CA2081890A1 - Cyclized and linear therapeutic peptides - Google Patents
Cyclized and linear therapeutic peptidesInfo
- Publication number
- CA2081890A1 CA2081890A1 CA002081890A CA2081890A CA2081890A1 CA 2081890 A1 CA2081890 A1 CA 2081890A1 CA 002081890 A CA002081890 A CA 002081890A CA 2081890 A CA2081890 A CA 2081890A CA 2081890 A1 CA2081890 A1 CA 2081890A1
- Authority
- CA
- Canada
- Prior art keywords
- amino acid
- therapeutic peptide
- ala
- phe
- peptide
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Abandoned
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
- C07K7/04—Linear peptides containing only normal peptide links
- C07K7/08—Linear peptides containing only normal peptide links having 12 to 20 amino acids
- C07K7/086—Bombesin; Related peptides
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
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- Chemical & Material Sciences (AREA)
- Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- General Health & Medical Sciences (AREA)
- Medicinal Chemistry (AREA)
- Life Sciences & Earth Sciences (AREA)
- General Chemical & Material Sciences (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Molecular Biology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Biochemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Biophysics (AREA)
- Genetics & Genomics (AREA)
- Pharmacology & Pharmacy (AREA)
- Animal Behavior & Ethology (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Peptides Or Proteins (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
Abstract
A therapeutic peptide comprising between seven and ten amino acid residues, inclusive, the peptide being an analog of one of the following naturally occuring peptides which terminate at the carboxy-terminus with a Met residue: (a) litorin; (b) the ten amino acid carboxy-terminal region of mammalian GRP, neuromedin B, or neuromedin C; and (c) the ten amino acid carboxy-terminal region of amphibian bombesin, said analog being an agonist of one of the naturally occurring peptides.
Description
~- WO91/17181 2 a 8 ~ PCT/US91/03265 ~, CYCLIZED AND LINEAR THERAPEUTIC PEPTIDES
Back~round of the Invention This inv~ention relates to peptides useful for treatment of benign or malignant proliferation of tissue.
The amphibian peptide bombesin, pGlu-Gln-Arg-Leu-Gly-Asn-Gln-Trp-Ala-Val-Gly-His-Leu-Met-NH2 :
~Anastasi et al., Experientia 27:166-167 (1971)), is closely related to the mammalian gastrin-releasing peptides (GRP), e.g., the porcine GRP, H2N-Ala-Pro-Val-Ser-Val-Gly-Gly-Gly-Thr-Val-Leu-Ala-Lys-Met-Tyr-Pro-Arg-Gly-Asn-His-Trp-Ala-Val-Gly-His-Leu-Met-(NH2) (McDonald et al., Biochem.
Biophys. Res. Commun. 90:227-233 (1979)) and human GRP, H2N-Val-Pro-Leu-Pro-Ala-Gly-Gly-Gly-Thr-Val-Leu-Thr-Lys-M
e t-Tyr-Pro-Arg-Gly-Asn-His-Trp-Ala-Val-Gly-His-Leu-Met ~ (NH2). Bombesin has been found to be a growlth factor i ~or a number of human cancer cell lines, including small-cell lung carcinoma (SCLC), and has been detected in human breast and prostate cancer ~Haveman et al., eds. Recent Results }~ Cancer Research - Pe~tide Hormones in u~a Cancer, Springer-Verlag, New York:1986). A number of these cancers are Xnown to secrete peptide hormones related to GRP or bombesin.
Consequently, antagonists to bombesin have been proposed ~ as agents for the treatment of these cancers.
,l Cuttitta et al. demonstrated that a specific monoclonal antibody to bombesin inhibited in vivo the growth of a human small-cell lung cancer cell line xenografted to nude mice (Cuttitta et al., Cancer Survey 4:707-727 (1985)). In 3T3 murine fibroblasts which are responsive to the mitotic effect of bombesin, Zachary l and Rozengurt observed that a substance P antagonist ;~ (Spantide) acted as a bombesin antagonist (Zachary et al., Proc. Natl. Acad. Sci. (USA), 82:7616-7620 (1985)).
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SUB~TIT~T~ SHEET
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Back~round of the Invention This inv~ention relates to peptides useful for treatment of benign or malignant proliferation of tissue.
The amphibian peptide bombesin, pGlu-Gln-Arg-Leu-Gly-Asn-Gln-Trp-Ala-Val-Gly-His-Leu-Met-NH2 :
~Anastasi et al., Experientia 27:166-167 (1971)), is closely related to the mammalian gastrin-releasing peptides (GRP), e.g., the porcine GRP, H2N-Ala-Pro-Val-Ser-Val-Gly-Gly-Gly-Thr-Val-Leu-Ala-Lys-Met-Tyr-Pro-Arg-Gly-Asn-His-Trp-Ala-Val-Gly-His-Leu-Met-(NH2) (McDonald et al., Biochem.
Biophys. Res. Commun. 90:227-233 (1979)) and human GRP, H2N-Val-Pro-Leu-Pro-Ala-Gly-Gly-Gly-Thr-Val-Leu-Thr-Lys-M
e t-Tyr-Pro-Arg-Gly-Asn-His-Trp-Ala-Val-Gly-His-Leu-Met ~ (NH2). Bombesin has been found to be a growlth factor i ~or a number of human cancer cell lines, including small-cell lung carcinoma (SCLC), and has been detected in human breast and prostate cancer ~Haveman et al., eds. Recent Results }~ Cancer Research - Pe~tide Hormones in u~a Cancer, Springer-Verlag, New York:1986). A number of these cancers are Xnown to secrete peptide hormones related to GRP or bombesin.
Consequently, antagonists to bombesin have been proposed ~ as agents for the treatment of these cancers.
,l Cuttitta et al. demonstrated that a specific monoclonal antibody to bombesin inhibited in vivo the growth of a human small-cell lung cancer cell line xenografted to nude mice (Cuttitta et al., Cancer Survey 4:707-727 (1985)). In 3T3 murine fibroblasts which are responsive to the mitotic effect of bombesin, Zachary l and Rozengurt observed that a substance P antagonist ;~ (Spantide) acted as a bombesin antagonist (Zachary et al., Proc. Natl. Acad. Sci. (USA), 82:7616-7620 (1985)).
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SUB~TIT~T~ SHEET
... .. -.. - - , . .. - . ~ . . , . .. . .. ~ .. . . . . - . .. . ... .
.;. . . . .. .... . . .. - ., . ,.. ..... . . .. .. .. .. . . .. ... ~. ...... . . . . .. ... . .. ... . . . .
2 Q ~ PCT/US91/0326;~
Heinz-Erian et al. replaced His at position 12 in bombesin with D-Phe and observed bombesin antagonist activity in dispersed acini from guinea pig pancreas (Heinz-Erian et al., Am. J. of Physiol. 2S2:G439-G442 (1987)). Rivier reported work directed toward restricting the conformational freedom of the bioactive C-terminal decapeptide of bombesin by incorporating intramolecular disulfide bridges; however, Rivier mentioned that, so far, bombesin analogs with this modification fail to exhibit any antagonist activity (Rivier et al., "Competitive Antagonists of Peptide Hormones," in Abstracts of the International Symposium on Bombesin-Like Peptides in Health and Disease, Rome, Italy (October, 1987).
Bombesin exhibits both direct and indirect effects on the gastrointestinal tract, including the release of hormones and the stimulation of pancreatic, gastric, and intestinal secretion and of intestinal mobility.
Gastrin and cholecystokinin (CCK) which are released by bombesin, have béen shown to play a role in the maintenance of normal gastrointestinal mucosa as well as in augmenting growth of normal and neoplastic tissues.
The growth of xenografted human colon and stomach carcinomas in nude mice has been stimulated by the , 25 administration of gastrin and later inhibited with the addition of secretin (Tanake et al., 1986, Tokaku J.
Exp. Ned. 148:459j and the growth of MC-26 murine colon carcinoma, which possesses gastrin receptors is ~ stimulated by pentagastrin (Winsett et al., 1980, d~ 30 Surgery 99:302, and inhibited by proglumide, a x-~ gastrin-receptor antagonist, Beauchamp et al., 1985, Ann. Surg. 202:303. Bombesin has been found to act concurrently as both a trophic agent for normal host ;~ pancreas and a growth inhibitory agent in xenografted ... .
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WO91/17181 2 0 3 1 ~ ~ ~ PCT/US9l,0326~
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human pancreatic tumor tissue, Alexander et al., 1988, Pancreas 3:247.
Abbreviations (uncommon3: ~
cyclohexyl-Ala = CHx-Ala (cyclohexyl alanine) ,.~-H
,. C CH2 identifying group ;
, CH
NH2--CH--C02H; : ' pGlu = H2C---CH-COOH (pyroglutamic acid);
, H2C NH
r C
~ Nle = H2N-CH-COOH (norleucine) ;~ (CH2)3-CH3 'Ss Pal - 3-pyridyl-alanine D-Cpa - D-para-chloro-phenylalanine HyPro = hydroxyproline Nal - naphthylalanine .
Sar = sarcosine F5-Phe s penta-fluoro-phenylalanine R = right (D) configuration; S = left (L) configuration; A.
~ 25 racemate = equal mix of R and S
.1 l-methyl-His; 3-methyl-His = methyl (CH3) group on -~-.j~ nitrogen at positions l or 3 of Histidine: -;;
.~ .
, ~ ~ C__NlH+ ., ,~j 30 3 C--N H
HCH
;~ NH2----CH----COOH .' "~: Summarv of the Invention -- :
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~` SUBSIITUTE SHET
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WO91/1~181 2 ~ , 9 ~ PCT/US91/0326 The invention features a linear or a cyclic therapeutic peptide, which includes between seven and ten amino acid residues, inclusive, and which is an . analog of one of the following naturally occurring -:
peptides which terminate at the carboxy-terminus with a : Met residue: (a) litorin; (b) the ten amino acid carboxy-terminal region of mammalian GRP, neuromedin B, or neuromedin C; and (c) the ten amino acid carboxy-terminal region of amphibian bombesin, which is an analog of the formula:
R
AO_Al--A2--A3--A4 -AS -A6 -A7--A3 -A9 -R3 R
wherein . 15 A = Gly, D- or L- isomer of any of pGlu, Nle, a-aminobutyric acid, Ala, Val, Gln, Asn, Leu, Ile, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, CH3), Trp, or ~-Nal, or is deleted;
l = the D- or L-isomer of any of pGlu, Nle, j 20 a-aminobutyric acid, Ala, Val, Gln, Asn, Leu, Ile, p-X-Phe (where X - H, F, Cl, Br, NO2, OH, or CH3), Asp, Glu, F5-Phe, Trp, ~-Nal, Cys, Lys, or is deleted;
A2 = Gly, D- or L- isomer of any of pGlu, Ala, Val, :~ 25 Gln, Asn, Leu, Ile, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Trp, ~-Nal, Asp, Glu, His, l-methyl-His 3-methyl-His, Cys, Lys, or is i~ deleted; A3 = the D- or L-isomer of any of p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), ~-Nal, or Trp;
A~ = Ala, Val, Gln, Asn, Gly, Leu, Ile, Nle, : a-aminobutyric acid, p-X-phe (where X = H, F, Cl, Br, NO2, OH, or ,CH3), Trp, or ~-Nal;
.
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~ ` SUBSI~ TE S~'~T
;. - . ' ~ ' ,;'. .'' . :' . ,: , ' ', , "' " ' . :
W091/17181 2 ~ PCT/US91/03265 A5 = Gln, Asn, Gly, Ala, Leu, Ile, Nle, a-aminobutyric acid, Val, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Trp, Thr, or ~-Nal; .
A6 = Sar, Gly or the D-isomer of any Ala, N-methyl-Ala, Val, Gln, Asn, Leu, Ile, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Trp, Cys, or ~-Nal; :.
A7 - l-methyl-His, 3-methyl-His, His, Lys, Asp, or Glu;
A8 = Leu, Ile, Val, Nle, a-aminobutyric acid, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or C~3), Trp, Thr, ~-Nal, Lys, Asp, Glu, CHx-Ala, or Cys;
A9 = L-isomer of any of Met, Met-oxide, Leu, Ile, Nle, :: .
a-aminobutyric acid, p-X-Phe (where X = H, F, Cl, -Br, N02, OH, or CH3), Trp, ~-Nal, CHx-Ala, or ~i : Cys;
` each Rl and R2, independently, is H, Cl_l2 alkyl, C7_l0 .-p y lkyl, COEl ~where El is Cl_20 alkyl, C3 20 alkenyl C3_20 alkinyl, phenyl, naphthyl, or C7_l0 phenylalkyl), or Cl-Cl2 acyl, and Rl and R2 are bonded to the N-terminal amino acid o~ the peptide; provided that when one of Rl ..
.~ or R2 is COEl, the other must be H; and R3 is H, NH2, Z Cl_l2 alkyl, C~_lO phenylalkyl, or C3_20 naphthylalkyl;
`~ and further provided that, if Ao is present, Al cannot be pGlu; and, if A or Al is present, A2 cannot be pGlu;
~1 and further provided that, when A is deleted and Al is -` pGlu, Rl must be H and R2 must be the portion of Glu that .
1 forms the imine ring in pGlu; and further provided that, .~ where A is deleted and Al is not pGlu, Al may be bonded to A9, or where A and Al are deleted and A2 is not pGlu, A2 may be bonded to A9, or where A, Al and A2 are deleted, A3 can be bonded to A9 to form a cyclized ~ .
peptide; and provided that where AO is deleted and Al is Asp or Glu, or where A and Al are deleted and A2 is Asp .;~ .
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SU~S~ T~ SHEET
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WO91/17181 ~t ~ Pcr/usg1/o3265a~
or Glu, either Al or A2 can be bonded with A7 or A8, where A7 or A8 is Lys, or where A is deleted and Al is Lys or A0 and Al are deleted and A2 is Lys, either Al or A2 can be bonded to A7 or A8, where A7 or A8 is Asp or Glu; and further provided that either one of Al or A~ can be Cys and can be bonded through a disulfide bridge with either A8 or A9, provided that either one of A8 or A9 can .: be Cys and can be bonded through a disulfide bridge with either Al or A2; and further provided that where A and Al are deleted and A6 is D-Ala, A8-A9 cannot be Leu-Met-NH2; or a pharmaceutically acceptable salt .
thereof.
i~ In the generic formula given above, when either of Rl or R2 is an aliphatio, aromatic, or lipophilic group, the n vlvo activity can be long lasting, and -1. delivery of the compounds of the invention to the target tissue can be facilitated.
More pre~erably, the analog of the generic formula above is of the formula:
A = pGlu, Gly, D-Phe, or is deleted;
.~ Al = pGlu, D-Phe, D-Ala, D-~-Nal, D-Cpa, D-Asn, Cys, :: or i8 deleted;
~ A2 , pGlu, Asn, Gln, His, l-methyl-His, 3-methyl-His, ... . .
Cys, or is deleted;
2S A3 = Trp;
.~ A4 = Ala;
,.,r A5 = Val;
Ah = Sar, Gly, D-Phe, or D-Ala;
A7 = His;
. 30 A = Leu, Phe, Chx-Ala, or Cys;
-~ A9 = L-isomer of any of Met, Leu, Ile, Nle, Phe, or ;, Cys.
~` Examples of preferred peptide analogs are~
pGlu-Gln-Trp-Ala-Val-Gly-His-Leu-Leu-NH2;
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``` SUB~ITUTF S~ET
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- ` --. - - . , ~ . ..... ... ... ... . .. ..... ..... ....... .... ... .
... .. . . ~ . . . ..
. . . - . . . . . . . . . ` . . . . .
~ wO g-/17181 2 0 8 ~ ~ 9 ~ PCT/US91/03265 ~, D-Phe-Gln-Trp-Ala-Val-Gly-His-Leu-Leu-NH2;
D-Phe-Gln-Trp-Ala-Val-Gly-His-Leu-Met-NH2;
D-Cpa-Gln-Trp-Ala-Val-Gly-His-Leu-Met-NH2;
D-Cpa-Gln-Trp-Ala-Val-Gly-His-Leu-Leu-NH2;
D-Phe-Gln-Trp-Ala-Val-D-Ala-His-Leu-Leu-NH2;
D-Phe-Gln-Trp-Ala-Val-D-Ala-His-Leu-Met-NH2;
' D-Cpa-Gln-Trp-Ala-Val-D-Ala-His-Leu-Met-NH2; . :
; pGlu-Gln-Trp-Ala-Val-Gly-His-Phe-Leu-NH2; :
D-Phe-Gln-Trp-Ala-Val-Gly-His-Phe-Leu-NH2;
D-Phe-Gln-Trp-Ala-Val-D-Ala-His-Phe-Met-NH ; ~:
.~ 2 ...
D-Phe-Gln-Trp-Ala-Val-D-Ala-His-Phe-Leu-NH2; !',:' D-Phe-Gln-Trp-Ala-Val-Gly-His-CHx-Ala-Leu-NH2;
D-Phe-Gln-Trp-Ala-Val-Gly-His-Leu-Nle-NH2;
:~ D-Phe-Gln-Trp-Ala-Val-D-Ala-His-Leu-Nle-NH2; ~-.
D-Phe-Gln-Trp-Ala-Val-Gly-His-Phe-Nle-NH2;
D-Phe-Gln-Trp-Ala-Val-D-Ala-His-Phe-Nle-NH2;
Ac-His-Trp-Ala-Val-D-A~la-His-Leu ~ :
-Leu-NH2 cyclo-D-Phe-Gln-Trp-Ala-Val-Gly-His-Leu-Leu;
~: :
.~ 20 D-Cys-Asn-Trp-Ala-Val-Gly-His-Leu-Cys-NH2;
" .
i;~ cyclo-His-Trp-Ala-Val-Gly-His-Leu-Met;
, .' ' j~l Cys-Trp-Ala-Val-Gly-His-Leu-Cys-NH2; :. .-~`3 `~ :
~I, cyclo-D-Phe-Gln-Trp-Ala-Val-Gly-His-Leu-Met; ..
; cyclo-D-Phe-His-Trp-Ala-Val-Gly-His-Leu-Met;
. .~ .
25 and cyclo-Trp-Ala-Val-Gly-His-Leu-Met;
. .~. .
~, :. -. ,;~
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~ SUBSTI~IuTE SHEET
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., , .. ;~.` i .. , . .... . , , . .,, ....... , . ; ...... ,, . , , ., .. . , , .,,, - , ,.
..... ... .. .. .. - . , ., . - ,` . ..... . ;. . , ~; .... . , .. ~ . ,- .
WO91/17181 2 0 318 g ~ PCT/US91/03265 Modification of a natural amino acid or substitution of a synthetic amino acid, examples of which are (but are not limited to) pGlu, Nle, Pal, D-Cpa, Met-oxide, Nal, Sar, l-methyl-His, or 3-methyl-His, at one or more of positions A, Al, A6, or A9 of an analog, or cyclization of the analog result in enhanced stability; i.e., protection of the amino terminus from exopeptidase degradation and of the internal portion of the peptide from endopeptidase degradation. The carboxy-terminal dipeptide sequence of some analogs of the invention corresponds to the natural ~ C-terminal sequence of bombesin or bombesin-related -~ analogs.
The invention also features a linear or a cyclic therapeutic peptide, which includes between seven and ten amino acid residues, inclusive, and which is an analog of one of the following naturally occurring - peptides which terminate at the carboxy-terminus with a Met residue: ~a) litorin; (b) the ten amino acid carboxy-terminal region of mammalian GRP, neuromedin B, or neuromedin C; and (c) the ten amino acid ~b carboxy-terminal region of amphibian bombesin, and the analog is an agonist of one of these naturally occurring peptides.
~ 25 In preferred embodiments, the analog may be an ;~ agonist or a partial agonist of the naturally occurring ,- biologically active peptide; preferably, the analog is -~
;~ at least 25%, more preferably 50~ or 75%, homologous with a region of the naturally occurring peptide. As used herein, an "agonist" mimics or enhances the - biological effect of the natural peptide on its target cell and a "partial agonist" mimics or enhances the ~-~ biological effect of the natural peptide, but to a lesser extent than an agonist. Biological effect, as used herein, is measured by the effect of the natural ~, .. .
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SUBSi'Til~E SHEET
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~WO91/17181 ~ PCT/US91/03265 .. ' ~, g :' ,:
peptide in one of two systems: an in vitro pancreatic amylase release assay and an in vitro 3T3 fibroblast cell division system, ~oth of which are described in European Patent Application 88308916.6, hereby -;
incorporated by reference. An agonist will stimulate ;~
the effect of the natural peptide on either amylase release from pancreatic cells or fibroblast cell division by 100%, whereas a partial agonist will have a lesser stimulatory effect, i.e., ranging between 0-99%.
, .
Peptides of the invention are useful for treating non-malignant proliferative disease in a human patient, e.g., the proliferation of smooth muscle. Peptides of the invention are also useful for treating cancer in a human patient, particularly for the treatment of prostatic, colon, breast, pancreatic, or lung cancer.
In addition, peptides of the invention may be used to suppress appetite, to stimulate pancreatic secretion, or to suppress a craving for alcohol.
Analogs of the invention can be provided in the ~1, 20 ~orm of pharmaceutically acceptable salts. Examples of ;' pre~erred 5alts are those with therapeutically ;~ acceptable organic acids, e.g., acetic, lactic, maleic, - citric, malic, ascorbic, succinic, benzoic, salicylic, methanesulfonic, toluene sulfonic, trifluoroacetic, or pamoic acid, as well as polymeric acids such as tannic acid or carboxymethyl cellulose, and salts with -inorganic acids such as the hydrohalic acids,e.g., , hydrochloric acid, sulfuric acid or phosphoric acid.
Other features and advantages of the invention will be apparent from the following description of the -i~ preferred embodiments thereof, and from the claims.
-~ Descri~tion of the Preferred Embodiments Before describing preferred embodiments of the ~, invention, the drawing will briefly be described.
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S~BSTITU~E SHEET
W091/17181 ~ PCT/US91/0326a Drawina Fig. 1 is the amino acid sequences of naturally occurring peptides of which peptides of the invention are analogs.
we now describe the structure, synthesis, and use of the preferred embodiments of the invention.
Structure Peptides of the invention are derived from one of the sequences shown in Fig. 1, which represent the sequences, or portions thereof, of naturally-occurring peptides. Bombesin, neuromedin B, neuromedin C, litorin, and GRP analogs of the invention are described in Coy et al., U.S. Patent Application Serial No.
502,438, filed March 30, 1990, which is a --continuation-in-part of U.S. Patent Application Serial No. 397,169, filed August 21, 1989, which is a s continuation-in-part of U.S. Patent Application Serial r No. 376,555, filed July 7, 1989, and U.S. Patent ~ Application Serial Nu~ber 394,727, filed August 16, -j 20 1989, both of which are continuation-in-parts of U.S. ~5 Patent Application Serial No. 317,941, filed March 2, 1989, which is a continuation-in-part of U.S. Patent Application Serial Number 282,328, filed December 9, 1988, which in turn is a continuation-in-part of U.S.
Patent Application Serial No. 257,998, filed October 14, 1988, which in turn is a continuation-in-part of U.S.
~ Patent Application Serial No. 248,771, filed September 'i~ 23, 1988, which in turn is a continuation-in-part of Coy et al., U.S. Patent Application Serial No. 207,759, '~ 30 filed June 16, 1988j which in turn is a -~'~ continuation-in-part of Coy et al., U.S. Patent Application Serial No. 204,171, filed June 8, 1988, ~ -which in turn is a continuation-in-part of Coy et al., U.S. Patent Application Serial No. 173,311, filed March i , , .
~ 35 25, 1988, which in turn is a continuation-in-part of Coy '~
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SUBSli~UT~ ~IEET
i WO91/17181 2 0 g 1 ~ ~ ~ PCT/US91/0326~
et al. U.S. Patent Application Serial No. 100,571, filed - September 24, 1987; all of which are assigned to the same assignee and hereby incorporated by reference; or -as described in Zachary et al., Proc. Nat. Aca. Sci.
82:7616, 1985; Heimbrook et al., "Synthetic Peptides:
Approaches to Biological Problems", UCLA Symposium on Mol. and Cell. Biol. New Series, Vol. 86, ed. Tam and ; Kaiser; Heinz-Erian et al., Am. J. Physiol. G439, 1986;
Martinez et al., J. Med. Chem. 28:1874, 1985; Gargosky ; 10 et al., Biochem. J. 247:427, 1987; Dubreuil et al., Drug ' Design and Delivery, Vol 2:49, Harwood Academic Publishers, GB, 1987; Heikkila et al., J. Biol. Chem.
262:16456, 1987; Caranikas et al., J. Med. Chem.
25:1313, 1982; Saeed et al., 1989, Peptides 10:597;
i 15 Rosell et al., Trends in Pharmacological Sciences 3:211, 1982; Lundberg et al., Proc. Nat. Aca. Sci. ~0:1120, 1983; Engberg et al., Nature 293:222, 1984; Mizrahi et al., Euro. J. Pharma. 82:101, 1982; Leander et al., Nature 294:467, 1981; Woll et al., Biochem. Biophys.
Res. Comm. 155:359, 1988; Rivier et al., Biochem.
~; 17:1766, 1978; Cuttitta et al., Cancer Surveys 4:707, 1985; Aumelas et al., Int. J. Peptide Res. 30:596, 1987;
all of which are hereby incorporated by re~erence.
Svnthesis of analos The synthesis of the bombesin agonist, BIM-26187, D-Phe-Gln-Trp-Ala-Val-Gly-His-Leu-Leu-NH2 follows.
Other bombesin or ~RP agonists can be prepared by making appropriate modifications of the following synthetic . method.
1) Incorporation of alpha-t-butoxycarbonyl (BOC)-~ leucine on 4-methyl benzhydrylamine.
-~ 4-methyl benzhydrylamine-polystyrene resin (Bachem, Inc.) (0.72-meq/g) in the chloride ion form is placed in the reaction vessel of an ACT200 peptide ~ 35 synthesizer (Advanced Chem Tech, Inc.) programmed to .~ ' . . .
S~BS~ lTESHEEI
WO91/17181 2 a ~ PCT/US91/0326~j~
perform the following reaction cycle: (a) methylene chloride; (b) l0~ triethylamine in chloroform; (c) ; methylene chloride; and (d) dimethylformide.
The neutralized resin is mixed with `. 5 alpha-t-butoxycarbonyl (BOC)-leucine and diisopropylcarbodiimide (3 molar eq each) in methylene `
chlrodie for l hour. The resulting amino acid resin is washed on the synthesizer with dimethylformamide and treated with 5% acetic anhydride in dimethylformamide `
for 5 min. Then it is washed with dimethylformamide and methylene chloride.
2) Couplings of the remaining amino acids.
The peptide synthesizer is programmed to perform ` the following reaction cycle: (a) methylene chloride;
(b) 33~ trifluoroacetic acid (TFA) in methylene chloride - (2 times for 5 and 25 min. each); (c) methylene chloride; (d) isopropyl alcohol; (e) 10% triethylamine `
in chloroform; and (f) methylene chloride.
i The following amino acids (3 molar eq.) are then `
~, 20 coupled successively by the same procedure: BOC-Leu, BoC-His (tosyl), BOC-Gly, BOC-Val, BOC-Ala, BOC-Trp, BOC-Gln (coupled in the presence of' l eq.
' hydroxybenzotriazole), BOC-D-Phe (coupled in the h, presence of l eq. hydroxybenzotriazole). The completed ` 25 resin is then washed with methanol and air dried.
The peptide resin described above (l.4l g) is ~ mixed with anisole (5 ml), dithioerythreitol (50mg), and '3~ anhydrous hydrogen fluoride (25 ml) at 0c for one hour.
' Excess hydrogen fluoride is evaporated rapidly under a stream of dry nitrogen, and the residue is washed in ether. Crude peptide i5 dissolved in l00ml of 4M acetic acid and the solution is then evaporated under reduced pressure. The crude peptide is dissoIved in minimum volume of methanol/water and triturated with ethyl acetate. The triturated peptide is applied to a column .~, .,,~ -:
~UeST~TU~E SHEEI
... ' . . ` ... ~.. - .` i . . .. . ~. ` .... . . . . .......... .. ~ . . .
.,,.. .. ` ... . .. ~ ~ . . .
WO91/17181 2 ~ PCT/US91/03265 . .
(9.4mm I.D. x 50 cm) of octadecylcilane-silica (Whatman Partisil l0 ODS - 2M9). The peptide is eluted with a linear gradient of 20-80% of 50/50 0.1%
TFA/Acetronitrile i 0.1% TFA in water. Fractions are :
examined by analytical high performance liquid ; -chromatography and appropriate fractions are evaporated to a small volume, which is further lyophilized, gives 65mg of the product as a colorless powder.
Other compounds including D-Cpa1, CHx-Ala8, or Nle 9 can be prepared as above.
Peptides of the invention may be cyclized as :
follows.
Crude peptide acid obtained from peptide-resin , ester by HF cleavage is dissolved in DMF (0.1%-1%
concentration), treaed with condensing agent (e.g., BOP
reagent, DEPC, DPPA, or any other condensing agent) followed by base ~e.g., triethylamine, diisopropylethylamine) at room temperature for 1-3 days.
Solvent is removed in vacuum to dryness. The residue is ,~ 20 purified by HPLC, according to conventional procedures.
The cyclization of, for example, cycIo[D-Phel, Leu8, `2 ~eu9]Litorin, in which D-Phel is covalently linked to Leu9, is accompli9hed according to the above procedure ~ using Benzotriazol-l-yloxytris(dimethylamine)phosphonium hexafluorophosphate a the BOP reagent, diethylcyanophosphonate as the DEPC reagent, and diphenylphosphoryalazide as the DPPA reagent.
Mechanism of Action Analogs of the invention may prevent or inhibit the growth of cancer cells, or may prevent the ~- proliferation of non-malignant tissue, by acting as agonists or partial agonists; i.e., the analog may fully or partially mimic or enhance the biological effect of the natural peptide on a target cell. One possible mechanism of analog inhibition of growth of cancer cells . ~ .
! .
SUBSTlTUTE ~HFET
~ ~ $~
.-: . .: . . , . " , . . ., - ...
~, . . . . . . . . . .
WO9l/17181 2 ~3 ~1~ 3 ~3 PCT/US91/03265 j - is suggested in Bunn et al. (1990, Proc. Nat. Aca. Sci.
87:2162), in which calcium ion flux was measured in CHO
cells after administration of one or more neuropeptides.
Bunn et al. observed a desensitization of cells to the neuropeptide after administration of the second dose, i.e., after administration of the first dose and the -return of calcium concentration to resting values, administration of a second dose of an identical peptide resulted in no calcium flux. However, when the second dose was administered using a different peptide, a new calcium flux occurred. Thus different peptides may trigger different calcium flux pathways. The results of Bunn et al. show that desensitization to the neuropeptide may occur both in cancerous and normal ; 15 tissue, thus suggesting that an agonist may suppress growth in a tumor cell by a similar mechanism.
` Use Analogs of the invention are useful for treating colon, prostatic, breast, pancreatic, or lung cancer, ` 20 for preventing the proliferation of smooth muscle, to -'J suppress appetite, to stimulate pancreatic secretion, or to 5uppress a craving for alcohol. Analogs of the invention ars administered to a mammal, particularly a human, in one of the traditional modes (e.g., orally, j 25 parenterally, transdermally, or transmucosally), in a sustained release formulation using a biodegradable biocompatible polymer, or by on-site delivery using ~' micelles, gels and liposomes, or rectally (e.g., by suppository or enema). The analogs can be administered ~ 30 to a human patient in a dosage of 0.25 mg/kg/day to 5 '~ mg/kg/day.
-~ Other embodiments are within the following . ~ .
~ claims. -:`, ` ' :
':
'.
. ~ :
` SliBST~TE SHET
.. .. .. .. - . . . .
Heinz-Erian et al. replaced His at position 12 in bombesin with D-Phe and observed bombesin antagonist activity in dispersed acini from guinea pig pancreas (Heinz-Erian et al., Am. J. of Physiol. 2S2:G439-G442 (1987)). Rivier reported work directed toward restricting the conformational freedom of the bioactive C-terminal decapeptide of bombesin by incorporating intramolecular disulfide bridges; however, Rivier mentioned that, so far, bombesin analogs with this modification fail to exhibit any antagonist activity (Rivier et al., "Competitive Antagonists of Peptide Hormones," in Abstracts of the International Symposium on Bombesin-Like Peptides in Health and Disease, Rome, Italy (October, 1987).
Bombesin exhibits both direct and indirect effects on the gastrointestinal tract, including the release of hormones and the stimulation of pancreatic, gastric, and intestinal secretion and of intestinal mobility.
Gastrin and cholecystokinin (CCK) which are released by bombesin, have béen shown to play a role in the maintenance of normal gastrointestinal mucosa as well as in augmenting growth of normal and neoplastic tissues.
The growth of xenografted human colon and stomach carcinomas in nude mice has been stimulated by the , 25 administration of gastrin and later inhibited with the addition of secretin (Tanake et al., 1986, Tokaku J.
Exp. Ned. 148:459j and the growth of MC-26 murine colon carcinoma, which possesses gastrin receptors is ~ stimulated by pentagastrin (Winsett et al., 1980, d~ 30 Surgery 99:302, and inhibited by proglumide, a x-~ gastrin-receptor antagonist, Beauchamp et al., 1985, Ann. Surg. 202:303. Bombesin has been found to act concurrently as both a trophic agent for normal host ;~ pancreas and a growth inhibitory agent in xenografted ... .
;' .
~ .
~.j .
SUBSTl, UTL SHE~
o~ " . ~, . ,,, . . , ~ ,, ,, ~, , , . ". .,~ . ; . " , ~ , ., ", ,.
WO91/17181 2 0 3 1 ~ ~ ~ PCT/US9l,0326~
~: .
: 3 ;
human pancreatic tumor tissue, Alexander et al., 1988, Pancreas 3:247.
Abbreviations (uncommon3: ~
cyclohexyl-Ala = CHx-Ala (cyclohexyl alanine) ,.~-H
,. C CH2 identifying group ;
, CH
NH2--CH--C02H; : ' pGlu = H2C---CH-COOH (pyroglutamic acid);
, H2C NH
r C
~ Nle = H2N-CH-COOH (norleucine) ;~ (CH2)3-CH3 'Ss Pal - 3-pyridyl-alanine D-Cpa - D-para-chloro-phenylalanine HyPro = hydroxyproline Nal - naphthylalanine .
Sar = sarcosine F5-Phe s penta-fluoro-phenylalanine R = right (D) configuration; S = left (L) configuration; A.
~ 25 racemate = equal mix of R and S
.1 l-methyl-His; 3-methyl-His = methyl (CH3) group on -~-.j~ nitrogen at positions l or 3 of Histidine: -;;
.~ .
, ~ ~ C__NlH+ ., ,~j 30 3 C--N H
HCH
;~ NH2----CH----COOH .' "~: Summarv of the Invention -- :
.. . .
~$
.... . . .
~` SUBSIITUTE SHET
.,.,"~"~",".,,.,.,~ "` ~,,-,.,..~, . ..
WO91/1~181 2 ~ , 9 ~ PCT/US91/0326 The invention features a linear or a cyclic therapeutic peptide, which includes between seven and ten amino acid residues, inclusive, and which is an . analog of one of the following naturally occurring -:
peptides which terminate at the carboxy-terminus with a : Met residue: (a) litorin; (b) the ten amino acid carboxy-terminal region of mammalian GRP, neuromedin B, or neuromedin C; and (c) the ten amino acid carboxy-terminal region of amphibian bombesin, which is an analog of the formula:
R
AO_Al--A2--A3--A4 -AS -A6 -A7--A3 -A9 -R3 R
wherein . 15 A = Gly, D- or L- isomer of any of pGlu, Nle, a-aminobutyric acid, Ala, Val, Gln, Asn, Leu, Ile, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, CH3), Trp, or ~-Nal, or is deleted;
l = the D- or L-isomer of any of pGlu, Nle, j 20 a-aminobutyric acid, Ala, Val, Gln, Asn, Leu, Ile, p-X-Phe (where X - H, F, Cl, Br, NO2, OH, or CH3), Asp, Glu, F5-Phe, Trp, ~-Nal, Cys, Lys, or is deleted;
A2 = Gly, D- or L- isomer of any of pGlu, Ala, Val, :~ 25 Gln, Asn, Leu, Ile, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Trp, ~-Nal, Asp, Glu, His, l-methyl-His 3-methyl-His, Cys, Lys, or is i~ deleted; A3 = the D- or L-isomer of any of p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), ~-Nal, or Trp;
A~ = Ala, Val, Gln, Asn, Gly, Leu, Ile, Nle, : a-aminobutyric acid, p-X-phe (where X = H, F, Cl, Br, NO2, OH, or ,CH3), Trp, or ~-Nal;
.
.; .
~ , :
. ~ .
~ ` SUBSI~ TE S~'~T
;. - . ' ~ ' ,;'. .'' . :' . ,: , ' ', , "' " ' . :
W091/17181 2 ~ PCT/US91/03265 A5 = Gln, Asn, Gly, Ala, Leu, Ile, Nle, a-aminobutyric acid, Val, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Trp, Thr, or ~-Nal; .
A6 = Sar, Gly or the D-isomer of any Ala, N-methyl-Ala, Val, Gln, Asn, Leu, Ile, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Trp, Cys, or ~-Nal; :.
A7 - l-methyl-His, 3-methyl-His, His, Lys, Asp, or Glu;
A8 = Leu, Ile, Val, Nle, a-aminobutyric acid, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or C~3), Trp, Thr, ~-Nal, Lys, Asp, Glu, CHx-Ala, or Cys;
A9 = L-isomer of any of Met, Met-oxide, Leu, Ile, Nle, :: .
a-aminobutyric acid, p-X-Phe (where X = H, F, Cl, -Br, N02, OH, or CH3), Trp, ~-Nal, CHx-Ala, or ~i : Cys;
` each Rl and R2, independently, is H, Cl_l2 alkyl, C7_l0 .-p y lkyl, COEl ~where El is Cl_20 alkyl, C3 20 alkenyl C3_20 alkinyl, phenyl, naphthyl, or C7_l0 phenylalkyl), or Cl-Cl2 acyl, and Rl and R2 are bonded to the N-terminal amino acid o~ the peptide; provided that when one of Rl ..
.~ or R2 is COEl, the other must be H; and R3 is H, NH2, Z Cl_l2 alkyl, C~_lO phenylalkyl, or C3_20 naphthylalkyl;
`~ and further provided that, if Ao is present, Al cannot be pGlu; and, if A or Al is present, A2 cannot be pGlu;
~1 and further provided that, when A is deleted and Al is -` pGlu, Rl must be H and R2 must be the portion of Glu that .
1 forms the imine ring in pGlu; and further provided that, .~ where A is deleted and Al is not pGlu, Al may be bonded to A9, or where A and Al are deleted and A2 is not pGlu, A2 may be bonded to A9, or where A, Al and A2 are deleted, A3 can be bonded to A9 to form a cyclized ~ .
peptide; and provided that where AO is deleted and Al is Asp or Glu, or where A and Al are deleted and A2 is Asp .;~ .
`` .:
SU~S~ T~ SHEET
. ... .. . , .. . ... ., .. .... .. . ,; .. . .. .. . . . . .... ... . .. . .
.. ,. ~ . , . . . . ~ ., . . .. . . . ~ .
WO91/17181 ~t ~ Pcr/usg1/o3265a~
or Glu, either Al or A2 can be bonded with A7 or A8, where A7 or A8 is Lys, or where A is deleted and Al is Lys or A0 and Al are deleted and A2 is Lys, either Al or A2 can be bonded to A7 or A8, where A7 or A8 is Asp or Glu; and further provided that either one of Al or A~ can be Cys and can be bonded through a disulfide bridge with either A8 or A9, provided that either one of A8 or A9 can .: be Cys and can be bonded through a disulfide bridge with either Al or A2; and further provided that where A and Al are deleted and A6 is D-Ala, A8-A9 cannot be Leu-Met-NH2; or a pharmaceutically acceptable salt .
thereof.
i~ In the generic formula given above, when either of Rl or R2 is an aliphatio, aromatic, or lipophilic group, the n vlvo activity can be long lasting, and -1. delivery of the compounds of the invention to the target tissue can be facilitated.
More pre~erably, the analog of the generic formula above is of the formula:
A = pGlu, Gly, D-Phe, or is deleted;
.~ Al = pGlu, D-Phe, D-Ala, D-~-Nal, D-Cpa, D-Asn, Cys, :: or i8 deleted;
~ A2 , pGlu, Asn, Gln, His, l-methyl-His, 3-methyl-His, ... . .
Cys, or is deleted;
2S A3 = Trp;
.~ A4 = Ala;
,.,r A5 = Val;
Ah = Sar, Gly, D-Phe, or D-Ala;
A7 = His;
. 30 A = Leu, Phe, Chx-Ala, or Cys;
-~ A9 = L-isomer of any of Met, Leu, Ile, Nle, Phe, or ;, Cys.
~` Examples of preferred peptide analogs are~
pGlu-Gln-Trp-Ala-Val-Gly-His-Leu-Leu-NH2;
. ' .
.:
``` SUB~ITUTF S~ET
. .
~ . .. ... ~ ... .. .. , ., . ~ . . . .. . . . . .. .. .
- ` --. - - . , ~ . ..... ... ... ... . .. ..... ..... ....... .... ... .
... .. . . ~ . . . ..
. . . - . . . . . . . . . ` . . . . .
~ wO g-/17181 2 0 8 ~ ~ 9 ~ PCT/US91/03265 ~, D-Phe-Gln-Trp-Ala-Val-Gly-His-Leu-Leu-NH2;
D-Phe-Gln-Trp-Ala-Val-Gly-His-Leu-Met-NH2;
D-Cpa-Gln-Trp-Ala-Val-Gly-His-Leu-Met-NH2;
D-Cpa-Gln-Trp-Ala-Val-Gly-His-Leu-Leu-NH2;
D-Phe-Gln-Trp-Ala-Val-D-Ala-His-Leu-Leu-NH2;
D-Phe-Gln-Trp-Ala-Val-D-Ala-His-Leu-Met-NH2;
' D-Cpa-Gln-Trp-Ala-Val-D-Ala-His-Leu-Met-NH2; . :
; pGlu-Gln-Trp-Ala-Val-Gly-His-Phe-Leu-NH2; :
D-Phe-Gln-Trp-Ala-Val-Gly-His-Phe-Leu-NH2;
D-Phe-Gln-Trp-Ala-Val-D-Ala-His-Phe-Met-NH ; ~:
.~ 2 ...
D-Phe-Gln-Trp-Ala-Val-D-Ala-His-Phe-Leu-NH2; !',:' D-Phe-Gln-Trp-Ala-Val-Gly-His-CHx-Ala-Leu-NH2;
D-Phe-Gln-Trp-Ala-Val-Gly-His-Leu-Nle-NH2;
:~ D-Phe-Gln-Trp-Ala-Val-D-Ala-His-Leu-Nle-NH2; ~-.
D-Phe-Gln-Trp-Ala-Val-Gly-His-Phe-Nle-NH2;
D-Phe-Gln-Trp-Ala-Val-D-Ala-His-Phe-Nle-NH2;
Ac-His-Trp-Ala-Val-D-A~la-His-Leu ~ :
-Leu-NH2 cyclo-D-Phe-Gln-Trp-Ala-Val-Gly-His-Leu-Leu;
~: :
.~ 20 D-Cys-Asn-Trp-Ala-Val-Gly-His-Leu-Cys-NH2;
" .
i;~ cyclo-His-Trp-Ala-Val-Gly-His-Leu-Met;
, .' ' j~l Cys-Trp-Ala-Val-Gly-His-Leu-Cys-NH2; :. .-~`3 `~ :
~I, cyclo-D-Phe-Gln-Trp-Ala-Val-Gly-His-Leu-Met; ..
; cyclo-D-Phe-His-Trp-Ala-Val-Gly-His-Leu-Met;
. .~ .
25 and cyclo-Trp-Ala-Val-Gly-His-Leu-Met;
. .~. .
~, :. -. ,;~
.,~, ~ ' .
~ SUBSTI~IuTE SHEET
, . ~.. ., . ` , ~ ., " , i ., . , , , ., . ~ . ;, .. .,, . ;,, .. , `, . . . .. . . . . . .
., , .. ;~.` i .. , . .... . , , . .,, ....... , . ; ...... ,, . , , ., .. . , , .,,, - , ,.
..... ... .. .. .. - . , ., . - ,` . ..... . ;. . , ~; .... . , .. ~ . ,- .
WO91/17181 2 0 318 g ~ PCT/US91/03265 Modification of a natural amino acid or substitution of a synthetic amino acid, examples of which are (but are not limited to) pGlu, Nle, Pal, D-Cpa, Met-oxide, Nal, Sar, l-methyl-His, or 3-methyl-His, at one or more of positions A, Al, A6, or A9 of an analog, or cyclization of the analog result in enhanced stability; i.e., protection of the amino terminus from exopeptidase degradation and of the internal portion of the peptide from endopeptidase degradation. The carboxy-terminal dipeptide sequence of some analogs of the invention corresponds to the natural ~ C-terminal sequence of bombesin or bombesin-related -~ analogs.
The invention also features a linear or a cyclic therapeutic peptide, which includes between seven and ten amino acid residues, inclusive, and which is an analog of one of the following naturally occurring - peptides which terminate at the carboxy-terminus with a Met residue: ~a) litorin; (b) the ten amino acid carboxy-terminal region of mammalian GRP, neuromedin B, or neuromedin C; and (c) the ten amino acid ~b carboxy-terminal region of amphibian bombesin, and the analog is an agonist of one of these naturally occurring peptides.
~ 25 In preferred embodiments, the analog may be an ;~ agonist or a partial agonist of the naturally occurring ,- biologically active peptide; preferably, the analog is -~
;~ at least 25%, more preferably 50~ or 75%, homologous with a region of the naturally occurring peptide. As used herein, an "agonist" mimics or enhances the - biological effect of the natural peptide on its target cell and a "partial agonist" mimics or enhances the ~-~ biological effect of the natural peptide, but to a lesser extent than an agonist. Biological effect, as used herein, is measured by the effect of the natural ~, .. .
-.
.' ~.'-.: .
SUBSi'Til~E SHEET
.
~WO91/17181 ~ PCT/US91/03265 .. ' ~, g :' ,:
peptide in one of two systems: an in vitro pancreatic amylase release assay and an in vitro 3T3 fibroblast cell division system, ~oth of which are described in European Patent Application 88308916.6, hereby -;
incorporated by reference. An agonist will stimulate ;~
the effect of the natural peptide on either amylase release from pancreatic cells or fibroblast cell division by 100%, whereas a partial agonist will have a lesser stimulatory effect, i.e., ranging between 0-99%.
, .
Peptides of the invention are useful for treating non-malignant proliferative disease in a human patient, e.g., the proliferation of smooth muscle. Peptides of the invention are also useful for treating cancer in a human patient, particularly for the treatment of prostatic, colon, breast, pancreatic, or lung cancer.
In addition, peptides of the invention may be used to suppress appetite, to stimulate pancreatic secretion, or to suppress a craving for alcohol.
Analogs of the invention can be provided in the ~1, 20 ~orm of pharmaceutically acceptable salts. Examples of ;' pre~erred 5alts are those with therapeutically ;~ acceptable organic acids, e.g., acetic, lactic, maleic, - citric, malic, ascorbic, succinic, benzoic, salicylic, methanesulfonic, toluene sulfonic, trifluoroacetic, or pamoic acid, as well as polymeric acids such as tannic acid or carboxymethyl cellulose, and salts with -inorganic acids such as the hydrohalic acids,e.g., , hydrochloric acid, sulfuric acid or phosphoric acid.
Other features and advantages of the invention will be apparent from the following description of the -i~ preferred embodiments thereof, and from the claims.
-~ Descri~tion of the Preferred Embodiments Before describing preferred embodiments of the ~, invention, the drawing will briefly be described.
~,.j .
.... .
S~BSTITU~E SHEET
W091/17181 ~ PCT/US91/0326a Drawina Fig. 1 is the amino acid sequences of naturally occurring peptides of which peptides of the invention are analogs.
we now describe the structure, synthesis, and use of the preferred embodiments of the invention.
Structure Peptides of the invention are derived from one of the sequences shown in Fig. 1, which represent the sequences, or portions thereof, of naturally-occurring peptides. Bombesin, neuromedin B, neuromedin C, litorin, and GRP analogs of the invention are described in Coy et al., U.S. Patent Application Serial No.
502,438, filed March 30, 1990, which is a --continuation-in-part of U.S. Patent Application Serial No. 397,169, filed August 21, 1989, which is a s continuation-in-part of U.S. Patent Application Serial r No. 376,555, filed July 7, 1989, and U.S. Patent ~ Application Serial Nu~ber 394,727, filed August 16, -j 20 1989, both of which are continuation-in-parts of U.S. ~5 Patent Application Serial No. 317,941, filed March 2, 1989, which is a continuation-in-part of U.S. Patent Application Serial Number 282,328, filed December 9, 1988, which in turn is a continuation-in-part of U.S.
Patent Application Serial No. 257,998, filed October 14, 1988, which in turn is a continuation-in-part of U.S.
~ Patent Application Serial No. 248,771, filed September 'i~ 23, 1988, which in turn is a continuation-in-part of Coy et al., U.S. Patent Application Serial No. 207,759, '~ 30 filed June 16, 1988j which in turn is a -~'~ continuation-in-part of Coy et al., U.S. Patent Application Serial No. 204,171, filed June 8, 1988, ~ -which in turn is a continuation-in-part of Coy et al., U.S. Patent Application Serial No. 173,311, filed March i , , .
~ 35 25, 1988, which in turn is a continuation-in-part of Coy '~
.~ .
SUBSli~UT~ ~IEET
i WO91/17181 2 0 g 1 ~ ~ ~ PCT/US91/0326~
et al. U.S. Patent Application Serial No. 100,571, filed - September 24, 1987; all of which are assigned to the same assignee and hereby incorporated by reference; or -as described in Zachary et al., Proc. Nat. Aca. Sci.
82:7616, 1985; Heimbrook et al., "Synthetic Peptides:
Approaches to Biological Problems", UCLA Symposium on Mol. and Cell. Biol. New Series, Vol. 86, ed. Tam and ; Kaiser; Heinz-Erian et al., Am. J. Physiol. G439, 1986;
Martinez et al., J. Med. Chem. 28:1874, 1985; Gargosky ; 10 et al., Biochem. J. 247:427, 1987; Dubreuil et al., Drug ' Design and Delivery, Vol 2:49, Harwood Academic Publishers, GB, 1987; Heikkila et al., J. Biol. Chem.
262:16456, 1987; Caranikas et al., J. Med. Chem.
25:1313, 1982; Saeed et al., 1989, Peptides 10:597;
i 15 Rosell et al., Trends in Pharmacological Sciences 3:211, 1982; Lundberg et al., Proc. Nat. Aca. Sci. ~0:1120, 1983; Engberg et al., Nature 293:222, 1984; Mizrahi et al., Euro. J. Pharma. 82:101, 1982; Leander et al., Nature 294:467, 1981; Woll et al., Biochem. Biophys.
Res. Comm. 155:359, 1988; Rivier et al., Biochem.
~; 17:1766, 1978; Cuttitta et al., Cancer Surveys 4:707, 1985; Aumelas et al., Int. J. Peptide Res. 30:596, 1987;
all of which are hereby incorporated by re~erence.
Svnthesis of analos The synthesis of the bombesin agonist, BIM-26187, D-Phe-Gln-Trp-Ala-Val-Gly-His-Leu-Leu-NH2 follows.
Other bombesin or ~RP agonists can be prepared by making appropriate modifications of the following synthetic . method.
1) Incorporation of alpha-t-butoxycarbonyl (BOC)-~ leucine on 4-methyl benzhydrylamine.
-~ 4-methyl benzhydrylamine-polystyrene resin (Bachem, Inc.) (0.72-meq/g) in the chloride ion form is placed in the reaction vessel of an ACT200 peptide ~ 35 synthesizer (Advanced Chem Tech, Inc.) programmed to .~ ' . . .
S~BS~ lTESHEEI
WO91/17181 2 a ~ PCT/US91/0326~j~
perform the following reaction cycle: (a) methylene chloride; (b) l0~ triethylamine in chloroform; (c) ; methylene chloride; and (d) dimethylformide.
The neutralized resin is mixed with `. 5 alpha-t-butoxycarbonyl (BOC)-leucine and diisopropylcarbodiimide (3 molar eq each) in methylene `
chlrodie for l hour. The resulting amino acid resin is washed on the synthesizer with dimethylformamide and treated with 5% acetic anhydride in dimethylformamide `
for 5 min. Then it is washed with dimethylformamide and methylene chloride.
2) Couplings of the remaining amino acids.
The peptide synthesizer is programmed to perform ` the following reaction cycle: (a) methylene chloride;
(b) 33~ trifluoroacetic acid (TFA) in methylene chloride - (2 times for 5 and 25 min. each); (c) methylene chloride; (d) isopropyl alcohol; (e) 10% triethylamine `
in chloroform; and (f) methylene chloride.
i The following amino acids (3 molar eq.) are then `
~, 20 coupled successively by the same procedure: BOC-Leu, BoC-His (tosyl), BOC-Gly, BOC-Val, BOC-Ala, BOC-Trp, BOC-Gln (coupled in the presence of' l eq.
' hydroxybenzotriazole), BOC-D-Phe (coupled in the h, presence of l eq. hydroxybenzotriazole). The completed ` 25 resin is then washed with methanol and air dried.
The peptide resin described above (l.4l g) is ~ mixed with anisole (5 ml), dithioerythreitol (50mg), and '3~ anhydrous hydrogen fluoride (25 ml) at 0c for one hour.
' Excess hydrogen fluoride is evaporated rapidly under a stream of dry nitrogen, and the residue is washed in ether. Crude peptide i5 dissolved in l00ml of 4M acetic acid and the solution is then evaporated under reduced pressure. The crude peptide is dissoIved in minimum volume of methanol/water and triturated with ethyl acetate. The triturated peptide is applied to a column .~, .,,~ -:
~UeST~TU~E SHEEI
... ' . . ` ... ~.. - .` i . . .. . ~. ` .... . . . . .......... .. ~ . . .
.,,.. .. ` ... . .. ~ ~ . . .
WO91/17181 2 ~ PCT/US91/03265 . .
(9.4mm I.D. x 50 cm) of octadecylcilane-silica (Whatman Partisil l0 ODS - 2M9). The peptide is eluted with a linear gradient of 20-80% of 50/50 0.1%
TFA/Acetronitrile i 0.1% TFA in water. Fractions are :
examined by analytical high performance liquid ; -chromatography and appropriate fractions are evaporated to a small volume, which is further lyophilized, gives 65mg of the product as a colorless powder.
Other compounds including D-Cpa1, CHx-Ala8, or Nle 9 can be prepared as above.
Peptides of the invention may be cyclized as :
follows.
Crude peptide acid obtained from peptide-resin , ester by HF cleavage is dissolved in DMF (0.1%-1%
concentration), treaed with condensing agent (e.g., BOP
reagent, DEPC, DPPA, or any other condensing agent) followed by base ~e.g., triethylamine, diisopropylethylamine) at room temperature for 1-3 days.
Solvent is removed in vacuum to dryness. The residue is ,~ 20 purified by HPLC, according to conventional procedures.
The cyclization of, for example, cycIo[D-Phel, Leu8, `2 ~eu9]Litorin, in which D-Phel is covalently linked to Leu9, is accompli9hed according to the above procedure ~ using Benzotriazol-l-yloxytris(dimethylamine)phosphonium hexafluorophosphate a the BOP reagent, diethylcyanophosphonate as the DEPC reagent, and diphenylphosphoryalazide as the DPPA reagent.
Mechanism of Action Analogs of the invention may prevent or inhibit the growth of cancer cells, or may prevent the ~- proliferation of non-malignant tissue, by acting as agonists or partial agonists; i.e., the analog may fully or partially mimic or enhance the biological effect of the natural peptide on a target cell. One possible mechanism of analog inhibition of growth of cancer cells . ~ .
! .
SUBSTlTUTE ~HFET
~ ~ $~
.-: . .: . . , . " , . . ., - ...
~, . . . . . . . . . .
WO9l/17181 2 ~3 ~1~ 3 ~3 PCT/US91/03265 j - is suggested in Bunn et al. (1990, Proc. Nat. Aca. Sci.
87:2162), in which calcium ion flux was measured in CHO
cells after administration of one or more neuropeptides.
Bunn et al. observed a desensitization of cells to the neuropeptide after administration of the second dose, i.e., after administration of the first dose and the -return of calcium concentration to resting values, administration of a second dose of an identical peptide resulted in no calcium flux. However, when the second dose was administered using a different peptide, a new calcium flux occurred. Thus different peptides may trigger different calcium flux pathways. The results of Bunn et al. show that desensitization to the neuropeptide may occur both in cancerous and normal ; 15 tissue, thus suggesting that an agonist may suppress growth in a tumor cell by a similar mechanism.
` Use Analogs of the invention are useful for treating colon, prostatic, breast, pancreatic, or lung cancer, ` 20 for preventing the proliferation of smooth muscle, to -'J suppress appetite, to stimulate pancreatic secretion, or to 5uppress a craving for alcohol. Analogs of the invention ars administered to a mammal, particularly a human, in one of the traditional modes (e.g., orally, j 25 parenterally, transdermally, or transmucosally), in a sustained release formulation using a biodegradable biocompatible polymer, or by on-site delivery using ~' micelles, gels and liposomes, or rectally (e.g., by suppository or enema). The analogs can be administered ~ 30 to a human patient in a dosage of 0.25 mg/kg/day to 5 '~ mg/kg/day.
-~ Other embodiments are within the following . ~ .
~ claims. -:`, ` ' :
':
'.
. ~ :
` SliBST~TE SHET
.. .. .. .. - . . . .
Claims (33)
1. A therapeutic peptide comprising between seven and ten amino acid residues, inclusive, said peptide being an analog of one of the following naturally occurring peptides which terminate at the carboxy-terminus with a Met residue: (a) litorin; (b) the ten amino acid carboxy-terminal region of mammalian GRP, neuromedin B, or neuromedin C; and (c) the ten amino acid carboxy-terminal region of amphibian bombesin, said analog being of the formula:
wherein A0 = Gly, D- or L- isomer of any of pGlu, Nle, a-aminobutyric acid, Ala, Val, Gln, Asn, Leu, Ile, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, CH3), Trp, or .beta.-Nal, or is deleted;
A1 = the D- or L-isomer of any of pGlu, Nle, a-aminobutyric acid, Ala, Val, Gln, Asn, Leu, Ile, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Asp, Glu, F5-Phe, Trp, .beta.-Nal, Cys, Lys, or is deleted;
A2 = Gly, D- or L- isomer of any of pGlu, Ala, Val, Gln, Asn, Leu, Ile, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Trp, .beta.-Nal, Asp, Glu, His, 1-methyl-His 3-methyl-His, Cys, Lys, or is deleted;
A3 = the D- or L-isomer of any of p-X-Phe (where X =
H, F, Cl, Br, NO2, OH, or CH3), .beta.-Nal, or Trp;
A4 = Ala, Val, Gln, Asn, Gly, Leu, Ile, Nle, a-aminobutyric acid, p-X-phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Trp, or .beta.-Nal;
A5 = Gln, Asn, Gly, Ala, Leu, Ile, Nle, a-aminobutyric acid, Val, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Trp, Thr, or .beta.-Nal;
A6 = Sar, Gly or the D-isomer of any Ala, N-methyl-Ala, Val, Gln, Asn, Leu, Ile, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Trp, Cys, or .beta.-Nal;
A7 = 1-methyl-His, 3-methyl-His, His, Lys, Asp, or Glu;
A8 = Leu, Ile, Val, Nle, a-aminobutyric acid, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Trp, Thr, .beta.-Nal, Lys, Asp, Glu, CHx-Ala, or Cys;
A9 = L-isomer of any of Met, Met-oxide, Leu, Ile, Nle, a-aminobutyric acid, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Trp, .beta.-Nal, CHx-Ala, or Cys;
each R1 and R2, independently, is H, C1-12 alkyl, C7-10 phenylalkyl, COE1 (where E1 is C1-20 alkyl, C3-20 alkenyl, C3-20 alkinyl, phenyl, naphthyl, or C7-10 phenylalkyl), or C1-C12 acyl, and R1 and R2 are bonded to the N-terminal amino acid of the peptide; provided that when one of R1 or R2 is COE1, the other must be H; and R3 is H, NH2, C1-12 alkyl, C7-10 phenylalkyl, or C3-20 naphthylalkyl;
and further provided that, if A0 is present, A1 cannot be pGlu; and, if A0 or A1 is present, A2 cannot be pGlu;
and further provided that, when A0 is deleted and A1 is pGlu, R1 must be H and R2 must be the portion of Glu that forms the imine ring in pGlu; and further provided that, where A0 is deleted and A1 is not pGlu, A1 may be bonded to A9, or where A0 and A1 are deleted and A2 is not pGlu, A2 may be bonded to A9, or where A0, A1 and A2 are deleted, A3 may be bonded to A9; and provided that where A0 is deleted and A1 is Asp or Glu, or where A0 and A1 are deleted and A2 is Asp or Glu, either Al or A2 can be bonded with A7 or A8, where A7 or A8 is Lys, or where A0 is deleted and A1 is Lys or A0 and A1 are deleted and A2 is Lys, either A1 or A2 can be bonded to A7 or A8, where A7 or A8 is Asp or Glu; and further provided that either one of A1 or A2 can be Cys and can be bonded through a disulfide bridge with either A8 or A9, provided that either one of A8 or A9 can be Cys and can be bonded through a disulfide bridge with either A1 or A2; and further provided that where A0 and A1 are deleted and A6 is D-Ala, A8-A9 cannot be Leu-Met-NH2; or a pharmaceutically acceptable salt thereof.
wherein A0 = Gly, D- or L- isomer of any of pGlu, Nle, a-aminobutyric acid, Ala, Val, Gln, Asn, Leu, Ile, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, CH3), Trp, or .beta.-Nal, or is deleted;
A1 = the D- or L-isomer of any of pGlu, Nle, a-aminobutyric acid, Ala, Val, Gln, Asn, Leu, Ile, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Asp, Glu, F5-Phe, Trp, .beta.-Nal, Cys, Lys, or is deleted;
A2 = Gly, D- or L- isomer of any of pGlu, Ala, Val, Gln, Asn, Leu, Ile, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Trp, .beta.-Nal, Asp, Glu, His, 1-methyl-His 3-methyl-His, Cys, Lys, or is deleted;
A3 = the D- or L-isomer of any of p-X-Phe (where X =
H, F, Cl, Br, NO2, OH, or CH3), .beta.-Nal, or Trp;
A4 = Ala, Val, Gln, Asn, Gly, Leu, Ile, Nle, a-aminobutyric acid, p-X-phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Trp, or .beta.-Nal;
A5 = Gln, Asn, Gly, Ala, Leu, Ile, Nle, a-aminobutyric acid, Val, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Trp, Thr, or .beta.-Nal;
A6 = Sar, Gly or the D-isomer of any Ala, N-methyl-Ala, Val, Gln, Asn, Leu, Ile, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Trp, Cys, or .beta.-Nal;
A7 = 1-methyl-His, 3-methyl-His, His, Lys, Asp, or Glu;
A8 = Leu, Ile, Val, Nle, a-aminobutyric acid, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Trp, Thr, .beta.-Nal, Lys, Asp, Glu, CHx-Ala, or Cys;
A9 = L-isomer of any of Met, Met-oxide, Leu, Ile, Nle, a-aminobutyric acid, p-X-Phe (where X = H, F, Cl, Br, NO2, OH, or CH3), Trp, .beta.-Nal, CHx-Ala, or Cys;
each R1 and R2, independently, is H, C1-12 alkyl, C7-10 phenylalkyl, COE1 (where E1 is C1-20 alkyl, C3-20 alkenyl, C3-20 alkinyl, phenyl, naphthyl, or C7-10 phenylalkyl), or C1-C12 acyl, and R1 and R2 are bonded to the N-terminal amino acid of the peptide; provided that when one of R1 or R2 is COE1, the other must be H; and R3 is H, NH2, C1-12 alkyl, C7-10 phenylalkyl, or C3-20 naphthylalkyl;
and further provided that, if A0 is present, A1 cannot be pGlu; and, if A0 or A1 is present, A2 cannot be pGlu;
and further provided that, when A0 is deleted and A1 is pGlu, R1 must be H and R2 must be the portion of Glu that forms the imine ring in pGlu; and further provided that, where A0 is deleted and A1 is not pGlu, A1 may be bonded to A9, or where A0 and A1 are deleted and A2 is not pGlu, A2 may be bonded to A9, or where A0, A1 and A2 are deleted, A3 may be bonded to A9; and provided that where A0 is deleted and A1 is Asp or Glu, or where A0 and A1 are deleted and A2 is Asp or Glu, either Al or A2 can be bonded with A7 or A8, where A7 or A8 is Lys, or where A0 is deleted and A1 is Lys or A0 and A1 are deleted and A2 is Lys, either A1 or A2 can be bonded to A7 or A8, where A7 or A8 is Asp or Glu; and further provided that either one of A1 or A2 can be Cys and can be bonded through a disulfide bridge with either A8 or A9, provided that either one of A8 or A9 can be Cys and can be bonded through a disulfide bridge with either A1 or A2; and further provided that where A0 and A1 are deleted and A6 is D-Ala, A8-A9 cannot be Leu-Met-NH2; or a pharmaceutically acceptable salt thereof.
2. The therapeutic peptide of claim 1, comprising the formula:
A0 = pGlu, Gly, D-Phe, or is deleted;
A1 = pGlu, D-Phe, D-Ala, D-.beta.-Nal, D-Cpa, D-Asn, Cys, or is deleted;
A2 = pGlu, Asn, Gln, His, 1-methyl-His, 3-methyl-His, Cys, or is deleted;
A3 = Trp;
A4 = Ala;
A5 = Val;
A6 = Sar, Gly, D-Phe, or D-Ala;
A7 = His;
A8 = Leu, Phe, Chx-Ala, or Cys;
A9 = L-isomer of any of Met, Leu, Ile, Nle, Phe, or Cys.
A0 = pGlu, Gly, D-Phe, or is deleted;
A1 = pGlu, D-Phe, D-Ala, D-.beta.-Nal, D-Cpa, D-Asn, Cys, or is deleted;
A2 = pGlu, Asn, Gln, His, 1-methyl-His, 3-methyl-His, Cys, or is deleted;
A3 = Trp;
A4 = Ala;
A5 = Val;
A6 = Sar, Gly, D-Phe, or D-Ala;
A7 = His;
A8 = Leu, Phe, Chx-Ala, or Cys;
A9 = L-isomer of any of Met, Leu, Ile, Nle, Phe, or Cys.
3. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
4. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
5. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
6. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
7. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
8. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
9. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
10. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
11. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
12. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
13. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
14. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
15. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
16. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
17. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
18. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
19. A therapeutic peptide comprising the amino acid formula:
.
.
20. A therapeutic peptide comprising the amino acid formula:
.
.
21. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
22. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
23. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
24. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
25. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
26. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
27. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
28. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
29. The therapeutic peptide of claim 2, comprising the amino acid formula:
.
.
30. A therapeutic peptide comprising between seven and ten amino acid residues, inclusive, said peptide being an analog of one of the following naturally occurring peptides which terminate at the carboxy-terminus with a Met residue: (a) litorin; (b) the ten amino acid carboxy-terminal region of mammalian GRP, neuromedin B, or neuromedin C; and (c) the ten amino acid carboxy-terminal region of amphibian bombesin, said analog being an agonist of one of said naturally occurring peptides.
31. The therapeutic peptide of claim 30, wherein said peptide is a partial agonist of said naturally occurring biologically active peptide.
32. The therapeutic peptide of claim 30, said analog being at least 25% homologous with said naturally occurring peptide.
33. The therapeutic peptide of claim 32, said analog being at least 50% homologous with said naturally occurring peptide.
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
US52022690A | 1990-05-09 | 1990-05-09 | |
US520,226 | 1990-05-09 |
Publications (1)
Publication Number | Publication Date |
---|---|
CA2081890A1 true CA2081890A1 (en) | 1991-11-10 |
Family
ID=24071694
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CA002081890A Abandoned CA2081890A1 (en) | 1990-05-09 | 1991-05-09 | Cyclized and linear therapeutic peptides |
Country Status (5)
Country | Link |
---|---|
EP (1) | EP0531342A1 (en) |
JP (1) | JPH05506862A (en) |
CA (1) | CA2081890A1 (en) |
HU (1) | HUT62604A (en) |
WO (1) | WO1991017181A1 (en) |
Families Citing this family (10)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US6541610B1 (en) * | 1989-09-05 | 2003-04-01 | Immunex Corporation | Fusion proteins comprising tumor necrosis factor receptor |
US5767236A (en) * | 1990-05-09 | 1998-06-16 | Biomeasure, Inc. | Linear therapeutic peptides |
US5834433A (en) * | 1990-07-26 | 1998-11-10 | Merrell Pharmaceuticals Inc. | Compounds and pharmaceutical uses of peptides of bombesin and GRP |
JPH07505865A (en) * | 1992-02-07 | 1995-06-29 | メレルダウファーマスーティカルズ インコーポレイテッド | Phenylalanine analogs of bombesin |
ES2124496T3 (en) * | 1995-03-13 | 1999-02-01 | Biomeasure Inc | BOMBESIN ANALOGS. |
DE10044989A1 (en) | 2000-09-11 | 2002-03-21 | Bayer Ag | Liquid sulfur-containing oligosiloxanes and their use in rubber compounds |
US8709998B2 (en) | 2003-04-22 | 2014-04-29 | Ipsen Pharma S.A.S. | Peptide vectors |
KR101642363B1 (en) | 2008-06-12 | 2016-07-25 | 입센 바이오이노베이션 리미티드 | Suppression of neuroendocrine diseases |
CN102083451A (en) | 2008-06-12 | 2011-06-01 | 赛恩泰新公司 | Suppression of cancers |
GB0820970D0 (en) | 2008-11-17 | 2008-12-24 | Syntaxin Ltd | Suppression of cancer |
Family Cites Families (7)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US4207311A (en) * | 1977-07-18 | 1980-06-10 | The Salk Institute For Biological Studies | Peptides having analgesic and thermoregulative properties |
US4331661A (en) * | 1980-10-03 | 1982-05-25 | The Salk Institute For Biological Studies | Bombesin analogs |
GB8427651D0 (en) * | 1984-11-01 | 1984-12-05 | Beecham Group Plc | Compounds |
US4803261A (en) * | 1986-06-27 | 1989-02-07 | The Administrators Of The Tulane Educational Fund | Method for synthesizing a peptide containing a non-peptide |
GB8813356D0 (en) * | 1988-06-06 | 1988-07-13 | Ici Plc | Polypeptide compounds |
GB2231051B (en) * | 1989-05-05 | 1992-12-16 | Erba Carlo Spa | Bombesin antagonists |
CA2042027A1 (en) * | 1989-09-15 | 1991-03-16 | Arthur E. Bogden | Treatment of colon cancer |
-
1991
- 1991-05-09 EP EP91909510A patent/EP0531342A1/en not_active Withdrawn
- 1991-05-09 HU HU9203481A patent/HUT62604A/en unknown
- 1991-05-09 WO PCT/US1991/003265 patent/WO1991017181A1/en not_active Application Discontinuation
- 1991-05-09 JP JP91509428A patent/JPH05506862A/en active Pending
- 1991-05-09 CA CA002081890A patent/CA2081890A1/en not_active Abandoned
Also Published As
Publication number | Publication date |
---|---|
EP0531342A4 (en) | 1994-04-06 |
JPH05506862A (en) | 1993-10-07 |
HU9203481D0 (en) | 1993-01-28 |
HUT62604A (en) | 1993-05-28 |
EP0531342A1 (en) | 1993-03-17 |
WO1991017181A1 (en) | 1991-11-14 |
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