USOO5811463A United States Patent (19) 11 Patent Number: 5,811,463 Legzdins et al. (45) Date of Patent: Sep. 22, 1998

54 COMPOSITIONS AND METHOD FOR 4,748,160 5/1988 Bennion et al...... 514/82 RELAXING SMOOTH MUSCLES 5,366,997 11/1994 Keefer et al...... 514/611 OTHER PUBLICATIONS (75) Inventors: Peter Legzdins, West Vancouver; Catherine C. Y. Pang, Vancouver; Butler, Anthony R. and Williams, D. Lyn H., “The Physi Michael J. Shaw, Cowansville, all of ological Role of ,” Chem. Soc. Rev. Canada 22(4):233–241, 1993. Clarke, M. J. and Gaul, J. B., “Chemistry Relevant to the Assignee: University of British Columbia, Biological Effects of Nitric Oxide and Metallonitrosyls,” Vancouver, Canada Struct. Bonding (Berlin) 81: 147-181, 1993. Primary Examiner Deborah Lambkin Appl. No.: 770.559 Attorney, Agent, or Firm Seed and Berry LLP Filed: Dec. 20, 1996 57 ABSTRACT Related U.S. Application Data Compositions and methods for relaxing Smooth muscle in a warm-blooded animal are provided, comprising the Step of 63 Continuation of Ser. No. 132,632, Oct. 6, 1993, Pat. No. administering to the animal a transition-metal nitrosylcom 5,631,284. plex. In one aspect, the transition-metal nitrosyl complex is 51 Int. Cl." ...... A61K 31/13; A61K 31/04; represented by the formula LaM(NO),X, where L is a A61K 31/28 two-electron Lewis base or L is a Six-electron Lewis base, 52) U.S. Cl...... 514/645; 514/740, 514/505; M is a Group 6 or 8 transition-metal, and when y is 1, X is 514/492 , and when y is 2, X is a halide or 58 Field of Search ...... 514/505, 492, pseudohalide. In another aspect, the transition-metal nitrosyl 514/645, 740; 564/300, 301; 424/646, 649, complex is represented by the formula M(NO)2X, where 655 X is a halide or pseudohalide, and when M is a Group 6 transition-metal, y is 2, and when M is a Group 8 transition 56) References Cited metal, y is 1. Methods are also described for treating hypertension, angina pectoris, congestive heart disease, and U.S. PATENT DOCUMENTS impotence utilizing pharmaceutical compositions compris 3,422,144 1/1969 Hoffman et al...... 564/300 ing the above-described transition-metal nitrosyl complexes. 3,480,653 11/1969 Pande et al...... 564/301 4,571,391 2/1986 Riley et al...... 514/4 14 Claims, 2 Drawing Sheets U.S. Patent Sep. 22, 1998 Sheet 1 of 2 5,811,463

20 - O CpCr(NO), C -20 - O CpMo(NO), C 2 C s -60 - SS -100 -

-140 -12 - 10 -8 -6 -4 -2 CONCENTRATION (M), LOG Fig. 1A

20 O CpCr(NO)Cl

o S 5 O

1 E-1 O 100 000 E4 1E5 DOSE (ug/kg), LOG Fig. 1B U.S. Patent Sep. 22, 1998 Sheet 2 of 2 5,811,463

NCUBATON FOR 1 h 20 20 INCUBATON FOR 1 h T

O WEHICLE O WEHICLE

2. -20 O CpCr(NO), Cl 2. -20 O NITROGLYCERN C C c s 3 -60+ is -60 t t SS SS - 100 -100

-140 -140 - 1 -9 -7 -5 -3 - 11 -9 -7 -5 -3 CpCr(NO), CI (M), LOG CpCr(NO), CI (M), LOG Fig. 2A Fig. 2C

NCUBATON FOR 1 h NCUBATON FOR 1 h 20 O WEHICLE 20 O WEHICLE s O NTROGLYCERN ONTROGLYCERN -20 2. -20 C Z t C 3 -60+ - t 3-60 SS SS -100 - 100

-140 -140 -9 -7 -5 -5 -1 - 10 -9 -8 - 7 -6 -5 (M), LOG (M), LOG Fig. 2B Fig. 2D 5,811,463 1 2 COMPOSITIONS AND METHOD FOR Common drugs which are used for this purpose include RELAXING SMOOTH MUSCLES diuretics (hydrochlorothiazide, chlorthalidone), Sym patholytics (clonidine, atenolol, prazosin), vasodilators CROSS-REFERENCE TO RELATED (minoxidil, Sodium nitroprusside), angiotensin converting APPLICATION inhibitors (, enalapril), and calcium entry blockers (, nitrendipine). The Selection of a spe This application is a continuation of U.S. patent applica cific drug depends upon the Severity of the disease, the tion Ser. No. 08/132,632, filed Oct. 6, 1993, U.S. Pat. No. patient's age and lifestyle, Side effects, cost, and concomi 5,631,284. tant diseases and therapies. TECHNICAL FIELD 1O One approach to the treatment of hypertension is to effect the reduction of elevated blood pressure through blood The present invention relates generally to compositions vessel dilation, which results in an increase of blood vessel and methods of relaxing Smooth muscles and, more diameter and concomitant blood pressure reduction. One Specifically, to compositions and methods which utilize class of compounds which are commonly utilized to dilate transition-metal nitrosyl compounds as Smooth muscle 15 blood vessels are , which have been utilized relaxants. for the relief of anginal attacks and the treatment of hyper tension. Clinically useful nitrovasodilators include organic BACKGROUND OF THE INVENTION and Such as amyl , glyceryl trinitrate Muscle is divided into three types: Skeletal, cardiac, and (nitroglycerin), , erythrityl tetranitrate, Smooth. Briefly, Smooth muscle is found in all muscles and pentaerythritol as well as an inorganic nitric oxide where contraction is involuntary, except for heart muscle. donor, Sodium nitroprusside. Generally, Smooth muscle is composed of elongated, Nitroglycerin and Sodium nitroprusside are the most com Spindle-shaped, nucleated cells arranged parallel to one monly used nitrovasodilators. Nitroglycerin is therapeuti another and to the long axis of the muscle. cally useful for the treatment of angina pectoris and gas Smooth muscle is classified both functionally, and ana 25 trointestinal Spasm, while Sodium nitroprusside is primarily tomically. When classified functionally, it is broken into two utilized for the treatment of hypertensive emergencies. groups: multi-unit, and Single-unit Smooth muscle. Briefly, Nitroglycerin may either be administered at the time of multi-unit Smooth muscle is activated by nerves which cause anginal attack or prophylactically in anticipation of exercise contraction of independent muscle units, and thus is not or StreSS. Although nitroglycerin is effective in relief of acute Spontaneously active. Examples of multi-unit muscle may attack, continual exposure to nitroglycerin and other organic be found in large arteries and veins, the urinary bladder, and nitrates results in a reduction in the ability of these com the iris and ciliary muscles of the eye. In contrast to pounds to vasodilate blood vessels. Because of the tolerance multi-unit Smooth muscle, Single- unit Smooth muscle is that develops from their chronic use, these nitrovasodilators usually Spontaneously active, and contains tight junctions have limited applicability. which facilitate the contraction of the muscle as a single 35 Sodium nitroprusside is likewise effective in lowering unit. Examples of Single-unit Smooth muscle may be found blood preSSure, although its use is primarily limited to in the vasculature (Small veins, Small arteries, and hypertensive emergencies due to its efficacy and toxicity. In arterioles), the bile duct, and the walls of the gastrointestinal particular, the infusion of Sodium nitroprusside at high doses and urinogenital Systems (gut, ureter, and uterus). 40 or for long periods of time (over 24 hours) is associated with When Smooth muscle is anatomically classified, it is the toxicities resulting from the accumulation of cyanide and generally divided into two groups: vascular and non or thiocyanate. vascular. Briefly, Vascular Smooth muscle consists only of Accordingly, there is a need in the art for physiologically blood vessels (arteries, arterioles, and veins), whereas non acceptable Smooth muscle relaxants which do not result in vascular muscle includes all other types of Smooth muscle 45 the development of tolerance and do not have toxic side (e.g., gastrointestinal (stomach, duodenum, ileum, jejunum, effects. The present invention fulfills these needs, and caecum, and colon), Spleen, trachea/bronchus, Seminal further, provides other, related advantages. vesicle, ductus deferens, corpus cavernoSum, biliary tract, ureter, and uterus). SUMMARY OF THE INVENTION Many diseases are characterized by the inability of 50 Briefly stated, the present invention provides methods for Smooth muscle to relax. For example, hypertension is a relaxing Smooth muscle in a warm-blooded animal, com disease found in 15%-20% of all adults in the United States. prising the Step of administering to the animal a therapeu This condition is characterized by persistently high arterial tically effective amount of a transition-metal nitrosyl com blood pressure which, if left untreated, may lead to other plex. Within one aspect of the invention, the transition-metal Serious cardiovascular diseases Such as heart failure, 55 nitrosyl complex is represented by the formula LM(NO) ischemia, renal failure, and Stroke. Hypertension is one of X, where L is a two-electron Lewis base or La is a the most important public health problems facing developed six-electron Lewis base, M is a Group 6 or 8 transition countries. metal, and when y is 2, X is either a halide or pseudohalide, Because of the health risk associated with untreated and when y is 1, X2 is either L or XL. hypertension, the importance of treating those who Suffer 60 Within another aspect of the invention, methods are from elevated blood pressure has been recognized. Clinical provided for relaxing Smooth muscle in a warm-blooded trials have shown that appropriate pharmacological treat animal, comprising administering to the animal a transition ment of patients with moderate or Severe hypertension metal nitrosyl complex represented by the formula M(NO) reduces the risk of Stroke, renal failure, and congestive heart 2X, where M is a Group 6 or 8 transition-metal, X is either failure. Based upon these Studies, patients are now com 65 a halide or pseudohalide, and when M is a Group 6 monly treated with antihypertensive drugs in order to lower transition-metal, y is 2, and when M is a Group 8 transition diastolic blood pressure (“DBP”) to levels below 90 mmHg. metal, y is 1. 5,811,463 3 4 A further aspect of the present invention provide methods LM(NO), Xs, wherein L is a two-electron Lewis base or La for treating hypertension in a warm-blooded animal, com is a six-electron Lewis base, wherein M is a transition-metal, prising administering to the animal a therapeutically effec wherein X is a halide or a pseudohalide, and wherein ify is tive amount of a transition-metal nitrosyl complex repre 1, X is carbon monoxide and if y is 2, X is a halide or sented by either the formula LM(NO), Xs, or M(NO) pseudohalide. AS used herein, the term “transition-metal 2X) as described above. nitrosyl complex refers to a composition in which a Within another aspect of the present invention, methods transition-metal is coordinatively complexed with for treating angina pectoris in a warm-blooded animal are wherein at least one is nitric oxide. The term provided, comprising administering to the animal a thera "nitroSyl” refers to the nitric oxide ligand. A transition-metal peutically effective amount of a transition-metal nitrosyl is defined as any metal that is a member of Periodic Groups complex represented by either the formula LM(NO), Xs, or 3-12. Preferred transition-metals include Groups 6 and 8 M(NO).X.), as described above is disclosed. metals, chromium, molybdenum, tungsten, and iron, A further aspect of the present invention provides meth ruthenium, osmium, respectively. A more preferred embodi ods for treating congestive heart failure in a warm-blooded ment of the present invention comprises transition-metals animal, comprising administering to the animal a therapeu 15 from Group 6. As represented by the formula LaM(NO), X tically effective amount of a transition-metal nitroSyl com y, the transition-metal ligands are L, NO (nitric oxide), and X wherein- L is a two-electron Lewis base or L is a plex represented by either the formula LM(NO),X, or Six-electron Lewis base and X is a halide or a pseudohalide, M(NO)2X, as described above. or alternatively, a two-electron Lewis base, L. AS used Within a further aspect of the present invention, methods herein, the term “Lewis base' refers to any chemical Species for treating impotence in a warm-blooded animal are which is an electron pair donor. Two-electron Lewis bases provided, comprising administering to the animal a thera are those bases which may donated a single pair of electrons. peutically effective amount of a transition-metal nitrosyl Suitable two-electron Lewis bases include bases which bear complex represented by either the formula LM(NO), Xs, or atoms from Periodic Groups 15 and 16. Lewis bases from M(NO)2X, as described above is disclosed. 25 Group 15 contain nitrogen, phosphorous, arsenic, antimony Another aspect of the present invention provides pharma or bismuth atoms as electron pair donors. Preferable Lewis ceutical compositions comprising a transition-metal nitrosyl base from Group 15 contain nitrogen, phosphorous, and complex and a pharmaceutically acceptable carrier or dilu antimony, and more preferably, nitrogen or phosphorous. ent. The transition-metal nitrosyl complex of the pharma Nitrogen containing Lewis bases include ammonia and its ceutical composition may be represented by either the carbon Substituted derivatives including primary, Secondary, formula LaM(NO),X, or M(NO).X.), as described and tertiary amines and aromatic amines including pyridine above. These pharmaceutical compositions may be utilized and pyrrole as well as Saturated analogs, piperidine and in the methods described above, as well as in a variety of in pyrrolidine. Other preferable nitrogen containing Lewis Vitro and in Vivo assays which are discussed in more detail bases include amino acids and their derivatives. Phospho below and in the Examples. 35 rous containing Lewis bases include phosphine and its These and other aspects of the present invention will carbon Substituted derivatives including primary, Secondary, become evident upon reference to the following detailed and tertiary phosphines as well as alkoxy Substituted deriva description and attached drawings. tives including primary, Secondary, and tertiary phosphites. BRIEF DESCRIPTION OF DRAWINGS Lewis bases from Group 16 contain oxygen, Sulfur, or 40 Selenium atoms as electron pair donors. Preferable Lewis FIG. 1 is a graph which illustrates the effects of CpCr bases from Group 16 contain oxygen or Sulfur. Oxygen (NO)C1 and CpMo(NO)C1 on the relaxation of containing Lewis bases include water and alcohols and phenylephrine-preconstricted rat aortic rings (1A) and the Sulfur containing bases include hydrogen Sulfide and thiols. change in mean arterial blood pressure in conscious rats Carbon monoxide is also a two-electron Lewis base. The (1B). 45 transition-metal nitrosyl complex may also have a six FIG. 2 is a graph which illustrates the effects of pre electron Lewis base as ligand. As represented by LM(NO) incubation with vehicle, CpCr(NO)Cl, and nitroglycerin on Xs, La may represent a six-electron Lewis base. Six Subsequent relaxation responses to Cp Cr(NO), Cl, electron Lewis bases include baseS Such cyclopentadienyl nitroglycerin, and Sodium nitroprusside of phenylephrine anion and derivatives. In a preferred embodiment, the Six preconstricted rat aortic rings. 50 electron Lewis base is permethylcyclopentadienyl anion, and in a most preferred embodiment, the Six-electron Lewis DETAILED DESCRIPTION OF THE base is cyclopentadienyl anion. The transition-metal nitrosyl INVENTION complex may also bear ligand X wherein X may be carbon AS noted above, the present invention is directed toward monoxide where y is 1, and wherein X may be either a halide transition-metal nitrosyl complexes which are effective in 55 or pseudohalide where y is 2. Suitable halides include relaxing Smooth muscle in Vivo, without the development of fluoride, chloride, bromide, and iodide. Preferred halides tolerance associated with organic nitrites and nitrates Such as include chloride and iodide. Suitable pseudohalides include nitroglycerin, and without the toxic Side effects of Sodium nitrite, , and cyanide anion. nitroprusside. AS Smooth muscle relaxants, these transition In a preferred embodiment of the invention, the nitrosyl complexes are useful as pharmaceutical com 60 metal nitroSyl complex may bear two nitrosyl ligands. In positions for the treatment of a variety of conditions wherein particular, the transition-metal nitrosyl complex represented it is desired to relax Smooth muscles, including for example, by the general formula LM(NO), Xs, where y is 2 becomes hypertension, angina pectoris, congestive heart failure, and LM(NO)X. In a most preferred embodiment, the six impotence, as well as a variety of in vitro and in Vivo assays electron Lewis base, L, is cyclopentadienyl anion; X may described below. 65 be selected from the group consisting of chlorides and nitro, The transition-metal nitrosyl complexes of the present and the transition-metal, M, may be selected from the group invention may be generally represented by the formula consisting of chromium, molybdenum, and tungsten. 5,811,463 S 6 In another preferred embodiment, the transition-metal Sublingual, intramuscular, or oral), and in a manner which is may bear a Single nitrosyl ligand. The transition-metal in accordance with accepted practices, Such as disclosed in nitrosyl complex represented by the general formula LM Remington's Pharmaceutical Sciences, Gennaro, Ed., Mack (NO), Xs, where y is 1 becomes LM(NO)X. In a most Publishing Co., Easton, Pa. 1990 (which is incorporated preferred embodiment, the Six-electron Lewis base, L, is 5 herein by reference in its entirety). In addition, pharmaceu cyclopentadienyl anion; X is carbon monoxide, and the tical compositions of the present invention may be placed transition-metal, M, may be Selected from the group con within containers, along with packaging material which Sisting of chromium, molybdenum, and tungsten. indicates that the pharmaceutical composition may be uti lized to relax Smooth muscles. In yet another embodiment, the transition-metal nitrosyl AS noted above, the present invention also provides complex bears a Single nitrosyl ligand and is represented by methods for relaxing Smooth muscle in a warm-blooded the formula LM(NO)X where X may be XL. Such a animal (e.g., humans, chimps, cows, goats, pigs, dogs, cats, transition-metal nitroSyl complex is represented by the for rats or mice) comprising the Step of administering to the mula LM(NO)XL. In a most preferred embodiment the animal a therapeutically effective amount of a transition Six-electron Lewis, base, L, is cyclopentadienyl anion; X is metal nitrosyl complex as described above. Briefly, the iodide, the two-electron Lewis base, L, may be Selected 15 efficacy (and therapeutically effective amount) of a particu from the group consisting of pyridine, piperidine, and tri lar transition-metal nitroSyl complex may be readily deter methylphosphite; and the transition-metal, M, may be mined by both in vitro and in Vivo techniques. See generally Selected from the group consisting of chromium, Blattner et al., Experiments on Isolated Smooth Muscle molybdenum, and tungsten. Preparations, Hugo Sachs Elektronik KG, 1980 Within another aspect of the present invention, transition (incorporated herein by reference in its entirety). For metal nitrosyl complexes of the present invention may be example, within one embodiment, a Small animal Such as a represented by the formula M(NO)2X, wherein M is a rat, rabbit, or guinea-pig is Sacrificed and eXSanguinated. Group 6 or 8 transition metal, X is a halide or pseudohalide, The required tissue (e.g., artery, vein, Stomach, ileum, and when M is a Group 6 transition-metal, y is 2, and when jejunum, caecum, colon, uterus, Spleen, trachea, Seminal 25 vesicle, ductus deferens, or corpus cavernosum) is removed M is a Group 8 transition-metal, y is 1. and placed in an oxygenated physiological Solution (e.g., Some of the transition-metal nitrosyl complexes of the Krebs or Tyrode solution). This tissue is then cut such that present invention are Soluble in water and may be formu one end is attached to the bottom of a muscle bath, and the lated as solutions in normal saline (0.9% sodium chloride). other end to a force-displacement transducer (for example, Water-insoluble complexes or complexes which have diffi Grass FT-03-C, Quincy, Mass.) for isometric recording with culty dissolving in water may be dissolved in 100% dim a physiological recorder (Grass, Model 79G). An appropri ethylsulfoxide (DMSO), 60% DMSO in water, or 1:1 ate pre-load (0.5 to 10 grams, depending on the tissue) is DMSO:Tween 80 followed by dilution with normal saline. then applied, and the tissue equilibrated in the bath for one Some complexes may require 10–20 minutes Sonication to hour prior to the start of the study. effect Solution. 35 The measurement of Smooth muscle contractile response The transition-metal nitrosyl complexes of the present may then be performed essentially as described by Pang and invention may be formulated as pharmaceutical composi Sutter in Blood Vessels 17:293–301, 1980 (incorporated tions Suitable for administration to a warm-blooded animal. herein by reference in its entirety). Briefly, a dose-response Such formulations may contain an effective amount of the curve of an agonist (e.g., phenylephrine, norepinephrine) is transition- metal nitrosyl complex as well as one or more 40 first obtained by measuring contractile response of the pharmaceutically acceptable carriers and diluents. More muscle as a function of agonist concentration. Response to Specifically, the pharmaceutical compositions of the present the agonist is then measured beginning with the addition of invention may be administered as liquids, emulsions, or the lowest dose of agonist to the bath. Each dose is equili Suspensions containing acceptable diluents Such as alcohols brated in the bath for 2 to 6 minutes in order to assure a and Sterile water containing Sugars, Salts, preservatives, 45 Steady State response, prior to proceeding to the next higher Suspending agents, emulsifying agents, or administered as dose. lotions, creams, or gels containing acceptable diluents, Measurement of the effectiveness of Smooth muscle relax preservatives, or carriers to impart the desired texture, ant compositions may then be determined by measurement consistency, and Viscosity. Such acceptable diluents and of the Smooth muscle relaxation response, in a manner carriers are familiar to those skilled in the art, and include 50 analogous to contractile response. See generally Wang, Poon (but are not limited to) emulsifying agents Such as non-ionic and Pang, J. Pharm. Exp. Ther. 265:112-119, 1993 ethoxylated and non-ethoxylated Surfactants, fatty alcohols, (incorporated herein by reference in its entirety). Briefly, fatty acids, preserving agents, wax esters, triglyceride esters, tissue is first contracted with an agonist Such as Steroid alcohols, phospholipids Such as lecithin and phenylephrine, usually at a concentration which results in cephalin, polyhydric alcohol esters, fatty acid esters, hydro 55 90% of the tissue's maximum contractile response (referred philic lanolin derivatives, hydrophobic beeswax derivatives, to EC). At the steady state phase of the contractile hydrocarbon oils (Such as palm oil, coconut oil, and mineral response to the agonist, a cumulative dose-response curve oil), cocoa butter waxes, Silicon oils, pH balancers, and for the Smooth muscle relaxant may be determined. For each cellulose derivatives. The present invention may be admin concentration of relaxant, a steady State is achieved and the istered as tablets or capsules which may contain Suitable 60 relaxation response measured. The effective concentration diluents, or administered as inhalations, sprays or aerosols (EC) of a relaxant is the concentration that produces relax which may contain propellant, or aqueous or oily vehicles, ation of a contraction previously induced by an agonist. For or administered as Suppositories which may contain cocoa example, the ECso of a relaxant is the effective concentration butter, glycerinated gelatin or other Suitable bases. of the relaxant required to relax a pre-contracted muscle to Preferably, pharmaceutical compositions of the present 65 50% of its maximum contractile response. invention are formulated for the particular method of admin AS described in more detail below, transition-metal istration (e.g., intravenous, topical, pulmonary, rectal, nitrosyl complexes of the present invention have been 5,811,463 7 8 demonstrated to be effective in the relaxation of Smooth nitrosyl complex as described above. The effectiveness of muscle in vitro. In particular, a Series of transition-metal the transition-metal nitrosyl complexes described above for nitrosyl complexes were tested for their ability to relax the treatment of impotence may readily be evaluated through pre-constricted rat aortic rings in Vitro. All of the transition both in vitro and in Vivo investigation. More Specifically, in metal nitrosyl complexes completely and dose-dependently Vitro evaluation may be accomplished utilizing corpus cav relaxed phenylephrine pre-constricted aortae with efficacies ernoSum excised from the penis of a rabbit, which is then similar to that of sodium nitroprusside. The results of the in dissected free from the tunica albuginea, cut into Strips and Vitro relaxation experiments are presented in Example 5 and mounted at resting tension. Phenylephrine-precontracted Table 1. (See Example 5 for experimental detail.) Strips may be used in procedures similar to those described Likewise, in Vivo tests may also be readily developed and above, and in the examples, in order to construct dose utilized in order to determine the efficacy (and therapeuti response curves for the transition-metal nitrosyl complexes. cally effective amount) of the transition-metal nitrosyl com Similarly, in vivo studies may likewise be utilized in order plexes described above. For example, within one aspect of to test the efficacy of the transition-metal nitrosyl complexes the present invention, a Series of transition-metal nitrosyl described above in the treatment of impotence. Briefly, complexes were tested in vivo for their ability to reduce the 15 previous in Vivo Studies involving the Systemic or central mean arterial blood pressure of conscious rats in Vivo. administration of low doses of apomorphine and other Briefly, as described in greater detail below in Example 6, dopaminergic agonists, have shown that penile erection and conscious, unrestrained rats were intravenous bolus injected yawning can be reliably induced in rats (Heaton et al., J. with the transition-metal nitrosyl complexes. Five of the Urol. 145:1099-102, 1991). Therefore, in vivo studies may complexes were determined to dose-dependently reduce be utilized in order to investigate the utility of the transition mean arterial pressure with maximum depressor effects metal nitrosyl complexes of the present invention to facili which ranged from -36 to -59 mm Hg. Maximum depressor tate apomorphine-induced penile erection in rats. For effects of Sodium nitroprusSide and nitroglycerin under example, within one embodiment of the invention rats are Similar conditions were -58 and -56 mm Hg, respectively. individually placed in Plexiglas cages which have a mirror Three of the complexes exhibited potencies Similar to, or 25 arranged in an oblique position to enhance observation. greater than, that of nitroglycerin, and two of the complexes Pretreatment of the rats with a transition-metal nitrosyl were found to be more potent than Sodium nitroprusSide. complex may then be utilized to generate dose-response The results of the effects of the transition-metal nitrosyl curves for apomorphine-induced rat penile erections. complexes on the depression of mean arterial pressure in Within another aspect of the present invention, methods conscious rats in Vivo are Summarized in Example 5 and are provided for enhancing learning and memory in warm Table 1. blooded animals, comprising the Step of administering to a In addition to determining efficacy of transition-metal warm-blooded animal one of the above-described transition nitrosyl complexes in either in vitro or in Vivo tests, it is also metal nitrosyl complexes. The efficacy of Such complexes in generally preferably to determine whether the composition enhancing memory and learning may be readily ascertained induces tolerance. For example, within one embodiment of 35 through a variety of tests, including, for example, the Morris the invention, induction of tolerance by the transition-metal water maze performance test for rats. See Morris, Learn nitrosyl complex described above may be readily deter Motiv: 12:239-60, 1981; Decker and Majchrzak, Psychop mined by exposing rat aortic rings to a transition-metal harmacol. 107:530-34, 1992. Briefly, rats are given daily nitrosyl complex prior to relaxation determinations. More injections of various doses of transition-metal nitrosylcom Specifically, as described in greater detail 10 below in 40 plexes at a time just prior to Swimming Sessions. The rats are Example 7, intact rat aortic rings were pre-incubated with a then trained Several times daily over a period of Several days typical transition- metal nitrosyl complex for one hour prior in a pool filled with warm milk. In each training trial, the rats to examination of its concentration-relaxation effect in are given a short period to find a Submerged escape platform phenylephrine-preconstricted conditions. Unlike the relax placed in one of the quadrants of the pool. Those not locating ation response of nitroglycerin, the relaxation response of 45 the platform will be guided to it. The mean eScape latency the transition-metal nitrosyl complexes was not diminished on each training day may be determined as an index of by pre-exposure of the aortic ring to the complex. The results information acquisition. The amount of time the rat Spends indicate that tolerance develops less rapidly with the in the training quadrant where the escape platform had been transition-metal nitrosyl complexes of the present invention located during training will be indicative of acquired spatial than with nitroglycerin. 50 information. The effect of transition-metal nitrosyl com AS noted above, transition-metal nitrosyl complexes of plexes on acquired spatial information may reflect the utility the present invention may be readily administered to a of these complexes in learning and memory. warm-blooded animal in order to relax a variety of Smooth Transition-metal nitrosyl complexes of the present inven muscles. For example, within one aspect of the present tion have demonstrated beneficial vasodilation properties invention, therapeutically effective amounts of a transition 55 through their capacity to relax blood vessels in Vitro and metal nitrosyl complex may be administered to a warm depress blood pressure in Vivo. Furthermore, the complexes blooded animal in order to relax vascular Smooth muscles. are as effective in Vasodilation as existing nitrovasodilators, Through Such methods, transition-metal nitrosyl complexes nitroglycerin and Sodium nitroprusside. Additionally, the may be utilized in order to treat a variety of Vascular complexes do not appear to cause the adverse effect of disorders, including for example, hypertension, angina 60 tolerance associated with nitroglycerin nor toxicity associ pectoris, and congestive heart failure. ated with either nitroglycerin or Sodium nitroprusside. The present invention also provides methods for relaxing The following examples are offered by way of illustration, nonvascular Smooth muscle through the administration of and not by way of limitation. the transition-metal nitrosyl complexes described above. For example, within one aspect of the present invention methods 65 EXAMPLES are provided for treating impotence, comprising the Step of Examples 1-4 describe the preparation of the transition administering to a warm-blooded animal a transition-metal metal nitrosyl complexes. All manipulations of compounds 5,811,463 10 described in Examples 14 were performed under an atmo vacuo. Sublimation of the resulting brown residue at Sphere of prepurified dinitrogen or argon. All Solvents were 50°-60° C./0.005 torr onto a water-cooled probe for three distilled from appropriate drying agents and degassed prior days affords (m-CH-)W(CO) (NO) (13.6 g., 84% yield). to use. Standard Schlenk and glove-box techniques were The corresponding chromium and molybdenum com used throughout. Unless otherwise indicated, all other reac plexes are obtained similarly in yields of 60 and 93% tions and manipulations were carried out in a well ventilated respectively. The compounds are orange to orange-red Solids fume hood. In these examples, the following abbreviations readily Soluble in organic Solvents. The Solids are stable in are used: THF, tetrahydrofuran; CHCl, dichloromethane; air for short periods of time and indefinitely under dinitro IR, infrared spectrum; V refers to frequency; EIMS, electron impact mass spectroscopy; P" refers to parention; H NMR, gen. proton nuclear magnetic resonance, CDCls, deuteriochloro Analysis calculated for CHCr(CO) (NO): C, 41.39; H, form; mp, melting point, FAB, fast atom bombardment; 2.48; N, 6.90. Found: C, 41.40; H, 2.60; N, 6.70. IR P(OMe), trimethylphosphite, C5H1N, pip, piperidine; (CHCl). v. 2020, 1945; vy, 1680 cm. EIMS (probe CHN, py, pyridine. temperature 120° C): m/z. 203 P). Examples 5-7 present in vitro and in vivo results of the 15 Analysis calculated for CHMo(CO) (NO): C, 34.03; H, evaluation of the efficiency of the transition-metal nitrosyl 2.04; N, 5.67. Found: C, 34.28; H, 2.24; N, 5.54. IR complexes in relaxing blood vessels and lowering blood (CHCl). v. 2020, 1937; v, 1663 cm. EIMS (probe pressure. Sprague-Dawley rats (300–400g) were used for all temperature 120° C): m/z 249 P”). experiments. Each group consisted of 1-6 rats for the in vivo Analysis calculated for CHW(CO) (NO): C, 25.10; H, 1.50; N, 4.18. Found: C, 25.29; H, 1.70; N, 4.13. IR experiments and 3–6 aortic rings for the in Vitro experi (CHCl): vi, 2010, 1925; vy, 1655 cm. EIMS (probe ments. Each aortic ring was from a different rat. temperature 120° C): m/z. 319 P”). EXAMPLE 1. B. Preparation of Dicarbonyl(m- permethylcyclopentadienyl)nitrosyl Complexes of Molyb The Synthesis of Transition-Metal Nitrosyl 25 denum and Tungsten Complexes with Formula LM(NO), X The permethylated complexes, (m-C.Mes)M(CO) (NO) Example 1 presents the Syntheses of the transition-metal (M=Mo, W) are prepared in a manner similar to that of their nitrosyl complexes generally represented by the formula cyclopentadienyl analogues except that LiC5Me5) is used LM(NO),X. where y is 1: LM(NO)X. instead of NaCH), and that higher temperatures (100° C.) A. Preparation of Dicarbonyl(m-cyclopentadienyl) are required to effect their Sublimation. nitrosyl Complexes of Chromium, Molybdenum, and Tung Analysis calculated for CoHMo(CO) (NO): C, 36.17; Sten H, 4.55; N, 4.22. Found: C, 36.17; H, 4.60; N, 4.30. IR Briefly, a 200-mL, three-necked flask is fitted with a (CH,Cl): va, 1659 cm. EIMS (probe temperature 120° nitrogen inlet and Stirrer and is thoroughly flushed with C.); m/z 333 IP"). H NMR (CDC1) d 1.95 (s). prepurified dinitrogen. A dry THF solution (50 mL) con 35 Analysis calculated for CHW(CO) (NO): C, 22.93; taining 4.18 g of Sodium cyclopentadienide, NaCHs is H, 1.92; N, 5.34. Found: C, 22.96; H, 1.87; N, 5.26. IR syringed into the flask, and Cr(CO) (11.00 g, 50.0 mmol) (CH,Cl): V, 1690 cm. EIMS (probe temperature 120° and 100 mL of di-n-butyl ether are added. (In the case of C.); m/z 421 IP"). molybdenum and tungsten the di-n-butyl ether is not added.) 40 The flask is then equipped with a Leibig condenser and the EXAMPLE 2 reaction mixture is refluxed with Vigorous stirring for 12 h. The Synthesis of Transition-Metal Nitrosyl During this time, the reaction vessel is Shaken occasionally Complexes with Formula LM(NO)X to reintroduce any sublimed Cr(CO) into the refluxing This example presents the Synthesis of the transition reaction mixture. The final reaction mixture is allowed to 45 cool to room temperature and filtered, and the pale-yellow metal nitrosyl complexes generally represented by the for solid thus collected is washed with diethyl ether (3x10 mL), mula LM(NO),X, where y is 2: LaM(NO).X. and dried under nitrogen. The excess Cr(CO) and any A. Preparation of Chloro(m-cyclopentadienyl)dinitrosyl di-n-butyl ether remaining in this Solid are removed by Complexes of Chromium, Molybdenum, and Tungsten Sublimation at 90° C./0.005 torr onto a water-cooled probe. 50 Approximately 2–3 mL of nitrosyl chloride is condensed The NaI(m-C5H5)M(CO), complexes of molybdenum and into a 5-mL graduated cold trap held at -78 C. It is then tungsten are freed of any unreacted heXacarbonyl in a allowed to melt So that its Volume can be measured, and it Similar fashion, and all three Sodium Salts are used without is distilled under static vacuum into a 100-mL, two-necked further purification. flask held at -78 C. until just prior to use when it is allowed The preparations of all three (m-CH3)M(CO) (NO) 55 to warm to room temperature. complexes from their corresponding (m-CH3)M(CO)4)- All three (m-C5H5)M(NO)C1 complexes are prepared in anions are similar. The experimental procedure using the a similar manner, but to achieve maximum yields of the tungsten complex dicarbonyl(m-cyclopentadienyl) chromium and molybdenum compounds, the reactions nitrosyltungsten, is given below. should be performed at -78 C. The molybdenum complex A300-mL, three-necked flask is equipped with a nitrogen 60 (chloro(m-cyclopentadienyl)nitrosylmolybdenum), on the inlet, an addition funnel, and a Stirrer. It is charged with other hand, can be obtained in excellent yields even at room NaI(m-CH-)W(CO) (17.3 g, 48.5 mmol) and THF (120 temperature as disclosed below. mL). A THF solution (50 mL) containing Diazald A 200-mL, three-necked flask, equipped with a Stirrer, a (N-methyl-N--p-toluenesulfonamide, Aldrich nitrogen inlet, and an addition funnel, is charged with Chemical Co.) (10.4 g., 48.6 mmol) is syringed into the 65 (m-C5H5)Mo(CO) (NO) (6.5g, 26 mmol) and CHCl (100 addition funnel. The solution of Diazald is added dropwise mL). The nitrosyl chloride Solution (typically containing 2.3 over a period of 15 minutes and the solvent is removed in mL (50 mmol) of nitrosyl chloride in 30 mL of CHCl) is 5,811,463 11 12 added dropwise to the Stirred reaction mixture. Gas evolu A. Preparation of Iodo(m-cyclopentadienyl) tion occurs and the orange Solution becomes dark green. The nitrosylchromium Dimer reaction is monitored by and the To a stirred orange solution of CpCr(CO) (NO) (2.03 g, nitrosyl chloride solution is added until the carbonyl absorp 10.0 mmol) is added solid I (1.24g, 4.90 mmol). Reaction tions of the initial organometallic reactant have disappeared. 5 occurs after 5 min as evidenced by gas evolution and a color It is extremely important that a Stoichiometric amount of change of the reaction mixture to green-brown. After being nitrosyl chloride (CINO) be used, since even a slight excess Stirred for 1 h to ensure completion of the reaction, the of CINO reduces significantly the yields of the desired mixture is taken to dryneSS under reduced pressure. Crys products, especially in the cases of the chromium and tallization of the residue from CHCl2/hexanes affords dark tungsten complexes. Conversely, if an insufficient quantity green CpCr(NO)I. (2.35 g, 88% yield) of CINO is added, the separation of any unreacted (m- Analysis calculated for CHCr(NO)I: C, 21.92; H, CH3)M(CO)2(NO) from the desired chloronitrosyl com 1.84; N, 5.11; I, 46.33. Found: C, 22.00; H, 1.77; N, 5.00; pleX is difficult. The final reaction mixture is concentrated in I, 46.08. IR (CHCl). v., 1673 cm. EIMS (probe tem vacuo to approximately 30 mL, and is filtered through a perature 120° C): m/z 548 (P. mp 119° C. (dec). short (3x5 cm) Florisil column. The column is washed with 15 CHCl until the washings are colorless, and the combined B. Preparation of Iodo(m-cyclopentadienyl)nitrosyl(L) filtrates are then concentrated in vacuo to a volume of 30 chromium (L=P(OMe), pip, py) mL. Dry hexanes are added until crystallization appears to Olive green CpCr(NO)I) (0.27 g., 0.50 mmol) is dis be complete; approximately 100-125 mL of hexanes are solved in CHCl (20 mL). To the stirred solution, 0.081 mL required. The resulting green crystals are collected by (1.0 mmol) of pyridine is added by syringe. The color filtration, washed with hexane (2x15 mL) and dried under changes from dark green to a brighter green over the period dinitrogen to obtain analytically pure (m-C5H5)Mo(NO)Cl of 30 min. The solution is reduced in volume to approxi (6.0 g, 89% yield). The chromium and tungsten compounds mately 10 mL, and then filtered through alumina (1.5x3 cm) are obtained similarly (except that the reaction flask is Supported on a medium-porosity frit. The column is washed maintained at -78° C. during the addition of the CINO) in 25 with CHCl until the washings are colorless. Hexanes (30 yields of 77 and 72%, respectively. mL) are added, and the Solution is reduced in Volume until Analysis calculated for CHCr(NO)Cl: C, 28.26; crystallization is initiated. The mixture is then cooled to H.2.37; N, 13.18; C1, 16.68. Found: C, 28.2; H, 2.55; N, -30 C. overnight to complete crystallization. The solvent is 12.86; C1, 16.99. IR (CHCl). v., 1816, 1711 cm. EIMS removed by cannulation, and the dark green crystals are (probe temperature 120° C): m/z 212 P. mp 144° C. washed with pentane (3x15 mL). This solid is dried in vacuo (dec). to obtain 0.32 g (90% yield) of CpCr(NO)(py)I. Analysis calculated for CHMo(NO)Cl: C, 23.42; H, Analysis calculated for CHNOCrI: C, 34.02; H, 2.85; 1.96; N, 10.92. Found: C, 23.53; H, 1.90; N, 10.70. IR N, 7.93. Found: C, 34.15; H, 2.82; N, 7.80. IR (Nujol mull): (CHCl). v., 1759, 1665 cm. EIMS (probe temperature vo, 1651 cm. FAB mass spectrum: m/z. 353 IP"). 120° C): m/z 258 LP"). mp 116° C. 35 The preparation of CpCr(NO)(L)I where L=piperidine Analysis calculated for CHW(NO)Cl: C, 17.44; H, and L=P(OMe) is analogous to above where L=pyridine. 1.46; N, 8.13; C1, 10.29. Found: C, 17.68; H, 1.68; N, 8.10; Data for CpCr(NO)P(OMe)I: 0.35g, 88% yield. Analy Cl, 10.26. IR (CHCl). v., 1733, 1650 cm. EIMS (probe sis calculated for CHNOPCrI: C, 24.13; H, 3.54; N, temperature 120° C): m/z 343 LP "P 127° C. (dec). 3.52. Found: C, 24.19; H, 3.54; N, 3.49. IR (Nujol mull): B. Preparation of Nitrito(m-cyclopentadienyl) 40 vo, 1650 cm. FAB mass spectrum: m/z 398 IP"). dinitrosylchromium Data for CpCr(NO)(pip)I. Analysis calculated for CpCr(NO)C1 (1.0 g, 4.7 mmol) (Cp=m-C5H5) is dis CHNOCrI: C, 33.44; H, 4.49; N, 7.80. Found: C, 33.23; solved in 50 mL distilled water. A distilled water solution (25 H, 4.46; N, 7.70. IR (Nujol mull): vs. 1651 cm. FAB mass mL) of AgNO (0.72 g, 4.7 mmol) is added to the stirred, 45 spectrum: m/z. 359 P. green solution of CpCr(NO)Cl. A flocculent white solid forms immediately. Stirring is continued for 30 min, and EXAMPLE 4 then the mixture is filtered to remove the white Solid. The The Synthesis of Transition-Metal Nitrosyl filtered solution is extracted with 3x50 mL of CHCl2. The Complexes with Formula M(NO),X), extracts are combined, dried with anhydrous NaSO, and 50 filtered. The Solvent is removed in vacuo until the Solution This example presents the Syntheses of transition-metal reaches a volume of 20 mL. Twenty mL of hexanes are then nitrosyl complexes generally represented by the formula added, and the mixture is cooled to -30° C. to induce M(NO).X.), when M is a Group 6 transition metal, y is 2, crystallization. This procedure results in the formation of and when M is a Group 8 transition metal, y is 1. analytically pure CpGr(NO) (ONO) (0.51 g, 49% yield). 55 A. Preparation of Dichloro Dinitrosyl Complexes of Analysis calculated for CHCr(NO)(ONO): C, 26.92; Molybdenum and Tungsten H, 2.26; N, 18.83. Found: C, 27.21; H, 2.15; N, 18.76. IR A 500-mL, three-necked round-bottom flask is equipped (CH,Cl). v., 1825, 1719 cm. H NMR (CDC1) d 5.78 with a gas inlet and an addition funnel, and is charged with (s). EIMS (probe temperature 120° C): m/z 223 P"). powdered W(CO) (17.6 g., 50.0 mmol). The entire system 60 is then thoroughly purged with prepurified dinitrogen and EXAMPLE 3 CHCl (90 mL) is added. The addition funnel is charged The Synthesis of Transition-Metal Nitrosyl Complexes with a CHCl solution of CINO (6 mL, 120 mmol). 6 mL with Formula LM(NO)XL of the CINO Solution is added to the flask, and a small This example presents the Syntheses of the transition amount of air (0.50 mL) is added by syringe to initiate the metal nitrosyl complexes generally represented by the for 65 reaction. The addition of the CINO Solution is carried out in mula LM(NO), X, where y is 1 and X is XL: LaM(NO) a dropwise manner, the reaction being monitored by IR XL. spectroscopy. When the IR band at 1977 cm due to the 5,811,463 13 14 W(CO) starting material has disappeared, the addition is EXAMPLE 5 halted, and the Solution is allowed to stir for a further 30 min. Relaxation of Blood Vessels Utilizing Transition The green precipitate which has formed is collected on a Metal Nitrosyl Complexes glass frit, washed with CH2Cl and dried in vacuo to obtain Example 5 presents the results of the evaluation of the W(NO),Cl) (14.52 g, 92% yield) as an olive green efficacy of the transition metal nitrosyl complexes in blood powder. vessel relaxation. In these in vitro experiments, phenyleph Analysis calculated for WNOCl: C, 0.00; H, 0.00; N, rine pre-constricted rat aortae were Subjected to increasing 8.90; C1, 22.52. Found: C, 0.23; H, 0.00; N, 8.88; C1, 21.97. doses of the transition-metal nitrosyl complexes, and the IR (Nujol mull): vs. 1794, 1680 cm. relaxation of the rat aortae measured. Concentration The preparation of Mo(NO)2Cl2 is analogous to response (relaxation) curves were prepared, and the ability W(NO),Cl), above. Analysis calculated for MoNOCl: of the transition-metal nitrosyl complexes to relax the blood N, 12.4; Cl, 31.3; Mo, 42.3. Found: N, 12.7; C1, 32.2, Mo, vessels were compared and related to nitroglycerin and 41.1. IR (Nujol mull): v., 1805, 1690 cm. 91% yield Sodium nitroprusside. The results of these experiments are based on MO(CO). reported in terms of ECs (effective concentration to pro 15 duce a 50% relaxation response) and E, (percent relax B. Preparation of Iodo(dinitrosyl)iron Dimer ation of phenylephrine-induced contraction) and are tabu Dry acetone (250 mL) and 33.5 g (0.3 mol) of iron powder lated in Table 1. are added to a 1000-mL three-necked round-bottom flask Briefly, rats were sacrificed by a blow on the head equipped with a gas inlet, a 500-mL pressure-equalizing followed by eXSanguination. The thoracic aorta was addition funnel equipped with a bubbler, and a mechanical removed and cleared of connective tissues. Four ring Seg Stirrer. A Solution containing 38.1 g (0.1 mol) of I2 in Et2O ments of 0.5 cm length each were prepared from one aorta (300 mL) is then added through the addition funnel dropwise and Suspended in Separate organ baths. Each ring was over a period of 45-74 min. After the addition is complete, connected to a Grass FT-03-C force-displacement transducer NO gas is introduced into the flask at a rate sufficient to (Quincy, Mass.) for isometric recording with a pre-load of 1 maintain a flow of gas of about one bubble per Second 25 g and was equilibrated for 1 hr (with 3 washouts) in normal through the bubbler. Stirring must be sufficiently vigorous to Krebs solution (pH 7.4) at 37° C. with a gas mixture of 95% ensure efficient uptake of the NO gas. The end of the O and 5% CO. The Krebs solution had the following reaction is marked by the cessation of the absorption of nitric composition (10M): NaCl, 118; glucose, 11; KCl, 4.7; CaCl, 2.5; NaHCO, 25; KHPO, 1.2: MgCl6HO, 1.2. oxide as determined by the use of the bubbler. Phenylephrine (10M, EC-o) was added to the baths and The solvent is removed in vacuo. When the black residue at 15–20 min later, at the Steady State phase of the contractile is completely dry, the desired product is Sublimed onto a response to phenylephrine, a cumulative concentration water-cooled probe. The flask should be heated slowly (2 response curve of Sodium nitroprusside or a metal nitrosyl C/min) to 90° C. Explosive decomposition can take place if complex was constructed using sequential doubling of the the heating rate is excessive. The product can be Scraped off drug concentration. Each concentration of a drug was left in the probe in an inert atmosphere drybox and Stored under 35 the bath for 2-4 minto ensure a plateau response is reached. dinitrogen. The time taken to complete each concentration-response Analysis calculated for Fe(NO)II: Fe, 23.0; I, 52.3. curve was approximately 20 min. FIG. 1A shows Found: Fe, 23.4; I 51.8. IR (Nujol mull): v., 1807, 1766 concentration-response curves of CpCr(NO)Cl and CpMo cm (NO)Cl.

TABLE 1. The effects of metal nitrosyl complexes on relaxation of pre-constricted aortic rings in Vitro and depression of mean arterial pressure in conscious rats in Vivo In vitro In vivo ECso Emax EDso Emax Toxic dose Entry Name n (uM) (%) n (ug/kg) (mm Hg) (mg/kg) 1 Na nitroprusside 6 0.052 -128 6 8.4-58 10,000 2 Nitroglycerin 6 O.30 -104 6 200 -56 3 CpCr(NO)Cl 6 O.11 -118 6 4.4-42 2,000? 4 CpMo(NO),Cl 6 17 -113 6 400 -59 25,600P 5 CpW(NO),CI 6 2.0 -122 2 2976 -36 40,000 6 CpCr(NO)(pip)I 5 0.45 -110 - - - 7 CpW(NO)(CO), 3 2.5 –107 3 - biphasic 500 8 Mo(NO)Cl. 6 81 -119 2 >5120 no effect - 9 W(NO)Cl. 6 335 -108 1 >5120 no effect - 10 Fe(NO).I2 6 20 -108 2 554 -51 5120° 11 CpCr(NO)(ONO) 6 0.1 -98 2 1.5 -45 128Oa "denotes dose which caused convulsion denotes dose which caused death denotes dose which caused neither convulsion nor death. 'denotes depressor followed by pressor response.

65 5,811,463 15 16 EXAMPLE 6 between nitroglycerin and CpCr(NO)Cl. See FIG. 2C. Pre-exposure to nitroglycerin Slightly, but Statistically Sig Depression of Arterial Blood Pressure Utilizing nificantly (p<0.05), increased the ECso of sodium Transition-Metal Nitrosyl Complexes nitroprusside, Suggesting a Small degree of cross-tolerance Example 6 describes the results of the determination of between these two drugs. See FIG. 2D. the efficacy of the transition-metal nitrosyl complexes in From the foregoing, it will be appreciated that, although specific embodiments of the invention have been described reducing blood pressure of conscious, unrestrained rats. In herein for purposes of illustration, various modifications these in Vivo experiments, the depression of rat arterial may be made without deviating from the Spirit and Scope of preSSure was measured as a function of transition-metal the invention. Accordingly, the invention is not limited nitrosyl complex dose. Dose-response (pressure depression) except by the appended claims. curves were constructed and the capacities of the transition We claim: metal nitroSyl complexes to reduce arterial pressure were 1. A method for relaxing Smooth muscle in a warm compared and related to nitroglycerin and Sodium nitroprus blooded animal, comprising administering to Said animal a Side. The results of these experiments are reported in terms therapeutically effective amount of a transition-metal of EDs (effective dose to produce a 50% pressure depres 15 nitrosyl complex represented by the formula Sion response) and E (maximum decrease in blood LM(NO), X pressure) and are Summarized in Example 5, Table 1. wherein L is a two-electron Lewis base or L is a six The rats were anaesthetized with halothane (4% in air for electron Lewis base, M is a Group 6 or 8 transition-metal, induction and 1.5% in air for Surgery). A polyethylene wherein y is 1 or 2, X is either a halide or pseudohalide, and cannula (PE50) was inserted into the left iliac artery for the when y is 1, Xis either Lor XL. measurement of mean arterial pressure (MAP) by a pressure 2. A method for treating hypertension in a warm-blooded transducer (P23DB, Gould Statham, Calif.). APE50 cannula animal, comprising administering to Said animal a therapeu was also inserted into the iliac vein for the administration of tically effective amount of a transition-metal nitrosyl com the vehicle or a drug. The rats were then put into a Small cage plex represented by the formula and given>6 hr to recover from the effects of anaesthesia and 25 LM(NO), X surgery before use. The rats were allowed to move freely wherein L is a two-electron Lewis base or L is a six during the recovery phase as well as during the conduction electron Lewis base, M is a Group 6 or 8 transition-metal, of the experiments as described in Wang and Pang, Br. J. wherein V is I or 2. and when y is 2, X is either a halide or Pharmacol. 103:2004-8, 1991. Only one dose-response pseudohalide, and when y is 1, X2 is either L or XL. curve of a metal nitrosyl complex, Sodium nitroprusSide, 3. A method for treating angina pectoris in a warm nitroglycerin or vehicle (0.9% NaCl solution or dimethyl blooded animal, comprising administering to Said animal a Sulfoxide Solution) was constructed in each rat. All drugs or therapeutically effective amount of a transition-metal vehicle were intravenous bolus injected at dose-intervals of nitrosyl complex represented by the formula 3-7 min, to allow the recovery of depressor responses, LM(NO), X 35 wherein L is a two-electron Lewis base or L is a six before the injection of the next higher dose. electron Lewis base, M is a Group 6 or 8 transition-metal, Five of the eight metal nitrosyl complexes tested were wherein V is 1 or 2, X is either a halide or pseudohalide, and found to lower blood pressure in a dose-related manner. The when y is 1, X is either L or XL. maximum depressor responses of Some of the metal nitrosyl 4. A method for treating congestive heart failure in a complexes were Similar to those of nitroglycerin and Sodium 40 warm-blooded animal, comprising administering to Said nitroprusside. The potencies (inverse of EDso) of Some of animal a therapeutically effective amount of a transition the metal nitrosyl complexes were similar to or greater than metal nitrosyl complex represented by the formula those of nitroglycerin and sodium nitroprusside. FIG. 1B shows concentration-response curves of CpCr(NO)Cl and LM(NO), X CpMo(NO)Cl. wherein L is a two-electron Lewis base or L is a six 45 electron Lewis base, M is a Group 6 or 8 transition-metal, EXAMPLE 7 wherein V is 1 or 2, X is either a halide or pseudohalide, and when y is 1, X is either L or XL. Example 7 presents the results of pre-exposing the rat 5. A method for treating impotence in a warm-blooded aortae to the transition-metal nitrosyl complexes to evaluate animal, comprising administering to Said animal a therapeu the development of tolerance. In these experiments, the rat 50 tically effective amount of a transition-metal nitrosyl com aortic rings were pretreated with a transition-metal nitrosyl plex represented by the formula complex prior to the measurement of dose-dependent relax LM(NO), X ation as described above in Example 5. wherein L is a two-electron Lewis base or L is a six In this study, the vehicles, CpCr(NO)C1 (10M), or electron Lewis base, M is a Group 6 or 8 transition-metal, nitroglycerin (3x10"M) was added to the baths for 1 h, 55 wherein y is 1 or 2 and when y is 2, X is either a halide or followed by the washout of drug or vehicle, preconstriction pseudohalide, and when y is 1, X is either L or XL. with phenylephrine and the construction of a concentration 6. A method for relaxing Smooth muscle in a warm relaxation curve of CpCr(NO)Cl. blooded animal, comprising administering to Said animal a The results show that the pre-incubation with CpCr(NO) therapeutically effective amount of a transition-metal Cl did not alter the ECso or E of the compound (see FIG. 60 nitrosyl complex represented by the formula 2A), Suggesting that tolerance to the tested drug does not M(NO).X.), readily develop. Pre-incubation with nitroglycerin shifted wherein M is a Group 6 or 8 transition-metal, X is either a the nitroglycerin dose-response curve to the right with ECso halide or pseudohalide, and when M is a Group 6 transition Significantly increased and E, unaltered. See FIG. 2B. metal, y is 2, and when M is a Group 8 transition-metal, y This shows that tolerance develops to nitroglycerin. Pre 65 is 1. exposure to nitroglycerin did not alter the ECso or E of 7. A method for treating hypertension in a warm-blooded CpCr(NO)2Cl, showing that no cross-tolerance exists animal, comprising administering to Said animal a therapeu 5,811,463 17 18 tically effective amount of a transition-metal nitroSyl com tically effective amount of a transition-metal nitrosyl com plex represented by the formula plex represented by the formula M(NO).X.), M(NO).X.), wherein M is a Group 6 or 8 transition-metal, X is either a wherein M is a Group 6 or 8 transition-metal, X is either a halide or pseudohalide, and when M is a Group 6 transition halide or pseudohalide, and when M is a Group 6 transition metal, y is 2, and when M is a Group 8 transition-metal, y metal, y is 2, and when M is a Group 8 transition-metal, y is 1. is 1. 8. A method for treating angina pectoris in a warm 11. A pharmaceutical composition, comprising a blooded animal, comprising administering to Said animal a transition-metal nitrosyl complex represented by the for therapeutically effective amount of a transition-metal mula nitrosyl complex represented by the formula M(NO).X.), M(NO).X.), wherein M is a Group 6 or 8 transition-metal, X is either a wherein M is a Group 6 or 8 transition-metal, X is either a halide or pseudohalide, and when M is a Group 6 transition halide or pseudohalide, and when M is a Group 6 transition metal, y is 2, and when M is a Group 8 transition-metal, y metal, y is 2, and when M is a Group 8 transition-metal, y 15 is 1. is 1, and a pharmaceutically acceptable carrier or diluent. 9. A method for treating congestive heart failure in a 12. The pharmaceutical composition of claim 11 wherein warm-blooded animal, comprising administering to Said X is a halide Selected from the group consisting of fluoride, animal a therapeutically effective amount of a transition chloride, bromide and iodide. metal nitrosyl complex represented by the formula 13. The pharmaceutical composition of claim 11 wherein M(NO).X.), X is a pseudohalide Selected from the group consisting of wherein M is a Group 6 or 8 transition-metal, X is either a nitrite, nitrate, and cyanide anion. halide or pseudohalide, and when M is a Group 6 transition 14. The pharmaceutical composition of claim 11 wherein metal, y is 2, and when M is a Group 8 transition-metal, y the transition-metal nitrosyl complex is iodo(dinitrosyl)iron is 1. 25 dimer. 10. A method for treating impotence in a warm-blooded animal, comprising administering to Said animal a therapeu