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  • A61K38/00—Medicinal preparations containing peptides
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  • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
  • A61K38/22—Hormones
  • A61K38/25—Growth hormone-releasing factor [GH-RF] (Somatoliberin)
• • A—HUMAN NECESSITIES
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  • A61K38/22—Hormones
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  • A61K38/00—Medicinal preparations containing peptides
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  • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
  • A61K38/22—Hormones
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  • C07C237/00—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by amino groups
  • C07C237/02—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by amino groups having the carbon atoms of the carboxamide groups bound to acyclic carbon atoms of the carbon skeleton
  • C07C237/22—Carboxylic acid amides, the carbon skeleton of the acid part being further substituted by amino groups having the carbon atoms of the carboxamide groups bound to acyclic carbon atoms of the carbon skeleton having nitrogen atoms of amino groups bound to the carbon skeleton of the acid part, further acylated
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  • C07C275/20—Derivatives of urea, i.e. compounds containing any of the groups, the nitrogen atoms not being part of nitro or nitroso groups having nitrogen atoms of urea groups bound to acyclic carbon atoms of an unsaturated carbon skeleton
  • C07C275/24—Derivatives of urea, i.e. compounds containing any of the groups, the nitrogen atoms not being part of nitro or nitroso groups having nitrogen atoms of urea groups bound to acyclic carbon atoms of an unsaturated carbon skeleton containing six-membered aromatic rings
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  • C07—ORGANIC CHEMISTRY
  • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
  • C07C323/00—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups
  • C07C323/50—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and carboxyl groups bound to the same carbon skeleton
  • C07C323/51—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and carboxyl groups bound to the same carbon skeleton having the sulfur atoms of the thio groups bound to acyclic carbon atoms of the carbon skeleton
  • C07C323/60—Thiols, sulfides, hydropolysulfides or polysulfides substituted by halogen, oxygen or nitrogen atoms, or by sulfur atoms not being part of thio groups containing thio groups and carboxyl groups bound to the same carbon skeleton having the sulfur atoms of the thio groups bound to acyclic carbon atoms of the carbon skeleton with the carbon atom of at least one of the carboxyl groups bound to nitrogen atoms
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  • C07D209/00—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom
  • C07D209/02—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom condensed with one carbocyclic ring
  • C07D209/04—Indoles; Hydrogenated indoles
  • C07D209/10—Indoles; Hydrogenated indoles with substituted hydrocarbon radicals attached to carbon atoms of the hetero ring
  • C07D209/14—Radicals substituted by nitrogen atoms, not forming part of a nitro radical
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  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D209/00—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom
  • C07D209/02—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom condensed with one carbocyclic ring
  • C07D209/04—Indoles; Hydrogenated indoles
  • C07D209/10—Indoles; Hydrogenated indoles with substituted hydrocarbon radicals attached to carbon atoms of the hetero ring
  • C07D209/18—Radicals substituted by carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals
  • C07D209/20—Radicals substituted by carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals substituted additionally by nitrogen atoms, e.g. tryptophane
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  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D403/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00
  • C07D403/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings
  • C07D403/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings linked by a chain containing hetero atoms as chain links
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  • C07K—PEPTIDES
  • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
  • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
  • C07K5/06—Dipeptides
  • C07K5/06008—Dipeptides with the first amino acid being neutral
  • C07K5/06017—Dipeptides with the first amino acid being neutral and aliphatic
  • C07K5/06026—Dipeptides with the first amino acid being neutral and aliphatic the side chain containing 0 or 1 carbon atom, i.e. Gly or Ala
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  • C07K—PEPTIDES
  • C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
  • C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
  • C07K5/06—Dipeptides
  • C07K5/06191—Dipeptides containing heteroatoms different from O, S, or N

Definitions

• Growth hormone which is secreted from the pituitary, stimulates growth of all tissues of the body that are capable of growing.
• growth hormone is known to have the following basic effects on the metabolic process of the body:
• a deficiency in growth hormone secretion can result in various medical disorders, such as dwarfism.
• growth hormone Various ways are known to release growth hormone. For example, chemicals such as arginine, L-3,4-dihydroxyphenylalanine (L-DOPA), glucagon, vasopressin, and insulin induced hypoglycemia, as well as activities such as sleep and exercise, indirectly cause growth hormone to be released from the pituitary by acting in some fashion on the hypothalamus perhaps either to decrease somatostatin secretion or to increase the secretion of the known secretagogue growth hormone releasing factor (GRF) or an unknown endogenous growth hormone- releasing hormone or all of these.
• L-DOPA L-3,4-dihydroxyphenylalanine
• GRF growth hormone releasing factor
• the source of growth hormone was the extraction of the pituitary glands of cadavers. This resulted in a very expensive product and carried with it the risk that a disease associated with the source of the pituitary gland could be transmitted to the recipient of the growth hormone. Recently, recombinant growth hormone has become available which, while no longer carrying any risk of disease transmission, is still a very expensive product which must be given by injection or by a nasal spray.
• the instant invention covers certain substituted dipeptide analogs which have the ability to stimulate the release of natural or endogenous growth hormone.
• the compounds thus have the ability to be used to treat conditions which require the stimulation of growth hormone production or secretion such as in humans with a deficiency of natural growth hormone or in animals used for food production where the stimulation of growth hormone will result in a larger, more productive animal.
• a still further object of this invention is to describe compositions containing the substituted dipeptide analogs for the use of treating humans and animals so as to increase the level of growth hormone secretions. Further objects will become apparent from a reading of the following description. DESCRIPTION OF THE INVENTION
• n 0 or 1
• p 0 to 6;
• q 0 to 4.
• w is 0 or 1 ;
• n 0 to 2;
• R 1 , R 2 , R 1a , R 2a , R 1 b , and R 2b are independently hydrogen, halogen, C 1 -C 7 alkyl, C 1 -C 3 perfluoroalkyl, C 1 -C 3 perfluoroalkoxy, -S(O) m- R 7a , cyano, nitro, R 7b O(CH 2 ) v -, R 7b COO(CH 2 ) v -, R 7b OCO-(CH 2 ) v -, R 4 R 5 N(CH 2 ) v -, R 7b CON(R 4 )(CH 2 ) v -, R 4 R 5 NCO(CH 2 ) v -, R 4 R 5- NCOO(CH 2 ) v -, phenyl or substituted phenyl where the substituenis are from 1 to 3 of halogen, C 1 -C 6 alkyl, C 1 -C 6 alkoxy, or
• R 3a and R 3b are independently hydrogen, R 9 , C 1 -C 6 alkyl substituted with R 9 , phenyl substituted with R 9 , or phenoxy substituted with R 9 ;
• R 7b O(CH 2 ) v -, R 7b COO(CH 2 ) v -, R 7b OCO(CH 2 ) v -, R 7b CO(CH 2 ) v -, R 7b O(CH 2 ) v CO-, R 4b R 12c N(CH 2 ) v -, R 12a R 12b NCO(CH 2 ) v -,
• R 12a R 12b NCS(CH 2 ) v -, R 4b R 12a NN(R 12b )CO(CH 2 ) v -,
• R 4b R 12a NN(R 12b )CS(CH 2 ) v -, R 4b R 12 aNCON(R 12c )(CH 2 ) v -,
• R 13 is C 1 -C 3 perfluoroalkyl, C 1 -C 6 alkyl, substituted C 1 -C 6 alkyl, where the substituents are hydroxy, -NR 10 R 1 1 , carboxy, phenyl or substituted phenyl; phenyl or substituted phenyl where the substituents on the phenyl are from 1 to 3 of halogen, C 1 -C 6 alkyl, C 1 -C 6 alkoxy or hydroxy where R 10 and R 1 1 are as defined;
• R 14 is hydrogen, R 1 , R 2 independently disubstituted phenyl, C 1 -C 10 alkyl or substituted C 1 -C 10 alkyl where the substituents are from 1 to 3 of imidazolyl, indolyl, hydroxy, fluoro, S(O) m R 7a , C 1 -C 6 alkoxy, C 3 - C 7 cycloalkyl, R 1 , R 2 independently
• R 1 , R 2 independently disubstituted heterocycle, where the heterocycle is imidazole, thiophene, furan, pyrrole, oxazole, thiazole, triazole, tetrazole, pyridine, benzofuran, benzothiophene, benzimi- dazole, indole, 7-azaindole, oxindole or indazole; where R 1 , R 2 , R 10 and R 1 1 are as defined above;
• R 4 , R 4a , R 4b , R 5 and R 5a are independently hydrogen, phenyl, substituted phenyl, C 1 -C 10 alkyl, substituted C 1 -C 10 alkyl, C 3 -C 10 alkenyl, substituted C 3 -C 10 alkenyl, C 3 -C 10 alkynyl, or substituted C3- C 10 alkynyl where the substituents on the phenyl, alkyl, alkenyl or alkynyl are from 1 to 5 of hydroxy, C 1 -C 6 alkoxy, C 3 -C 7 cycloalkyl, fluoro, R 1 , R 2 independently disubstituted phenyl C 1 -C 3 alkoxy, R 1 , R 2 independently disubstituted phenyl, C 1 -C 20 -alkanoyloxy, C 1 -C 5 alkoxycarbonyl, carboxy, formyl, or -NR 10
• R 6 is hydrogen, C 1 -C 10 alkyl, phenyl or phenyl C 1 -C 10 alkyl;
• alkyl groups specified above are intended to include those alkyl groups of the designated length in either a straight or branched configuration.
• exemplary of such alkyl groups are methyl, ethyl, propyl, isopropyl, butyl, sec-butyl, tertiary butyl, pentyl, isopentyl, hexyl, isohexyl, and the like.
• alkoxy groups specified above are intended to include those alkoxy groups of the designated length in either a straight or branched configuration.
• exemplary of such alkoxy groups are methoxy, ethoxy, propoxy, isopropoxy, butoxy, isobutoxy, tertiary butoxy, pentoxy, isopentoxy, hexoxy, isohexoxy and the like.
• halogen is intended to include the halogen atom fluorine, chlorine, bromine and iodine.
• n is 0 or 1;
• p 0 to 4.
• q 0 to 2;
• w is 0 or 1 ; R 1 0
• R 7b COO(CH 2 ) v -, R 7b OCO(CH 2 ) v -, phenyl or substituted phenyl where the substituents are from 1 to 3 of halogen, C 1 -C 6 alkyl, C 1 -C 6 alkoxy, or hydroxy;
• R 7a and R 7b are independently hydrogen, C 1 -C 3 perfluoroalkyl, C 1 -C 6 alkyl, substituted C 1 -C 6 alkyl, where the substituents are phenyl; phenyl and v is 0 to 2;
• R 3a and R 3b are independently hydrogen, R 9 , C 1 -C 6 alkyl substituted with R 9 , phenyl substituted with R 9 , or phenoxy substituted with R 9 ;
• R 9 is as defined above;
• R 12a , R 12b and R 12c are independently R 5a , OR 5a , or COR 5a ;
• R 12a and R 12b , or R 12b and R 12c , or R 13 and R 12b or R 12a and R 4b can be taken together to form -(CH 2 ) r -B-(CH 2 ) s - where B is CHR 1 , O, S(O) m or NR 10 , m is 0, 1 or 2, r and s are independently 0 to 3,
• R 1 is as defined above and
• R 10 is hydrogen, C 1 -C 6 alkyl, phenyl C 1 -C 6 alkyl or C 1 -C 5 alkanoyl-C 1
• R 6 is hydrogen, C 1 -C 10 alkyl or phenyl C 1 -C 10 alkyl; A is
• R 8 , R 8a and R 8b are independently hydrogen, C 1 -C 10 alkyl, substituted C 1 -C 10 alkyl where the substituents are from 1 to 3 of imidazolyl, indolyl, hydroxy, fluoro, S(O) m R 7a , C 1 -C 6 alkoxy, R 1 , R 2
• R 8 and R 8a can be taken together to form -(CH 2 ) t - where t is 2 to 4; and R 8 and R 8a can independently be joined to one or both of R 4 and R 5 to form alkylene bridges between the terminal nitrogen and the alkyl portion of the A group wherein the bridge contains from 1 to 5 carbon atoms;
• n 0 or 1
• p 0 to 3;
• q 0 to 2;
• w is 0 or 1 ;
• n 0 or 1;
• R 1 , R 2 , R 1 a , R 2a , R 1b , and R 2b are independently hydrogen, halogen, C 1 -C 7 alkyl, C 1 -C 3 perfluoroalkyl, -S(O) m R 7a , R 7b O(CH 2 ) v -,
• R 7b COO(CH 2 ) v -, R 7b OCO(CH 2 ) v -, phenyl or substituted phenyl where the substituents are from 1 to 3 of halogen, C 1 -C 6 alkyl, C 1 -C 6 alkoxy, or hydroxy;
• R 7a and R 7b are independently hydrogen, C 1 -C 6 alkyl, substituted C 1 - C 6 alkyl, where the substituents are phenyl and v is 0 to 2;
• R 3a and R 3b are independently hydrogen, R 9 , C 1 -C 6 alkyl substituted with R 9 , phenyl substituted with R 9 , or phenoxy substituted with R 9 ;
• R 9 is as defined above; R 12a , R 12b and R 12c are independently R 5a or OR 5a .
• R 12a and R 12b , or R 12b and R 12c , or R 13 and R 12b or R 12a and R 4b can be taken together to form -(CH 2 ) r -B-(CH 2 ) s - where B is CHR 1 , O, S(O) m or NR 10 , m is 0, 1 or 2, r and s are independently 0 to 2,
• R 1 is as defined above and R 10 is hydrogen, C 1 -C 6 alkyl or C 1 -C 5 alkanoyl-C 1 -C 6 alkyl;
• R 13 is C 1 -C 6 alkyl, substituted C 1 -C 6 alkyl, where the substituents are phenyl or substituted phenyl; phenyl or substituted phenyl where the substituents on the phenyl are from 1 to 3 of
• R 6 is hydrogen or C 1 -C 10 alkyl
• R 8 , R 8a and R 8b are independently hydrogen, C 1 -C 10 alkyl, substituted C 1 -C 10 alkyl where the substituents are from 1 to 3 of imidazolyl, indolyl, hydroxy, fluoro, S(O)mR 7a , C 1 -C 6 alkoxy, R 1 substituted or R 1 , R 2 independently disubstituted phenyl, C 1 -C 5 -alkanoyloxy, C 1 -C 5 alkoxycarbonyl, carboxy; or R 8 and R 8a can be taken together to form -(CH 2 ) t - where t is 2; or R 8 and R 8a can independently be joined to one or both of R 4 and R 5 to form alkylene bridges between the terminal nitrogen and the alkyl portion of the A group wherein the bridge contains from 1 to 5 carbon atoms;
• n 0 or 1
• p 0 to 2;
• w 1;
• X is O, S(O)m
• R 9 is as defined above;
• R 12a , R 12b and R 12c are independently R 5a .
• R 12a and R 12b , or R 12b and R 12c , or R 13 and R 12b or R 12a and R 4b can be taken together to form -(CH 2 ) r -B-(CH 2 ) s - where B is CHR 1 , O, S(O) m or NR 10 , m is 0, 1 or 2, r and s are independently 0 to 2,
• R 1 is as defined above and R 10 is hydrogen, C 1 -C 6 alkyl or C 1 -C 5 alkanoyl C 1 -C 6 alkyl;
• R 13 is C 1 -C 6 alkyl, substituted C 1 -C 6 alkyl, where the substituents are phenyl or substituted phenyl; phenyl or substituted phenyl where the substituents on the phenyl are from 1 to 3 of halogen,
• R 6 is hydrogen
• R 8 , R 8a and R 8b are independently hydrogen, C 1 -C 10 alkyl, substituted C 1 -C 10 alkyl where the substituents are from 1 to 3 of imidazolyl, indolyl, hydroxy, fluoro, S(O)mR 7a , C 1 -C 6 alkoxy, R 1 substituted or R 1 , R 2 independently disubstituted phenyl, C 1 -C 5 -alkanoyloxy, C 1 -C 5 alkoxycarbonyl, carboxy; or R 8 and R 8a can be taken together to form -(CH 2 ) t - where t is 2; and R 8 and R 8a can independently be joined to one or both of R 4 and R 5 to form alkylene bridges between the teiminal nitrogen and the alkyl portion of the A group wherein the bridge contains from 1 to 5 carbon atoms;
• Representative preferred growth hormone releasing compounds of the present invention include the following:
• the compounds of the instant invention all have at least one asymmetric center as noted by the asterisk in the structural Formula I above. Additional asymmetric centers may be present on the molecule depending upon the nature of the various substituents on the molecule. Each such asymmetric center will produce two optical isomers and it is intended that all such optical isomers, as separated, pure or partially purified optical isomers or racemic mixtures thereof, be included within the ambit of the instant invention.
• the asymmetric center represented by the asterisk in Formula I it has been found that the compound in which the 3-amino substituent is above the plane of the structure, as seen in Formula la, is more active and thus more prefened over the compound in which the 3-amino substituent is below the plane of the structure.
• This center will be designated according to the R/S rules as either R or S depending upon the values of X, n, p and R 15 .
• the instant compounds are generally isolated in the form of their pharmaceutically acceptable acid addition salts, such as the salts derived from using inorganic and organic acids.
• acids are hydrochloric, nitric, sulfuric, phosphoric, formic, acetic, trifluoroacetic, propionic, maleic, succinic, malonic and the like.
• certain compounds containing an acidic function such as a carboxy can be isolated in the form of their inorganic salt in which the counterion can be selected from sodium, potassium, lithium, calcium, magnesium and the like, as well as from organic bases.
• Amino acid intermediates 1 are, in some cases, commercially available in the form of their N-t-butoxycarbonyl or N- benzyloxycarbonyl derivatives. These intermediates can also be prepared by a variety of methods described in the literature and familiar to one skilled in the art. For example, the Strecker synthesis may be employed for the construction of racemic amino acid intermediates. Resolution can be achieved by classical methods, for example separation of diastereomeric salts by fractional crystallization. Alternatively, a chiral amino acid synthesis may be employed using the procedures described by R.M. Williams and M.N. Im (J. Amer. Chem. Soc. , 1 13, 9276-9286, 1991.).
• Conversion of the free amino acid product to its N-t-butoxycarbonyl (BOC) derivative can be achieved by a number of methods known in the art, for example, treatment with di- t-butyl dicarbonate in an inert solvent such as methylene chloride.
• Benzyloxycarbonyl (CBz) protected derivatives are obtained by treatment of the amino acid with, for example, benzyl chloroformate.
• benzyl ester 2 is carried out by treatment with benzyl alcohol in the presence of a coupling agent, such as 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC), in methylene chloride with a catalytic amount of 4-dimethylaminopyridine.
• a coupling agent such as 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC)
• EDC 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride
• Attachment of the amino acid sidechain to intermediates of formula II is accomplished by the route shown in Scheme 2. Coupling is conveniently carried out by the use of an appropriately protected amino acid derivative, such as that illustrated by formula III, and a coupling reagent such as benzotriazol-1-yloxytris(dimethylamino)- phosphonium hexafluorophosphate ("BOP") in an inert solvent such as methylene chloride.
• BOP benzotriazol-1-yloxytris(dimethylamino)- phosphonium hexafluorophosphate
• the protecting group G must be selected to be compatible with the conditions employed for removal of the specific class of ester present in 4. Hence, as illustrated for the benzyl ester 4, G is taken as t- butoxycarbonyl. It may further be appreciated that other combinations of protecting group G and ester functionality may be employed; for example, the benzyloxycarbonyl protecting group is inert to the standard conditions of aqueous sodium hydroxide employed to
• the protected amino acid derivatives III are, in many cases, commercially available in t-butoxycarbonyl (BOC) or benzyloxy- carbonyl (CBz) forms.
• BOC t-butoxycarbonyl
• CBz benzyloxy- carbonyl
• G is t-butoxycarbonyl or benzyloxycarbonyl
• the substituted phenyl sidechains V are prepared from the conesponding alkylating agent VI by displacement of the leaving group Y with sodium azide as shown in Scheme 5. Reduction of the azide product by hydrogenation in the presence of a transition metal catalyst, or alternatively by reaction with triphenylphosphine followed by hydrolysis, gives the desired amine derivative 10. Conversion to the desired intermediate V is achieved by the aforementioned reductive alkylation procedure.
• Y is a leaving group
• an alternative route involves coupling of intermediate IV with R 14 NH 2 using one of the coupling reagents described previously, followed by alkylation of the amide bond with VI. Alkylation is carried out in an inert solvent, such as
• Alkylating agents VI are, in some cases, commercially available or may be prepared by the procedures described in the following schemes. SCHEME 6
• G is t-butoxycarbonyl or benzyloxycarbonyl
• Alkylating agents VI are, in some cases commercially available compounds or may be prepared by methods described in the literature and familiar to one skilled in the art.
• Compounds of formula I wherein R 3a or R 3b is a tetrazole (13) are prepared as described in Scheme 7 by reaction of IV with a suitably substituted intermediate 11 containing a nitrile as tetrazole precursor.
• Elaboration to the desired product 13 is carried out by treatment with trimethyltin azide in boiling toluene.
• Coupling of the carboxylic acid derivative 24 with R 4 R 5 NH is conveniently carried out by the use of a coupling reagent such as benzotriazol-1-yloxytris-(dimethylamino)phosphonium hexafluorophosphate ("BOP") in an inert solvent such as methylene chloride.
• BOP benzotriazol-1-yloxytris-(dimethylamino)phosphonium hexafluorophosphate
• an inert solvent such as methylene chloride.
• the requisite carboxylic acid precursors can be prepared as illustrated in Scheme 12 for the biphenyl compound 24.
• intermediate 32 alternative methods of reduction are indicated, such as chemical reduction with stannous chloride under acidic conditions.
• protecting group G in intermediate 32 must be compatible with the experimental conditions anticipated for reduction.
• intermediate 32 wherein G is t-butoxycarbonyl (BOC) are stable to the conditions of catalytic reduction employed in the conversion to 37.
• Intermediate 37 may also be further elaborated to new intermediate 38 by reductive alkylation with an aldehyde by the aforementioned procedures.
• Terminally disubstituted compounds 40 can be obtained directly by reaction of 37 with a disubstituted carbamoyl chloride in an inert solvent such as methylene chloride in the presence of triethylamine or 4-dimethylaminopyridine.
• monosubstituted compounds 41 wherein either R 4b or R 12a is hydrogen are obtained from 37 by reaction with an isocyanate as shown in Scheme 18.
• amine 32 is converted to an isocyanate 42 by treatment with phosgene or an equivalent reagent such as bis(trichloro- methyl)carbonate (triphosgene) as indicated in Scheme 19. Subsequent reaction of 42 with primary or secondary amines in an inert solvent such as methylene chloride gives the corresponding urea derivates 43 in good yield. Isocyanate 42 is also converted to substituted semi- carbazides 44 or hydroxy- or alkoxyureas 45 by reaction with substituted hydrazines or hydroxy- or alkoxy lamines, respectively.
• phosgene or an equivalent reagent such as bis(trichloro- methyl)carbonate (triphosgene) as indicated in Scheme 19.
• Isocyanate 42 is also converted to substituted semi- carbazides 44 or hydroxy- or alkoxyureas 45 by reaction with substituted hydrazines or hydroxy- or alkoxy lamines, respectively.
• Conversion of the carboxylic acid to the benzylamine derivative 52 can be achieved by a five-step sequence consisting of: 1) formation of a mixed anhydride with isobutyl chloroformate; 2) reduction with sodium borohydride to the benzyl alcohol; 3) formation of the mesylate with methanesulfonyl chloride; 4) formation of the azide by reaction with sodium azide, and finally, 5) reduction of the azide with tin(II) chloride.
• the benzylamine intermediate 52 can be further elaborated to 53 by the aforementioned reductive amination procedure.
• 53 G is benzyloxycarbonyl Reaction of amine 53 with the appropriate reagents to form urea-linked compounds 54 and 55 carbamate-linked compounds 56, and amide-linked structures 57 are illustrated in Scheme 22.
• Desilylation is carried out by treatment with tetra-n-butylammonium fluoride; conversion to the O-methanesulfonate 62 is achieved by reaction of the intermediate benzyl alcohol with methanesulfonic anhydride. Conversion to the requisite amine derivative V is achieved by the procedure described in Scheme 5.
• removal of benzyloxycarbonyl groups can also be achieved by treatment with a solution of hydrogen bromide in acetic acid.
• Catalytic hydrogenation is also employed in the removal of N- triphenylmethyl (trityl) protecting groups.
• Removal of t-butoxycarbonyl (BOC) protecting groups is carried out by treatment of a solution in a solvent such as methylene chloride or methanol, with a strong acid, such as hydrochloric acid or trifluoroacetic acid.
• the growth hormone releasing compounds of Formula I are useful in vitro as unique tools for understanding how growth hormone secretion is regulated at the pituitary level. This includes use in the evaluation of many factors thought or known to influence growth hormone secretion such as age, sex, nutritional factors, glucose, amino acids, fatty acids, as well as fasting and non-fasting states.
• the compounds of this invention can be used in the evaluation of how other hormones modify growth hormone releasing activity. For example, it has already been established that somatostatin inhibits growth hormone release.
• hormones that are important and in need of study as to their effect on growth hormone release include the gonadal hormones, e.g., testosterone, estradiol, and progesterone; the adrenal hormones, e.g., cortisol and other corticoids, pinephrine and norepinephrine; the pancreatic and gastrointestinal hormones, e.g., insulin, glucagon, gastrin, secretin; the vasoactive intestinal peptides, e.g., bombesin; and the thyroid hormones, e.g., thyroxine and
• the compounds of Formula I can also be employed to investigate the possible negative or positive feedback effects of some of the pituitary hormones, e.g., growth hormone and endorphin peptides, on the pituitary to modify growth hormone release. Of particular scientific importance is the use of these compounds to elucidate the subcellular mechanisms mediating the release of growth hormone.
• the compounds of Formula I can be administered to animals, including man, to release growth hormone in vivo.
• the compounds can be administered to commercially important animals such as swine, cattle, sheep and the like to accelerate and increase their rate and extent of growth, and to increase milk production in such animals.
• these compounds can be administered to humans in vivo as a diagnostic tool to directly determine whether the pituitary is capable of releasing growth hormone.
• the compounds of Formula I can be administered in vivo to children.
• Serum samples taken before and after such administration can be assayed for growth hormone. Comparison of the amounts of growth hormone in each of these samples would be a means for directly determining the ability of the patient's pituitary to release growth hormone.
• the present invention includes within its scope pharmaceutical compositions comprising, as an active ingredient, at least one of the compounds of Formula I in association with a pharmaceutical carrier or diluent.
• the active ingredient of the pharmaceutical compositions can comprise a growth promoting agent in addition to at least one of the compounds of Formula I or another composition which exhibits a different activity, e.g., an antibiotic or other
• Growth promoting agents include, but are not limited to, TRH, diethylstilbesterol, theophylline, enkephalins, E series prostaglan- dins, compounds disclosed in U.S. Patent No. 3,239,345, e.g., zeranol, and compounds disclosed in U.S. Patent No. 4,036,979, e.g., sulbenox or peptides disclosed in U.S. Patent No. 4,411,890.
• a still further use of the disclosed novel substituted dipeptide analogs is in combination with other growth hormone secretagogues such as GHRP-6, GHRP-1 as described in U.S. Patent Nos. 4,411,890; and publications WO 89/07110 and WO 89/07111 and B-HT920 or growth hormone releasing factor and its analogs or growth hormone and its analogs or somatomedins including IGF-1 and IGF-2.
• a still further use of the disclosed novel substituted dipeptide analogs is in combination with ⁇ 2 adrenergic agonists or ⁇ 3 adrenergic agonists in the treatment of obesity or in combination with parathyroid hormone or bisphosphonates, such as MK-217 (alendronate), in the treatment of osteoporosis.
• growth hormone As is well known to those skilled in the art, the known and potential uses of growth hormone are varied and multitudinous. Thus, the administration of the compounds of this invention for purposes of stimulating the release of endogenous growth hormone can have the same effects or uses as growth hormone itself.
• These varied uses of growth hormone may be summarized as follows: stimulating growth hormone release in elderly humans; Prevention of catabolic side effects of glucocorticoids, treatment of osteoporosis, stimulation of the immune system, treatment of retardation, acceleration of wound healing, accelerating bone fracture repair, treatment of growth retardation, treating renal failure or insufficiency resulting in growth retardation, treatment of physiological short stature, including growth hormone deficient children, treating short stature associated with chronic illness, treatment of obesity and growth retardation associated with obesity, treating growth retardation associated with Prader-Willi syndrome and Turner's syndrome; Accelerating the recovery and reducing
• the compounds of this invention can be administered by oral, parenteral (e.g., intramuscular, intraperitoneal, intravenous or subcutaneous injection, or implant), nasal, vaginal, rectal, sublingual, or topical routes of administration and can be formulated in dosage forms appropriate for each route of administration.
• parenteral e.g., intramuscular, intraperitoneal, intravenous or subcutaneous injection, or implant
• nasal, vaginal, rectal, sublingual, or topical routes of administration and can be formulated in dosage forms appropriate for each route of administration.
• Solid dosage forms for oral administration include capsules, tablets, pills, powders and granules.
• the active compound is admixed with at least one inert pharmaceutically acceptable carrier such as sucrose, lactose, or starch.
• Such dosage forms can also comprise, as is normal practice, additional substances other than inert diluents, e.g., lubricating agents such as magnesium stearate.
• the dosage forms may also comprise buffering agents. Tablets and pills can additionally be prepared with enteric coatings.
• Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups, the elixirs containing inert diluents commonly used in the art, such as water. Besides such inert diluents, compositions can also include adjuvants, such as wetting agents, emulsifying and suspending agents, and sweetening, flavoring, and perfuming agents.
• Preparations according to this invention for parenteral administration include sterile aqueous or non-aqueous solutions, suspensions, or emulsions.
• non-aqueous solvents or vehicles are propylene glycol, polyethylene glycol, vegetable oils, such as olive oil and corn oil, gelatin, and injectable organic esters such as ethyl oleate.
• Such dosage forms may also contain adjuvants such as preserving, wetting, emulsifying, and dispersing agents. They may be sterilized by, for example, filtration through a bacteria-retaining filter, by incorporating sterilizing agents into the compositions, by irradiating the compositions, or by heating the compositions. They can also be manufactured in the form of sterile solid compositions which can be dissolved in sterile water, or some other sterile injectable medium immediately before use.
• compositions for rectal or vaginal administration are preferably suppositories which may contain, in addition to the active substance, excipients such as cocoa butter or a suppository wax.
• compositions for nasal or sublingual administration are also prepared with standard excipients well known in the art.
• the dosage of active ingredient in the compositions of this invention may be varied; however, it is necessary that the amount of the active ingredient be such that a suitable dosage form is obtained.
• the selected dosage depends upon the desired therapeutic effect, on the route of administration, and on the duration of the treatment. Generally, dosage levels of between 0.0001 to 100 mg/kg. of body weight daily are administered to patients and animals, e.g., mammals, to obtain effective release of growth hormone.
• Zinc chloride (3.3 g, 24.3 mmol, 0.5 eq) was added to 15 mL of N,N-dimethylformamide in small portions while maintaining the temperature below 60°C.
• the suspension of zinc chloride was cooled to room temperature and treated with 5.0 g of benzonitrile (48.5 mmol, 1.0 eq) followed by 3.2 g of sodium azide (48.5 mmol, 1.0 eq).
• the heterogeneous mixture was heated at 115°C with agitation for 18 hours.
• the mixture was cooled to room temperature, water (30 mL) was added and the mixture acidified by the addition of 5.1 mL of concentrated hydrochloric acid.
• the mixture was cooled to 0°C and aged for one hour, then filtered and the filter cake washed with 15 mL of cold 0.1N HCl then dried at 60°C under vacuum to afford 6.38 g (43.7 mmol, 90%) of the product.
• Step B 5-Phenyl-2-trityltetrazole
• a solution of zinc chloride (6.3 g, 46.2 mmol, 0.6 eq) in 35 mL of tetrahydrofuran was dried over molecular sieves.
• 5-Phenyl-2- trityltetrazole (30.0 g, 77.3 mmol, 1.0 eq) was dissolved in 300 mL of dry tetrahydrofuran and the solution gently stirred while being degassed three times by alternating vacuum and nitrogen purges.
• the stirred solution was cooled to -15°C and treated slowly with 50.5 mL of 1.6 M n-butyllithium in hexane (80.0 mmol, 1.05 eq) so as to maintain the temperature below -5°C.
• the solution was maintained at -5 to -15°C for 1.5 hours then treated with the dried zinc chloride solution and allowed to warm to room temperature.
• the reaction mixture was stirred vigorously for 8 hours at room temperature then quenched by the slow addition of a solution of 10 mL of glacial acetic acid (1.6 mmol, 0.02 eq) in 60 mL of tetrahydrofuran at a rate so that the temperature was maintained below 40°C.
• the mixture was stirred for 30 minutes and 150 mL of 80% saturated aqueous sodium chloride was added; the reaction mixture was extracted for 30 minutes and the layers allowed to separate. The organic layer was removed and washed with 150 mL of 80% saturated aqueous sodium chloride buffered to pH >10 by the addition of ammonium hydroxide.
• the organic phase was removed and concentrated under vacuum to approximately 50 mL then 250 mL of acetonitrile was added. The mixture was again concentrated under vacuum to 50 mL and acetonitrile added to make the final volume 150 mL.
• the resulting slurry was cooled at 5°C for 1 hour then filtered and washed with 50 mL of cold acetonitrile followed by 150 mL of distilled water. The filter cake was air dried to a free flowing solid then further dried
• Step D N-Triphenylmethyl-5-[2-(4'-bromomethylbiphen4- yl]tetrazole
• Step E N-Triphenylmethyl-5-[2-(4'-azidomethylbiphen-4- yl]tetrazole
• Step F N-Triphenylmethyl-5-[2-(4'-aminomethylbiphen-4- yl]tetrazole
• Step G (R)- ⁇ -[t-Butoxycarbonylamino]-N-[[2'-(N-triphenyl- methyl-tetrazol-5-yl)[1,1'-biphenyl]-4-yl]methyl]benzene- butanamide
• Step F To a solution of 30.5 mg (0.11 mmol) of N-BOC-D- homophenylalanine in 1 mL of methylene chloride at room temperature under a nitrogen atmosphere was added 54 mg (0.11 mmol, 1 eq) of N- triphenylmethyl-5-[2-(4'aminomethylbiphen-4-yl)]tetrazole (Step F), 25 mg (0.13 mmol, 1.2 eq) of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and 13 mg (0.13 mmol, 1.2 eq) of triethylamine.
• Step H (R)- ⁇ -[t-Butoxycarbonylamin o]-N-[[2'-(1H-tetrazol-5- yl)[1 ,1 '-biphenyl]-4-yl ]methyl]-benzene-butanamide
• Step I (R)- ⁇ -Amino-N-[[2'-(1H-tetrazol-5-yl)[1,1 '-biphenyl]-
• Step J 2.2-Dimethylbutanedioic acid, 4-methyl ester
• 2,2-Dimethylsuccinic acid (20 g, 137 mmol) dissolved in 200 mL of absolute methanol at 0°C was treated dropwise with 2 mL of concentrated sulfuric acid. After the addition was complete, the mixture was allowed to warm to room temperature and stir for 16 hours. The mixture was concentrated under vacuum to 50 mL and slowly treated with 200 mL of saturated aqueous sodium bicarbonate. The mixture was washed with hexane (3x) and the aqueous layer removed and cooled in an ice bath. The mixture was acidified to pH 2 by slow addition of 6N HCl then extracted with ether (8x). The combined extracts were washed with brine, dried over magnesium sulfate, filtered and solvents removed under vacuum.
• Step K 3-Benzyloxycarbonylamino-3-methylbutanoic acid, methyl ester
• Step L 3-Benzyloxycarbonylamino-3-methylbutanoic acid
• a solution of 18.27 g (68.9 mmol) of 3-benzyloxycar- bonylamino-3-methylbutanoic acid methyl ester in 20 mL of methanol at room temperature was treated dropwise with 51 mL of 2N NaOH (102 mmol).
• the mixture was stined at room temperature for 16 hours then transferred to a separatory funnel and washed with hexane (3x).
• the aqueous layer was removed, cooled to 0°C and slowly acidified to pH 2 (paper) by dropwise addition of 6N HCl.
• Step M 3-Benzyloxycarbonylamino-3-methylbutanoic acid, N- hydroxysuccinimide ester
• Step N (R)- ⁇ -[(3-Benzyloxycarbonylamino-3-methyl-1-oxo- butyl)amino]-N-[[2'-(1H-tetrazol-5-yl)-[1 ,1 '-biphenyl]- 4yl]methyl]benzenebutanamide
• a solution of 12 mg (0.023 mmol) of the intermediate obtained in Step I in 0.5 mL of methylene chloride at room temperature was treated with 9.2 mg (0.027 mmol, 1.2 eq) of 3-benzyloxy- carbonylamino-3-methylbutanoic acid, N-hydroxysuccinimide ester (Step M) and 3.5 mg (0.027 mmol, 1.2 eq) of diisopropylethylamine.
• Step O (R)- ⁇ -[(3-Amino-3-methyl-1-oxobutyl)amino]-N-[[2'-(1H- tetrazol-5-yl)[1,1'-biphenyl]-4-yl]methyl]benzenebutana- mide. tri-fluoroacetate
• Step A (R)- ⁇ -[t-Butoxycarbonylamino]-N-[[2'-(N-triphenyl- memyl-tetrazol-5-yl)[1,1'-biphenyl]-4-yl]methyl]-(Nim- formyl)indole-3-propanamide
• Step B (R)-a-[t-Butoxycarbonylamino]-N-[[2'-(1H-tetrazol-5- yl)[1,1'-biphenyl]-4-yl]methyl]-(Nim-formyl)indole-3- propanamide
• Step C (R)- ⁇ -Amino-N-[[2'-(1H-tetrazol-5-yl)[1 ,1 '-biphenyl]-4- yl]methyl]-(Nim-formyl)indole-3-propanamide, trifluoroacetate
• a 3-neck 3L round bottom flask equipped with a magnetic stiner, thermometer, cold finger condenser and nitrogen bubbler was charged with 1L of ether.
• the flask was cooled to -65°C and into it was condensed 500-600 mL of isobutylene.
• the cold finger condenser was replaced with a dropping funnel and 200 mL (325 g, 2.30 mol) of chlorosulfonyl isocyanate was added dropwise over 1.5 hours.
• the mixture was maintained at -65°C for 1.5 hours then the dry ice/acetone cooling bath replaced with methanol/ice and the internal temperature slowly increased to -5°C at which time the reaction initiated and the internal temperature rose to 15°C with evolution of gas.
• the internal temperature remained at 15°C for several minutes then dropped back down to -5°C and the mixture stirred at -5°C for 1 hour.
• the internal temperature remained at 15°C for several minutes then dropped back down to -5°C and the mixture
• the reaction mixture was transferred to a 3-neck 12L round bottom flask fitted with a mechanical stiner and diluted with 2L of ether.
• the well-stined reaction mixture was treated with 2L of saturated aqueous sodium sulfite. After 1 hour, an additional 1L of saturated aqueous sodium sulfite was added followed by sufficient sodium bicarbonate to adjust the pH to approximately 7.
• the mixture was stirred another 30 minutes then the layers allowed to separate.
• the ether layer was removed and the aqueous layer reextracted with 2 x 1 L of ether.
• the combined ether extracts were washed once with 500 mL of saturated aqueous sodium bicarbonate and once with 500 mL of saturated aqueous sodium chloride.
• the ether layer was removed, dried over magnesium sulfate, filtered and concentrated under vacuum to give 33 g of a pale yellow oil.
• the aqueous layer was made basic by the addition of solid sodium bicarbonate and extracted with 3 x 1L of ether. The combined ether extracts were washed and dried as described above, then combined with the original 33 g of pale yellow oil and concentrated under vacuum to give 67.7 g of product. Further extraction of the aqueous layer with 4 x 1L of methylene chloride and washing and drying as before gave an additional 74.1 g of product. Still further extraction of the aqueous layer with 4 x 1L of methylene chloride gave an additional 21.9 g of product.
• reaction mixture was stined at room temperature overnight then diluted with 1L of methylene chloride and washed with 500 mL of saturated aqueous ammonium chloride, 500 mL of water, and 500 mL of saturated aqueous sodium chloride.
• the organic layer was separated, dried over magnesium sulfate, filtered and concentrated under vacuum to afford 180.3 g of crude product as an orange solid.
• the aqueous layer was acidified by the addition of 1L of saturated aqueous sodium bisulfate, then extracted with 1 x 1L and 2 x 500 mL of ether. The combined organic layer and ether extracts were washed with 500 mL of saturated aqueous sodium
• Step G (R)- ⁇ -[(3-t-Butoxycarbonylamino-3-methyl-1-oxo- butyl)amino]-N-[[2'-(1H-tetrazol-5-yl)-[1,1'-biphenyl]-4- yl ]methy] - ( N im -formyl)-indole-3-propanamide
• reaction mixture was evaporated to dryness under vacuum and the residue was dissolved in 1 mL of anisole and treated with 4 mL of trifluoroacetic acid.
• the mixture was stined at room temperature for 30 minutes the concentrated under vacuum.
• the residue was taken up in methanol and the solids removed by filtration.
• the filtrate was concentrated under vacuum; the residue was purified by reverse phase high pressure liquid chromatography on C18 to yield 31 mg (42%) of the product.
• Step H (R)- ⁇ -[(3-Amino-3-methyl-1-oxobutyl)amino]-N-[[2'-(1H- tetrazol-5-y1)[1,1'-biphenyl]-4-yl]methyl]-1H-indole-3- propanamide, trifluoroacetate
• Step A (R)-2-(t-Butoxycarbonylamino)-N-phenyl-butanamide Prepared from N-BOC-D-2-aminobutanoic acid and aniline by the method described in Example 1, Step G.
• 1 H NMR 200 MHz, CDCl 3 ): 1.00 (t, 6Hz, 3H), 1.45 (s, 9H), 1.82 (m, 2H), 4.30 (m, 1H), 5.59 (d, 7Hz, 1H), 7.00-7.62 (m, 5H), 8.90 (br s, 1H).
• Step B (R)-2-(t-Butoxycarbonylamino)-N-phenyl-N-[[2'-(N- triphenylmethyl-tetrazol-5-yl)-[1,1'-biphenyl]-4-yl]methyl]- butanamide
• the mixture was stirred at room temperature for 2 hours, then quenched by the addition of 2 mL of water.
• the mixture was extracted several times with ethyl acetate; the combined extracts were washed with 5% aqueous citric acid, saturated aqueous sodium bicarbonate then dried over magnesium sulfate, filtered and the filtrate dried under vacuum.
• the crude material was chromatographed on a flash silica column, eluting with hexane/ethyl acetate (1:1), to give 91 mg (48%) of the product.
• Step C (R)-2-(t-Butoxycarbonylamino)-N-phenyl-N-[[2'-(1H- tetrazol-5-yl)-[1,1'-biphenyl]-4-yllmethyl]butanamide
• Step D (R)-2-Amino-N-phenyl-N-[[2'-(1H-tetrazol-5-yl)[1,1'- biphenyl]-4-yl]methyl]butanamide, hvdrochloride
• Step E 3-t-Butoxycarbonylamino-3-methylbutanoic acid, N- hydroxysuccinimide ester
• Step F (R)-2-[(3-t-Butoxycarbonylamino-3-methyl-1-oxobutyl)- amino]-N-phenyl-N-[[2'-(1H-tetra-zol-5-yl)[1,1'-biphenyl]-
• Step G (R)-2-[(3-Amino-3-methyl-1-oxobutyl)amino]-N-phenyl-N- [[2'-(1H-tetrazol-5-yl)[1,1 '-biphenyl]-4-yl]methyl]- butanamide, trifluoroacetate
• Step A 2-t-Butoxycarbonylamino-N-[[2'-(N-triphenylmethyl- tetrazol-5-yl)[1,1'-biphenyl]-4-yl]-methyl]acetamide
• Step B 2-t-Butoxycarbonylamino-N-[[2'-(1H-tetrazol-5-yl)[1 ,1'- biphenyl]-4-yl]methyl]acetamide
• Step C 2-Amino-N-[[2'-(1H-tetrazol-5-yl)[1,1'-bi-phenyl]-4- yl]methyl]acetamide, hydrochloride
• Step D 2-[(3-Benzyloxycarbonylamino-3-methyl-1-oxo-butyl)- amino]-N-[[2'-(1H-tetrazol-5-yl)[1,1'-biphenyl]-4yl]- methyl]acetamide
• Step E 2-[(3-Amino-3-methyl-1-oxobutyl)amino]-N-[[2'-(1H- tetrazol-5-yl)[1 ,1'-biphenyl]-4-yl]methyl]acetamide, trifluoroacetate
• Step A N-(t-Butoxycarbonyl)-D-tryptophan benzyl ester
• Finely divided t-butoxycarbonyl-D-tryptophan (3 g, 10 mmol) was suspended in methylene chloride and benzyl alcohol (1.08 mL, 10 mmol) and 4-dimethylaminopyridine (0.12 g, 1 mmol) were added and stined at room temperature.
• Solid 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (1.92 g, 10 mmol) was then added in three roughly equal portions over 5 minutes. The reaction mixture was stined for 3 hours at room temperature during which time the reaction mixture became a homogeneous solution.
• the reaction mixture was poured into water (100 mL) and extracted with methylene chloride (2 x 50 mL). The combined methylene chloride layers were washed with 5% aqueous citric acid solution (100 mL) and 5% aqueous sodium bicarbonate solution (100 mL). The resulting methylene chloride layer was dried over magnesium sulfate, filtered and
• Step B D-Tryptophan benzyl ester
• N-(t-Butoxycarbonyl)-D-tryptophan benzyl ester (3.5 g, 8.87 mmol) was dissolved in methylene chloride (10 mL) and stined at room temperature and trifluoroacetic acid (20 mL) was added dropwise to the ester. The reaction mixture was stined at room temperature for one hour during which time the reaction darkened. The reaction mixture was directly evaporated under vacuum to give a white solid. This solid was dissolved in chloroform (100 mL) and washed with saturated aqueous sodium bicarbonate. The aqueous layer was extracted with chloroform (2 x 25 mL) and the combined chloroform layers were dried over potassium carbonate. Filtration and concentration of the chloroform solution under vacuum gave a pale yellow oil (3.14 g, 82%) which was mainly the desired product.
• Step C (R)- ⁇ -[(2-t-Butoxycarbonylamino-2-methyl-1-oxopropyl)- amino]-1H-indole-3-propanoic acid, benzyl ester
• the chloroform layer was separated and washed with 5% aqueous citric acid solution (25 mL) and 5% aqueous sodium bicarbonate solution (25 mL).
• the chloroform layer was dried over magnesium sulfate, filtered and evaporated under vacuum to afford a thick oily foam. Chromatography on silica gel using ethyl acetate/hexanes (2:3 v/v) afforded a yellow foam (0.822 g 50%).
• Step D (R)- ⁇ -[(2-t-Butoxycarbonylamino-2-methyl-1-oxopropyl)- amino]-1H-indole-3-propanoic acid
• the benzyl ester (0.82 g, 1.71 mmol) obtained in Step C and 10% palladium on carbon (150 mg) were stined together in ethyl acetate (5 mL).
• the solution was degassed and a hydrogen atmosphere introduced over the reactants using a balloon for 32 hours.
• the reaction products were isolated by filtering the reaction mixture through a Celite plug. The plug was washed with additional ethyl acetate (3 x 10 mL). The combined filtrates were evaporated under vacuum to afford the product (680 mg, 102%).
• Step E 4-Methyl-2'-nitro-1 ,1 '-biphenyl
• Step F 4-Bromomethyl-2'-nitro-1 ,1'-biphenyl
• Step G 4-Azidomethyl-2'-nitro-1 ,1'-biphenyl
• Step H 4-Aminomethyl-2'-nitro-1 ,1 '-biphenyl
• Step I (R)- ⁇ -[(2-t-Butoxycarbonylamino-2-methyl-1-oxopropyl)- amino]-N-[[(2'-nitro)[1,1'-biphenyl]-4-yl]methyl]-1H- indole-3-propanamide
• triethylamine (0.245 mL, 1.76 mmol) were dissolved in methylene chloride (8 mL) and stined at room temperature.
• Benzotriazolyl-1- yloxytris(dimethylamino)phosphonium hexafluorophosphate (388 mg, 0.87 mmol) was added in a single portion.
• the reaction mixture was stined togerther for 3.5 hours then the reaction was quenched by adding saturated aqueous sodium chloride (10 mL) and extracted with methylene chloride (3 x 20 mL). The combined extracts were dried over magnesium sulfate, filtered and evaporated under vacuum.
• Step J (R)- ⁇ -[(2-t-Butoxycarbonylamino-2-methyl-1-oxopropyl)- amino]-N-[[(2'-amino)[1,1'-biphenyl]-4-yl]methyl]-1H- indole-3-propanamide
• the amide (142 mg, 0.24 mmol) from Step I was dissolved in ethanol (5 mL) and 10% palladium on carbon (15 mg) was added.
• the ethanolic mixture was degassed and a hydrogen atmosphere introduced and maintained above the reaction mixture for 2.5 hours using a balloon.
• the hydrogenation catalyst was removed by filtration through a Celite pad. The pad was washed carefully with methylene chloride (4 x 5 mL). The combined filtrates were evaporated under vacuum to give a powdery white foam (124 mg, 92%).
• Step K (R)- ⁇ -[(2-t-Butoxycarbonylamino-2-methyl-1-oxopropyl)- amino]-N-[[2'-[[(methylamino)-carbonyl]amino][1 ,1 '- biphenyl]-4-yl]methyl]-1H-indole-3-propanamide
• Step L (R)- ⁇ -[(2-Amino-2-methyl-1-oxopropyl)amino]-N-[[2'-
• Step K The intermediate obtained in Step K (15 mg, 0.023 mmol) and anisole (0.01 mL, 0.09 mmol) were dissolved in methanol (0.5 mL) and hexanes (0.5 mL). To this solution 0.5 mL of 9 N aqueous hydrochloric acid was added. The reactants were stined at room temperature for 30 minutes then the hexane layer was removed using a pipette. The aqueous methanolic layer was evaporated at atmospheric pressure using a fast stream of nitrogen at room temperature. The solid material thus obtained was purified by reverse phase medium pressure liquid chromatography on C8, eluting with methanol/0.1% aqueous trifluoroacetic acid (85:15 v/v).

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• 1993
  • 1993-10-29 JP JP6512178A patent/JPH08503213A/ja active Pending
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  • 1993-10-29 EP EP94900505A patent/EP0669830A4/en not_active Withdrawn
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