Digitalis-like pregnanes. Cardiac and renal effects of a glycoside of 14~-hydr~x~progesterone'

J. F. TEMPLETON AND V. I". SASHI KUMAR Facubty of Pharmacy, University of Manitoba, Winnipeg, Man., Canada R3T 2N2

D. BOSE AND D. D. SMYTH Department of Internal Medicine and Department of Pharmacology and Therapeutics, Faculty of Medicine,

University sf Manitoba, 770 Banmtyne Avenue, Winnipeg, Man., Canada R3E OW3

W. S. KIM AND F. S. LABEL LA^ Department sf Pharmacology and Therapeutics, Faculq of Medicine, University of Manitoba,

770 Bannatyne Avenue, Winnipeg, Man., Gamda R3E O W

Received May 5, I988

TEMPLETON, J . F., KUMAW, V. P. S. , BOSE, D., SMYTH, D. D., B ~ M , R. S., and LABELLA, F. S. 1988. Digitalis-like pregnanes. Cardiac and renal effects of a glycoside of 14p-hydroxyprogesterone. Can. J . Phy siol. Pharmacol. 66: 1420- 1424.

The synthesis sf the glucoside, 3P-[(P-D-glucopyrmosyl)oxy]- 14-hydroxy- 14p-pregn-4-en-20-one, a 14p-hydrsxypro- gesterone glucoside (14P-OHP-glu), is described. This compound has an of 1 pM in a [3~~ouabain binding assay, and is about 10 times more potent than the aglycone. Like 14p-hydroxyprogesterone, the glucoside enhances contractility of isolated cardiac muscle. 14p-OHP-glu or ouabain, when infused at comparable doses into the renal artery of the anesthetized rat, markedly increases urine volume. Whereas ouabain significantly enhances urinary potassium excretion with little or no effect on sodium excretion, 14p-OHP-glu promotes a marked natriuresis with no significant effect on potassium excretion.

TEMPLETON, J. F., KUMAR, V. P. S., BOSE, D., SMYTH, D. B., KIM, R. S., et LABELLA, F. S. 1988. Digitalis-like pregmnes. Cardiac md renal effects of a glycoside of 14p-hydroxyprogesterone. Can. 9. Physiol. Phmacol. 66 : 1420- 1424.

Om ddcrit la synthhse du glucoside 3~-[(~-~-glucopyran6ssyl)oxy]-14-hydroxy-I4~-pregn-4-en-20-one, un l4a-hydroxy- progesterone glucoside (14P-OHP-glu). Ce composC a un ICS0 de 1 pM dam un dosage pour la fixation de [%]ouaba'ine et il est environ 10 fois plus puissant que l'aglycone. Comme la 14B-hydroxyprogestCrone, le glucsside stimule la contractilitd du muscle cardiaque isold. Bes perfusions de doses cornparables de 14B-OHP-glu ou d'ouabaiine dans I'apbkre dnale du rat anesth6siC augmentent nettement le volume umnaire. Alors que l'ouabayne stimule significativement 19excr6tion de potassium urinaire avec peu ou pas d'effet sup l'excr6tion de sodium, la 14P-OHP-glu favorise une natriurhse marquee sans effet significatif sur l'exaktion de potassium.

(Traduit par la revue]

Introduction The digitalis-derived steroid glycosides have been among the

most widely used drugs for two hundred years (Greef and Schadewaldt 1981). Digoxin is, today, the most favored digitdoid in the clinic, and is extensively used to treat certain cardiac dysfunctions (Hoffman and Bigger 1985). The cardiac glycosides bind with high affinity and high specificity to a Naf - and K+-transporting ATPase. It is generally accepted that inhibition of this enzyme by the cardiac glycosides initiates a series of events in cardiac muscle that Bead to increased contractility (Akera 198 1). In congestive heart failure, stimula- tion of urine formation occurs with cxdiotonic doses sf the cardiac glycosides and reverses many of the clinical signs and symptoms (Hoffman and Bigger 1985).

Certain derivatives of progesterone bind directly to the cardiac glycoside recognition site on Na-K ATBase and inhibit the enzyme and the sodium pump (Chow et a&. 1979; Kim et a / . 1980; LaBella et ak. 1985; Templeton et al. 1987 b), the latter a reflection of the functioning enzyme in cell membranes. In addition, crystallography reveals crucial spatial relationships between the active pmgnanes and the cardiac glycosides (LaBella et ab. 1985). We recently reported the synthesis of 14P-hydrox yprogesterone (Templeton et al. 1987a ; Bose et a&.

%upported by the Manitoba Heart Foundatisan and the Medical Research Council of Canada.

'career Investigator of the Medical Research Council sf Canada.

1988). This steroid is the first of the progesterone congeners to possess both the CID cis ring junction and the propensity to enhance cardiac contractility.

Glycosidation of the cadioactive digitalis steroids can alter significantly their solubility, metabolism and distribution, and phmacodynamics (Guntert and Linde B 98 1). Depending upon the individual steroid or sugar, the carbohydrate moiety may increase the potency of the compound profoundly. The clinical- ly useful digitaloids are glycosides. We now report the synthesis and some phmacslogical properties of the 3fl-D-glucoside derivative of 149-hydroxyprogesterone.

Experimental section Steroid synthesis 'H 13c NMR spectra were recorded on a Bmker AM 300 instrument

using tetrmethylsilane as internal standard. Preparations were moni- tored by TLC on silica gel (Merck type 60 H), and plates were developed in 25% ethyl acetate in hexane for the aglycone synthesis a d 2% ethanol in dichloromethane for the glucoside. Column chomatogmphy was carried out on silica gel (Merck type 60 H for TLC). Melting points are uncorrected. Chemical microanalyses for carbon and hydrogen are within "0.3% of the theoretical value.

3P, d4-Dkhydroq-14P -pregn-4-en-200~~e (2) The trio1 (4) (Templeton et ak. 1987a) (1.7 g, 5 rnmol) in dry

dimethylfomamide (DMF) (58 d) was treated with iirmndazole (0.9 g, 13.2 mmsl) followed by tert-butyldimethy1sily1 chloride (TBDMSiCl) (8.975 g, 6.5 m o l ) as described (Corey and Venkateswaalu 1972) for

Can

. J. P

hysi

ol. P

harm

acol

. Dow

nloa

ded

from

ww

w.n

rcre

sear

chpr

ess.

com

by

Uni

vers

ity o

f P.

E.I

. on

11/1

2/14

For

pers

onal

use

onl

y.

TEMPLETON ET AL. 1421

1 h to give a g u m y residue (1.83 g) which was dissolved in a mixture of dichloromethane (140 mL) and BMF (15 mL), and pyridinium dichromate (18 g, 26.6 mmol) was added witkt stirsing for 1 h. Filtration over silica gel using CH2C12 as eluent gave a residue (1.53 g) which was dissolved in tetrahydrofuran (THF) (100 mL) and treated with tetra-n-butylmmonium fluoride (TBAF) in THF as described (Corey and Venkateswaralu 1972) for 15 h to give a crystalline residue (1.33 g) which was purified by column chromatography over silica gel (20 g) in CH2C12 to yield, on elution with 4% acetone-CH2C12, 3g,14- dihydroxy- 14p-pregn-4-en-20-one (2) (0.68 g). Recrystallization from CW2CB2-acetone gave (2) (0.51 g, 30%) mp 200-202°C; 'W NMR (CDC13) 8: 8.99 (3H,s,C-13 CH3), 1.04 (3H,s,C-10 CH3), 2.23 (3H,s,C-20CH3), 2.90(1H,dd,J = 4.5,9Hz, 17 H),4.14(1H,m,wa = 20Hz,C-3H), 4.38 (lH,s,C-140H), 5.29 (lH,d,$ = 1.5 Hz,C-4H); I3C NMR(CX13) 8: 35.5(1), 29.5(2), 67.9(3), 123.4(4), 147.1(5), 32.3(6), 28.9(7), 40.3(8), 49.7(9), 37.5(10), 20.7(1 I), 39.1(12), 49.1(13), 84.7(14), 33.9(15), 24.9(16), 62.2(%7), 15.3(18), 19.1(19), 217.7(28), 33.4(2%). Anal. (C21H3203) C,H.

38-[(B -~-Glucopyranos)?l)o~]-44-hydroxgr-44P pregn-4-en-20-one (1 )

To a stirred solution of (2) (0.14 g, 0.42 m o l ) in dry benzene (20 mL) was added Fetizon's reagent (3 g), followed by Hg(CN)2 and HgBr2 (0.1 g each) under argon. Acetobromoglucose (0.75 g) (10 equiv.) in dry benzene (20 mL) was dripped in over 1 la and the mixture was left stking for 24 h. The mixture was filtered through a Celite pad and the filtrate was diluted with ether (200 mk), washed with aqueous NaHC03, water, dried over Na2S04, and evaporated to yield a residue that was dissolved in a mixture of methanol (20 mL), triethylamine (10 mL), and water (1 mL) and left under argon for 16 h (Brown et al. 1981). The solvents were evaporated under reduced pressure at room temperature and the residue was purified on a silica gel column (5 g) packed in ch1orofom. Elution with 5% methanol in chloroform gave the 3g-D-glucoside (1) (0.105 g). Recrystallization from acetone gave the pure glucoside (30 mg, 15%) mp 220-222°C. % HMR (1: 1 CDC13:CD30B) 8 :1.0 (3H,s,C-13 CH3), 1.06 (2H,s,C-10 CH3), 2.27 (3H,s,C-20CH3),2.94(1H,dd,J=4.1,8.2Hz, 17H),4.25(1H9~.~94 = 22Hz.C-3H),4.43 ( l H , d , $ = 7 . 8 H z , C - l ' H ) , 5 . 4 2 ( 1 H , s , w ~ = 4 Hz, C-4H); 13c NMR (1:l CDCI3:CD30D) 6: 35.3(1), 26.9(2), 75.7(3), 120.2(4), 147.3(5), 32.0(6), 28.5(7), 40.0(8), 49.7(9), 37.3(10), 20.3(11), 38.'7(12), - (13), 85.0(14), 33.5(15), 24.6(16), 62.0(17), 14.8(18), 18.5(19), 218.7(20), 32.7(21), 10%.8(C;), 73.4(C;), 76.3(C;), 70.1(Ci), 75.8(C;), 61.6(C;). Anal. (C29H4208) C,H.

Lithium tri-t-bu~o~al~rninum hydride reduction of di(-hydrsxy-B4P- pregn-4-ene-d,20-dione (7)

14g-Hydroxypmgesterone (7) (Templeton et al. 1987a) (10 mg, 0.032 m o l ) was stirred at room temperature with 90% lithium pai-t-butoxyalumjnum hydride (LTBA; 10 mg, 0.0.35 mmol) in dry ether (4 mL) for 10 min when no starting material remained by %LC. After ether extraction the 'H NMW spectrum of the crude product showed no methyl signal at 2.25 (C-20 CH3) but retained a strong signd at 5.74 (4-en-3-one H).

[3~]ouabain binding assay Hearts from pentobarbitd-anesthetized mongrel dogs were removed

and immediately immersed in Kiebs-Henseleit buffer (mM):NaCl, 118; KCI, 4.7; CaC12, 2.5; MgS04, 1.2; M2PQ4, 1.4; N d C 0 3 , 26; glucose, 11) equilibrated with 95% 0,:5% C02 at room temperature. Within 6Q f i n the ventricles were chopped with scissors and disintegrated with a Polytron (Brinkmann Instruments Inc.) in ice-cold 58 Tris buffer, pH 7.4. The suspension was passed bough a coarse screen to remove the bulk of connective tissue and the filtrate was centrifuged at 34 000 x g for 45 rnin at 4°C. 'Phe supmatant was discarded and the pellet was resuspended in the buffer for radioligand binding assay (see below) and stored at -20°C. Once thawed, the tissue suspension was used immediately or discarded. The buffer was 45 anWI Tris, 5 IPlM Tris-phosphate, 3 HnM Mg&112, pH 7.4. Duplicate assay tubs contained 2.0 nM ['H]ouabain in 0.1 mL buffer,

approximately 10 mg tissue (wet weight) in 8.1 mL buffer, and steroids (see below). Nonspecific binding (about 10% of total binding) was determined in the presence of 0.1 mM ouabain. Following incubation at O"C for 90 d n , assay tubes were centrifuged at 2700 X g for 25 min at 4"C, and the supernatants were aspirated off. Pellets were dissolved in 0.3 mL 2 M KOH, and 0.2-mL aliquots were dispensed into vials for liquid scintillation counting. Data were expressed as percent of specifically bound [3~]ouabain (i.e., the difference between total binding and nonspecific binding). The steroids dissolved in 5 FL of 95% ethanol were added to reaction tubes; this volume of ethanol alone had no effect on specific binding of [%]ouabain. Similar potencies were obtained for the steroids in the radioligand binding assay when the ethanolic solutions were evaporated to dryness in the reaction tube, the tissue suspension was added, and the tube was agitated on a vortex mixer for 1 min.

Measurement sf contractility of isolated cardiac tissue Dogs of either sex weighing between 5 and 12 kg were anesthetized

with sodium pentobarbital (35 rng/kg i .c .), and the heart was removed. Thin (<I m diameter) free-mnnhg tsakcdae fmm the fight ventricle were placed in a 10-mL vertical tissue bath containing Krebs-Henseleit solution and bubbled with 95% 0 2 and 5% C02; the bath was maintained at 37.0 2 O.2"C. The muscle was equilibrated for 1 h prior to the experiment after being tied to the base of a Perspex holder which had embedded electrodes for punctate stimulation. The opposite end of the tissue was connected to a Grass IT-03C isometric force transducer, and a Grass polygraph. A Pulsar 6i stimulator (F. Waer Co.) connected to a custom-built computer-controlled programmable pulse generator (Boyechko and Bose 1984) provided square-wave stimuli of 2080 ms duration to the trabecula. Stimulus amplitude was adjusted to about 10-20% above threshold. The resting tension was increased until the maximum active tension in response to electrical stimulation was evoked. The basic stimulus interval was 2080 ms.

In vivo renal function Male Sprague-Dawley rats (380-350 g) were uninephrectomiaed

(left kidney, ether anesthesia) 7-10 days prior to the experiment. On the day of the experiment, the rats were mesthetized with pentobarbital, tracheostomy was performed, and the animals were placed on a thermostatically controlled heating blanket (Haward Animal Blanket Control Unit) to maintain body temperature at 38°C. The jugular vein and carotid artery were cannulated for the infusion of saline (97 yUmin) and the recording of blood pressure, respectively. The remaining kidney was exposed by a flank incision and the ureter was emulated with BE-50 polyethylene tubing. A 30-gauge hypodermic needle was inserted into the aorta and advanced into the renal artery for infusion of vehicle or h g . After a 45-min stabilization period, urine was collected into a preweighed tube for 15 mh. Immediately after this collection, the infusion of the vehicle (saline), ouabain (0.19 mg/mL), or 149-OHP-glm (8.19 mg/mL) was begun and maintained for 10 min at 0.26 mL/min; during this infusion, urine was collected. Five additional 15-min wine collections were made. A plasma sample was obtained at the end of the experiment. Urine and plasma sodium and potassium concentrations were determined on a Beckman Klina Flame photo- meter. Creatinine concentrations in plasma and urine were measured with a Beckman creatinine analyzer model 2. Osmoldity was deter- mined with a micro-Osmette (Recision Systems).

Results and discussion The aglycone, 3f3,14-dihyQroxy- 14~-pregn-4-en-20-one (2)

(see scheme I), was prepared from 1463-hydroxyprogesterone (3) by utilizing the isomeric intermediate 3,14~,20-trial mix- ture (4) (Templeton ct st / . 1987a). Regioselective reaction of (4) with 1.3 equiv. of TBDMSiCl for 1 h gave the 3-silyBoxy derivative (5) (Corey and Venkateswaralu 8972). Neutral oxidation of (5) with pyridinium dichromate (Corey and Schmidt 1979) resulted in the formation of the 28-ketone (6). Besilylation with TBAF in THF (Corey and Venkateswaralu 1972) gave the epimeric 3-hydroxy mixture from which the

Can

. J. P

hysi

ol. P

harm

acol

. Dow

nloa

ded

from

ww

w.n

rcre

sear

chpr

ess.

com

by

Uni

vers

ity o

f P.

E.I

. on

11/1

2/14

For

pers

onal

use

onl

y.

CAN. J . PHYSIOL. PHAWMACOL. VOL. 66, 1988

SCHEME 8 . ( a ) See Templeton e ~ a l . 1987a; (b) t-butyldimethylsilyl chloride (TBDMSiCl), irnidazole, dimethylforaknide (DMF); R = TBDMSi; ( c ) pyridiniurn dichmmate, CW2C12, DMF; (4 tetra-n- butylammonium fluoride, tetrahydrofuran: ( e ) column-SiQ; lithium tri- t-butoxy aluminum hydride; (g) acetobrornoglucose, F6tizon5s reagent, Wg(@E9)2, HgBr2, and benzene; and Et3N, MeOH. and W 2 0 .

3(3,14-dihydroxy-14P-pregn-4-en-20-one (2) was separated and crystallized. This reaction sequence was carried out with- out the isolation of the individual intermediates because of the presence of stereoisomers. Reduction of 14P-hydroxy- progesterone (7) with lithium tri-t-butoxyaluminaam hydride gave more rapid reduction at C-20 and therefore could not be used to prepare 2. The attachment of P-D-glucose at C-3 was achieved by using the procedure of Brown et al. with FCtizon's reagent (Brown et ak. 198 1).

In the [3~]ouabain binding assay, 14P-OHP-glu has an ICso of B .3 pM, and is about BO times more potent thm the aglycone (Fig. 1). 1CSo values for ouabain, ouabagenin, and progesterone are 14 M, 0.25 pM, and 55 pM, respectively.

Progesterone interacts weakly (ICS8 of 55 ylM) and chlor- madinone acetate (CMA), a semisynthetic progestin, interacts potently (ICS0 of 0.3 pM) with the cardiac glycoside binding site on NA-K ATPase (La Bella et al. 1985). Although this interaction is associated with inhibition of the enzyme and would be expected to cause positive inotmpy in cardiac muscle, cardiodepression is the rule with progesterone and is the predominant myocardial response to CMA (LaBella et ak. 1985; Bose et a&. 1988; Weiland et al. 198'7; LaBella et al, 1988). These findings led to a suggestion that the cardiac glycoside "receptor" that mediates positive inotropy may not be Na-K ATPase (Lae l la et a&. 1979). Subsequently , this proposal was modified to include the possibility that the digitaloid pregnanes have a direct depressant effect on the myocardium which counteracts the positive inotropy directly linked to inhibition of the sodium pump (LaBella et ( P I . 1984). Thomas et al. (1980) equated the delayed onset of cardiodepression with the time period anticipated for the charged steroid hydrazone to permeate the cardiac cell and inhibit critical intmcellular processes. They noted that those steroid guanylhyglrazones, which elicit negative inotropy in isolated cardiac muscle, also inhibit mitochondria1 respiration. The negative inotropy could be reversed by increas- ing the oxygen flow in the muscle bath.

In the csudiodepressive progesterone and CMA, the steroid moiety differs from that of the cardiostimulant cardiac glyco- sides, specifically, in the configuration of the A/B and CID ring

Specific binding

(%I

0

bog concn. ( M I

FIG. 1 . Inhibition by various steroids of specific binding of [ 3 ~ ] ~ u a b a i n to cardiac ventricular tissue. Each point is the mean of five replicates and the entire displacement curves were generated several times with very similar results.

148 -0HPglu Concentration (YM)

FIG. 2. Dose-response relationship for 14B-OWB-glu on isometric contractile force in tmbecular muscle. Vertical bars indicate + SEM. n=4.

junctions; both junctions are cis (bent) in the cardiac glycosides and planar in progesterone and CMA. Introduction of a 14P-OH moiety into progesterone converts the C/D junction to the cis configuration and confers a positive inotropic action to that steroid. The glycoside of another pregnane with both A/B and C/D junctions in a cis configuration has been reported to enhance cardiac contractility in vivo a d to be associated with an effective dose : toxic dose ratio superior to that of digoxin (Halpryn et al. 198'7). This pregnane is equipotent to digoxin (Jmeau et ak. 1984; SchonfeBd et al. 1985). We have synthesized the 5P(H), 14P(OH)-andog sf progesterone ( N B cis and C/D cis) and have found its glycoside to be about one-twentieth as potent as digoxin md to elicit positive inotropy in isolated heart tissues (LaBella et a&. 1988).

14fb-OHB-glu consistently causes positive inotropic response in isolated canine, ventricular9 trabeculae muscle in a dose- dependent manner (Fig. 2). High concentrations elicit a

Can

. J. P

hysi

ol. P

harm

acol

. Dow

nloa

ded

from

ww

w.n

rcre

sear

chpr

ess.

com

by

Uni

vers

ity o

f P.

E.I

. on

11/1

2/14

For

pers

onal

use

onl

y.

TEMPLETON BT AL.

TABLE 1. Renal effects sf saline (vehicle), ouabain, and 14fl-BHP-glu hydrsxy- progesterone in the anesthetized rat

Saline Bmabain 14p-OHP-glu -

B l d pressure (mmHg) 107212 126+6 Creatinine clearance (mk/min) 1.39 1'0.36 1 -43 2 0.06 E N a (9%) 3.68+ 1.20 4.21 20.34 Osmolar clearance (pL/min) 89k 12 146+ 18* Free water clearance ( pL/min) -49+ 10 - 5 0 6 Urine osmolarity (mosmol/kg) 7202 92 447f 13* n 3 3

- - - - - - - - - - - - - - -- -

NOTE: Values are the means + SE obtained during the sixth collection period. excretion sf sodium. 1 mmHg = 133.3 Pa.

+ p < 0.05 versus the sdine group (ANOVA, Duncan's test).

FENa, fractional

CONTROL

FIG. 3. Isometric contraction of canine ventricular trabecula. Top panel: control. Bottom panel: 14p-OHP-glu, 5 X ~ o - ~ M ) . Normal stimulation interval of 2000 ms was interspersed with a rapid train (interval 250 ms; horizontal arrows) to initiate aftercontraction. Notice positive inotropic effect of I4P-OHP-glu on nomally driven contrac- tions and initiation of an aftercontraction (oblique arrow).

biphasic contractile response, an initial increase followed by a decrease. During the peak inotropic response and subsequent depression, it is possible to elicit spontaneous aftercontrac- tion(s) by rapid electrical stimulation of the muscle (Fig. 3). This is similar to the toxic responses to other digitalis com- pounds (Ferrier 1977; Chan et ak. 1987).

The effects of an intrarenal infusion of ouabain and 14P- hydroxyprogesterone glucoside were compared (Fig. 4; Table 1). Both compounds increased urine volume. Ouabain produced a marked increase in potassium excretion with little or no change in sodium excretion without significant changes in renal hernodynamics (blood pressure, creatinine clearance), suggest- ing an action at the tubular level. Our findings ace consistent with those of others who report natriuresis by ouabain in only 58% of the rats but increased potassium excretion in all, and ouabain inhibited rend reabsorption of potassium to a greater extent than that of ssdium (Duarte et ak. 197 1). In studies where ouabain elicited consistent natriuresis, rats had been subjected to a low potassium diet (Rmsey and Sachs 1867). 14P-OHB- glu increases sodium excretion significantly but has no influ-

P..' *

ColOectiow Period

FIG. 4. The effect of intrarenal infusions of saline vehicle (a), ouabain (El), and I4p-BHP-glu (A) on wine volume, sodium excretion (UN,V), md potassium excretion (UKV) in the anesthetized rat. Drug infusions were initiated immediately following the first urine collection and maintained for 10 min. There were thee animals in each group. Means f SE are shown. * p < 0.05 versus the control group.

Can

. J. P

hysi

ol. P

harm

acol

. Dow

nloa

ded

from

ww

w.n

rcre

sear

chpr

ess.

com

by

Uni

vers

ity o

f P.

E.I

. on

11/1

2/14

For

pers

onal

use

onl

y.

1424 CAN. J. PHYSIQL. PHARMACQL. VOL. 66. I988

ence on potassium excretion, a response profile opposite to that observed with ouabaiw.

As discussed above, the wider margin s f safety associated with the pregnanes' cardiac action reflects dissimilarity in the pharmacology of the two classes of compounds. Inhibition of metabolism by the pregnanes at an intracellular site is one possible mechanism underlying the cardiodepression and is supported by experimental data (Thomas et a1. 1980). Differ- ences between the paegnme and cadiac glycosides in funda- mental cellular actions should manifest themselves, no doubt, in the responses of the urinary system as well. The marked natriga~sis and absence of kd ims i s by the cadiotonic prepmes have practical implications and indicate, further, the potential clinical superiority of these agents as suggested in animal studies (Halpryn et 611. 1987; Jmeau et ak. 1984; Wieland et a&. 1987). The potential clinical advantage of the cardiotonic pregnanes over the cardiac glycosides has been reviewed recently (LaBella et a&. 1888).

A ~ M , T. 198 1. Effects of cardiac glycosides on N~' ,K+ - A T P ~ S ~ . 18 Handbook of Experimental Phmco logy . Part 1. Chapt. 14, Edited by K. Greef. Springer-Verlag , Berlin. pp. 287-336.

BBSE, B., ELLIOTT, D., KOBAYASWI, T., TEMPLETON, J. F., KUMAW, V. P. S., and LABELLA, F. S. 1988. 14P-Hydroxyprogesterone binds to the digitalis receptor, inhibits the sodium pump and enhances cardiac contractility. Br. J. Bhamacol. 93: 453-461.

BOYECHKO, Go , and BOSE, B. 1984. A versatile computer-controlled biological stimulus sequencer. 9. Phmacol. Methods, 12: 45-52.

BROWN, L., BOUTAGY, J., and THOMAS, R. 1981. Cardenolide analogues. II. Improved method for use of Fktizon's reagent in the synthesis of cardiac glycosides. hzneim. Fomch./Dmg Wes. 31: 1059- 1064.

CHAN, T., CHAU, T., and BOSE, B. 1987. Role of sarcoplasrnic reticulum in digitalis induced electrical and mechanical oscillations in the heart. Can. 9. Cadiol. 3: 197-204.

CHOW, E., M$M, R. S., LABELLA, F. S., and QUEEN, G. 1979. 0uabain receptor binding of hydroxyprogesterone derivatives. Br. J. Phmacol. 6'7: 345-452.

COREY, E. J., and SCHMIDT, G. 1979. Useful procedures for the oxidation of alcohols involving pyridinium dickomate in aprotic media. Tetrahedron Lett. 5: 399-402.

COREY, E. J., and VENKATESWARALU, A. 1972. Protection of hydroxyl groups as tert-butyldimethylsilyl derivatives. 9. Am. Chem. Soc. 94: 6190-6191.

DUARTE, C. G., CHQMETY, F., and G r ~ ~ a s c a , G. 1971. Effect of adloride, ouabain and furosemide on distal tubular function in the rat. Am. J. Physiol. 221: 632-639.

FERRJER, 6. R. 1997. Relationship between acetylstrophanthidin- induced aftercontractions and the strength of contraction of canine ventricular m p c d i u m . Circ . Res . 41: 622-629.

GREBF, K, , and SCHADEWALDT, H. 1981. Introduction and remarks on the history of cardiac glycosides. In Cardiac glycosides. Part 1. Edited by K. Greef. Springer-Verlag , Berlin. pp. 1 - 12.

GUNTER'~, T. W. , and EINDE, H. H. A. 1981. Chemistry and structure-activity relationships of cardioactive steroids, In Cardiac glycosides. Part 2. Chapt. 2. Edited by K. Greef. Springer-Verlag, Berlin. pp. 13-24.

HALPRYN, B. , FULTON, R., BANASEVICH, R., OGERAU, T., FENARD, S., BAGGIBNI, A., KOENIG, J . , and CWANG, Y. 1987. Activity of LND 623, a new inotropic steroid, in dogs. Pharmacologist, 29: 135.

HOFFMAN, B. F., and BIGGER, J. T. 1985. Digitalis and allied cardiac glycosides. In The phmacological basis of therapeutics. Chapt. 30. 7th ed. Edited by A. 6. Gilman, L. S. Goodman, T. W. Rall, and F. Murad. Macrnillan Publishing Co., New Yo&, Toronto, London. pp. 7 1 6-747.

S A ~ E A U , F. X., KOENIG, J. J., and FENAWD, S. 1984. A new inotmpic aminosteroid: END 623. Eur. Heart 9. S(Suppl. F): 309-3 14.

&M, R. S., LABELLA, F. S., ZUNZA, H., ZUNZA F., and TEMPLETON, J. F. 1980. Progesterone derivatives that bind to the digitalis receptor: structure activity relationships. Mol. Phmacol. 18: 395- 398.

LABELLA, F. S., BIHLER, I., and KIM, W. S. 1979. Progesterone derivative binds to the cardiac ouabain receptor and shows dissocia- tion between sodium pump inhibition and increased contractile force. Nature (London), 28: 57 1-573.

1984. Progesterone derivatives that bind to the digitalis receptor; effects on 8 6 ~ b uptake and contractility in the isolated guinea pig heart. Can. J . Physiol. Phmacol. 62: 1057- 1064.

LABELLA, F. S., BIHLER, I., TEMPLETON, J., KIM, R. S., HNATBW- ncH, M., a d ROHRER, D. 1985. Progesterone derivatives that bind to the digitalis receptor: effects on Na', K + - A T P ~ ~ ~ and isolated tissues. Fed. $roc. Fed. Am. Soc. Exp. Biol. 44: 2806-281 1.

LABELLA, F. S., TEMPLETON, J . F., SASHI KUMAR, V. P., and BOSE, D. 1988. Cardiotonic properties of progesterone derivatives that bind to the digitalis receptor. Trends Phmacol. Sci. In press.

RAMSEY, A. G . , and SACHS, G. 1967. ~ f f e c t of ouabain on Na' and K' excretion in rats. Roc. Soc. Exp. Biol. Med. 126: 294-2198.

SCHONPELD, W., WEILAND, J . , LINDIG, C., MASNYK, M., KABAT, Me M., K u ~ K , A., WICWA, J., and REPKE, K. R. H. 1985. f i e lead structure in cardiac glycosides is %fl, 14f3-an&ostane-3fl,14-diol. Naunyn-Schrniedekrg's Arch. P h m c o l . 329: 4 14-426.

TEMPLETON, J. F., KUMAR, V. P., COTE, D., BOSE, D., ELLIOTT, D., UM, W. S., and LABELLA, F. S. 1987a. Progesterone derivatives that bind to the digitalis receptor: synthesis of 14P-hydroxy- progesterone. A novel steroid with positive inotropic activity. 9. Med. Chem. 3: 1502-1504.

TEMPLETON, 9. F., SASHB KUMAR, V. P., KIM, R. S., and LABELLA, F. S. 1987b. Structure-activity relationships of progesterone deriva- tives that bind to the digitalis receptor: modifications in A and B rings. Steroids, 49: 383-396.

THOMAS, R., BROWN, Lo, BOUTAGY, S., and GELBART, A. 1980. The digitalis receptor: inferences from structure activity relationship studies. C k . Res. (Suppl. I) 46: 167- 172.

WEILANB, J., SCHWABE, K., HUBLEW, D., SCHONPELB, W., and WEPKE, K. R. H. 1987. Glycosidation of chlomadinol acetate alters its actions on M~'/K'-transporting ATPase and cardiac contractility: a contribution to the endogenous digitalis problem. J . Enzyme Inhib. 2: 31-36.

Can

. J. P

hysi

ol. P

harm

acol

. Dow

nloa

ded

from

ww

w.n

rcre

sear

chpr

ess.

com

by

Uni

vers

ity o

f P.

E.I

. on

11/1

2/14

For

pers

onal

use

onl

y.