WO1992003472A1 - Peptides containing amino-polyols as transition-state mimics - Google Patents

Abstract

The present invention provides novel peptides. These peptides have a novel non-cleavable transition state insert corresponding to the 10,11-position of the renin substrate (angiotensinogen) of formula (XL8). These peptides are useful as renin inhibitors and as inhibitors of retroviral proteases. Renin inhibitors are useful for the diagnosis and control of renin-dependent hypertension, congestive heart failure, renin-dependent hyperaldosterism, other renin-dependent cardiovascular disorders and ocular disorders. Inhibitors of retroviral proteases, such as the HIV-I protease, are useful for treating diseases caused by retroviruses, such as human acquired immunodeficiency disease syndrome (AIDS).

Description

PEPTIDES CONTAINING

AMINO-POLYOLS AS TRANSITION-STATE MIMICS

DESCRIPTION BACKGROUND OF THE INVENTION

The present invention provides novel compounds. More particularly, the present invention provides novel peptide analogs. The peptides of the present invention contain an amino-polyol moiety as a novel non-cleavable transition state insert corresponding to the 10,11-position of the renin substrate (angiotensinogen). These peptides are useful as renin inhibitors and as inhibitors of retroviral proteases. Renin inhibitors are useful for the diagnosis and control of renin-dependent hypertension, congestive heart failure, renin-dependent hyperaldosterism, and other renin-dependent cardiovascular disorders. Inhibitors of retroviral proteases, such as the HIV-I protease, are useful for treating diseases caused by retroviruses, such as human acquired immunodeficiency disease syndrome (AIDS).

Renin is an endopeptidase which specifically cleaves a particular peptide bond of its substrate (angiotensinogen), of which the N-terminal sequence in equine substrate is for example:

Renin

Asp-Arg-Val-Tyr-Ile-His-Pro-Phe-His-Leu-Leu-Val-Tyr-Ser- IA

1 2 3 4 5 6 7 8 9 10 11 12 13 14 as found by L.T. Skeggs, et al., J. Exper. Med. 106, 439 (1957). Human renin substrate has a different sequence as recently discovered by D.A. Tewkesbury, et al., Biochem. Biophys. Res. Comm. 99, 1311 (1981). It may be represented as follows:

Renin

-Val-Ile-His- 11 12 13 IB and having the sequence to the left of the arrow (1) being as designated in formula IA above.

Renin cleaves angiotensinogen to produce angiotensin I, which is converted to the potent pressor angiotensin II. A number of angiotensin I converting enzyme inhibitors are known to be useful in the treatment of hypertension. Inhibitors of renin are also useful in the treatment of hypertension.

A number of renin-inhibitory peptides have been disclosed. Thus, U.S. patent 4,424,207;

European published applications 45,665; 104,041; and 156,322; and European published application 0173481, published 5 March 1986; disclose certain peptides with the dipeptide at the 10,11-position containing an isostere bond. A number of statine derivatives stated to be renin inhibitors have been disclosed, see, e.g., European published applications 77,028; 81,783; 114,993; 156,319; and 156,321; and U.S. patents 4,478,826; 4,470,971; 4,479,941; and 4,485,099. Terminal disulfide cycles have also been disclosed in renin inhibiting peptides; see, e.g., U.S. patents 4,477,440 and 4,477,441. Aromatic and aliphatic amino acid residues at the 10,11 position of the renin substrate are disclosed in U.S. patents 4,478,827 and 4,455,303. C-terminal amide cycles are disclosed in U.S. patent 4,485,099 and European published applications 156,320 and 156,318. Certain tetrapeptides are disclosed in European publications 111,266 and 77,027. Further, European published application No. 118,223 discloses certain renin inhibiting peptide analogs where the 10-11 peptide link is replaced by a one to four atom carbon or carbon-nitrogen link. Additionally, Holladay, et al., in "Synthesis of Hydroxyethylene and Ketomethylene Dipeptide lsosteres", Tetrahedron Letters, Vol. 24, No. 41, pp. 4401-4404, 1983 disclose various intermediates in a process to prepare stereo-directed "ketomethylene" and "hydroxyethylene" dipeptide isosteric functional groups disclosed in the above noted U.S. Patent No. 4,424,207.

Additionally, published European Applications 45,161 and 53,017 disclose amide derivatives useful as inhibitors of angiotensin converting enzymes.

Certain dipeptide and tripeptides are disclosed in U.S. patents 4,514,332; 4,510,085; and 4,548,926 as well as in European published applications 128,762; 152,255; and 181,110. Pepstatin derived renin inhibitors have been disclosed in U.S. patent 4,481,192. Retro-inverso bond modifications at positions 10-11 have been disclosed in U.S. patent 4,560,505 and in European published applications 127,234 and 127,235. Derivatives of isosteric bond replacements at positions 10-11 have been disclosed in European published applications 143,746 and 144,209; and European published application 237202, published 16 September, 1987. Isosteric bond modifica- tions at positions 11-12 and 12-13 have been disclosed in European published application 179,352. Certain peptides containing 2-substituted statine analogues have been disclosed in European published application 157,409. Certain peptides containing 3-aminodeoxystatine have been disclosed in European published application 161,588. Certain peptides containing 1-amino-2-hydroxybutane derivatives at positions 10-11 have been disclosed in European published application 172,346. Certain peptides containing l-amino-2-hydroxypropane derivatives at positions 10-11 have been disclosed in European published application 172,347. Certain peptides containing N-terminal amide cycles have been disclosed in PCT application, Publication No. WO 87/05909, published 8 October 1987. Certain peptides containing dihalostatine have been disclosed in PCT application, Publication No. WO 86/06379, published 6 November 1986.

European published applications 156,322; 114,993; and 118,223; and PCT application,

Publication No. WO 87/02986, published 21 May 1987; European published application 0173481, published 5 March 1986; European published application 237202, published 16 September 1987; and PCT application, Publication No. WO 87/05909, published 8 October 1987; disclose hydroxamic acids or esters at the C-terminus.

INFORMATION DISCLOSURE

European patent application 189,203, published 30 July 1986, and European patent application 230,266, published 24 July 1987, disclose N-dihydroxyalkyl peptide derivatives which are useful as inhibitors of renin for treating hypertension. They specifically disclose the intermediate compound , (2S , 3R,4S)- 1 -amino-2 , 3-dihydroxy-4-tert-butoxy-carbonylamino-5- cyclohexylpentane, and the final compound Boc-Phe-His Amide of (2S,3R,4S)-1-(3- Methylbutylcarbonylamino)-2,3-dihydroxy-4-amino-5-cyclohexylpentane.

European patent application 184,855, published 18 June 1986, discloses new hydroxy carbonyl substiruted-statine peptide derivatives which are useful as inhibitors of renin for treating hypertension.

Derivatives of isosteric bond replacements at positions 10,11 as dihydroxy ethylene isosteres are disclosed in International Patent Application, PCT/US87/00291, published 11 September 1987 (Publication No. WO87/05302).

International Patent Application, PCT/US 87/03007, published 30 June 1988 (Publication No. WO88/04664) discloses renin inhibitory peptides having an epoxide or glycol moiety at the 10, 11-position.

International Application PCT/US90/03754, filed 9 July 1990, discloses peptides having a diamino glycol moiety as the non-cleavable transition state insert corresponding to the 10,11-position of the renin substrate (angiotensinogen). These peptides are disclosed as being useful as renin inhibitors and as inhibitors of retroviral proteases.

Ciba-Gigy has described a compound that is reported to exhibit good oral activity and it includes 3-t-butylsulfonyl-2-benzylpropionic acid as the N-terminal group. P. Buhlmayer et al., J. Med. Chem 31: 1839 (1988).

Derwent Abstracts, Accession Nos. 89-101394, 89-101395, 89-101396 and 89-101397, disclose compounds having an amino:diol alkyl moiety as the transition state insert which are useful as renin inhibitors in the treatment of hypertension.

SUMMARY OF THE INVENTION

The present invention particularly provides:

A peptide having a non-cleavable transition state insert corresponding to the 10,11 -positionof a renin substrate (angiotensinogen) of the formula XL₈

wherein R₁ is

(a) hydrogen,

(b) C₁-C₁₀ alkyl, (c) aryl,

(d) C₃-C₇ cycloalkyl,

(e) Het,

(f) C₁-C₃ alkoxy, or

(g) C₁-C₃ alkylthio;

wherein each R₂ is independently

(a) hydrogen,

(b) C₃-C₇ alkyl,

(c) -(CH₂)_p-aryl, or

(d) -(CH₂)_p-Het;

wherein p is 0 to 2 inclusive;

wherein q is three to six, inclusive;

wherein aryl is phenyl or naphthyl substituted by zero to three of the following:

(a) C₁-C₃ alkyl,

(b) hydroxy,

(c) C₁-C₃ alkoxy,

(d) halo,

(e) amino,

(f) mono- or di- C₁-C₃ alkylamino,

(g) -CHO,

(h) -COOH,

(i) -CONH₂,

(j) -CONH( C₁-C₅ alkyl),

(k) -nitro,

(l) mercapto,

(m) C₁-C₃ alkylthio,

(n) C₁-C₃ alkylsulfinyl,

(o) C₁-C₃ alkylsulfonyl,

(p) -N(R₈)- C₁-C₃ alkylsulfonyl,

(q) SO₃H,

(r) SO₂NH₂,

(s) -CN, or

(t) -CH₂NH₂;

wherein -Het is a 5- or 6-membered saturated or unsaturated ring containing from one to three heteroatoms selected from the group consisting of nitrogen, oxygen, and sulfur; and including any bicyclic group in which any of the above heterocyclic rings is fused to a benzene ring, which heterocyclic moiety is substituted with zero to three of the following:

(a) C₁-C₆ alkyl,

(b) hydroxy,

(c) trifluoromethyl,

(d) C₁-C₄ alkoxy,

(e) halo,

(f) aryl,

(g) aryl C₁-C₄ alkyl-,

(h) amino,

(i) mono- or di-( C₁-C₄ alkyl) amino, or

(j) C₁-C₅ alkanoyl;

wherein R₈ is

(a) hydrogen,

(b) C₃-C₇ alkyl,or

(c) (CH₂)_p-phenyl;

or a carboxy, amino or other reactive group protected form or a pharmaceutically acceptable acid or base addition salt thereof.

These compounds are shown in relation to the human renin substrate as follows:

6 7 8 9 10 11 12 13

-His Pro Phe His Leu Val He His-

X A₆ B₇ C₈ D₉ E₁₀ F₁₁ G₁₂ H₁₃ I₁₄ Z,

The present invention provides peptide inhibitors of renin and retroviral proteases which contain an amino-polyol moiety as a novel non-cleavable transition state insert corresponding to the 10,11 -position of the renin substrate (angiotensinogen).

By "renin inhibitory peptide" is meant a compound capable of inhibiting the renin enzyme in mammalian metabolism and having three or more amino acid residues linked by peptidic or pseudo-peptidic bonds.

By "a non-cleavable transition state insert" is meant a transition state insert which is not cleavable by a hydrolytic enzyme in mammalian metabolism. Renin inhibitory peptides having a variety of such transition state inserts, corresponding to the 10,11 -position of the renin substrate, are known in the art, including those disclosed in the following references, which are hereby incorporated by reference:

U.S. patent 4,424,207 (Szelke); European patent 104041A (Szelke); European patent application 144,290A (Ciba Geigy AG); European patent 0,156,322 (Merck); European patent 161- 588A (Merck); European pa nt 0,172,347 'Abbott); European patent 172-346-A (Abbott); European patent 156-318 (Merck); European patent 157-409 (Merck); European patent 152-255 (Sankyo); and U.S. patent 4,548,926 (Sankyo); and

PCT application, Publication No. WO 87/05302, published 11 September 1987; PCT application, Publication No. WO 87/05909, published 8 October 1987; PCT application,

Publication No. WO 86/06379, published 6 November 1986; PCT application, Publication No. WO

88/04664, published 30 June 1988; and European published application 0173481, published 5

March 1986; and

A. Spaltenstein, P. Carpino, F. Miyake and P.B. Hyskins, Tetrahedron Letters, 27:2095 (1986); D.H. Rich and M.S. Bernatowicz, J. Med. Chem., 25:791 (1982); Roger, J. Med. Chem., 28: 1062 (1985); D.M. Glick, et al., Biochemistry, 21:3746 (1982); D.H. Rich, Biochemistry, 24:3165 (1985); R.L. Johnson, J. Med. Chem., 25:605 (1982); R.L. Johnson and K. Verschovor, J. Med. Chem., 26:1457 (1983); R.L. Johnson, J. Med. Chem., 27:1351 (1984); P.A. Bartlett, et al., J. Am. Chem. Soc, 106:4282 (1984); and Peptides: Synthesis, Structure and Function (V.J. Hruby; D.H. Rich, eds.) Proc. 8th American Peptide Sym., Pierce Chemical Company, Rockford, 111., pp. 511-20; 587-590 (1983).

By "derivatives" of amino acids is meant the well known amino acid derivatives commonly employed in renin inhibitors as set forth in the references above.

Examples of the peptides of the present invention are represented by formula I. In formula I, the non-cleavable transition state insert, corresponding to the 10,11-position of the renin substrate, is designated E₁₀.

Most renin inhibitory peptides known in the art are highly lipophilic and have low water solubility. There is a need in this field for renin inhibitory peptides with increased hydrophilicity and thus higher water solubility. In general, peptides which are water soluble are readily adaptable to oral administration.

The peptides of the present invention meet this need by having a novel amino-polyol moiety as their non-cleavable transition state insert. This insert results in peptides having increased hydrophilicity and thus higher water solubility. Some of the peptides of the present invention also contain other modifications, s as a beta-valine group or a glucosamine carbohydrate group at the N-terminus, which further mproves their hyprophilicity and water solubility.

Another advantage of these peptides is their small size since the polyhydroxylated unit is part of the transition state insert. This unit is C-terminally truncated which further reduces the overall size of the final peptides. In the peptides of the present invention, tetraols are the preferred transition state analog inserts.

As is apparent to those of ordinary skill in the art, the renin inhibitory peptides of the present invention can occur in several diastereomeric forms, depending on the configuration around the asymmetric carbon atoms. All such diastereomeric forms are included within the scope of the present invention. Preferably, the stereochemistry of the amino acids corresponds to that of the naturally-occurring amino acids.

Renin inhibitory peptides commonly have protecting groups at the N-terminus and the C- terminus. These protecting groups are known in the polypeptide art. Examples of these protecting groups are given below. Any of these protecting groups are suitable for the renin inhibitory peptides of the present invention.

Examples of pharmaceutically acceptable acid addition salts include: acetate, adipate, alginate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, citrate, camphorate, camphorsulfonate, cyclopentanepropionate.digluconate, dodecylsulfate, ethane-sulfonate, fumarate, glucoheptanoate, glycerophosphate, hemisulfate, heptanoate, hexanoate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate, lactate, maleate, methanesulfonate, 2- naphthalenesulfonate, nicotinate, oxalate, palmoate, pectinate, persulfate, 3-phenylpropionate, picrate, pivalate, propionate, succinate, tartrate, thiocyanate, tosylate, and undecanoate.

The carbon atom content of various hydrocarbon-containing moieties is indicated by a prefix designating the minimum and maximum number of carbon atoms in the moiety, i.e., the prefix (C_i-C_j) indicates a moiety of the integer "i" to the integer "j" carbon atoms, inclusive. Thus (C₁-C₄)alkyl refers to alkyl of one to 4 carbon atoms, inclusive, or methyl, ethyl, propyl, butyl, and isomeric forms thereof. C₄-C₇cyclic amino indicates a monocyclic group containing one nitrogen and 4 to 7 carbon atoms.

Examples of (C₃-C₁₀)cycloalkyl, which include alkyl-substituted cycloalkyl containing a total of up to 10 total carbon atoms, are cyclopropyl, 2-methylcyclopropyl, 2,2-dimethylcyclopropyl, 2,3-diethylcyclopropyl, 2-butylcyclopropyl, cyclobutyl, 2-methylcyclobutyl,

3-propylcyclobutyl, cyclopentyl, 2,2-dimethylcyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclononyl, cyclodecyl and isomeric forms thereof.

Examples of aryl include phenyl, naphthyl, (o-, m-, or p-)tolyl, (o-, m-, or p-)ethylphenyl, 2-ethyl-tolyl, 4-ethyl-o-tolyl, 5-ethyl-m-tolyl, (o-, m-, or p-)propylphenyl, 2-propyl-(o-, m-, or p-)tolyl, 4-isopropyl-2,6-xylyl, 3-propyl-4-ethylphenyl, (2,3,4- 2,3,6-, or 2,4,5-)trimethylphenyl, (o-, m-, or p-)fluorophenyl, (o-, m-, or p-trifluoromethyl)phenyl, 4-fluoro-2,5-xylyl, (2,4-, 2,5-, 2,6-, 3,4-, or 3,5-)difluorophenyl, (o-, m-, or p-)chlorophenyl, 2-chloro-p-tolyl, (3-, 4-, 5- or 6-)chloro-o-tolyl, 4-chloro-2 -propyl phenyl, 2-isopropyl-4-chlorophenyl, 4-chloro-3-fluorophenyl, (3- or 4-)chloro-2 -fluorophenyl, (o-, m-, or p-)trifluoro-methyl phenyl, (o-, m-, or p-)ethoxyphenyl, (4- or 5-)chloro-2-methoxy-phenyl, and 2,4-dichloro(5- or

6-)methyl phenyl, and the like.

Examples of -Het include: 2-, 3-, or 4-pyridyl, imidazolyl, indolyl, Nⁱⁿ-formyl-indolyl,

Nⁱⁿ- C₁-C₅alkyl-C(0)-indolyl, 1 ,2,4-triazolyl, 2-, 4-, or 5-pyrimidinyl, 2- or 3-thienyl, piperidinyl, pyrryl, pyrrolinyl,pyrrolidinyl pyrazolyl, pyrazolinyl, pyrazolidinyl, imidazolinyl, imidazolidinyl, pyrazinyl,piperazinyl, pyridazinyl, oxazolyl, oxazolidinyl, isoxazolyl, isoxazolidinyl, morpholinyl, thiazolyl, thiazolidinyl, isothiazolyl, isothiazolidinyl, quinolinyl, isoquinolinyl, benzimidazolyl, benzothiazolyl, benzoxazolyl, furyl, thienyl, and benzothienyl. Each of these moieties may be substituted as noted above.

As would be generally recognized by those skilled in the art of organic chemistry, a heterocycle as defined herein for -Het would not be bonded through oxygen or sulfur or through nitrogen which is within a ring and part of a double bond.

Halo is halogen (fluoro, chloro, bromo, or iodo) or trifluoromethyl.

Examples of pharmaceutically acceptable cations include: pharmacologically acceptable metal cations, ammonium, amine cations, or quaternary ammonium cations. Especially preferred metal cations are those derived from the alkali metals, e.g., lithium, sodium, and potassium, and from the alkaline earth metals, e.g., magnesium and calcium, although cationic forms of other metals, e.g., aluminum, zinc, and iron are also within the scope of this invention. Pharmacologically acceptable amine cations are those derived from primary, secondary, or tertiary amines.

The novel peptides herein contain both natural and synthetic amino acid residues. These residues are depicted using standard amino acid abbreviations (see, e.g., IUPAC-IUB Joint Commission on Biochemical Nomenclature (JCBN), "Nomenclature and Symbolism for amino Acids and Peptides," Eur. J. Biochem. 138:9-37 (1984)) unless otherwise indicated.

The peptides of this invention are useful for treating any medical condition for which it is beneficial to reduce the levels of active circulating renin. Examples of such conditions include renin-dependent hypertension, hypertension, hypertension under treatment with another antihypertensive and/or a diuretic agent, congestiveheart failure, renin-dependent hyperaldosterism, angina, post-myocardial infarction, other renin-dependent cardiovascular disorders and ocular disorders. The renin-angiotension system may play a role in maintenance of intracellular homeostasis: see Clinical and Experimental Hypertension, 86, 1739-1742 (1984) at page 1740 under Discussion.

The peptides of the present invention are preferably orally administered to humans to effect renin inhibition for the purpose of favorably affecting blood pressure. For this purpos. , the compounds are administered from 0.1 mg to 100 mg per kg per dose, administered from 1 to 4 times daily. Equivalent dosages for other routes of administration are also employed. For example, renin-associated hypertension and hyperaldosteronism are effectively treated by the administration of from 0.5 to 50 milligrams of the compound per kilogram of body weight per day. The exact dose depends on the age, weight, and condition of the patient and on the frequency and route of administration. Such variations are with in the skill of the practitioner or can readily be determined.

The peptides of the present invention to effect renin inhibition may be in the form of pharmaceutically acceptable salts both those which can be produced from the free bases by methods well known in the art and those with which acids have pharmacologically acceptable conjugate bases.

Conventional forms and means for administering renin-inhibiting compounds may be employed and are described, e.g. , in U.S. Patent No. 4,424,207 which is incorporated by reference herein. Likewise, the amounts disclosed in the U.S. Patent No. 4,424,207 are examples applicable to the compounds of the present invention.

The peptides of the present invention to effect renin inhibition are preferably orally administered in the form of pharmacologically acceptable acid addition salts. Preferred pharmacologically acceptable salts for oral administration include the citrate and aspartate salts, although any pharmacologically acceptable salt is useful in this invention, including those listed above. These salts may be in hydrated or solvated form.

For renin inhibition, the peptides of the present invention may be administered topically, parenterally, by inhalation spray, or rectally in dosage unit formulations containing conventional non-toxic pharmaceutically acceptable carriers, adjuvants and vehicles. The term parenteral as used herein includes subcutaneous injections, intravenous, intramuscular, intrasternal injection or infusion techniques. In addition to the treatment of warm-blooded animals such as mice, rats, horses, dogs, cats, etc., the compounds of the invention are effective in the treatment of humans.

The pharmaceutical compositions of the peptides of the present invention for renin inhibition may be in the form of a sterile injectable preparation, for example as a sterile injectable aqueous or oleagenous suspension. This suspension may be formulated according to the known art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic parenterally-acceptable diluent or solvent, for example as a solution in 1,3-butanediol. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose any bland fixed oil may be employed including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid find use in the preparation of injectables.

The peptides of this invention may also be administered in the form of suppositories for rectal administration of the drug. These compositions can be prepared by mixing the drug with a suitable non-irritating excipient which is solid at ordinary temperatures but liquid at the rectal temperature and will therefore melt in the rectum to release the drug. Such materials are cocoa butter and polyethylene glycols. The peptides of this invention may e administered in combination with other agents used in antihypertensive therapy such as diuretics, a and/or β-adrenergic blocking agents, CNS-acting agents, adrenergic neuron blocking agents, vasodilators, angiotensin I converting enzyme inhibitors, and the like as described for example in published European patent application 156,318.

The present invention is also directed to combinations of the novel renin-inhibitory peptides of Formula I with one or more antihypertensive agents selected from the group consisting of diuretics, a and/or β-adrenergic blocking agents, CNS-acting agents, adrenergic neuron blocking agents, vasodilators, angiotensin I converting enzyme inhibitors, and other antihypertensive agents.

For example, the compounds of this invention can be given in combination with such compounds or salt or other derivative forms thereof as:

Diuretics: acetazolamide; amiloride; bendroflumethiazide; benzthia-zide; bumetanide; chlorothiazide; chlorthalidone; cyclothiazide; ethacrynic acid; furosemide; hydrochlorothiazide; hydroflumethiazide; indacrinone (racemic mixture, or as either the (+) or (-) enantiomer alone, or a manipulated ratio, e.g., 9:1 of said enantiomers, respectively); metolazone; methyclothiazide; muzolimine; polythiazide; quinethazone; sodium ethacryπ :e; sodium nitroprusside; spironolactone; ticrynaten; trimaterene; trichlormethiazide;

α-Adrenergic Blocking Agents: dibenamine; phentolamine; phenoxybenzamine; prazosin; tolazoline;

β-Adrenergic Blocking Agents: atenolol; metoprolol; nadolol; propranolol; timolol;

((±)-2-[3-(te/t-butylamino)-2-hydroxypropoxy]-2-furananilide) (ancarolol);

(2-acetyl-7-(2-hydroxy-3-isopropylaminopropoxy)benzofuran HCl)(befunolol);

((±)-1-(isopropylamino)-3-(p-(2-cyclopropylme-hoxyethyl)-phenoxy)-2-propranol HCl)(betaxolol);

(1-[(3,4-dimethoxyphenedιyl)amino]-3-(m-tolyloxy)-2-propanol HCl)(bevantolol);

(((±)-1-(4-((2-isopropoxyethoxy)methyl)phenoxy)-3-isopropylamino-2-propanol)

fumarate) (bisoprolol);

(4-(2-hydroxy-3-[4-(phenoxymethyl)-piperidino]-propoxy)-indole);

(carbazolyl-4-oxy-5,2-(2-medιoxyphenoxy)-ethylamino-2-propanol);

(1-((1 ,1-dimethylethyl)amino)-3-((2-methyl 'H-indol-4-yl)oxy)-2-pro-panol benzoate) (bopindolol);

(1-(2-exobicyclo[2.2.1]-hept-2-ylphenoxy)-3-[(1-methyledιyl)-amino]-2-propanol HCl) (bomaprolol);

(o-[2-hydroxy-3-[(2-indol-3-yl-1,1-dimethyledιy imino]propoxy]benzonitrile HCl) (bucindolol);

(α-[(tert.butylamino)me-hyl]-7-edιyl-2-benzofuranmedιanol) (bufur-alol);

(3-[3-acetyl-4-[3-(tert.butylamino)-2-hydroxypropyl]-phenyl]-1 ,1-diethylurea HCl) (celiprolol);

((±)-2-[2-[3-[(1 , 1-dimethylethyl)amino]-2-hydroxypropoxy]phenoxy]-N-methylacetamide HCl) (cetamolol);

(2-benzimidazolyl-phenyl(2-isopropylaminopropanol)); ((±)-3'-acetyi-4'-(2-hydroxy-3-isopropylaminopropoxy)-acetanilide HCl) (diacetolol);

(methyl-4-[2-hydroxy-3-[(1 -methylethyI)aminopropoxyl]]-benzene-propanoate HCl) (esmolol); (erythro-DL-1-(7-methylindan-4-yloxy)-3-isopropylaminobutan-2-ol);

(1-(tert.butylamino)-3-[0-(2-propynyloxy)phenoxy]-2-propanol (pargo-lol);

(1-(tert.butylamino)-3-[0-(6-hydrazino-3-pyridazinyl)phenoxy]-2-propanol di HCl) (prizidilol); ((-)-2-hydroxy-5-[(R)-1-hydroxy-2-[(R)-(1-medιyl-3-phenylpropyl)-amino]ethyl]benzamide); (4-hydroxy-9-[2-hydroxy-3-(isopropylamino)-propoxy]-7-methyl-5H-furo[3,2-g][1]-benzopyran-5- one) (iprocrolol);

((-)-5-(tert.butylamino)-2-hydroxypropoxy]-3,4-dihydro-1-(2H)-naphdιalenoneHCl)0evobunolol); (4-(2-hydroxy-3-isopropylamino-propoxy)-1,2-benzisodιiazole HCl);

(4-[3-(tert.butylamino)-2-hydroxypropoxy]-N-methylisocarbostyril HCl);

((±)-N-2-[4-(2-hydroxy-3-isopropylaminopropoxy)phenyl]ethyl-N'-isopropylurea) (pafenolol); (3-[[(2-trifluoroacetamido)edιyl]amino]-1-phenoxypropan-2-ol);

(N-(3-(o-chlorophenoxy)-2-hydroxypropyl)-N'-(4'-chloro-2,3-dihydro-3-oxo-5-pyridazinyl)- ethylenediamine);

((±)-N-[3-acetyl-4-[2-hydroxy-3-[(1-methylethyl)amino]propoxyphenyl]-butanamide)

(acebutolol);

((+)-4'-[3-(tert-butylamino)-2-hydroxypropoxy]spiro[cyclohexane-1,2'-indan]-1 '-one)

(spirendolol);

(7-[3-[[2-hydroxy-3-[(2-methylindol-4-yl)oxylpropyl]amino]butyl]thio-phylline) (teoprolol);

((±)-1-tert.butylamino-3-(thiochroman-8-yloxy)-2-propanol) (tertato-lol);

((±)-1-tert.butylamino-3-(2,3-xylyloxy)-2-propanol HCl) (xibenolol);

(8-[3-(tert.tberutylamino)-2-hydroxypropoxy]-5-methylcoumarin) (bucumo-lol);

(2-(3-(tert.butylamino)-2-hydroxy-propoxy)benzonitrile HCl) (bunitro-lol);

((±)-2'-[3-(tert-butylamino)-2-hydroxypropoxy-5'-fluorobutyrophenone) (butofiloloI);

(1 -(carbazol-4-yloxy)-3-(isopropylamino)-2-propanol) (carazolol);

(5-(3-tert.butylamino-2-hydroxy)propoxy-3,4-dihydrocarbotyril HCl) (carteolol);

(1-(tert.butylamino)-3-(2,5-dichlorophenoxy)-2-propanol) (clorano-lol);

(1-(inden-4(or 7)-yloxy)-3-(isopropylamino)-2-propanol HCl) (indeno-lol);

(1-isopropylamino-3-[(2-methylindol-4-yl)oxy]-2-propanol) (mepindo-lol);

(1-(4-acetoxy-2,3,5-trimemylphenoxy)-3-isopropylaminopropan-2-oI) (metipranolol);

(1-(isopropylamino)-3-(o-methoxyphenoxy)-3-[(1-me-hylethyl)amino]-2-propanol) (moprolol); ((1-tert.butylamino)-3-[(5,6,7,8-tetrahydro-cis-6,7-dihydroxy-1-naphthyl)oxy]-2-propanol) (nadolol); ((S)-1-(2-cyclopentylphenoxy)-3-[(1,1-dimethyledιyl)amino]-2-propanol sulfate (2: 1)) (penbutolol); (4'-[1-hydroxy-2-(amino)ethyl]methanesulfonanilide) (sotalol); (2-medιyl-3-[4-(2-hydroxy-3-tert.butylaminopropoxy)phenyl]-7-methoxy-isoquinolin-1-(2H)-one);

(1-(4-(2-(4-fluorophenyloxy)efhoxy)phenoxy)-3-isopropylamino-2-propanol HCl);

((-)-p-[3-[(3,4-dimethoxyphenethyl)amino]-2-hydroxypropoxy]-β-methyl-cinnamonitrile)

(pacrinolol);

((±)-2-(3'-tert.butylamino-2'-hydroxypropylthio)-4-(5^,-carbamoyl-2'-thienyl)thiazole HCI)

(arotinolol);

((+)-1-[p-[2-(cyclopropylmethoxy)edιoxy]phenoxy]-3-(isopropylamino)-2-propanol) (cicloprolol);

((±)-1-[(3-chloro-2-methylindol-4-yl)oxy]-3-[(2-phenoxyedιyl)amino]-2-propanol) (indopanolol);

((±)-6-[[2-[[3-(p-butoxyphenoxy)-2-hydroxypropyl]amino]ethyl]amino]-1,3-dimethyluracil) (pirepolol);

(4-(cyclohexylamino)-1-(1-naphdιolenyloxy)-2-butanol);

(1-phenyl-3-[2-[3-(2-cyanophenoxy)-2-hydroxypropyl]aminoethyl]hydan-toin HCl);

(3,4-dihydro-8-(2-hydroxy-3-isopropylaminopropoxy)-3-nitroxy-2H-1-benzopyran) (nipradolol);

Angiotensin I Converting Enzyme Inhibitors:

1-(3-mercapto-2-medιyl-1-oxopropyl)-L-proline (captopril);

(1-(4-ethoxycarbonyl-2,4(R,R)-dimethylbutanoyl)indoline-2(S)-car-boxylic acid);

(2-[2-[(1-(ethoxycarbonyl)-3-phenyl-propyl]amino]-1-oxopropyI]-1,2,3,4-tetrahydro-3-isoquinoline carboxylic acid);

((S)-1-[2-[(1-(ethoxycarbonyl)-3-phenylpropyl]amino]-1-oxopropyl]octahydro-1H-indole-2-carboxylic acid HCI);

(N-cyclopentyl-N-(3-(2,2-dimethyl- 1-oxopropyl)thiol-2-methyl-1-oxo-propyl)glycine) (pivalopril);

((2R,4R)-2-(2-hydroxyphenyl)-3-(3-mercaptopropionyl)-4-thiazolidine-carboxylic acid);

(1-(N-[1(S)-ethoxycarbonyl-3-phenylpropyl]-(S)-alanyl)-cis,syn-octa-hydroindol-2(S)-carboxylic acid HCI);

((-)-(S)-1-[(S)-3-mercapto-2-methyl-1-oxopropyl]indoline-2-carboxylic acid);

([1(S),4S]-1-[3-(benzoylthio)-2-methyl-1-oxopropyl]-4-phenylthio-L-proline;

(3-([1-ethoxycarbonyl-3-phenyl-(1S)-propyl]amino)-2,3,4,5-tetrahydro-2-oxo-1-(3S)-benzazepine-1-acetic acid HCI);

(N-(2-benzyl-3-mercaptopropanoyl)-S-ethyl-L-cysteine) and the S-methyl analogue;

(N-(1(S)-edιoxycarbonyl-3-phenylpropyl)-L-alanyl-L-proline maleate) (enalapril);

N-[1-(S)-carboxy-3-phenylpropyl]-L-alanyl-1-proline;

N²-[1-(S)-carboxy-3-phenylpropyl]-L-lysyl-L-proIine (lysinopril);

Other Antihypertensive Agents: aminophylline; cryptenamine acetates and tannates; deserpidine; meremethoxylline procaine; pargyline; tri-methaphan camsylate; and the like, as well as admixtures and combinations thereof.

Typically, the individual daily dosages for these combinations can range from about one- fifth of the minimally recommended clinical dosages to the maximum recommended levels for the entities when tiiey are given singly. Coadministration is most readily accomplished by combining the active ingredients into a suitable unit dosage form containing the proper dosages of each. Other methods of coadministration are, of course, possible.

Thus, the novel peptides of the present invention possess an excellent degree of activity in treating renin-associated hypertension and hyperaldosteronism.

Renin inhibitors have also been disclosed to control the rise in intraocular pressure associated with the use of steroidal anti-inflammatory drugs as described in International Application PCT/US86/02291 (International Publication Number WO 87/02581 dated 7 May 1987).

The peptides of the present invention are also useful as novel human retroviral protease inhibitory peptide analogs. Therefore, the peptides of the present invention inhibit retroviral proteases and thus inhibit the replication of the virus. They are useful for treating human patients infected with a human retrovirus, such as human immunodeficiency virus (strains of HIV-1 or HIV-2) or human T-cell leukemia viruses (HTLV-I or HTLV-II) which results in acquired immunodeficiency syndrome (AIDS) and/or related diseases.

The capsid and replicative enzymes (i.e. protease, reverse transcriptase, integrase) of retroviruses are translated from the viral gag and pol genes as polyproteins that are further processed by the viral protease (PR) to the mature proteins found in the viral capsid and necessary for viral functions and replication. If the PR is absent or nonfunctional, the virus cannot replicate. The retroviral PR, such as HIV-1 PR, has been found to be an aspartic protease with active site characteristics similar to those exhibited by the more complex aspartic protease, renin.

The term human retrovirus (HRV) includes human immunodeficiency virus type I, human immunodeficiency virus type II, or strains thereof, as well as human T cell leukemia virus 1 and

2 (HTLV-1 and HTLV-2) or strains apparent to one skilled in the art, which belong to the same or related viral families and which create similar physiological effects in humans as various human retroviruses.

Patients to be treated would be those individuals: 1) infected with one or more strains of a human retrovirus as determined by the presence of either measurable viral antibody or antigen in the serum and 2) in the case of HIV, having either a symptomatic AIDS defining infection such as i) disseminated histoplasmosis, ii) isopsoriasis, iii) bronchial and pulmonary candidiasis including pneumocystic pneumonia iv) non-Hodgkin's lymphoma or v) Kaposi's sarcoma and being less than sixty years old; or having an absolute CD4 lymphocyte count of less than 200/m³ in the peripheral blood. Treatment would consist of maintaining an inhibitory level of the peptide used according to this invention in the patient at all times and would continue until the occurrence of a second symptomatic AIDS defining infection indicates alternate therapy is needed.

More specifically, an example of one such human retrovirus is the human immunodeficiency virus (HIV, also known as HTLV-III or LAV) which has been recognized as the causative agent in human acquired immunodeficiency disease syndrome (AIDS), P. Duesberg, Proc. Natl. Acad. Sci. USA, 86:755 (1989). HIV contains a retro viral encoded protease, HIV-I protease, that cleaves the fusion polypeptides into the functional proteins of the mature virus particle, E.P. Lillehoj, et al., J. Virology, 62:3053 (1988); C. Debuck, et al., Proc. Natl. Acad. Sci., 84:8903 (1987). This enzyme, HIV-I protease, has been classified as an aspartyl protease and has a demonstrated homology to other aspartyl proteases such as renin, L.H. Pearl, et al., Nature 329:351 (1987); I. Katoh, et al., Nature 329:654 (1987). Inhibition of HIV-I protease blocks the replication of HIV and thus is useful in the treatment of human AIDS, E.D. Clerq, J. Med. Chem. 29:1561 (1986). Inhibitors of HIV-I protease are useful in the treatment of AIDS.

Pepstatin A, a general inhibitor of aspartyl proteases, has been disclosed as an inhibitor of

HIV-I protease, S. Seelmeier, et al., Proc. Natl. Acad. Sci. USA, 85:6612 (1986). Other substrate derived inhibitors containing reduced bond isosteres or statine at the scissle position have also been disclosed, M.L. Moore, et al, Biochem. Biophys, Res. Commun. 159:420 (1989); S. Billich, et al., J. Biol. Chem. 263:17905 (1988); Sandoz, D.E. 3812-576-A.

Thus, the peptides of the present invention are useful for treating diseases caused by retroviruses, such as human acquired immunodeficiency disease syndrome (AIDS), using dosages, forms and modes of administration equivalent to those described above for renin inhibition. Exact dosages, forms and mode of administration would be apparent to one of ordinary skill in the art such as a physician or pharmacologist.

The peptides of the present invention are also useful for treating non-human animals infected with a retrovirus, such as cats infected with feline leukemia virus. Other viruses that infect cats include, for example, feline infectious peritonitis virus, calicivirus, rabies virus, feline immunodeficiency virus, feline parvovirus (panleukopenia virus), and feline chlamydia. Exact dosages, forms and modes of administration of the peptides of the present invention to non-human animals would be apparent to one of ordinary skill in the art, such as a veterinarian.

The compounds of the present invention are prepared as depicted in the charts and as described more fully in the Preparations and Examples. In these charts, the variables are as defined above.

CHART A

Chart A describes the preparation of 1-cyclohexyl-2S-amino-3R,4,5,6-tetrahydroxyhexane in protected form which is incorporated into certain peptides of the present invention as the novel transition state analog insert.

The syntheses of this insert starts with the aldehyde of formula A-1, obtained from International Application, Publication No. WO 88/04664, published 30 June 1988. Reaction with carbomethoxymethylenetriphenylphosphorane gives the unsaturated ester of formula A-2. This compound is reduced with diisobutylaluminum hydride to the corresponding alcohol of formula A 3. Reaction with osmium tetroxide affords the m-hydroxylated products of formula A-5a (wherein X₁ and X₂ are each α-OH) and formula A-5b (wherein X₁ and X₂ are each β-OH), as a mixture of diastereomers. The compound of formula A-3 is also reacted with m-chloroperbenzoic acid to give the epoxides of formula A-4a and A-4b as a mixture of diastereomers which are hydrolyzed to a mixture of compounds of formula A-6a (wherein X₁ is α-OH and X is β-OH) and formula A- 6b (wherein X₂ is β-OH and X₂ is α-OH).

CHART B

Chart B illustrates the preparation of the novel peptides N-terr-butyloxycarbonyl-L- phenylaIanyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane of formula B-7a

(wherein X₁ and X₂ are each α-hydroxy) and N-tert-butyloxycarbonyl-L-phenylalanyl-L-histidyl- 2S-amino-1-cyclohexyl-3R,4S,5S,6-tertahydroxyhexane of formula B-7b (wherein X₁ and X₂ are each β-hydroxy) of the present invention.

The mixture of formula B-1a,b prepared as the compounds of formula A-5a,b in Chart A, is treated with potassium hydride and benzyl bromide to give the mixture of tri-benzyl ethers of formula B-2a (wherein Y₁ and Y₂ are each α-O-benzyl) and formula B-2b (wherein Y₁ and Y₂ are each β-O-benzyl). The acid labile protecting groups are removed with methanolic hydrogen chloride to give the mixture formula B-3a,b. Coupling with N-tert-butyloxycarbonyl-N^ιm-tosyl-L- histidine using diethylphosphoryl cyanide affords the compounds of formula B-4a,b which are readily separated. The compound of formula B-4a (wherein Y₁ and Y₂ are each α-O-benzyl) is treated with trifluoroacetic acid and the resulting amine is coupled with N-tert-butyloxycarbonyl-L-phenylalanine using diethylphosphoryl cyanide to give the compound of formula B-5a (wherein Y₁ and Y₂ are each α-O-benzyl). The tosyl protecting group is removed with 1-hydroxybenzotriazole to give the compound of formula B-6a (wherein Y₁ and Y₂ are each α-O-benzyl) which is then hydrogenolyzed using palladium catalyst to give the desired final compound of formula B-7a (wherein X₁ and X₂ are each α-hydroxy).

In a similar manner, the diastereomer of formula B-4b (wherein Y₁ and Y₂ are each β-O-benzyl) is converted in the sequence of reactions to the desired final compound of formula B-7b (wherein X₁ and X₂ are each β-hydroxy).

CHART C

Chart C illustrates the preparation of the novel peptides N-tert-butyloxycarbonyl-L- phenylalanyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,5S,6-tertahydroxyhexane of formula C-7a

(wherein X₁ is α-hydroxy and X₂ is β-hydroxy) and N-tert-butyloxycarbonyl-L-phenylalanyl-L- histidyl-2S-amino-1-cyclohexyl-3R,4S,5R,6-tetrahydroxyhexaneof formula C-7b(wherein X₁ is β-hydroxy and X₂ is α-hydroxy) of the present invention.

The mixture of formula C-1a,b prepared as the compounds of formula A-6a,b in Chart A, is treated with potassium hydride and benzyl bromide to give the mixture of tri-benzyl ethers of formula C-2a (wherein Y₁ is α-O-benzyl and Y₂ is β-O-benzyl) and formula C-2b (wherein Y₁ is β-O-benzyl and Y₂ is α-O-benzyl). The acid labile protecting groups are removed with methanolic hydrogen chloride to give a mixture of compounds of formula C-3a,b. Coupling with N-tert-butyIoxycarbonyl-N^im-tosyl-L-histidine using diethylphosphoryl cyanide affords a mixture of compounds of formula C-4a,b. This mixture is treated with trifluoroacetic acid and the resulting amines are coupled with N-tert-butyloxycarbonyl-L-phenylalanine using diethylphosphoryl cyanide to give a mixture of compounds of formula C-5a,b. The tosyl protecting group is removed with 1-hydroxybenzotriazole to give a mixture of compounds of formula C-6 a,b which is then hydrogenolyzed using palladium catalyst to give the desired compounds of formula C-7a (wherein X, is α-hydroxy and X₂ is β-hydroxy) and formula C-7b (wherein X₁ is β-hydroxy and X₂ is α-hydroxy) as two separable diastereomers.

CHART D

Chart D describes the preparation of 2S-amino-1-cyclohexyl-6-methyl-3R,4S,5,6-tetrahydroxyheptane in protected form which is used as a transition-state-analogue insert for the construction of certain peptides of the present invention.

The synthetic route begins from the allylic alcohol of formula D-1 which is prepared from the Grignard reaction between the corresponding aldehyde of formula A-1 of Chart A and 2-methyl-1-propenyl magnesium bromide. The olefin of formula D-1 is treated with a solution of osmium teroxide in t-butanol employing N-methylmorpholine N-oxide as an internal oxidation source to afford the cw-hydroxylated tri-ols of formula D-2. The alcohols are then protected as benzyl ethers by treatment with potassium hydride followed by addition of benzyl bromide. The resulting benzyl ethers of formula D-3 are reacted with methanolic hydrogen chloride to remove the acid labile groups. The free amine of formula D-4 is then utilized in the construction of other peptides of the present invention.

CHART E

Chart E illustrates the preparation of the peptide N-tert-butyloxycarbonyl-L-phenylalanyl-L-histidyl-2S-amino-1-cyclohexyl-6-methyl-3R,4R,5S,6-tetrahydroxyheptane of formula E-5 of the present invention.

The free amine of formula E-1, prepared as the compound of formula D-4 in Chart D is reacted with Boc-His(Ts)-OH employing diethylphosphoryl cyanide as the coupling agent and diisopropylethyl amine as the hindered base. The resulting peptide of formula E-2 is treated with trifluoroacetic acid to remove the tert-butyloxy carbonyl group and the resulting amine is coupled to Boc-Phe-OH to afford the peptide of formula E-3. The major isomer is separated from a mixture of minor components and is subsequently treated with 1-hydroxybenzotriazole to remove the tosyl protecting group from histidine. In the final synthetic step, the peptide of formula E-4 is subjected to hydrogenolysis with palladium black to afford the final peptide of formula E-5.

CHART F

Chart F describes the preparation of the novel peptide βVal-Phe-His-Y of formula F-3 (wherein Y is 2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane), also known as β-Valyl-L- phenylalanyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane, bis tri- fluoroacetate of the present invention.

The peptide of formula F-1, prepared as the compound of formula B-7a in Chart B, is deprotected with trifluoroacetic acid and the resulting amine is coupled to Boc-βVal-OH with diethyl cyanophosphonate to give the peptide of formula F-2. The final step is removal of the tert- butyloxy carbonyl group with trifluoracetic acid to give the desired peptide of formula F-3 which is isolated as the bis trifluoroacetate salt. In Chart F, Y is 2S-amino-1-cyclohexyl-3R,4R,5R,6- tetrahydroxyhexane.

CHART G

Chart G describes the preparation of the novel peptide Boc-Tyr(OMe)-His-Y of formula G-4 (wherein Y is 2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane), also known as N-tert- butyloxycarbonyl-O-methyl-L-tyrosyl-L-histidyl-2S-amino-l -cyclohexyl-3R,4R,5R,6- tetrahydroxyhexane of the present invention.

The peptide of formula G-1, prepared as the compounds of formula B-4a in Chart B, is deprotected with trifluoroacetic acid and the resulting amine is then coupled to Boc-Tyr(OMe) DCHA salt with DEPC and to afford the compound of formula G-2. The protecting groups must be removed to yield the desired final product. The tosyl group is removed with 1-hydroxybenzotriazole in methanol to give the compound of formula G-3. Finally, the benzyl protecting groups are removed by hydrogenolysis with palladium black to afford the desired compound of formula G-4.

In Chart G, Y is 2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane and X is 2S-amino-1-cyclohexyl-3R-hydroxy-4R,5R,6-tribenzyloxyhexane.

CHART H

Chart H describes the preparation of the novel peptide βVal-Tyr (OMe)-His-Y of formula H-5 (wherein Y is 2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane), also known as β-Valyl-O-methyl-L-tyrosyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane, bis trifluoroacetate of the present invention.

Coupling of the amine from the peptide of formula H-1, prepared as the compound of formula G-2 in Chart G, with Boc-βVal-OH gives the compound of formula H-2. The protecting groups must be removed to yield the desired peptide. The tosyl group is removed with 1-hydroxybenzotriazole to give the peptide of formula H-3. The benzyl protecting groups are removed by hydrogenolysis with palladium black to afford the compound of formula H-4. Finally, the Boc group is removed with trifluoroacetic acid to give the desired peptide of formula H-5 which is isolated as the bis trifluoroacetate salt.

In Chart H, Y is 2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane and X is 2S-amino-1-cycIohexyl-3R-hydroxy-4R,5R,6-tribenzyloxyhexane.

CHART I

Chart I describes the preparation of the novel peptide Glc-Pro-Tyr (OMe) His-Y of formula 1-4 (wherein Y is 2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane) or 2-Deoxy-D-glucopyranos-2-aminocarbonyl-L-prolyl-O-methyl-L-tyrosyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane of the present invention.

Coupling of the amine from the peptide of formula 1-1, prepared as the compound of formula G-2 in Chart G, with per acetylated glucosamine-Pro-OH gives the compound of formula 1-2. The protecting groups must be removed to yield the desired peptide. The tosyl group and the carbohydrate acetates are removed by treatment with methanol saturated with ammonia to give the compound of formula 1-3. Finally, the benzyl ethers are removed by hydrogenolysis with palladium black to give the desired compound of formula 1-4.

In Chart I, Y is Y is 2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane and X is 2S-amino-1-cyclohexyl-3R-hydroxy-4R,5R,6-tribenzyloxyhexane.

CHART J

Chart J describes the preparation of the novel peptides SPP-His-Y of formula J-4a,b (wherein Y is 2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane), also known as 2R-tert- Butylsulfonylmethyl-3-phenyl-propionyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane and 2S-tert-butylsulfonylmethyl-3-phenylpropionyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane of the present invention.

Coupling of the amine from the peptide of formula J-1, prepared as the compounds of formula B-4a of Chart B, with the epimeric mixture of tert-butylsulfonylmethyl-3-phenylpropionic acid (abbreviated as SPP) with DEPC affords the peptides of formula J-2a,b as a separable mixture. They are independently carried on to final products. The protecting groups must be removed to yield the desired peptides. The tosyl group is removed from the less polar peptide of formula J-2a by treatment with 1-hydroxybenzotriazole in methanol to afford the compound of formula J-3a. Finally, the benzyl ethers from the peptide of formula J-3a are removed by hydrogenolysis with palladium black to give the desired compound of formula J-4a.

Using the procedures for the preparation of J-4a, the more polar diastereomer of formula J-2b, is converted to the desired compound of formula J-4b, which is 2S-tert-butylsulfonylmethyl-3-phenylpropionyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane.

In Chart J, Y is 2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane and X is 2S-amino-1-cyclohexyl-3R-hydroxy-4R,5R,6-tribenzyloxyhexane. Using procedures analogous to those described in the charts above, the other peptides of the present invention may be prepared.

Generally, the renin inhibiting polypeptides may be prepared by solution phase peptide synthetic procedures analogous to those described hereinafter or to those methods known in the art. Appropriate protecting groups, reagents, and solvents for the solution phase method can be found in "The Peptides: Analysis, Synthesis, and Biology," Vols. 1-5, eds. E. Gross and T. Meienhofer, Academic Press, NY, 1979-1983; "The Practice of Peptide Synthesis", M. Bodansky and A. Bodansky, Springer-Verlag, New York, 1984; "The Principles of Peptide Synthesis", M. Bodansky, Springer-Verlag, New York, 1984. Thus, for example, the carboxylic moiety of N^α-t-butyloxycarbonyl (BOC)-substituted amino acid derivatives having suitable side chain protecting groups, if necessary, may be condensed with the amino functionality of a suitably protected amino acid or peptide using a conventional coupling protocol such as dicyclohexylcarbodiimide (DCC) and 1-hydroxybenzotriazole (HOBT) or diethylphosphoryl cyanide (DEPC) and triethylamine (Et₃N) in methylene chloride or dimethylformamide.

Following coupling reaction completion, the N^α-BOC moiety may be selectively removed with 50% trifluoroacetic acid with or without 2% anisole (v/v) in methylene chloride. Neutralization of the resultant trifluoroacetate salt may be accomplished with 10% diisopropylethylamine or sodium bicarbonate in methylene chloride.

Variations in the above description for starting materials, reactants, reaction conditions and required protecting groups to obtain other such N-alkylated compounds are known to an ordinarily skilled chemist or are readily available in the literature.

The compounds of the present invention may be in either free form or in protected form at one or more of the remaining (not previously protected) peptide, carboxyl, amino, hydroxy, or other reactive groups. The protecting groups may be any of those known in the polypeptide art. Examples of nitrogen and oxygen protection groups are set forth in T.W. Greene, Protecting Groups in Organic Synthesis, Wiley, New York, (1981); J.F.W. McOmie, ed. Protective Groups in Organic Chemistry, Plenum Press (1973); and J. Fuhrhop and G. Benzlin, Organic Synthesis, Verlag Chemie (1983). Included among the nitrogen protective groups are t-butoxycarbonyl (BOC), benzyloxycarbonyl, acetyl, allyl, phthalyl, benzyl, benzoyl, trityl and the like.

The following compounds of the present invention are preferred:

N-tert-Butyloxycarbonyl-L-phenylalanyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxy-hexane;

Boc-Tyr(OMe)-His-Y (wherein Y is 2S-amino-1-cyclohexyl-3R-4R,5R,6-tetrahydroxyhexane) or N-tert-Butyloxycarbonyl-O-memyl-L-tyrosyl-L-histidyl-2S-amino-1-cyclohexyl3R,4R,5R,6-tetrahydroxy-hexane;

SPP-His-Y (wherein Y is 2S-amino-1-cyclohexyl-3R-4R,5R,6-tetrahydroxyhexane) or 2S- tert-Butylsulfonylmethyl-3-phenyl-propionyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxy-hexane; and

N-tert-butyloxycarbonyl-L-phenylalanyl-L-histidyl-2S-amino-1-cyclohexyl-6-methyl-3R,4R,5S,6-tetrahydroxyheptane.

DESCRIPTION OF THE PREFERRED EMBODIMENTS

The following Preparations and Examples illustrate the present invention.

In the Preparations and Examples below and throughout this document:

¹H-NMR is nuclear magnetic resonance

BOC is t-butoxycarbonyl

BOPCL is bis (2-oxo-3-oxazolidinyl) phosphinic chloride

Bz is benzyl

C is centigrade

Cbz is benzyloxycarbonyl

CDCl₃ is deuteriochloroform

Celite is a filter aid

CVA is ChaΨ[CH(OH)CH₂]Val

DCC is dicyclohexylcarbodiimide

DEPC is diethylphosphoryl cyanide

EtOAc is ethyl acetate

g is grams

γ-Glu is γ-glutamic acid

His is histidine

N-MeHis is Nα-methyl histidine

HOBT is 1-hydroxybenzotriazole monohydrate

HPLC is high performance liquid chromatography

IR is infrared spectra

LVA is Leuø(CH(OH)CH₂)Val with the S configuration at C4 (the hydroxyl- bearing carbon atom)

M or mol is mole

Me is methyl

min is minute

mL is milliliter

MPLC is medium pressure liquid chromatography

MS is mass spectroscopy

Ph is phenyl

Phe is phenylalanine Pro is proline

RIP means a compound having the formula H-Pro-His-Phe-His-Phe-Phe-Val-Tyr- Lys-OH.2(CH₃C(O)OH).XH₂O which is a known renin-inhibiting peptide

SPP is the racemic acid, 2-t-butylsulfonylmedιyl-3-phenylpropionic acid

TEA is triethylamine

TFA is trifluoroacetic acid

THF is tetrahydrofuran

TLC is thin layer chromatography

Tos is p-toluenesulfonyl

TsoH is p-toluenesulfonic acid

Tyr is tyrosine

(OCH₃)Tyr is O-methyl tyrosine

β-Val is β-Valine.

The wedge-shape line indicates a bond which extends above the plane of the paper relative to the plane of the compound thereon.

The dotted line indicates a bond which extends below the plane of the paper relative to the plane of the compound thereon.

In the examples below, the renin inhibitory activity of the compounds of the present invention (IC₅₀'s) are determined using the in vitro test described in published European patent application 0 173 481, published 5 March 1986, pages 103-105, which are hereby incorporated by reference.

In the examples below, HPLC is high pressure liquid chromatography and k' is the partition ratio obtained. The solvent system used is indicated in parentheses after the partition ratio. HPLC is performed on a Perkin-Elmer Series IV liquid chromatograph operating at 1.5 mL/min through a Brownlee RP-18 10 micron column, with UV monitoring by a Kratos Spectroflow 773 detector. A Perkin-Elmer LCI-100 integrator is used for peak data. Solvent A is 90% by volume Burdick & Jackson water, 10% acetonitrile, and 0.1% trifluoroacetic acid; solvent B is 10% by volume Burdick & Jackson water, 90% acetonitrile, and 0.1 % trifluoroacetic acid.

Preparation 1 Methyll-[3-tert-Butyloxycarbonyl-4S-cyclohexyl-methyl-2,2-dimethyl-5R- oxazolidinylj-propenoate (Formula A-2) Refer to Chart A.

To a flame-dried flask under an argon atmosphere is charged 5.00 g of aldehyde of formula

A-1 in Chart A, 6.50 g of (carbethoxymethylene)-triphenylphosphorane and 150 mL of dry toluene. The solution is placed in an 80°C oil bath and heated for 20 hours. The suspension of a white solid in an orange solution is then cooled to room temperature and treated with diethyl ether. The mixture is filtered and the white solid washed with diethyl ether. The filtrate is concentrated under reduced pressure and the residue is triturated with diethyl ether, filtered and then concentrated. The trituration process is repeated and the resulting viscous orange oil is flash chromatographed on silica gel with 10% to 15% diethyl ether in hexane to afford 5.34 g of the title product as a clear, tan oil which solidifies upon standing.

Physical characteristics are as follows:

FAB HRMS: C₂₂H₃₈N₁O₅ (m+H)=396.2753.

Preparation 2 1-[3-tert-Butyloxycarbonyl-4S-cyclohexyl-methyl-2,2-dimethyl-5R- oxazolidinyl]-3-propenol (Formula A-3) Refer to Chart A.

To a stirring solution of 5.0 g of the title product of Preparation 1 in 25 mL of anhydrous diethyl ether under an argon atomsphere at -78°C is added 26 mL of diisobutylaluminum hydride in toluene (1.5 M) slowly via cannula. The first drop of the hydride solution causes a yellow color to appear which increases in intensity during the addition of the first equivalent. The color gradually decreased during the remainder of the addition period. After 1 hour, the solution is warmed to -60°C for 30 minutes and then recooled to -78°C. The reaction mixture is slowly treated with 5 mL of methanol and warmed to ambient temperature. The mixture is then treated with 80 mL of 1 M sodium potassium tartrate, stirred vigorously for 45 minutes, and then diluted with diethyl ether. The layers are separated and the aqueous layer is extracted with diethyl ether. The organic layers are combined, washed with brine, and then concentrated under reduced pressure. The residue is flash chromatographed on silica gel with 30% ethyl acetate in hexanes to afford 3.92 g of the title product as a pale tan oil which solidifies upon standing.

Physical characteristics are as follows:

FAB HRMS: C₂₀H₃₆N₁O₄ (m+H)=354.2628.

Preparation 3 1-[3-tert-ButyloxycarbonyI-4S-cyclohexyl-methyl-2,2-dimethyl-5R- oxazoliidinyl]-2,3-dihydroxy-1-propanol (Formula A-5a: X₁ and X₂ are each α-hydroxy; Formula A-5b: X₁ and X₂ are each β-hydroxy) Refer to

Chart A.

To a stirring solution of 1.40 g of the title product of Preparation 2 in 5.0 mL of acetone is added 5.0 mL of water dropwise. The cloudy mixture is placed under an argon atomsphere and then treated with 734 mg of N-methy norpholine N-oxide dihydrate. After 15 minutes, 0.40 mL (1 M in tert-butanol) of osmium tetroxide is added. The reaction mixture becomes black then gradually changes to dark green finally resulting in a light green suspension with a voluminous precipitant. The reaction mixture is vigorously stirred at ambient temperature overnight.

During the reaction time, the mixture becomes difficult to stir and is then diluted with dichloromethane. The reaction is treated with 10% aqueous sodium bisulfite, filtered through Celite, and the filter cake washed with dichloromethane-methanol. The layers are separated and the aqueous layer is repeatedly extracted with dichloromethane. The combined organic layers are dried (magnesium sulfate) and then concentrated under reduced pressure. The resulting white solid is adsorbed onto silica gel with methanol and the solvent is removed under high vacuum. The material is chromatographed on silica gel with 75% ethyl acetate in hexane to afford 1.09 g of the tide products.

Physical characteristics are as follows:

The proposed structure was supported by ¹H NMR

Preparation 4 1-Benzyloxy-[3-tert-butyloxycarbonyl-4S-cyclohexyl-methyl-2,2-dimedιyl- 5R-oxazolidinyl]-2,3-dibenzyloxypropane (Formula B-2a: Y₁ and Y₂ are each α-O-benzyl; Formula B-2b: Y₁ and Y₂ are each β-O-benzyl) Refer to Chart B.

To a flask containing 170 mg of the title products of Preparation 3 as a suspension in 2 mL of dry tetrahydrofuran at 0°C under an argon atomsphere is added 245 mg (35% by weight in oil) of washed, oil-free potassium hydride suspended in 1 mL of dry tetrahydrofuran. The initial light green suspension becomes more cloudy and deepened in color as the mixture is warmed to ambient temperature. After a short time at room temperature, 0.55 mL of benzyl bromide is added and the reaction mixture is left to stir overnight. The cloudy suspension is diluted with dichloromethane and partitioned with saturated aqueous sodium bicarbonate. The aqueous layer is extracted with dichloromethane and the combined organic layers are washed with brine, dried (magnesium sulfate), and then concentrated under reduced pressure. The residue is chromatographed on silica gel with 10% ethyl acetate in hexane to afford 284 mg of the title products as a pale oil.

Physical Characteristics are as follows:

FAB HRMS: C₄₁H₅₆NO₆(m+H)=658.4149.

Preparation 5 2S-amino-1-cyclohexyl-3R-hydroxy-4,5,6-tribenzyloxyhexane

(Formula B-3a: Y₁ and Y₂ are each α-O-benzyl; Formula B-3b: Y₁ and Y₂ are each β-O-benzyl) Refer to Chart B.

To a flask containing 4.0 mL of methanol at 0°C is slowly added 0.25 mL of acetyl chloride and tightly capped. After 15 minutes, the solution is warmed to ambient temperature. After an additional 15 minutes, the methanolic hydrogen chloride is added to flask containing 284 mg of the title products of Preparation 4. After 3 hours, the solution is treated with 0.85 g of solid sodium bicarbonate in portions to control gas evolution. The resulting suspension is stirred for 30 minutes and the volatiles are then removed with the aid of a stream of nitrogen gas. The resulting white residue is triturated with dichloromethane, filtered with dichloromethane washings, and then concentrated under reduced pressure. The resulting pale oil is used directly in the next reaction and no further purification is performed.

Physical characteristics are as follows:

The proposed structure was supported by ¹H NMR. Preparation 6 N-tert-Butyloxycarbonyl-N^im-tosyl-L-histidyl-2S-amino-1-cyclohexyl-3R- hydroxy-4,5,6-tribenzyloxyhexane (Formula B-4a: Y₁ and Y₂ are each α- O-benzyl; Formula B-4b: Y₁ and Y₂ are each β-O-benzyl) Refer to Chart

B.

To a stirred solution of 211 mg of the tide products of Preparation 5 and 220 mg of Boc- His(Ts)-OH in 3 ml of dry dichloromethane is added 120 uL of diisopropylethylamine, followed by 80 uL of DEPC. The solution is stirred overnight and then concentrated under reduced pressure. Chromatography of the residue on silica with 30% to 50% ethyl acetate in hexane provides 205 mg (0.225 mmol) of the less polar title product of formula B-4a as a glassy solid and 80 mg of the more polar title product of formula B-4b as a glassy solid.

Physical Characteristics for the title product of formula B-4a are as follows:

FAB HRMS: C₅₁H₆₅N₄O₉S (m+H)-=909.4454.

Physical Chacteristics for the title product of formula B-4b are as follows:

FAB HRMS: C₅₁H₆₅N₄O₉S (m+H)=909.4436.

Preparation 7 N-tert-Butyloxycarbonyl-L-phenylalanyl-N^im-tosyl-L-histidyl-2S-amino-1- cyclohexyl-3R-hydroxy-4,5,6-tribenzyloxyhexane (Formula B-5a: Y₁ and Y₂ are each α-O-benzyl) Refer to Chart B.

A solution of 205 mg of the less polar title product formula B-4a of Preparation 6 in 3 mL of 1:1 TFA-dichloromethane is allowed to stand for 1.5 hour at room temperature, men diluted with more dichloromethane and slowly added to excess stirred, saturated aqueous sodium bicarbonate. After 30 minutes, the aqueous layer is extracted with additional dichloromethane and the combined organic extracts are dried (magnesium sulfate) and are then concentrated under reduced pressure to give the free amine.

To a stirred solution of 178 mg of the amine and 80 mg of Boc-Phe-OH in 2.0 ml of dry dichloromethane is added 65 uL of diisopropylethylamine, followed by 45 uL of DEPC. The solution is stirred overnight at ambient temperature and then concentrated under reduced pressure.

Flash chromatography of the residue on silica gel with 50% ethyl acetate in hexane provides 210 mg of the title product.

Physical characteristics are as follows:

FAB HRMS: C₅₀H₇₄N₅O₁₀S (m+H)= 1056.519.

Preparation 8 N-tert-Butyloxycarbonyl-L-phenylalanyl-N^im-tosyl-L-histidyl-2S-amino-1- cyclohexyl-3R-hydroxy-4,5,6-tribenzyloxyhexane (Formula B-5b: Y₁ and

Y₂ are each β-O-benzyl) Refer to Chart B.

A solution of 80 mg of the more polar title product of formula B-4b of Preparation 6 in 1 mL of 1:1 TFA-dichloromethane is allowed to stand for 1.5 hour at room temperature, then diluted with more dichloromethane and slowly added to excess stirred, saturated aqueous sodium bicarbonate. After 30 minutes, the aqueous layer is extracted with additional dichloromethane and the combined organic extracts are dried (magnesium sulfate) and are then concentrated under reduced pressure to give the free amine.

To a stirred solution of 66 mg of the amine and 30 mg of Boc-Phe-OH in 0.8 ml of dry dichloromethane is added 25 uL of diisopropylethylamine, followed by 17 uL of DEPC. The solution is stirred overnight at ambient temperature and then concentrated under reduced pressure.

Chromatography of the residue on silica gel with 50% ethyl acetate in hexanes provides 76 mg of the title product.

Physical characteristics are as follows:

FAB HRMS: C₆₀H₇₄N₅O₁₀S (m+H)= 1056.

Preparation 9 N-tert-Butyloxycarbonyl-L-phenylalanyl-L-histidyl-2S-amino-1-cyclohexyl- 3R-hydroxy-4R,5R,6-tribenzyloxyhexane (Formula B-6a: Y₁ and Y₂ are each α-O-benzyl) Refer to Chart B

A solution of 210 mg of the title product of Preparation 7 and 127 mg of HOBT hydrate in 2.0 mL of methanol is allowed to stir at room temperature overnight. The reaction mixture is then concentrated under reduced pressure and the residue chromatographed on silica gel with 4% methanol saturated with ammonia in dichloromethane to afford 157 mg of the title product.

Physical characterics are as follows:

FAB HRMS: C₅₃H₆₈N₅O₈ (m+H)= 902.5061.

Preparation 10 N-tert-Butyloxycarbonyl-L-phenylalanyl-L-histidyl-2S-amino-1-cyclohexyl- 3R-hydroxy-4S,5S,6-tribenzyloxyhexane (Formula B-6b: Y₁ and Y₂ are each β-O-benzyl) Refer to Chart B.

A solution of 76 mg of the title product of Preparation 8 and 44 mg of HOBT hydrate in 0.7 mL of methanol is allowed to stir at room temperature overnight. The reaction mixture is then concentrated under reduced pressure and the residue chromatographed on silica gel with 4% methanol saturated with ammonia in dichloromethane to afford 64 mg of the title product.

Physical characteristics are as follows:

FAB HRMS: C₅₃H₆₈N₅O₈ (m+H)= 902.5044.

Example 1 N-tert-Butyloxycarbonyl-L-phenylalanyl-L-histidyl-2S-amino-1-cyclohexyl- 3R,4R,5R,6-tetrahydroxyhexane (Formula B-7a: X₁ and X₂ are each α- hydroxy) Refer to Chart B.

To a solution of 109 mg of the title product of Preparation 9 in 1 mL of acetic acid is added 200 mg of palladium black catalyst. The suspension is placed on a Parr apparatus at 50 psi of hydrogen gas and allowed to shake overnight. The reaction mixture is diluted with dichloromethane/methanol and then filtered through Celite with dichloromethane/methanol to pure methanol washings [Note: washing the catalyst under these conditions can cause an exothermic reaction to occur, therefore suitable precautions should be taken]. The filtrates are concentrated under reduced pressure and the residual acetic acid azeotroped with the aid of toluene. The resulting white solid is chromatographed on silica gel with 10% to 20% methanol saturated with ammonia in dichloromethane to afford 68 mg of the tide product. The product is lyophilized with the aid of methanol as a co-solvent to afford a white solid.

Physical characteristics are as follows:

FAB HRMS: C₃₂H₄₉N₅O₈ (m+H)=632

HPLC (60:40, A:B) (225 nm) k'=4.65.

IC₅₀ value of 6.8 nM.

Example 2 N-tert-Butyloxycarbonyl-L-phenylalanyl-L-histidyl-2S-amino-1-cyclohexyl- 3R,4S,5S,6-tertahydroxyhexane (Formula B-7b: X₁ and X₂ are each β- hydroxy) Refer to Chart B.

To a solution of 64 mg of the title product of Preparation 10 in 2 mL of acetic acid is added 200 mg of palladium black catalyst. The suspension is placed in a high pressure bomb at 1200 psi of hydrogen gas and allowed to stir overnight. The reaction mixture is diluted with dichloromethane and then filtered through Celite with dichloromethane/methanol to pure methanol washings. The filtrates are concentrated under reduced pressure and the residual acetic acid azeotroped with the aid of toluene. The resulting white solid is chromatographed on silica gel with

10% to 20% methanol saturated with ammonia in dichloromethane to afford 9 mg of the title product. The product is lyophilized with the aid of methanol as a co-solvent to afford white, fluffy solid.

Physical characteristics are as follows:

FAB HRMS: C₃₂H₄₉N₅O₈ (m+H) =632.

HPLC (60:40, A:B) (225 nm) k'=4.94

9% renin inhibition at 10^-8 M.

Preparation 11 1-[3-tert-ButyIoxycarbonyl-4S-cyclohexyl-methyl-2,2-dimethyl-5R- oxazolidinyl]-1,2-epoxy-3-hydroxypropane (Formula A-4a: α-epoxide;

Formula A-4b: β-epoxide). Refer to Chart A.

To a stirring solution of 1.21 g of the title product of Preparation 2 in dry dichloromethane is added 0.71 g (70%) of m-chloroperbenzoic acid and 0.50 g of solid sodium bicarbonate as a buffer. The reaction mixture is vigorously stirred under an argon atomsphere for 15 hours. The mixture is then treated with 1.7 g of solid sodium sulfite and the suspension left to stir at ambient temperature. After 1 hour, the reaction mixture is filtered and the filtrate is concentrated under reduced pressure. The resulting white foam is flash chromatographed on silica gel with 25% to 30% ethyl acetate in hexane to afford 1.26 g of the title products as a clear, colorless oil which solidifies upon standing. Physical characteristics are as follows:

FAB HRMS: C₂₀H₃₆NO₅ (m+H)= 370.2571.

Preparation 12 1-[3-tert-Butyloxycarbonyl-4S-cyclohexyl-methyl-2,2-dimethyl-5R oxazolidinyl]-2,3-dihydroxy-1-propanol (Formula A-6a: X₁ is α-hydroxy and X₂ is β-hydroxy; Formula A-6b:X₁ is β-hydroxy and X₂ is α- hydroxy) Refer to Chart A.

To a stirring solution of 1.25 g of the tide products of Preparation 11 in 10 mL of dioxane is added 3.5 mL (40% in water) of tetrabutylammonium hydroxide. The reaction mixture immediately begins to turn yellow. The flask is equipped with a reflux condenser and then placed in an 110°C oil bath. After 38 hours, the pale brown solution is cooled to ambient temperature, diluted with dichloromethane and then partitioned with pH=7 phosphate buffer. The aqueous layer is extracted with dicholoromethane and the organic layers are combined, washed with brine, dried (magnesium sulfate) and then concentrated under reduced pressure. The residue is flash chromatographed with 30% ethyl acetate in hexane to afford 546 mg of the title products as a pale yellow solid.

Physical characteristics are as follows:

FAB HRMS: C₂₀H₃₈NO₆ (m+H)=388.2698.

Preparation 13 1-Benzyloxy-[3-tert-butyloxycarbonyl-4S-cyclohexyl-methyl-2,2-dimedιyl- 5R-oxazolidinyl]-2,3-dibenzyloxy-propane (Formula C-2a: Y₁ is α-O- benzyl and Y₂ is β-O-benzyl; Formula C-2b: Y₁ is β-O-benzyl and Y₂ is α-O-benzyl) Refer to Chart C.

To a stirring suspension of 485 mg of the title products of Preparation 12 in 1.25 mL of dry tetrahydrofuran is added 300 mg of sodium hydride (50% by weight in oil) as a suspension in tetrahydrofuran (2 mL and 3 x 1 mL rinses). After the initial evolution of gas, the suspension is left to stir under an argon atomsphere. After 10 minutes, 0.75 mL of benzyl bromide is added and the mixture left to stir at ambient temperature. After 2.5 days, the reaction mixture is diluted with dichloromethane and pardoned against saturated aqueous sodium bicarbonate. The aqueous layer is extracted with dichloromethane and the combined organic layers are washed with brine, dried (magnesium sulfate) and then concentrated under reduced pressure. The residue is flash chromatographed on silica gel with 10% to 50% ethyl acetate in hexane to afford 530 mg of the title products as a crystalline solid.

Physical characteristics are as follows:

FAB HRMS: C₄₁H₅₆NO₆ (m+H)=658.4118.

Preparation 14 2S-amino-1-cyclohexyl-3R-hydroxy-4,5,6-tribenzyloxyhexane(FormulaC- 3a: Y-, is α-O-benzyl and Y₂ is β-O-benzyl; Formula C-3b: Y₁ is β-O- benzyl and Y₂ is α-O-benzyl) Refer to Chart C. To a flask containing 8.0 mL of methanol at 0°C is slowly added 0.51 mL of acetyl chloride and is then tightly capped. After 15 minutes, the solution is warmed to ambient temperature. After an additional 15 minutes, the methanolic hydrogen chloride solution is added to a flask containing 0.53 g of the title products of Prepartion 13. After 3 hours, the solution is treated with 0.90 g of solid sodium bicarbonate in portions to control gas evolution. The resulting suspension is stirred for 30 minutes and the volatiles are removed with the aid of a stream of nitrogen gas. The white residue is triturated with dichloromethane, filtered with dichloromethane washings, and then concentrated under reduced pressure. The resulting pale foam is used direcdy in the next reaction and no further purification is performed.

Physical characteristics are as follows:

The proposed structure was supported by ¹H NMR.

Preparation 15 N-tert-Butyloxycarbonyl-N^im-tosyl-L-histidyl-2S-amino-1-cyclohexyl-3R- hydroxy-4,5,6-tribenzyloxyhexane (Formula C-4a: Y₁ is α-O-benzyl and Y₂ is β-O-benzyl; Formula C-4b: Y₁ is S-O-benzyl and Y₂ is α-O-benzyl) Refer to Chart C.

To a stirred solution of 160 mg of the title products of Preparation 14 and 170 mg of Boc-His(Ts)-OH in 3 ml of dry dichloromethane is added 90 uL of diisopropylethylamine, followed by 65 uL of DEPC. The solution is stirred overnight and then concentrated under reduced pressure. Gravity chromatography of the residue on silica gel with 30% to 50% ethyl acetate in hexane provides 205 mg of the title products as a pale viscous oil.

Physical Characteristics are as follows:

FAB HRMS: C₅₁H₆₅N₄O₉S (m+H)= 909.4498.

Preparation 16 N-tert-Butyloxycarbonyl-L-phenylalanyl-N^im-tosyl-L-histidyl-2S-amino-1- cyclohexyl-3R-hydroxy-4,5,6-tribenzyloxyhexane (Formula C-5a: Y₁ is α- O-benzyl and Y₂ is β-O-benzyl; Formula C-5b: Y- is β-O-benzyl and Y₂ is α-O-benzyl) Refer to Chart C.

A solution of 205 mg of the title products of Preparation 15 in 3 mL of 1:1 TFA-dichloromethane is allowed to stir for one hour at room temperature, then diluted with more dichloromethane and slowly added to excess stirred, saturated aqueous sodium bicarbonate. After 30 minutes, the aqueous layer is extracted with add.tional dichloromethane and the combined organic extracts are dried (magnesium sulfate) and are then concentrated under reduced pressure to give the free amines.

To a stirred solution of 140 mg of the amines and 65 mg of Boc-Phe-OH in 1.5 ml of dry dichloromethane is added 55 uL of diisopropylethylamine, followed by 37 uL of DEPC. The solution is stirred for 3 days and was then concentrated under reduced pressure. Chromatography of the residue on silica gel with 30% to 50% ethyl acetate in hexane provides 157 mg of the title products.

Physical characteristics are as follows:

FAB HRMS: C₆₀H₇₄N₅O₁₀S (m+H)= 1056.514.

Preparation 17 N-terr-Butyloxycarbonyl-L-phenylalanyl-N^imtosyl-L-histidyl-2S-amino-1- cyclohexyl-3R-hydroxy-4,5,6-tribenzyloxyhexane (Formula C-6a: Y₁ is α-O-benzyl and Y₂ is β-O-benzyl; Formula C-6b: Y₁ is β-O-benzyl and

Y₂ is α-O-benzyl) Refer to Chart C.

A solution of 157 mg of the title products of Preparation 16 and 90 mg of HOBT hydrate in 1.25 mL of methanol is allowed to stir at room temperature. After 15 hours, the reaction mixture is concentrated under reduced pressure and the residue chromatographed on silica gel with

3% to 4% methanol saturated with ammonia in dichloromethane to afford 127 mg of the title products.

Physical characteristics are as follows:

FAB HRMS: C₅₃H₆₈N₅O₈ (m+H)=902.5106.

Example 3 N-tert-Butyloxycarbonyl-L-phenylalanyl-L-histidyl-2S-amino-1-cyclohexyl- 3R,4R,5S,6-tetrahydroxyhexane (Formula C-7a: X₁ is α-hydroxy and X₂ is β-hydroxy) and N-tert-butyloxycarbonyl-L-phenylalanyl-L-histidyl-2S- amino-1-cyclohexyl-3R,4S,5R,6-tetrahydroxyhexane (Formula C-7b: X₁ is β-hydroxy and X₂ is α-hydroxy). Refer to Chart C.

To a solution of 45 mg of the title products of Preparation 17 in a small amount of acetic acid is added 105 mg of palladium black catalyst. The suspension is placed in a high pressure bomb at 1250 psi of hydrogen gas and allowed to stir overnight. The reaction mixture is removed from the bomb and diluted with dichloromethane/methanol. The mixture is filtered through Celite with dichloromethane/methanol to pure methanol washings. The filtrates are concentrated under reduced pressure and the residual acetic acid azeotroped with the aid of toluene. The resulting white solid is chromatographed on silica gel with 5% to 20% methanol saturated with ammonia in dichloromethane to afford 6 mg of pure less polar isomer title product of formula C-7a, 6 mg of more polar title product of formula C-7b and 7 mg of mixed products. The products are lyophilized with the aid of methanol as a co-solvent to afford white, fluffy solids.

Physical characteristics of the title product of formula C-7a are as follows:

FAB HRMS: C₃₂H₅₀N₅O₈ (m+H)= 632.3690.

HPLC (60:40, A:B) (225 nm) k'=6.06

13% renin inhibition at 10^-8 M.

Physical Characteristics of the title product of formula C-7b are as follows:

FAB HRMS: C₃₂H₅₀N₅O₈ (m+H)= 632. HPLC (60:40, A:B) (225 nm) k'=4.41

24% renin inhibition at 10^-8 M.

Preparation 18 1R-[3-rert-Butyloxycarbonyl-4S-cyclohexyl-methyl-2,2-dimethyl-5R- oxazolidinyl]-3-medιyl-2,3-dihydroxy-1-butanol (Formula D-2) Refer to Chart D.

To a stirring solution of 495 mg of allylic alcohol of formula D-l in 1.5 mL of acetone is added 1.5 mL of water dropwise. The cloudy mixture is placed under an argon atmosphere and then treated with 291 mg of N-methylmorpholine N-oxide dihydrate. After 15 minutes, 0.15 mL (1 M in re/τ-butanol) of osmium tetroxide is added. The reaction mixture becomes black then gradually changes to dark green finally resulting in a light green suspension. The reaction mixture is vigorously stirred at ambient temperature overnight. During the reaction time, the mixture becomes difficult to stir and is then diluted with dichloromethane. The reaction is treated with solid sodium bisulfite and allowed to stir for 15 minutes. The layers are separated ana the aqueous layer is repeatedly extracted with dichloromethane. The combined organic layers are dried (magnesium sulfate) and then concentrated under reduced pressure. The resulting tan solid is flash chromatographed on silica gel with 50% ethyl acetate in hexane to afford 380 mg of the title products.

Physical Characteristics are as follows:

The proposed structure was supported by ¹H NMR.

Preparation 19 1R-[3-tert-Butyloxycarbonyl-4S-cyclohexyl-methyl-2,2-dimethyl-5R- oxazolidinyl]-3-medιyl-1-2,3-tribenzyloxybutane (Formula D-3) Refer to

Chart D.

To a flask containing 124 mg of the tide products of Preparation 18 as a suspension in 0.5 mL of dry tetrahydrofuran at 0°C under an argon atmosphere is added 174 mg (35% by weight in oil) of washed, oil-fre potassium hydride suspended in 1 mL of dry tetrahydrofuran with 2 × 0.25 mL rinses. The initial light green suspension becomes gel-like and unstirrable. The paste is diluted with an additional 2 mL of dry terahydrofuran. The resulting slurry is treated with 0.7 mL of benzyl bromide and the reaction mixture is left to stir and warm to room temperature overnight.

The pale, cloudy suspension is diluted with dichloromethane and slowly treated with 1 mL of methanol to contn. as evolution. After a short time the reaction mixture is partitioned against half-saturated aqueous sodium bicarbonate. The aqueous layer is extracted with dichloromethane and the combined organic layers are washed with brine, dried (magnesium sulfate), and then concentrated under reduced pressure. The residue is flash chromatographed on silica gel with 5% to 10% ethyl acetate in hexane to afford 227 mg of the title product as a pale oil.

Physical characteristics are as follows:

FAB HRMS: C₄₃H₅₉NO₆ (m+H)=686. Preparation 20 2S-amino-1-cyclohexyl-3R-hydroxy-6-medιyMR,5,6-tribenzyloxyheptane (Formula D-4) Refer to Chart D.

To a flask containing 3.0 mL of methanol at 0°C is slowly added 0.20 mL of acetyl chloride and tighdy capped. After 15 minutes, the solution is warmed to ambient temperature. After an additional 15 minutes, the methanolic hydrogen chloride is added to a flask containing 227 mg of the title product of Preparation 19. After 3 hours, the solution is treated with solid sodium bicarbonate in portions to control gas evolution. The resulting suspension is stirred for 30 minutes and the volatiles are then removed with the aid of a stream of nitrogen gas. The resulting white residue is triturated with dichloromethane, filtered with dichloromethane washings, and then concentrated under reduced pressure. The resulting white solid is used directly in the next reaction and no further purification is performed.

Physical characteristics are as follows:

The proposed structure was supported by ¹H NMR.

Preparation 21 N-tert-Butyloxycarbonyl-N^im-tosyl-L-histidyl- 2S-amino-1-cyclohexyl-SR- hydroxy-6-medιyl-4R,5,6-tribenzyloxyheptane (Formula E-2) Refer to Chart E.

To a stirred solution of 168 mg of the title product of Preparation 20 and 171 mg of Boc- His(Ts)-OH in 3 ml of dry dichloromethane is added 93 uL of diisopropylethylamine, followed by 63 uL (0.42 mmol) of DEPC. The solution is stirred for 2 days and then concentrated under reduced pressure. Chromatography of the residue on silica with 10% to 20% ethyl acetate in dichloromethane provides 138 mg of the title product as a pale foam.

Physical characteristics are as follows:

FAB HRMS: C₅₃H₆₈N₄O ₉S (m+H) =937.4781.

Preparation 22 N-tert-Butyloxycarbonyl-L-phenylalanyl-N^im-tosyl-L-histidyl-2S-amino-1- cyclohexyl-3R-hydroxy-6-methyl-4R,5S,6-tribenzyloxyheptane (Formula

E-3) Refer to Chart E.

A solution of 138 mg of the title product of Preperation 21 in 1.5 mL of 1:1 TFA-dichloromethane is allowed to stir for 1 hour at room temperature, then diluted with more dichloromethane and slowly added to excess stirred, saturated aqueous sodium bicarbonate. After 30 minutes, the aqueous layer is extracted with additional dichloromethane and the combined organic extracts are dried (magnesium sulfate) and are then concentrated under reduced pressure to give the free amine. To a stirred solution of 121 mg of the amine and 55 mg of Boc-Phe-OH in 1.5 ml of dry dichloromethane is added 46 uL of diisopropylethylamine, followed by 31 uL of DEPC. The solution is stirred overnight at ambient temperature and then concentrated under reduced pressure. Chromatography of the residue on silica gel with ethyl acetate in dichloromethane provides 74 mg of the title product as the major isomer. Physical Characteristics are as follows:

FAB HRMS: C₆₂H₇₇N₅O ₁₀S (m+H) = 1084.547.

Preparation 23 N-tert-Butyloxycarbonyl-L-phenylalanyl-L-histidyl-2S-amino-1 -cyclohexyl- 3R-hydroxy-6-medιyl-4R,5S,6-tribenzyloxyhexane (Formula E-4) Refer to Chart E.

A solution of 74 mg of the title product of Preparation 22 and 42 mg of HOBT hydrate in 1.0 mL of methanol is allowed to stir at room temperature overnight. The reaction mixture is then concentrated under reduced pressure and the residue gravity chromatographed on silica gel with

2% to 6% methanol saturated with ammonia in dichloromethane to afford 57 mg of the tide product.

Physical Characteristics are as follows:

FAB HRMS: C₅₅H₇₁N₅O₈ (m+H)=930.5371.

Example 4 N-tert-Butyloxycarbonyl-L-phenylalanyl-L-histidyl-2S-amino-1-cyclohexyl- 6-methyl-3R,4R,5S,6-tetrahydroxyheptane (Formula E-5) Refer to Chart E.

To a solution of 57 mg of the title product of Preparation 23 in a small amount of acetic acid is added 100 mg of palladium black catalyst. The suspension is placed on a Parr apparatus at 50 psi of hydrogen gas and allowed to shake overnight. The reaction mixture is diluted with dichloromethane/methanol and then filtered through Celite with dichloromethane/methanol to pure methanol washings [Note: washing the catalyst under these conditions can cause an exothermic reaction to occur, therefore suitable precautions should be taken]. The filtrates are concentrated under reduced pressure and the residual acetic acid azeotroped with the aid of toluene. The resulting white solid is chromatographed on silica gel with 10% to 30% methanol saturated with ammonia in dichloromethane to afford 37 mg of the title product as a white solid.

Physical Characteristics are as follows:

FAB HRMS: C₃₄H₅₃N₅O₈ (m+H)=660.3°61.

HPLC (60:40, A:B) (225 nm) k'=6.25

IC₅₀ Value of 0.7 nM.

Preparation 24 Boc-βVal-Phe-His-Y (Formula F-2: Y is 2S-amino-1-cyclohexyl-3R- 4R,5R,6-tetrahydroxyhexane Refer to Chart F.

To a flask containing 0.7 mL of methanol at 0°C is slowly added 0.041 mL of acetyl chloride and then tightly capped. After 15 minutes, the solution is warmed to room temperature and stirred for an additional 15 minutes. The methanolic hydrogen chloride is then added to a vial containing 42 mg of the title product of Example 1 and the acidic flask is rinsed with 0.3 mL of additional methanol before addition to the compound vial. The peptide is slow to dissolve and a cloudy mixture results. After 2 hours, the reaction mixture is diluted with dichloromethane and thin layer chromatography suggests a significant amount of unreacted starting material is stil present. The mixture is concentrated by treatment with a stream of nitrogen gas to remove the solvents.

The resulting residue is treated with 1 mL of a solution of 1 : 1 trifluoroacetic acid-dichloromethane and allowed to stir at room temperature for 1.25 hours. The mixture is then slowly dripped into 60 mL of a stirring solution of 1:2 diethyl ether-hexane. The resulting cloudy suspension is stirred for 0.5 hours, divided into two centrifuge tubes, spun down, decanted, and finally dried. The resulting tan solid is used directly in the next reaction and no further purification is performed. The structure was supported by ¹H-NMR.

To a stirred solution of 45 mg of H-Phe-His-Y and 14 mg of Boc-βVal-OH in 0.5 ml of dimethylformamide is added 36 μL of diisopropylethylamine, followed by 10 uL of DEPC. The reaction mixture is stirred overnight and then chromatographed on silica gel with 2% to 20% methanol saturated with ammonia in dichloromethane to afford 14 mg of the title product as a white solid.

Physical characteristics are as follows:

FAB HRMS: C₃₇H₅₈N₆O₉(m+H)=731.4318.

Example 5 βVal-Phe-His-Y (Formula F-3: Y is 2S-amino-1-cyclohexyI-3R-4R,5R,6- tetrahydroxyhexane) or j8-Valyl-L-phenylalanyl-L-histidyl-2S-amino-1- cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane,bis trifluoroacetate. Refer to Chart F.

A solution of 15 mg of the title product of Preparation 24 in 1.0 mL of 1:1 TFA-dichloromethane is allowed to stir for 1 hour and is then dripped slowly into 50 mL of a rapidly stirred 1:2 ether-hexane solution. The precipitated solid is spun down, washed twice with 1:2 ether-hexane, and dried under air followed by both house and high vacuum to afford 13 mg of the title product as a white solid.

Physical characteristics are as follows:

FAB HRMS: C₃₄H₅₆N₇O₅ (Sm + H) = 631.3810.

HPLC (10/90 A/B; 225 nm) k'=6.33.

21% renin inhibition at 10^-8 M.

Preparation 25 Boc-Tyr(OMe)-His(Ts)-X (Formula G-2: X is2S-amino-1-cyclohexyl-3R- hydroxy-4R,5R,6-tribenzyloxyhexane) Refer to Chart G.

To a flask containing of 810 mg of the title products B-4a of Preparation 6 is added 10 mL of a solution of 1 : 1 trifluoroactice acid-dichloromethane, tightly capped and allowed to stir at room temperature. After 1.5 hours, the reaction mixture is diluted with dichloromethane and then added slowly to 120 mL of a saturated aqueous solution of sodium bicarbonate. The resulting mixture is stirred for 15 minutes, separated and the aqueous layer extracted with several portions of dichloromethane. The combined organic phase is dried (magnesium sulfate) and then concentrated to give the free amine.

To a stirred solution of 245 mg of H-His(Ts)-X and 149 mg of Boc-Tyr(OMe)-OH dicyclohexylamine salt in 3 ml of dichloromethane is added 75 uL of diisopropylethylamine, followed by 48 uL of DEPC. The reaction mixture is stirred overnight and then chromatographed on silica gel with 50% ethyl acetate in hexanes to afford 233 mg of the title as a pale foam.

Physical characteristics are as follows:

FAB-HRMS: C₆₁H₇₅N₅O₁₁S (m+H)= 1086.529.

Preparation 26 Boc-Tyr(OMe)-His-X (Formula G-3: X is 2S-amino-1-cyclohexyl-3R- hydroxy-4R,5R,6-tribenzyloxyhexane) Refer to Chart G.

A solution of 112 mg of the title product of Preparation 25 and 69 mg of HOBT hydrate in 1 mL of methanol is allowed to stir overnight. The solution is then concentrated under a stream of nitrogen gas and the resulting residue is gravity chromatographed on silica with 4% to 5% methanol (saturated with ammonia) in dichloromethane. The purification affords 89 mg of the title product as a glassy solid.

Physical characteristics are as follows:

FAB-HRMS: C₅₄H₆₉N₅O₉ (m+H)=932.5157.

Example 6 Boc-Tyr(OMe)-His-Y (Formula G-4: Y is 2S-amino-1-cyclohexyl-3R- 4R,5R,6-tetrahydroxyhexane) or N-tert-Butyloxycarbonyl-O-methyl-L- tyrosyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane.

Refer to Chart G.

To a solution of 89 mg of the title product of Preparation 26 in a small amount of acetic acid is added 180 mg of palladium black catalyst. The suspension is placed on a Parr apparatus at 50 psi of hydrogen gas and allowed to shake overnight. The reaction mixture is diluted with dichloromethane/methanol and then filtered through Celite with dichloromethane/methanol to pure methanol washings [Note: washing the catalyst under these conditions can cause an exothermic reaction to occur, therefore suitable precautions should be taken]. The filtrates are concentrated under reduced pressure and the residual acetic acid azeotroped with the aid of toluene. The resulting white solid is chromatographed on silica gel with 10% to 20% methanol saturated with ammonia in dichloromethane to afford 60 mg of the title product as a white solid. The solid is lyophilized with the aid of methanol to a fluffy white compound.

Physical characteristics are as follows:

FAB HRMS: C₃₃H₅₁N₅O₉ (m+H)=662.3771.

IC₅₀ value of 5.3 nM.

HPLC (60:40, A:B) (225 nm) k'=3.87 Preparation 27 Boc-βVal-Tyr(OMe)-His(Ts)-X (Formula H-2: X is 2S-amino-1- cyclohexyl-3R-hydroxy-4R,5R,6-tribenzyloxyhexane) Refer to Chart H. To a flask containing of 333 mg of the tide product of Preparation 25 is added 3 mL of a solution of 1:1 trifluoroacetic acid-dichloromethane, then tightly capped and allowed to stir at room temperature. After 1 hour, the reaction mixture is diluted with dichloromethane and then added slowly to a saturated aqueous solution of sodium bicarbonate. The resulting mixture is stirred for 30 minutes, separated and the aqueous layer extracted with several portions of dichloromethane. The combined organic phase is dried (magnesium sulfate) and then concentrated to give the free amine.

To a stirred solution of 121 mg of H-Tyr(OMe)-His(Ts)-X and 32 mg of Boc-βVal-OH in 1 ml of dichloromethane is added 33 uL of diisopropylethylamine, followed by 23 uL of DEPC. The reaction mixture is stirred overnight, then chromatographed on silica with 5% methanol in dichloromethane to afford 140 mg of the title product as a pale foam.

Physical characteristics are as follows:

FAB-HRMS: C₆₆H₈₄N₆O₁₂S (m+H)= 1185.

Preparation 28 Boc-βVal-Tyr(OMe)-His-X (Formula H-3: X is 2S-amino-1-cyclohexyl- 3R-hydroxy-4R,5R,6-tribenzyloxyhexane) Refer to Chart H.

A solution of 140 mg of the title product of Preparation 27 and 75 mg of HOBT hydrate in 1 mL of methanol is allowed to stir overnight and is then concentrated under reduced pressure. Chromatography of the residue on silica with 5% methanol (saturated with ammonia) in dichloromethane affords 125 mg of the title product as a white foam.

Physical characteristics are as follows:

FAB-HRMS: C₅₉H₇₈N₆O₁₀ (m+H)= 1031.582.

Prepartion 29 Boc-βVal-Tyr(OMe)-His-Y (Formula H-4: Y is 2S-amino-1-cyclohexyl- 3R-4S,5S,6-tetrahydroxyhexane). Refer to Chart H.

To a solution of 125 mg of the title product of Preparation 28 in a small amount of acetic acid is added 375 mg of palladium black catalyst. The suspension is placed on a high pressure bomb under 1250 psi of hydrogen gas and allowed to stir overnight. The reaction mixture is diluted with dichloromethane/methanol and then filtered through Celite with dichloromethane/methanol to pure methanol washings. The filtrates are concentrated under reduced pressure and the residual acetic acid azeotroped with the aid of toluene. The resulting white solid is chromatographed on silica gel with 10% to 20% methanol saturated with ammonia in dichloromethane to afford 64 mg of the title product as a white solid.

Physical characteristics are as follows:

FAB HRMS: C₃₈H₆₀N₆O₁₀ (m+H) =761.4442. Example 7 βVal-Tyr(OMe)-His-Y (Formula H-5: Y is 2S-amino-1-cyclohexyl-3R- 4R,5R,6-tetrahydroxyhexane)orβ-Valyl-O-methyl-L-tyrosyl-L-histidyl-2S- amino-1-cyclohexyI-3R,4R,5R,6-tetrahydroxyhexane, bis trifluoroacetate. Refer to Chart H.

A solution of 64 mg of the title product of Preparation 29 in 1 mL of 1:1 TFA-dichloromethane is allowed to stir for 1.5 hours and is then dripped slowly into 40 ml of rapidly stirred 1:2 diethyl ether-hexane. The solution begins to turn cloudy and the white suspension is allowed to form over 0.5 h. The precipitated solid is spun down, washed twice with 1:2 etherhexane, and carefully dried under both house and high vacuum to afford 67 mg of the title product as a white solid.

Physical characteristics are as follows:

FAB HRMS: C₃₃H₅₂N₆O₈ (m+H) =661.3942.

HPLC (10/90 A/B; 225 nm) k'= 7.56.

21% renin inhibition at 10^-8 M.

Preparation 30 Glc(per Ac)-Pro-Tyr(OMe)-His(Ts)-X (Formula 1-2: X is 2S-amino-1- cyclohexyl-3R-hydroxy-4R,5R,6-tribenzyloxyhexane ethers). Refer to

Chart I.

To a stirred solution of 161 mg of the deprotected title product of Preparation 25 and 108 mg of Glc(per Ac)-Pro-OH in 1.5 ml of dichloromethane is added 48 uL of diisopropylethylamine, followed by 34 uL of DEPC. The reaction mixture is stirred overnight and then chromatographed on silica with 5% methanol in dichloromethane to afford 240 mg of the title product as a white foam.

Physical characteristics are as follows:

FAB-HRMS: C₇₆H₉₃N₇O₂₀S (m+H)= 1456.

Preparation 31 Glc-Pro-Tyr(OMe)-His-X (Formula 1-3: X is 2S-amino-1-cyclohexyl-3R- hydroxy-4R,5R,6-tribenzyloxyhexane) Refer to Chart I.

A solution of 240 mg of the title product of Preparation 30 in 2 mL of methanol saturated with ammonia is allowed to stir overnight and is then concentrated under reduced pressure. Chromatography of the residue on silica with 10% to 20% methanol (saturated with ammonia) in dichloromethane affords 101 mg of the title proc ;t as a white solid.

Physical characteristics are as follows:

The proposed structure was supported by ^-NMR

Example 8 Glc-Pro-Tyr(OMe)-His-Y (Formula 1-4: Y is 2S-amino-1-cyclohexyl-3R- 4R,5R,6-tetrahydroxyhexane) or 2-Deoxy-D-glucopyranos-2-aminocarbonyl-L-prolyl-O-medιyl-L-tyrosyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,- 5R,6-tetrahydroxyhexane. Refer to Chart I. To a solution of 101 mg of the title product of Preparation 31 in a small amount of acetic acid is added 200 mg of palladium black catalyst. The suspension is placed on a Parr apparatus at 50 psi of hydrogen gas and allowed to shake for 18 hours. The reaction mixture is diluted with dichloromethane/methanol and then filtered through Celite with dichloromethane/methanol to pure methanol washings. The filtrates are concentrated under reduced pressure and the residual acetic acid azeotroped with the aid of toluene. The resulting white solid is gravity chromatographed on silica gel with 20% to 50% methanol saturated with ammonia in dichloromethane to afford 32 mg of the title product as a pale solid.

Physical characteristics are as follows:

FAB HRMS: C₄₀H₆₁N₇O₁₄ (m+H) =864.4399.

HPLC (10:90, A:B) (225 nm) k' = 8.94.

21% renin inhibition at 10^-8 M.

Preparation 32 SPP-His(Ts)-X (Formulas J-2a,b: X is 2S-amino-1-cyclohexyl-3R- hydroxy-4R,5R,6-tribenzyloxyhexane) Refer to Chart J.

To a stirred solution of 97 mg of the deprotected title products of Preparation 6 and 49 mg of 2-tert-butylsulfonylmedιyl-3-phenyl-propionic acid in 1 ml of dichloromethane is added 38 uL of diisopropylethylamine, followed by 24 uL of DEPC. The reaction mixture is stirred overnight and then chromatographed on silica with 20% to 40% ethyl acetate in dichloromethane to afford 69 mg of the title product of formula J-2a as a pale oil and 53 mg of the title product of formula J-2b also as a pale oil.

Physical characteristics for the compound of formula J-2a are as follows:

FAB-HRMS: C₆₀H₇₄N₄O₁₀S₂(m + H)= 1075.484

Physical characteristics for the compound of formula J-2b are as follows:

FAB-HRMS: C₆₀H₇₄N₄O₁₀S₂(m+H)= 1075.489

Preparation 33 SPP-His-X (Formula J-3a: X is 2S-amino-1-cyclohexyl-3R-hydroxy- 4R,5R,6-tribenzyloxyhexane) Refer to Chart J.

A solution of 69 mg of the title product J-2a of Preparation 32 and 39 mg of HOBT hydrate in 0.65 mL of methanol is allowed to stir overnight and is then concentrated under reduced pressure. Chromatography of the residue on silica with 2% to 6% methanol (saturated with ammonia) in dichloromethane affords 45 mg of the title product as a pale glassy solid.

Physical characteristics are as follows:

FAB-HRMS: C₅₃H₆₈N₄O₈S (m+H)=921.4823

Example 9 SPP-His-Y (Formula J-4a: Y is 2S-amino-1-cyclohexyl-3R-4R,5R,6- tetrahydroxyhexane)or2R-tert-butylsulfonylmethyl-3-phenyl-propionyl-L- histidyl-2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane. Refer to Chart J. To a solution of 45 mg of the title product of Preparation 33 in a small amount of acetic acid is added 44 mg of palladium black catalyst. The suspension is placed on a Parr apparatus at 45 psi of hydrogen gas and allowed to shake for 3 days. The reaction mixture is diluted with dichloromethane/methanol and then filtered through Celite with dichloromethane/methanol to pure methanol washings. The filtrates are concentrated under reduced pressure and the residual acetic acid azeotroped with the aid of toluene. The resulting white solid is chromatographed on silica gel with 10% to 20% methanol saturated with ammonia in dichloromethane to afford 22 mg of the title product as a white solid.

Physical characteristics are as follows:

FAB HRMS: C₃₂H₅₀N₄O₈S (m+H)=651.3465.

34% renin inhibition at 10^-7 M.

Preparation 34 SPP-His-X (Formula J-3b: X is 2S-amino-1-cyclohexyl-3R-hydroxy- 4R,5R,6-tribenzyloxyhexane). Refer to Chart J.

A solution of 53 mg of the title product of formula J-2b of Preparation 32 and 30 mg (0.196 mmol) of HOBT hydrate in 0.5 mL of methanol is allowed to stir overnight and is then concentrated under reduced pressure. Chromatography of the residue on silica with 2% to 6% methanol (saturated with ammonia) in dichloromethane affords 28 mg of the title product as a white glassy solid.

Physical characteristics are as follows:

FAB-HRMS: C₅₃H₆₈N₄O₈S (m+H) =921.4823.

Example 10 SPP-His-Y (Formula J-4b: Y is 2S-amino-1-cyclohexyl-3R-4R,5R,6- tetrahydroxyhexane)or2S-tert-butylsulfonylmethyl-3-phenyl-propionyl-L- histidyl-2S-amino-1 -cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane. Refer to

Chart J.

To a solution of 28 mg of the title product of Preparation 34 in a small amount of acetic acid is added 24 mg of palladium black catalyst. The suspension is placed on a Parr apparatus at 45 psi of hydrogen gas and allowed to shake for 3 days. The reaction mixture is diluted with dichloromethane/methanol and then filtered tiirough Celite with dichloromethane/methanol to pure methanol washings. The filtrates are concentrated under reduced pressure and the residual acetic acid azeotroped with the aid of toluene. The resulting white sc'id is gravity chromatographed on silica gel with 10% to 20% methanol saturated with ammonia in dichloromethane to afford 18 mg of the title product as a white solid.

Physical characteristics are as follows:

FAB HRMS: C₃₂H₅₀N₄O₈S (m+H)=651.3465.

IC₅₀ value of 3.8 nM.

Examples 11-16 Following chemical processes and procedures analogous to those described above and using starting materials and reactants which are commercially available or readily prepared by methods known in the art, the following compounds of the present invention, having the indicated physical characteristics, are prepared:

Example 11 N-tert-Butyloxycarbonyl-L-phenylalanyl-N-methyl-L-histidyl-2S-amino-1- cyclohexyl-3R,4S,5S,6-tetrahydroxy-hexane.

FAB-HRMS: C₃₃H₅₂N₅O₈ (m + H) = 646.3811.

HPLC (A): 14.62 min.

0% renin inhibition at 10^-8M;Expected to have activity at a higher dose.

Example 12 N-tert-Butyloxcarbonyl-L-phenylalanyl-N-meΛyl-L-histidyl-2S-amino-1- cyclohexyl-3R,4R,5R,6-tetrahydroxy-hexane.

FAB-HRMS: C₃₃H₅₂N₅O₈ (m + H) = 646.

HPLC (A): 11.26 min.

9% renin inhibiton at 10^-8M.

Example 13 2R-Hydroxy-3-(tetrahydro-2-naphdιyl)-propanoyl-L-histidyl-2S-amino-1- cyclohexyl-3R,4R,5R,6-tetrahydroxy-hexane.

FAB-HRMS: C₃₁H₄₇N₄O₇ (m + H) = 587.3453.

HPLC (B): 20.62 min.

8% renin inhibition at 10^-8M.

Example 14 N-tert-Butyloxycarbonyl-L-phenylalanyl-N-methyl-L-histidyl-2S-amino-1- cyclohexyl-3R,4S,5R,6-tetrahydroxy-hexane.

FAB-HRMS: C₃₃H₅₂N₅O₈ (m + H) = 640.3805.

HPLC (A): 11.36 min.

15% renin inhibition at 10^-8M.

Example 15 Tetrahydro-1-naphthoxyacetyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,5- R,6-tetrahydroxy-heptane.

FAB-HRMS: C₃₀H₄₄N₄O₇ (m + H) = 572.

Example 16 Tetrahydro-1-naphdιoxyacetyl-L-histidyl-2S-amino-1-cyclohexyl-6-methyl- 3R,4R,5S,6-tetrahydroxy-heptane.

FAB-HRMS: C₃₂H₄₈N₄O₇ (m + H) = 601.3642

For the HPLC's in examples 11-16 above, the retention time is given and the solvent system used is indicated in parentheses. Solvent A is 60% of 90:10 = water: acetonitrile, 0.1% trifluoroacetic acid; and 40% of 10:90 = water: acetonitrile, 0.1% trifluoroacetic acid. Solvent

B is 70% of 90:10 = water: acetonitrile, 0.1% trifluoroacetic acid; and 30% of 10:90 = water: acetonitrile, 0;1 % trifluoroacetic acid.

*

Claims

1. A peptide having a non-cleavable transition state insert corresponding to the 10, 11 -position of a renin substrate (angiotensinogen) of the formula XL₈

in R₁ is

(a) hydrogen,

(b) C₁-C₁₀ alkyl,

(c) aryl,

(d) C₃-C₇ cycloalkyl,

(e) Het,

(f) C₁-C₃ alkoxy, or

(g) C₁-C₃ alkylthio;

wherein each R₂ is independently

(a) hydrogen,

(b) C₁-C₅ alkyl,

(c) -(CH₂)_p-aryl, or

(d) -(CH₂)_p-Het;

wherein p is O to 2 inclusive;

wherein q is three to six, inclusive;

wherein aryl is phenyl or naphthyl substituted by zero to three of the following:

(a) C₁-C₃ alkyl,

(b) hydroxy,

(c) C₁-C₃ alkoxy,

(d) halo,

(e) amino,

(0 mono- or di-C₁-C₃ alkylamino,

(g) -CHO,

(h) -COOH,

(i) -CONH₂,

(j) -CONH( C₁-C₅ alkyl),

(k) -nitro,

(l) mercapto,

(m) C₁-C₃ alkylthio, (n) C₁-C₃ alkylsulfinyl,

(o) C₁-C₃ alkylsulfonyl,

(p) -N(R₈)- C₁-C₃ alkylsulfonyl,

(q) SO₃H,

(r) SO₂NH₂,

(s) -CN, or

(t) -CH₂NH₂;

wherein -Het is a 5- or 6-membered saturated or unsaturated ring containing from one to three heteroatoms selected from the group consisting of nitrogen, oxygen, and sulfur; and including any bicyclic group in which any of the above heterocyclic rings is fused to a benzene ring, which heterocyclic moiety is substituted with zero to three of the following:

(a) C₁-C₆ alkyl,

(b) hydroxy,

(c) trifluoromethyl,

(d) C₁-C₄ alkoxy,

(e) halo,

(f) aryl,

(g) aryl C₁-C₄ alkyl-,

(h) amino,

(i ) mono- or di-(C₁-C₄ alkyl) amino, or

(j) C₁-C₅ alkanoyl;

wherein R₈ is

(a) hydrogen,

(b) C₁-C₅ alkyl,or

(c) (CH₂)_p-phenyl;

or a carboxy, amino or other reactive group protected form or a pharmaceutically acceptable acid or base addition salt thereof.

2. In a peptide, the improvement which comprises inclusion in the peptide of a non-cleavable transition state insert corresponding to the 10,11-position of a renin substrate (angiotensinogen) of the formula XL₈

wherein R₁ is

(a) hydrogen, (b) C₁-C₁₀ alkyl,

(c) aryl,

(d) C₃-C₇ cycloalkyl,

(e) Het,

(f) C₁-C₃ alkoxy, or

(g) C₁-C₃ alkylthio;

wherein each R₂ is independently

(a) hydrogen,

(b) C₁-C₅ alkyl,

(c) -(CH₂)_p-aryl, or

(d) -(CH₂)_p-Het;

wherein p is O to 2 inclusive;

wherein q is three to six, inclusive;

wherein aryl is phenyl or naphthyl substituted by zero to three of the following:

(a) C₁-C₃ alkyl,

(b) hydroxy,

(c) C₁-C₃ alkoxy,

(d) halo,

(e) amino,

(f) mono- or di-C₁-C₃ alkylamino,

(g) -CHO,

(h) -COOH,

(i) -CONH₂,

(j) -CONH( C₁-C₅ alkyl),

(k) -nitro,

(l) mercapto,

(m) C₃-C₇ alkylthio,

(n) C₁-C₃ alkylsulfinyl,

(o) C₁-C₃ alkylsulfonyl,

(p) -N(R₈)- C₁-C₃ alkylsulfonyl,

(q) SO₃H,

(r) SO₂NH₂,

(s) -CN, or

(t) -CH₂NH₂;

wherein -Het is a 5- or 6-membered saturated or unsaturated ring containing from one to three heteroatoms selected from the group consisting of nitrogen, oxygen, and sulfur; and including any bicyclic group in which any of the above heterocyclic rings is fused to a benzene ring, which heterocyclic moiety is substituted with zero to three of the following:

(a) C₁-C₆ alkyl,

(b) hydroxy,

(c) trifluoromethyl,

(d) C₁-C₄ alkoxy,

(e) halo,

(f) aryl,

(g) aryl C₁-C₄ alkyl-,

(h) amino,

(i) mono- or di-(C₁-C₄ alkyl) amino, or

(j) C₁-C₅ alkanoyl;

wherein R₈ is

(a) hydrogen,

(b) C₁-C₅ alkyl,or

(c) (CH₂)p-phenyl;

or a carboxy, amino or other reactive group protected form or a pharmaceutically acceptable acid or base addition salt thereof.

3. The peptide of claim 1 of the formula I

X₁-B₇-C₈-D₉-E₁₀ I wherein X₁ is

(a) hydrogen,

(b) Y₁-(CH₂)_n-C(O)-,

(c) (HOCH₂)₃C-NH-C(O)-,

(d) (HOCH₂)₂CH-NH-C(O)-,

(f) HOH₂C(CHOH)_nCH(CH₂OH)-NH-C(O)-,

(g) NaO₃S(CH₂)₂N(CH₃)C(O)(CH₂)₆C(O)-,

(h) R₅C(O)-, (i) R₅-O-C(O)-,

(j) Z₁-C(O)-,

(k) tetrahydro-naphdιyl-(CH₂)_n-CH(OH)-C(O)-, or

(l) tetrahydro-naphthyl-O-(CH₂)_n-C(O)-;

wherein B₇ is absent or a divalent moiety of the formula XL_b;

wherein C₈ is absent or a divalent moiety of the formula XL₁, XL₂ or XL₃;

or wherein C₈ is a monovalent moiety of the formula XL_1a or XL_2a when X₁ and B₇ are absent;

wherein D₉ is absent or a divalent moiety of the formula XL₄:

or wherein C₈-D₉ is XL₇, or when X₁ and B₇ are absent, XL_7a;

wherein E₁₀ is a monovalent moiety of the formula XL₈;

wherein W₁ is H or OH;

wherein Y₁ is

(a) -NH₂,

(b) -CO₂H,

(c) -C(NH₂)(CH₃)₂,

(d) -CH(NH₂)(CO₂H), or

(e) -OP(O)(OH)₂;

wherein Z₁ is -Het bonded via nitrogen; wherein Q₁ is

(a) -CH₂-,

(b) -CH(OH)-,

(c) -O-, or

(d) -S-;

wherein M₁ is -C(O)- or -CH₂-;

wherein m is i 1 or 2;

wherein n is 0 to 5, inclusive;

wherein p is 0 to 2, inclusive;

wherein q is 3 to 6, inclusive;

wherein R₁ is

(a) hydrogen,

(b) C₁-C₁₀ alkyl,

(c) aryl,

(d) C₃-C₇ cycloalkyl,

(e) Het,

(f) C₁-C₃ alkoxy, or

(g) C₁-C₃ alkylthio;

wherein each R₂ is independently

(a) hydrogen,

(b) C₁-C₅ alkyl,

(c) -(CH₂)_p-aryl, or

(d) -(CH₂)_p-Het;

wherein R₃ is

(a) hydrogen,

(b) C₁-C₁₀ alkyl,

(c) -(CH₂)_p-aryl,

(d) -(CH₂)_p-Het, (e) -(CH₂)_p-C₃-C₇ cycloalkyl, or

(f) 1- or 2-adamantyl;

wherein R₄ is

(a) hydrogen,

(b) C₁-C₁₀ alkyl,

(c) -(CH₂)_p-aryl,

(d) -(CH₂)_p-Het, or

(e) -(CH₂)_p-C₃-C₇ cycloalkyl; wherein R₅ is

(a) C₁-C₁₀ alkyl,

(b) C₃-C₇ cycloalkyl,

(c) aryl,

(d) Het,

(e) aryl C₁-C₅ alkyl, or

(f) Het C₁-C₅ alkyl;

wherein R₆ is

(a) hydrogen,

(b) C₁-C₅ alkyl,

(c) -(CH₂)_p-aryl, or

(d) -(CH₂)_p-Het;

wherein R₇ is

(a) hydrogen,

(b) C₁-C₁₀ alkyl,

(c) hydroxy,

(d) amino C₁-C₄ alkyl,

(e) guanidinyl C₁-C₄ alkyl,

(f) aryl,

(g) Het,

(h) methylthio,

(i) -(CH₂)_p-C₃-C₇ cycloalkyl, or

(j) amino,

wherein R₈ is

(a) hydrogen,

(b) C₁-C₅ alkyl, or

(c) -(CH₂)_p-phenyl;

wherein R₉ is (a) C₁-C₁₀ alkyl, or

(b) aryl;

wherein aryl is phenyl or naphthyl substituted by zero to three of the following:

(a) C₁-C₃ alkyl,

(b) hydroxy,

(c) C₁-C₃ alkoxy,

(d) halo,

(e) amino,

(f) mono- or di- C₁-C₃ alkylamino,

(g) -CHO,

(h) -COOH,

(i) -CONH₂,

(j) -CONH( C₁-C₅ alkyl)

(k) -nitro,

(i) mercapto,

(m) C₁-C₃ alkylthio,

(n) C₁-C₃ alkylsulfinyl,

(o) C₁-C₃ alkylsulfonyl,

(p) -N(R₈)- C₁-C₅ alkylsulfonyl,

(q) SO₃H,

(r) SO₂NH₂,

(s) -CN, or

(t) -CH₂NH₂;

wherein -Het is a 5- or 6-membered saturated or unsaturated ring containing from one to three heteroatoms selected from the group consisting of nitrogen, oxygen and sulfur; and including any bicyclic group in which any of the above heterocyclic rings is fused to a benzene ring, which heterocyclic moiety is substituted with zero to three of the following:

(a) C₁-C₆ alkyl,

(b) hydroxy,

(c) trifluoromethyl,

(d) C₁-C₄ alkoxy,

(e) halo,

(f) aryl,

(g) aryl C₁-C₄ alkyl-,

(h) amino,

(i) mono- or di ( C₁-C₄ alkyl)amino, or (j) C₁-C₅ alkanoyl;

or a carboxy, amino or other reactive group protected form thereof;

or a pharmaceutically acceptable acid or base addition salt thereof.

4. A peptide of claim 3

wherein X₁ is

(a) t-butyl-O-C(O)-,

(b) t-butyl-CH₂-C(O)-,

(c) tetrahydro-naphthyl-CH₂-CH(OH)-C(O)-,

(d) tetrahydro-naphthyl-O-CH₂-C(O)-,

(e)

(f) β-valine;

wherein B₇ is

(a) absent, or

(b) Pro;

wherein C₈ is

(a) absent,

(b) Phe,

(c) (OCH₃)-Tyr, or

(d) when X_χ and B₇ are absent, t-butyl-SO₂-CH₂-CH-(CH₂-phenyl)-C(O)-; wherein D₉ is

(a) His, or

(b) (N-Me) His;

wherein R₁ is cyclohexyl;

wherein R₂ is

(a) hydrogen, or

(b) methyl;

wherein q is three;

or a pharmaceutically acceptable salt thereof.

5. A peptide of claim 4 selected from the group consisting of:

N-tert-butyloxycarbonyI-L-phenylalanyl-L-histidyI-2S-amino-1-cyclohexyl-3R,4R,5R,6- tetrahydroxyhexane;

N-tert-butyloxycarbonyl-L-phenylalanyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4S,5S,6- tertahydroxyhexane;

N-tert-butyloxycarbonyl-L-phenylalanyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,5S,6- tertahydroxyhexane;

N-tert-butyloxycarbonyl-L-phenylalanyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4S,5R,6- tetrahydroxyhexane;

βVal-Phe-His-Y (wherein Y is 2S-amino-1-cyclohexyl-3R-4R,5R,6-tetrahydroxyhexane) or β-Valyl-L-phenylalanyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane,bis trifluoroacetate;

Boc-Tyr(OMe)-His-Y (wherein Y is 2S-amino-1-cyclohexyl-3R-4R,5R,6-tetrahydroxy- hexane) or N-tert-butyloxycarbonyl-O-methyl-L-tyrosyl-L-histidyl-2S-amino-1-cyclohexyl- 3R,4R,5R,6-tetrahydroxyhexane;

/3Val-Tyr(OMe)-His-Y (wherein Y is 2S-amino-1-cyclohexyl-3R-4R,5R,6-tetrahydroxy-hexane)orβ-Valyl-O-methyl-L-tyrosyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane, bis trifluoroacetate;

Glc-Pro-Tyr(OMe)-His-Y (wherein Y is 2S-amino-1-cyclohexyl-3R-4R,5R,6-tetrahydroxy-hexane) or 2-Deoxy-D-glucopyranos-2-aminocarbonyl-L-prolyI-O-methyl-L-tyrosyl-L-histidyl-2S-amino-1-cycohexyl-3R,4R,5R,6-tetrahydroxyhexane;

SPP-His-Y (wherein Y is 2S-amino-1-cyclohexyl-3R-4R,5R,6-tetrahydroxyhexane) or 2R-tert-butylsulfonylmethyl-3-phenyl-propionyl-L-histidyI-2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane;

SPP-His-Y (wherein Y is 2S-amino-1-cyclohexyl-3R-4R,5R,6-tetrahydroxyhexane) or 2S-tere-butylsulfonylmethyl-3-phenyl-propionyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxyhexane;

N-tert-Butyloxycarbonyl-L-phenylalanyl-N-methyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4S,5S,6-tetrahydroxy-hexane;

N-tert-Butyloxycarbonyl-L-phenylaIanyl-N-methyl-L-histidyl-2S-amino-1-cyclohexyl- 3R,4R,5R,6-tetrahydroxy-hexane;

2R-Hydroxy-3-(tetrahydro-2-naphthyl)-propanoyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxy-hexane;

N-tert-Butyloxycarbonyl-L-phenylalanyl-N-methyl-L-histidyI-2S-amino-1-cyclohexyl-3R,4S,5R,6-tetrahydroxy-hexane; Tetrahydro-1-naphdιoxyacetyl-L-histidyl-2S-amino-1-cyclohexyl-3R,4R,5R,6-tetrahydroxy-hexane;

Tetrahydro-1-naphthoxyacetyl-L-histidyl-2S-amino-1-cyclohexyl-6-methyl-3R,4R,5S,6-tetrahydroxy-heptane; and

N-tert-ButyIoxycarbonyl-L-phenylalanyI-L-histidyl-2S-amino-1-cyclohexyl-6 methyl- 3R,4R,5S,6-tetrahydroxyheptane.