz)rCOOH, KOtBu/THF, -10 OC; (h) DBU,
`ICH(CH&/acetone, room temperature.
`9 is reacted further with dimethyl (2-oxo-4-phenylbutyl)-
`phosphonate using the Wadsworth-Emmons method to
`give lO.l*Jg The resulting crystalline @unsaturated
`ketone was reduced stereoselectivity with lithium tri-sec-
`butylborohydride (Lithium Selectride)2°*21 at -120 “C to
`fumiah a 7r3 mixture of 15S-i~omer 11 m d 15R-isomer 12.
`Alternatively, sodium borohydride in the preaence of
`cerium chloride heptahydrate in methanol at room tem-
`perature can be used to reduce the enone with lower
`selectivity. The epimers are separated by flash column
`chromatography on silica gel to give 11 as a solid
`(crystallized from ethanol) in 45% yield. This is reduced
`under hydrogen atmosphere in ethanol using Pd-C as
`
`catalyst in the presence of sodium h y d r o ~ i d e ~ ~ l ~ ~ to give
`compound 13 as an oil in quantitative yield. The phe-
`nylbenzoyl group is removed wing potassium carbonate
`in methanol, affording compound 15 as an oil, which was
`purified by column chromatography on silica gel using
`EtOAc as eluent. The lactone 15 is treated with diisobu-
`tylaluminum hydride (DIBAL)24126 in dry THF at -78 OC
`to give the lactol 17 as a solid in good yield. The Wittig
`reaction26 with (4-carboxybutyl)triphenylphosphonium
`bromide and potassium tert-butoxide in THF to afford
`the acid Sa. This is further reacted without isolation with
`isopropyl iodide and DBU in acetone to give the ester Sb,
`which is isolated by gradient flash column chromatography
`using first dichloromethane then ethanol in dichlo-
`romethane (5.0-7.5%) to give the desired product 5b as
`an oil. Compound 6b (Scheme 11) is prepared in a similar
`way using the 1SR isomer 12.
`Pharmacology
`The compounds were tested for IOP-reducing effect,
`effect on the pupil, ocular irritation, and conjunctival
`
`Concentration (mol/l)
`Figure 2. Effect of the test compounds on the cat iridialsphincter
`muscle. Each point represents the mean of four values obtained
`on four different preparations. Bars indicate SEM. The ECw
`vdues€af eompmmds ln4a ara 8.4 x lV,?.1 X 1W,7.1 X l&W,
`3.0 X 10-8, 3.6 X lo4, and 2.4 X 10” M, respectively.
`hyperemia. IOP was measured in cynomolgus monkeys.
`The cat eye was used as a model for ocular irritation.
`Conjunctival hyperemia (surface hyperemia of the eye)
`was evaluated in albino rabbits. The activity of the test
`compounds was also tested in a muscle bath using isolated
`cat iris sphincters. This muscle expresses predominantly
`FP receptor^.^'
`Compound 3a exerted the strongest contractile effect
`on the feline iris sphincter muscle with an ECm = 7.1 X
`lO-lo M. Interestingly, this compound exhibited higher
`potency than the naturally occuring ligand (la) (Figure
`2). Inversion of the 15-hydroxyl group decreased the
`activity in all the analogues.
`Compounds lb, 3b, and Sb effectively reduced IOP in
`monkeys while the 15-epimers had much weaker effects
`(Table I). All compounds, in particular lb, 3b, and Sb,
`decreased the pupil diameter of the cat, while in monkeys
`a slight increase in pupil diameter was observed with some
`of the phenyl-substituted compounds.
`None of the phenyl-substituted analogues (compounds
`3b-6b) caused irritation, whereas l b and 2b had a clear-
`cut irritating effect on feline eyes (Table I). While all
`analogues caused some conjunctival hyperemia, this side
`effect was much less pronounced with the phenyl-sub-
`stituted analogues, ae compared to the isopropyl ester of
`PGF2,1 (compound lb; Figure 3).
`
`Micro Labs Exhibit 1047-2
`
`

`

`Antigkaucoma Phenyl-Substituted Prostaglandins
`
`Table I. Maximum Reduction of the Intraocular Pressure in
`Normotensive Monkeys and Maximum Irritation of Cat Eyes
`aftar Topical Adminietration of 3 rcg of Compounds lb-6b
`(n = 6)
`
`maximum reduction
`maximum
`irritation of cat eyee
`of IOP in monkeys
`compound
`(arbitrary unit@
`no.
`(mmHg)"
`2.5 * 0.3***
`2.5 & O.O***
`lb
`2.3 i 0.1***
`0.7 t 0.5
`2b
`1.9 * 1.0
`0.2 t 0.2b
`2.9 1.0*
`3b
`0.0 t 0.0b
`4b
`1.0 * 0.2**
`2.6 i O.4**
`Sb
`0.0 0.0
`6b
`0.0 0.0
`p < 0.001 (matched pair t-test).
`0 (*) p < 0.05; (**) p < 0.01; (I**)
`*Dose = 5 rcg in cats.
`
`0.01
`
`0.03
`
`0.1
`
`0.3
`Dose ( P a
`Figure 3. Maximum difference in hyperemia (experimental eye -
`control eye) of rabbit eyes treated with the test compounds. Each
`point representa the mean derived from six animals.
`
`1
`
`3
`
`10
`
`Conclusions
`Replacement of the C l g m fragment in PGF2, and ita
`13,lddihydro derivative with a phenyl group affords
`analogues with unique properties, in that they exert ocular
`hypotensive effects with greatly reduced side effectswhen
`applied topically to the eye. The 13,lbdihydro derivative
`Sb is a nearly ideal ocular hypotensive compound that
`lacks significant irritative and conjunctival hyperemic side
`effects while retaining full ocular hypotensive potency in
`primates. We speculate that the phenyl ring imposes a
`conformational change on the omega chain that enables
`discrimination between yet undefined prostaglandin re-
`ceptor subtypes.
`Experimental Section
`Chemistry. All chemical reagents were commercially avail-
`able. Melting pointa were determined in open glass capillaries
`on a Buchi apparatus. Flash column chromatography was
`performed on silica gel 60,230400 mesh (E. Merck). Thin-layer
`chromatography (TLC) was carried out on silica gel 60 F245 glass
`plates and visualized by spraying the plate with 3 % cupric acetate
`in 15% phosphoric acid or 5% phosphomolybdic acid in ethanol,
`followed by heating. *H and 1 4 2 NMR spectra were recorded in
`CDClaor CDJOD,usingtetr~ethy~ilaneasaninte~staadard,
`with a JEOL-200 or a Varian-600 instrument. Chemical shifta
`are expreased as & values. All the spectra were in accordance
`with the assigned structures. Specific rotations were measured
`with a Perkin-Elmer 241 polarimeter. Analytical HPLC was
`performed on a Nucleosil CIS column (7 pm, 250 x 4 mm) in a
`gradient system using phosphate buffer pH 2.5/CH&N as the
`mobile phase, flow rate 1.8 mL/min. Preparative HPLC was
`performed on a Ghn-303 system using a silica gel column 3 x
`30cm, the mobile phase consisting of 6-10 % ethanol in n-hexane
`
`Journal of Medicinal Chemietry, 1993, Vol. 36, No. 2 246
`at a flow rate of 15 mL/min. Spectrophotometric detection waa
`performed at 200 nm in both cases. Elemental analytw were
`performed by Mikrokemi AB, Uppsala and were within 0.4% of
`the calculated value.
`17-Phenyl- 18,19,2O-trinor-PGF& Isopropyl Enter (3b).
`DBU (0.37 g, 2.43 "01)
`was added dropwine to a stirred solution
`in acetone (16 mL) at 0 OC. The
`of So (0.40 g, 0.81 "01)
`was allowed to warm to room temperature whereupon itwpropyl
`was added dropwise. After b e i i stirred
`iodide (0.41 g, 2.43 "01)
`for 10 h (TLC monitoring), the reaction was quenchedwith water,
`the mixture was extracted with EtOAc (50 mL), and the extzact
`was washed with brine (20 mL), citric acid 3% (30 mL), and
`f i i y sodium hydrogen carbonate 5 % (2 X 20 mL). After drying
`with anhydrous sodium sulfate, the solvent was removed in vacuo
`and the residual oil was chromatographed on silica gel wing
`EtOAc as eluent. This afforded 0.34 g (80%) of the title
`compound as a colorless oil: R/ = 0.24 (EtOAc); [a]% = +33.49O
`(C 0.74, CHsCN); 'H NMR (CDCh) 6 1.2 (6,6 H (C&)l), 1.48
`(m, 1H), 1.64(m,2H), 1.74-1.78(dd, 1H),1.86 (m,2H), 1.78-1.9
`(m, 2 H), 2.08 (m, 2 H), 2.23 (t, 2 H), 2.24 (m, 2 H), 2.27 (m, 1
`H), 2.3 (m, 1 H), 2.62-2.72 (m, 2 H), 3.9 (m, 1 H), 4.07 (t, 1 H),
`4.13 (m, 1 H), 4.9 (m, 1 H), 5.43-5.59 (m, 4 H), 7.1-7.3 (m, 5 H);
`IaC NMR (CDCh) 6 173.37, 141.9, 128.3, 125.7, 134.6 (C db),
`132.03 (C db), 129.6 (C db), 129.04 (C db), 78.01, 72.83, 71.50,
`67.55, 55.6, 42.9, 38.70, 33.90, 31.70, 26.50, 25.50, 24.80, 21.76,
`21.75.
`Enter (4b).
`17-Phenyl-1~-18,19~O-trinor-PG~~I~p~~yl
`To a stirred solution of 3b (0.24 g, 0.55 m o l ) in dioxane (8.0
`mL) was added portionwise DDQ (0.51 g, 2.2 "01)
`whereupon
`the mixture became brown. After b e i i stirred at room tem-
`perature for 24 h (TLC monitoring), the mixture wm ritered,
`the precipitate was washed with ether (3 X 50 mL), and the
`combined organic phase was washed with water (2 X 20 mL), 1
`M NaOH (3 X 30 mL), and brine (2 X 20 mL). Drying with
`sodium sulfate and concentrating in vacuo gave an oily residue
`which after chromatography on silica gel using ether as eluent
`gave 180 mg (76%) of the enone 7. This was dholved in
`&chloromethane (5 mL) and added dropwise to a suapension of
`sodium borohydride (0.01 g, 0.25 m o l ) and cerium chloride
`in methanol-CH&ls 2:l (12
`heptahydrate (0.05 g, 0.13 "01)
`mL). After 30 min, the mixture was quenched with saturated
`ammonium chloride, extracted with EtOAc (2 X 30 mL), and
`dried with sodium sulfate. The solvent was removed in vacuo,
`and the residue was chromatographed on silica gel, achieving
`separation of the isomers 3b 67.5 mg (28% ) and 4b 98.6 mg (41 7% )
`which was obtained as a colorless oil Rf = 0.32 (EtOAc); [ul%
`= +42.32' (15 mL) (C = 2.1, CHsCN); 'H NMR (CDCb) 6 1.2 (a,
`6 H), 1.46 (m, 1 H), 1.65 (m, 2 H), 1.74-1.78 (dd, 1 H), 1.86 (m,
`2 H), 1.78-1.90 (m, 2 HI, 2.15 (m, 2 HI, 2.25 (t, 2 H), 2.64-2.77
`(m, 2 H), 3.93 (m, 1 H), 4.15 (t, 1 HI, 4.16 (m, 1 HI, 5.0 (m, 1 H),
`5.39-5.44(m,2H),5.61-5.65(m,2H),7.1-7.3(m,6H);1FNMR
`(CDCh) 6 173.37,141.9,128.3,125.7,134.6 (Cdb), 132.03 (Cdb),
`129.6 (Cdb), 129.04 (Cdb), 78.01,72.83, 71.50,67.55,55.6,42.9,
`38.70, 33.90, 31.70, 26.50, 25.50, 24.80, 21.76, 21.75.
`Dimethyl (2-Os~~phenylbutyl)phorphonots. To a stirred
`suspension of sodium hydride (7.20 g, 300 "01)
`previowly
`washedwithn-penteneindryTHF(2M)mL)atroomtsmperakve
`was added dropwiw a solution of dimethyl (2-oxopropy1)-
`phoephonate(47.5g,285.9mmol)inTHF (1lOmL). Thereaction
`mixture was stitred for 2 h, then cooled in an ice bath and treated
`with a solution of n-BuLi (22 g, 340 "01)
`in hexane, cawing
`a dark brown solution to be formed. S t i r r i i was continued for
`2 h at 0 "C, followed by dropwise addition of benzyl bromide
`(53.8 g, 314.6 "01)
`in THF (50 mL). T h e reaction mixture WM
`gradually warmed to room temperature and after 3 h (TLC
`monitoring),itwasquenchedwith lMHCl(20mL). Themixture
`was poured into ice-water (200 mL) and extracted with CHCb
`(2 x 150 mL), the organic layers were collected, w a d d with
`brine (150 mL), and chromatographed on silica gel using CH&la
`and EtOAc successively as eluent, furnishing 43.4 g (68%) of a
`(CDCh) 6 2.9 (m, 4 H, (CH2)9), 3.04-3.14 (d, 2 H), 3.69-3.79 (d,
`OCHs, 6 H), 7.11-7.38 (m, 5 H); lSC NMR (CDCb) & 29.14,39.92,
`42.48, 45.17, 52.70.
`
`slightly yellow oil: Rj - 0.23 (EtOAcacetone l:l), 'H NMR
`
`Micro Labs Exhibit 1047-3
`
`

`

`Resul et al.
`
`246 Journal of Medicinal Chemistry, 1993, Vol. 36, No. 2
`(lS,SR,GR,7R)-6-Formy1-7-[ (4-phenylbenzoyl)oxy]-2-
`71.53 (CH-CHCHOH), 54.04,42.66, 38.71, 37.55, 34.83,31.57.
`osabicyclo[ 3.3.01octan-3-one (9). To a solution of the alcohol
`Anal. (C31H~05) C, H.
`8 (15.0 g, 42.56 mmol) in DME (100 mL), cooled to 18 'C, were
`(lS$R,6R,7R)-6-[ (3R)-3-Hy~ry-S-phenyl-l-pentyl)-7-[
`(4-
`phenylbenzoyl)ory]-2-oxabicyclo[3J.O]octan-3-one (13). To
`added dicyclohexylcarbodiimide (DCC) (26.3 g, 127.69 mmol)
`a suspension of 10% Pd/C (0.4 g) in 1 M NaOH (1.0 mL, 0.4
`and phosphoric acid (1.43 mL, 21.28 mmol). The temperature
`mmol) and ethanol (15 mL) was added a solution of 11 (1.94 g,
`of the reaction mixture was kept below 25 'C for 30 min. The
`4.01 mmol) in ethanol (6.0 mL). The solution was stirred under
`reaction mixture was stirred at room temperature for additional
`hydrogen atmosphere for 6 h (TLC monitoring) and quenched
`2 h (TLC monitoring) and the precipitate was removed by
`with 1 M HC1. The catalyst was removed by filtration through
`filtration and washed with ether (2 X 50 mL). The combined
`a Celite pad, washed with ethanol absolute (20 mL). The solvent
`organic layer was washed with water (50 mL) and brine (2 X 50
`mL), the aqueous solution was extracted with ether (100 mL),
`was removed in vacuo. The resulting oil was dissolved in EtOAc
`(110 mL) and washed with brine 15% (30 mL). The water phase
`and the organic layers were collected and dried with sodium
`sulfate, filtered, and used directly for the next step: TLC Rf =
`was washed with EtOAc (30 mL). The combined organic extracts
`0.42 (silica gel, EtOAc-toluene 2:l).
`were dried with sodium sulfate and filtered. The solvent was
`removed in vacuo giving 13 as a colorless oil. Chromatography
`(1S,SR,6R,7R)-6-( 3-Oxo-6-phenyl- lE-pentenyl)-7-[ (4-phe-
`on silica gel using EtOAc as eluent yielded 1.51 g (78.4%): Rf =
`nylbenzoyl)oxy]-2-oxabicyclo[3.3.0]octan-3-one (10). To a
`0.44 (silica gel, EtOAc); [(YIzoD = 69.62' (c = 0.8, CH3CN); lH
`stirred suspension of NaH (3.08 g, 128.3 mmol), prewashed with
`NMR (CDC13) S 1.40 (m, 1 H), 1.60 (m, 4 H), 1.78 (m, 2 H), 2.15
`n-pentane, in DME (150 mL) under nitrogen was added dropwise
`(m, 1 H), 2.40 (m, 2 H), 2.50 (dd, 1 H), 2.63 (m, 2 H), 2.78 (m,
`dimethyl (2-oxo-4-phenylbuty1)phoaphonate (28.52 g, 113.3
`1 H), 2.90 (m, 1 H), 3.14 (m, CHOH), 5.08 (m, 1 H), 5.28 (9, 1 H),
`mmol) in DME (100 mL) and stirred vigorously for 1 h at room
`7.18 (m, 3 H), 7.26 (m, 2 H), 7.38 (m, 1 H), 7.45 (m, 2 H), 7.60
`temperature. The mixture was then cooled to -10 'C, and a
`(m, 2 H), 7.67 (m, 2 H), 8.50 (m, 2 H); 13C NMR (CDCb) S 176.81
`solution of the crude aldehyde 9 was added dropwise. After 30
`(C6HrC=O), 165.86 (lacetone C-O), 145.91, 141.72, 139.85,
`min at 0 "C and 2 h a t room temperature (TLC monitoring), the
`130.12, 128.85, 128.41, 128.35, 128.32, 128.11, 127.21, 127.12,
`reaction mixture was neutralized with AcOH, the solvent was
`125.89, 84.44 (CHzCHOCO), 80.13 (CsHlCOOCH), 70.88
`removed, and to the residue was added EtOAc (200 mL). The
`(CHzCHOH), 52.67,43.57,39.07,37.80,36.28,35.12,32.00,29.37.
`solution was washed with water (50 mL) and brine (50 mL). The
`organic layer was dried over anhydrous sodium sulfate. The
`Anal. (C31H3206) C, H.
`(lS,SR,6R,7R)-&[ (3S)-3-Hydroxy-5-phenyl- l-pentyl]-7-[ (4-
`solvent was removed in vacuo, the oil was stirred with ether (100
`phenylbenzoyl)olry]-29.abicyclo[3J.O]oc~-3-one (14). This
`mL) and the resulting white precipitate was filtered off and
`was prepared as compound 13 from the lactone 12 giving a
`washed with cold ether, giving a white crystalline substance: mp
`colorless oil yield 1.25 g (65 % ). The compound was used directly
`134-135.5 'c; yield 24.0 g (58.5%); [CY]~D = -116' (C = 1.26,
`for the next step, Rf = 0.47 (silica gel EtOAc).
`CH3CN); 'H NMR (CDCl3) 6 2.9 (m, 8 H), 5.1 (t, 1 H), 5.3 (9, 1
`H), 6.2 (d, 1 H), 6.7 (dd, 1 H), 7.1-7.6 (m, 10 HI, 8.1 (d, 4 HI; 13C
`(lS$R,6R,7R)-6-[ (3R)-3-Hydroxy-S-phenyl- l-pentyl]-7(il)-
`NMR (CDCl3) S 198.49 (CH-CHCO), 175.68 (c$H&o), 146.19,
`hydroxy-2-oxabicyclo[3.3.0]octan-3-one (15). To a solution
`of the lactone 13 (1.6 g, 3.1 mmol) in methanol (15 mL) was
`142.95 (CH-CHCO), 140.79, 139.75, 131.36 (CH-CHCO),
`added potassium carbonate (0.28 g, 1.98 mmol) and the mixture
`130.17, 128.89, 128.46, 128.34, 128.31, 128.21, 127.87, 127.22,
`127.17,126.15,83.11,78.50,54.07,42.56,42.44,37.80,34.90,29.96.
`stirred at ambient temperature for 6 h (TLC monitoring). The
`mixture was neutralized with 1 N HC1 and extracted with EtOAc
`Anal. (C31H&) C, H.
`(2 x 30 mL). The organic phase was dried on anhydrous sodium
`(lS,SR,GR,?R)-6-[ (3S)-3-Hydroxy-5-phenyl-l-pen~nyl]-7-
`[ (4-phenylbenzoyl)oxy]-2-oxabicyclo[3.3.O]octan-3-one (1 1).
`sulfate and evaporated to dryness. The crude product was
`chromatographed (silica gel EtOAc-acetone 1:l). The title
`Toa stirred solution of lithium tri-sec-butylborohydride (Lithium
`compound 15 was obtained as a colorless oil: yield 0.77 g (85% );
`Selectride) (5.14 g, 27.05 mmol) in THF-ether 1:2 (60 mL) at
`R\= 0.19 (EtOAc); [CY]~D = -17.60' (C = 0.34, CH3CN); 'H NMR
`-130 "C under nitrogen was added dropwise a solution of the
`(CDC13) S 1.28 (m, 1 H), 1.54 (m, 3 HI, 1.78 (m, 3 H), 2.15 (m,
`enone 10 (13.0 g, 27.05 "01)
`in THF (20 mL) cooled to -75/-78
`1 H), 2.28 (m, 1 H), 2.46 (m, 1 H), 2.51 (m, 1 H), 2.67 (m, 1 H),
`"C. The reaction mixture was stirred for 1 h (TLC monitoring)
`2.78 (m, 2 H), 3.60 (m, CHzCHOHCHz), 3.97 (m, CHOH), 4.92
`and then quenched with saturated ammonium chloride. The
`(m, CHOC=O), 7.18 (m, 3 H), 7.28 (m, 2 H); 13C NMR (CDCl3)
`temperature was raised to 0 'C, water (20 mL) was added, and
`6 177.55 (lactone C 4 ) , 141.81, 128.42, 128.33, 125.89, 83.88
`the mixture was diluted with EtOAc (80 mL). The organic layer
`(CHpCHOCO), 77.46 (CHzCHOH), 71.28 (CHzCHOHCHz),53.94,
`was separated and washed with brine, dried on anhydrous sodium
`43.22, 40.52, 39.08, 35.96, 35.20,32.03,28.95. Anal. (ClaHz404)
`sulfate, concentrated in vacuo, and chromatographed twice on
`silica gel using toluene-EtOAc 21 and 1:l successively as eluent,
`C, H.
`( lS,SR,6R,7R)-6-[ (3S)-3-Hydroxy-S-pbenyl-l-pentyl]-7(Rf-
`furnishing 11 as a white crystalline product: mp 108-110 'C;
`yield 6.8g (52% ); R, = 0.33 (silica gel, EtOAc-toluene 2:l); [CYIz0D
`hydroxy-2-oxabicyclo[3.3.O]octan-3-one ( 16). Prepared as
`compound 15 from the lactone 14. The product was chromato-
`= -101.59O (c = 0.69, CHaCN); 'H NMR (CDC13) 6 1.84 (m, 2 H),
`graphed on silica gel using ethyl acetate as eluent. This gave a
`2.25 (dq, 1 H), 2.5 (dd, 1 H), 2.60-2.90 (m, 6 H), 4.14 (m, CHOH),
`colorless oil: yield 0.75 g (83.6%); [(YIpoD = -8.26' (c = 0.79,
`5.06 (m, 1 H), 5.27 (9, 1 H), 5.59-5.72 (m, CH-CH), 7.10-7.28
`CH3CN); R, = 0.18 (EtOAc); 'H NMR (CDCb) 6 1.40 (m, 1 HI,
`(m, 6 HI, 7.38-7.48 (m, 2 H), 7.58-7.67 (m, 4 HI, 8.02-8.08 (m,
`1.54 (m, 2 H), 1.78 (m, 3 H), 2.15 (m, 1 HI, 2.28 (m, 1 HI, 2.48
`2 H); 13C NMR (CDCl3) S 176.24 (C&C=O), 165.89 (lactone
`(m, 2 H), 2.66 (m, 1 HI, 2.78 (m, 2 HI, 3.60 (m, CHpCHOHCHz),
`C 4 ) , 146.05, 141.50, 139.85, 136.02 (CH=CH), 130.14, 128.89
`3.97 (m, CHOH), 4.92 (m, CHOC=O), 7.18 (m, 3 H), 7.28 (m, 2
`(CH-CH), 128.71, 128.35, 128.21, 128.18, 125.92, 83.19
`H); 13C NMR (CDC13) 6 177.60 (lactone C d ) , 141.82, 128.41,
`(CHzCHOCO), 78.99 (C~HICOOCH), 71.39 (CH--CHCHOH),
`128.32, 125.87, 83.90 (CHpCHOCO), 77.27 (CHzCHOH), 71.06
`54.03, 42.72, 38.67, 37.58, 34.89, 31.56. Anal. (C31Hw06) C, H.
`(CHpCHOHCHz), 53.54,43.01,40.47, 39.16, 35.92, 34.90, 32.00,
`(lS,SR,BR,7R)-C[ (3R)-3-Hydroxy-5-phenyl- 1-pentenyll-7-
`28.80. Anal. (CleHuO4) C, H.
`[ (4-phenylbenzoyl)oxy]-2-oxabicyclo[3.3.O]octan-3-one ( 12).
`(lS,SR,6R,7R)-6-[ (3R)-3-Hydrolry-S-phenyl- l-pentyl]-7-hy-
`The preparation of this compound was achieved by chromato-
`droxy-2-oxabicyclo[3.3.0]octan-3-one (17). A solution of di-
`graphic separation of the isomers obtained by following the
`isobutylaluminium hydride (DIBAL) (1.53 g, 10.78 mmol) in dry
`procedure described for the preparation of compound 11. This
`gave a colorless oil: yield 4.3 g (33.1%); R, = 0.25 (silica gel,
`toluene (6.2 mL) was added dropwise to a stirred solution of the
`lactone 15 (0.82 g, 2.70 mmol) in dry THF (22 mL) at -72/-80
`EtOAc-toluene 2:l); [aI2O~ = -109.29' (c = 0.7, CH3CN); lH
`NMR (CDCld S 1.85 (m, 2 H), 2.24 (dq, 1 H), 2.52 (dd, 1 H),
`OC. After 1 h (TLC monitoring), the reaction mixture was
`quenched with methanol (5 mL) and was warmed to room
`2.58-2.90 (m, 6 HI, 4.12 (m, CHOH), 5.04 (m, 1 H), 5.26 (9, 1 H),
`temperature followed by addition of water (50 mL) and 1 M HCl
`5.56-5.72 (m, C H e H ) , 7.10-7.28 (m, 6 H), 7.38-7.48 (m, 2 H),
`(50 mL), and extracted with EtOAc (2 X 50 mL). The organic
`7.58-7.67 (m, 4 H), 8.02-8.08 (m, 2 H); 13C NMR (CDC13) S 176.27
`phase was dried with sodium sulfate and filtered, and triethyl-
`165.83 (lactone C=O), 146.01,141.46,139.79,136.16
`(C6H4-),
`amine (0.1 mL) was added to stabilize the triol. The solvent was
`(CH-CH), 130.11, 128.89 (CH-CH), 128.87, 128.78, 128.35,
`128.31,128.16,125.85,83.13 (CHpCHOCO), 78.92 (C6H&OOCH),
`removed in vacuo, the residue was chromatographed on silicagel
`
`Micro Labs Exhibit 1047-4
`
`

`

`Antiglaucoma Phenyl-Substituted Prostaglandins
`
`Journal of Medicinal Chemistry, 1993, Vol. 36, No. 2 247
`Pharmacology. The analogues were used as free acidsa in
`using EtOAc and EtOAc-acetone 1:1, respectively, as eluent to
`the in vitro tests and as isopropyl esters in the In vivo testa to
`give a white crystalline product: yield 0.54 g (65% 1; mp 102-104
`OC; R, = 0.31 (EtOAc-acetone 1:l); [a]%D = -22.08'
`enhance penetration into the eye.29 The compounds were
`(c = 0.18,
`THF); 1H NMR (CD30D) 6 1.24 (m, 1 H), 1.48 (m, 2 H), 1.58 (m,
`dissolved in a vehicle containing 0.9% sodium chloride and 0.5 7%
`polysorbate 80 for the in vivo experiments. The amount of drug
`2 H), 1.72 (m, 2 H), 1.94 (m, 1 H), 2.04 (m, 1 H), 2.26 (m, 2 H),
`2.64(m,lH),2.77(m,lH),3.54(m,lH),3.74(m,lH),4.42-4.56
`used in various experiments corresponds to free acid equivalents.
`(dq, 1 H), 5.38-5.54 (dd, 1 H), 7.12-7.26 (m, 5 HI; I3C NMR
`For the in vitro experiments the test substances were dissolved
`in 0.6 M NaHC03 solution titrated to pH = 7.4 using 1 M NaH2-
`(CD30D) 6 144.19, 129.88, 129.78, 127.15, 102.39, 101.57, 84.79,
`81.97, 79.93, 79.73, 72.52, 55.39, 55.19, 47.30,44.0, 42.59,42.22,
`PO4 to a final concentration of 0.1 mol/L. Further dilutions were
`41.98,40.75, 40.70, 36.97,33.52,31.49, 30.62. Anal. (C18H2604)
`made with 0.9% NaCl.
`C, H.
`IOP, Pupil Diameter, and Irritation in the Cat Eye.
`(lS,SR,6R,7R)-6-[ (3S)-3-Hyclroxy-5-phenyl-l-penty1]-7(R)-
`Domestic female cats weighing 2-3 kg and specially trained for
`hydroxy-2-oxabicyclo[ 3.3.01octan-3-one (18). Prepared as
`IOP measurements were used. One eye of each animal was
`compound 17 from the lactone 16. The product was chromato-
`topically treated with the drug (drop size 20 pL) and the
`graphed on silica gel using ethyl acetate and ethyl acetawacetone
`contralateral eye received the vehicle solution. The intraocular
`1:l as eluent. This gave a colorless oil: yield 0.34 g (83% 1; [(YI2OD
`pressure was measured using a pneumatonometer (Digilab
`= -13.23" (c = 0.4, CH,CN); R, = 0.32 (EtOAc-acetone 1:l); lH
`Modular One, Bio-Rad) under local anaesthesia with oxibup-
`NMR (CDC13) 6 1.32-1.68 (m, 4 H), 1.74 (m, 2 H), 1.96 (m, 1 H),
`rocain. The horizontal diameter of the pupil was measured with
`2.12 (m, 2 H), 2.14-2.34 (m, 4 H), 2.64 (m, 1 H), 2.66 (m, 1 H),
`a millimeter ruler under constant illumination conditions.
`2.78 (m, 1 H), 3.Nk3.94 (dq, 1 H), 4.55-4.66 (m, 1 H), 5.46-5.64
`Measurements of IOP and pupil diameter were performed before
`(dd, 1 H), 7.15-7.20 (m, 3 H), 7.25-7.29 (m, 2 H); 13C NMR (CDCl3)
`drug treatment and 1,3,6, and 23 h after treatment. The ocular
`6 142.01,128.37,128.35,125.79,100.99,99.92,85.97,81.87,79.63,
`irritation was evaluated from the behavior of the animals, in
`79.02,71.03,70.94,54.66,54.21,47.71,46.16,42.49,41.28,41.17,
`particular the degree of lid closure during the first hour after
`40.29, 39.14,39.10,35.26, 35.20,32.07,32.05,30.78,29.29,29.25.
`drug treatment. An arbitrary scale from 0 to 3 was used, 0
`Anal. (C18H2604) C, H.
`indicating absence of irritation and 3 complete closure of the
`13,14-Dihydro-17-phenyl-18,19,20-trinor-PGF~a
`(Sa). To
`lids.
`a stirred suspension of (4-carboxybutyl)triphenylphosphonium
`IOP and Pupil Diameter in the Monkey Eye. Cynomolgus
`bromide (2.32 g, 5.22 mmol) in THF (20 mL) under nitrogen at
`monkeys specially trained for IOP measurements were used. The
`0-5 "C was added potassium tert-butoxide (1.18 g, 10.44 mmol)
`animals were sedated with ketamine (2-3 mg/kg body weight)
`and the mixture stirred for 30 min at room temperature. To the
`for transportation from the animal housing facility to the
`resultant red-orange solution of ylide at -15/-10 OC was added
`laboratory and placed in specially designed chairs. Oxibuprocain
`the lactoll7 (0.4 g, 1.30 mmol) in THF (5 mL), and the mixture
`was used for local anaesthesia. IOP was measured with a
`was stirred for 3-4 h (TLC monitoring). The reaction mixture
`pneumatonometer (Digilab Modular One) calibrated for monkey
`was diluted with water (20 mL) and washed with ether (4 X 40
`eyes30 and the pupil diameter was measured with a millimeter
`mL). The water layer was acidified with 5% citric acid to pH
`ruler. The drug was applied to one eye (drop size 10 pL) and the
`4 and extracted with EtOAc (2 X 40 mL). The organic phase was
`contralateral eye received the vehicle solution. Measurements
`washed with brine (30 mL), dried on sodium sulfate, and filtered.
`were performed before treatment and 1, 2, 4, and 6 h after
`The solvent was removed in vacuo, and the slurry 5a was used
`directly without isolation for the next step, RI = 0.27 (EtOAc).
`treatment.
`Conjunctival Hyperemia in the Rabbit. Albino rabbits
`
`(15s)- 13,14-Dihydro-17-pheny1-18,19,20-trinor-PGFz, (6a).
`(New Zealand White, 2-3 kg) were used for evaluation of
`Prepared as compound 5a from the lactol18. The product was
`conjunctival hyperemia. Color photographs of the drug-treated
`used directly without isolation for the next step.
`and control eyes were taken using a camera equipped with a
`13,14-Dihydro-17-pheny1-18,19,20-trinor-PGF~
`Isopropyl
`Medical Nikkor lens (magnification 2X) before treatment, and
`Eater (5b). DBU (1.2 g, 7.98 mmol) was added dropwise to a
`1,2,3, and 4 h after treatment. The photographs were used for
`stirred solution of the crude product 15 (0.52 g, 1.32 mmol) in
`semiquantitative evaluation of conjunctival hyperemia using an
`acetone (15 mL) at 0 "C. The mixture was allowed to warm to
`arbitrary scale from 0 to 5 (0 = totally pale conjunctiva, 1 =
`room temperature when isopropyl iodide (1.13 g, 6.70 mmol) was
`vessels normal, 2 = mild hyperemia, 3 = moderate hyperemia,
`added dropwise. After 4 h (TLC monitoring), the mixture was
`4 = severe hyperemia, 5 = severe hyperemia with chemosis).
`diluted with EtOAc (100 mL), washed with brine (30 mL), citric
`acid 3% (2 X 25 mL), and sodium hydrogen carbonate 5% (2 X
`Contraction of Cat Iris Sphincter in Vitro. Functional
`25 mL), and dried over anhydrous sodium sulfate. The solvent
`receptor studies were performed using iris sphincter muscles from
`was removed in vacuo and the residue was chromatographed on
`cat eyes. The eyes were either used directly after enucleation or
`silica gel using a gradient elution with dichloromethane, dichlo-
`stored in ice-cold saline overnight. The iris sphincter muscles
`romethane-ethanol10:0.5, and dichloromethane-ethanol10:0.75
`were prepared, cut in halves, and mounted in thermostated (37
`successively. This afforded a colorless oil: yield 0.19 g (38%);
`"C) tissue baths with oxygenated modified Kreb's solution
`containing indomethacin (2.8 X lo4 M), atropine (lo-' M), and
`R,= 0.32 (EtOAc); [aI2O~ = +31.57' (C = 0.91, CHsCN); 'H NMR
`(CDC13) 6 1.2 @,6 H), 1.32-1.42 (m, 3 H), 1.62 (m, 2 H), 1.69 (m,
`propranolol (lo4 M). A resting tension of 150 mg was applied,
`2 H), 1.79 (m, 2 H), 2.12 (m, 2 H), 2.22-2.33 (m, 2 H), 2.28 (t, 2
`and the contractile force was measured after cumulative dosing
`H),2.45 (d,lH),2.65-2.78(dm,2 H),3.65 (m,CH2CHOHCHd,
`of prostaglandin analogues in free acid form. The interval
`3.94 (m, CHzCHOH), 4.16 (m, CH2CHOH), 5.0 (sept, 1 H), 5.38
`between doses was approximately 5-10 min, which was the time
`(m, db), 5.47 (m, db), 7.19-7.27 (dm, Ar); 13C NMR (CDC13) 8
`required to reach a stable level of contraction. Force transducers
`173.46 (C=O), 142.84, 128.4, 125.8, 129.6 (C5), 129.3 (C6), 78.8
`for measurement of isometric contraction (Grass FT30C) con-
`(Cll), 74.79 (C9), 71.33 (C15), 67.65, 53.91 (C12), 51.94 (C8),
`nected to a polygraph (Grass Model 7) were used for registration
`42.57 (ClO), 39.10 (C16), 35.84 (C14), 34.07 (C2), 32.14 (C17),
`of the response. For each tissue sample the maximal response
`29.69 (C13), 26.97 (C7), 26.67 (C4), 24.96 ((331, 21.86.
`was normalized to 100%. The mean response from four different
`(15S)-13,14-Dihydro-17-phenyl- 18,19,2O-trinor-PGF~. Iso-
`preparations was calcu