CA2226177C - Analogs of parathyroid hormone - Google Patents
Analogs of parathyroid hormone Download PDFInfo
- Publication number
- CA2226177C CA2226177C CA2226177A CA2226177A CA2226177C CA 2226177 C CA2226177 C CA 2226177C CA 2226177 A CA2226177 A CA 2226177A CA 2226177 A CA2226177 A CA 2226177A CA 2226177 C CA2226177 C CA 2226177C
- Authority
- CA
- Canada
- Prior art keywords
- cha
- aib
- leu
- hpth
- phe
- 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.)
- Expired - Lifetime
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- 125000002777 acetyl group Chemical group [H]C([H])([H])C(*)=O 0.000 description 1
- 125000002252 acyl group Chemical group 0.000 description 1
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- 125000001797 benzyl group Chemical group [H]C1=C([H])C([H])=C(C([H])=C1[H])C([H])([H])* 0.000 description 1
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- 230000037396 body weight Effects 0.000 description 1
- BBBFJLBPOGFECG-VJVYQDLKSA-N calcitonin Chemical compound N([C@H](C(=O)N[C@@H](CC(C)C)C(=O)NCC(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CO)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC=1NC=NC=1)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H]([C@@H](C)O)C(=O)NCC(=O)N[C@@H](CO)C(=O)NCC(=O)N[C@@H]([C@@H](C)O)C(=O)N1[C@@H](CCC1)C(N)=O)C(C)C)C(=O)[C@@H]1CSSC[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CO)C(=O)N[C@@H]([C@@H](C)O)C(=O)N1 BBBFJLBPOGFECG-VJVYQDLKSA-N 0.000 description 1
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- BGRWYRAHAFMIBJ-UHFFFAOYSA-N diisopropylcarbodiimide Natural products CC(C)NC(=O)NC(C)C BGRWYRAHAFMIBJ-UHFFFAOYSA-N 0.000 description 1
- 239000003937 drug carrier Substances 0.000 description 1
- 230000002124 endocrine Effects 0.000 description 1
- DNJIEGIFACGWOD-UHFFFAOYSA-N ethyl mercaptane Natural products CCS DNJIEGIFACGWOD-UHFFFAOYSA-N 0.000 description 1
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- 125000002485 formyl group Chemical group [H]C(*)=O 0.000 description 1
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- 229940088597 hormone Drugs 0.000 description 1
- 229910000040 hydrogen fluoride Inorganic materials 0.000 description 1
- NPZTUJOABDZTLV-UHFFFAOYSA-N hydroxybenzotriazole Substances O=C1C=CC=C2NNN=C12 NPZTUJOABDZTLV-UHFFFAOYSA-N 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
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- 230000000968 intestinal effect Effects 0.000 description 1
- 238000007918 intramuscular administration Methods 0.000 description 1
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- ZLNQQNXFFQJAID-UHFFFAOYSA-L magnesium carbonate Chemical compound [Mg+2].[O-]C([O-])=O ZLNQQNXFFQJAID-UHFFFAOYSA-L 0.000 description 1
- 239000001095 magnesium carbonate Substances 0.000 description 1
- 229910000021 magnesium carbonate Inorganic materials 0.000 description 1
- VZCYOOQTPOCHFL-UPHRSURJSA-N maleic acid Chemical compound OC(=O)\C=C/C(O)=O VZCYOOQTPOCHFL-UPHRSURJSA-N 0.000 description 1
- UZKWTJUDCOPSNM-UHFFFAOYSA-N methoxybenzene Substances CCCCOC=C UZKWTJUDCOPSNM-UHFFFAOYSA-N 0.000 description 1
- CSHFHJNMIMPJST-HOTGVXAUSA-N methyl (2s)-2-[[(2s)-2-[[2-[(2-aminoacetyl)amino]acetyl]amino]-3-phenylpropanoyl]amino]-4-methylpentanoate Chemical compound NCC(=O)NCC(=O)N[C@H](C(=O)N[C@@H](CC(C)C)C(=O)OC)CC1=CC=CC=C1 CSHFHJNMIMPJST-HOTGVXAUSA-N 0.000 description 1
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- 201000008968 osteosarcoma Diseases 0.000 description 1
- WLJNZVDCPSBLRP-UHFFFAOYSA-N pamoic acid Chemical compound C1=CC=C2C(CC=3C4=CC=CC=C4C=C(C=3O)C(=O)O)=C(O)C(C(O)=O)=CC2=C1 WLJNZVDCPSBLRP-UHFFFAOYSA-N 0.000 description 1
- 210000002990 parathyroid gland Anatomy 0.000 description 1
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/575—Hormones
- C07K14/635—Parathyroid hormone, i.e. parathormone; Parathyroid hormone-related peptides
-
- 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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- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Endocrinology (AREA)
- Life Sciences & Earth Sciences (AREA)
- Organic Chemistry (AREA)
- Biophysics (AREA)
- Gastroenterology & Hepatology (AREA)
- Biochemistry (AREA)
- Zoology (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Medicinal Chemistry (AREA)
- Molecular Biology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Toxicology (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Peptides Or Proteins (AREA)
Abstract
Peptide variants of fragment (1-34) of parathyroid hormone, in which at least one of the amino acid residues at positions 7, 11, 23, 24, 27, 28 and 31 is cyclohexylalanine, or at least one of the amino acid residues at positions 3, 16, 17, 18, 19 and 34 is .alpha.-aminoisobutyric acid; or, alternatively, at least the amino acid residue at position 1 is .alpha.,.beta.-diaminopropionic acid, the amino acid residue at position 27 is homoarginine, or the amino acid residue at position 31 is norleucine.
Description
CA 02226l77 l997-l2-30 W O 97/02834 PCTrUS96/11292 ANALOGS OF PARATHYROID HORNONE
Backqround of the Invention n Parathyroid hormone ("PTH") is a polypeptide 5 produced by the parathyroid glands. The mature circulating form of the hormone is comprised of 84 amino acid residues. The biological action of PTH can be reproduced by a peptide fragment of its N-terminus (e.g.
amino acid residues 1 through 34). Parathyroid hormone-10 related protein ("PTHrP") is a 139 to 173 aminoacid-protein with N-terminal homology to PTH. PTHrP
shares many of the biological effects of PTH including binding to a common PTH/PTHrP receptor. Tregear, et al., Endocrinol., 93:1349 (1983). PTH peptides from many 15 different sources, e.g., human, bovine, rat, chicken, have been characterized. Nissenson, et al., Receptor, 3:193 (1993).
PTH has been shown to both improve bone mass and quality. Dempster, et al., Endocrine Rev., 14:690 (1993); and Riggs, Amer. J. Med., 91 (Suppl. 5B):37S
(1991). The anabolic effect of intermittently a~m;n;~tered PTH has been observed in osteoporotic men and women either with or without concurrent antiresorptive therapy. Slovik, et al., J. Bone Miner.
25 Res., 1:377 (1986); Reeve, et al., Br. Med. J., 301:314 (1990); and Hesch, R-D., et al., Calcif. Tissue Int'l, 44:176 (1989).
Summary of the Invention In one aspect, the invention relates to peptide 30 variants of PTH(1-34) of the following generic formula:
CA 02226l77 l997-l2-30 W O 97/02834 PCTrUS96/11292 R
1~
V 1 A Glu-A5-Gln-A7-A8-His-Asn-Al1 A12 Ly 15 A16 A17 Al8 - Al9 - Arg - A2l - Glu - A23 - A24 - Arg - Lys - A27 - A
wherein A1 is Ser, Ala, or Dap;
o A3 is Ser, Thr, or Aib;
A5 is Leu, Nle, Ile, Cha, ~-Nal, Trp, Pal, Phe or p-X-Phe, in which X is OH, a halogen, or CH3;
A7 is Leu, Nle, Ile, Cha, ~-Nal, Trp, Pal, Phe, or p-X-Phe in which X is OH, a halogen, or CH3;
A8 is Met, Nva, Leu, Val, Ile, Cha, or Nle;
All is Leu, Nle, Ile, Cha, ~-Nal, Trp, Pal, Phe or p-X-Phe in which X is OH, a halogen, or CH3;
A12 is Gly or Aib;
A15 is Leu, Nle, Ile, Cha, ~-Nal, Trp, Pal, Phe, 20 or p-X-Phe in which X is OH, a halogen, or CH3;
A16 is Ser, Asn, Ala, or Aib;
A17 is Ser, Thr, or Aib;
A18 is Met, Nva, Leu, Val, Ile, Nle, Cha, or Aib;
Alg is Glu or Aib;
25 A21 is Val, Cha, or Met;
A23 is Trp or Cha;
A24 is Leu or Cha;
A27 is Lys, Aib, Leu, hArg, Gln, or Cha;
A28 is Leu or Cha;
30 A30 is Asp or Lys;
A31 is Val, Nle, Cha, or deleted;
A32 is His or deleted;
A33 is Asn or deleted;
A34 is Phe, Tyr, Amp, Aib, or deleted;
W O 97/02834 PCT~US96!11292 each of R1 and R2 is, independently, H, Cl_l2 alkyl, C2_12 alkenyl, C7_20 phenylalkyl, Cll_20 napthylalkyl, Cl_l2 hydroxyalkyl, C2_12 hydroxyalkenyl, C7_ 20 hydroxyphenylalkyl, or Cll_20 hydroxynapthylalkyl; or one and only one of Rl and R2 is COE1 in which El is Cl_l2 alkyl, C2_12 alkenyl, C7_20 phenylalkyl, Cll_20 napthylalkyl, C1_12 hydroxyalkyl, C2_12 hydroxyalkenyl, C7_ 20 hydroxy-phenylalkyl, or Cll_20 hydroxynapthylalkyl; and R3 is OH, NH2, Cl_l2 alkoxy, or NH-Y-CH2-Z in which o Y is a C1_12 hydrocarbon moiety and Z is H, OH, C02H, or CoNH2;
provided that (i) at least one of A5, A7, A8, A11, Al5~ A18, A21, A23, A24, A27, A28, and A31 is Cha, or at least one of A3, A12, Al6, Al7, A18, Alg, and A34 is Aib;
lS or that (ii) at least Al is Dap, A7 is ~-Nal, Trp, Pal, Phe, or p-X-Phe, A15 is ~-Nal, Trp, Pal, Phe, or p-X-Phe, A27 is hArg, or A31 is Nle; or a pharmaceutically acceptable salt thereof.
A subset of the compounds covered by the above 20 formula are those in which at least one of A5, A7, All, Als~ Al8, A2l, A23, A24, A27, A28, and A31 is Cha. For example, A3 is Ser; A5 is Ile; A7 is Leu or Cha; A8 is Met, Nva, Leu, Val, Ile, or Nle; A11 is Leu or Cha; A12 is Gly; A15 is Leu or Cha; A16 is Asn or Aib; A17 is Ser; A18 2s is Met or Nle; A21 is Val; A27 is Lys, hArg, or Cha; A32 is His; A31 is Val, Nle, or Cha; A33 is Asn; A34 is Phe, Tyr, Amp, or Aib; Rl is H; R2 is H; and R3 is NH2;
provided that at least one of A5, A7, A8, All, Al5, A18, A2l, A23, A24, A27, A28, and A31 is Cha, or at least one of 30 A3~ A12, A16, A17, Al8, Alg, and A34 is Aib. If desired, at least one of A7 and A11 can be Cha; or at least one of A1s~ A23, A24, A27, A28, and A31 is Cha.
In another subset, at least one of A3, A12, A16, A17, A18, Alg, and A34 is Aib. For example, A3 is Ser or 3s Aib; A5 is Ile; A7 is Leu or Cha; A8 is Met, Nva, Leu, CA 02226l77 l997-l2-30 W O 97/02834 PCT~US96/11292 Val, Ile, or Nle; All is Leu or Cha; A15 is Leu or Cha, A16 is Asn or Aib; A18 is Met, Aib, or Nle; A21 is Val; A27 is Lys, Aib, Leu, hArg, or Cha; A31 is Val, Nle, or Cha;
A32 is His; A33 is Asn; A34 is Phe, Tyr, Amp, or Aib; R1 5 is H; R2 is H; and R3 is NH2; provided that at least one A5~ A7~ A8, A11~ A15, A18, A21, A23~ A24~ A27, A25, and A31 is Cha, or at least one of A3, A12, A16~ A17, A18, A19, and A34 is Aib. If desired, at least one of A7 and A11 can be Cha; or at least one of A1s, A23, A24~ A27~ A28~ and o A31 is Cha.
In a still further subset, at least one of A5, A7, A8, Al1, A15, A18~ A21~ A23, A24, A27, A28, and A31 is Cha, or at least one of A3, A12, A16, A17, A18~ A1s~ and A34 is Aib. For example, A3 is Ser or Aib; A5 is Ile; A7 is Leu 15 or Cha; A8 is Met, Nva, Leu, Val, Ile, or Nle; All is Leu or Cha; A15 is Leu or Cha; A16 is Asn or Aib; A18 is Met, Aib, or Nle; A21 is Val; A27 is Lys, Aib, Leu, hArg, or Cha; A31 is Val, Nle, or Cha; A32 is His; A33 is Asn; A34 is Phe, Tye, Amp, or Aib; R1 is H; R2 is H; and R3 is NH2.
20 If desired, at least one of A7 and All is Cha and at least one of A16, Alg, and A34 is Aib; or at least one of A24, A28, and A31 is Cha and at least one of A16 and A17 is Aib.
In yet another subset, at least one of A1 is Dap, A7 is ~-Nal, Trp, Pal, Phe or p-X-Phe, A13 is ~-Nal, Trp, 25 Pal, Phe, or p-X-Phe. For example, Al is Ser, Gly, or Dap; A3 is Ser or Aib; A8 is Met, Nva, Leu, Val, Ile, or Nle; A16 is Asn or Aib; A18 is Met, Aib, or Nle; A21 is Val; A27 is Lys, Aib, Leu, hArg, or Cha; A31 is Val, Nle, or Cha; A32 is His; A33 is Asn; A34 is Phe, Tyr, Amp, or 30 Aib; Rl is H; R2 is H; and R3 is NH2 The following are examples of the peptide of this invention as covered by the above formula: [Cha7]hPTH(1-34)NH2; [Chall]hPTH(1-34)NH2; [Chal5]hPTH(1-34)NH2; [cha7 ( 34)NH2i [Cha7~ 11, Nle8, 18 Tyr34]h~5 [Cha23]hPTH(1-34)NH2; [Cha24]hPTH(1-34)NH2; [Nle3~ 18, WO 97/02834 PCTrUS96/11292 Cha27]hPTH(1--34)NH2; [Cha28]hPTH(1--34)NH2; [Cha31]hPTH(l--34)NH2; [Cha27]hPTH(1--34)NH2; [Cha27~ 29]hPTH(1--34)NH2;
[Cha28]bPTH(1-34)NH2; [Cha28]rPTH(1-34)NH2; [Cha24~ 28~
31]hPTH(1--34)NH2; [Aibl6]hPTH(1--34)NH2; [Aibl9]hPTH(l--34)NH2; [Aib34]hPTH(1--34)NH2; [Aib16~ 19]hPTH(1--34)NH2;
[Aibl6' 19, 34]bPTH(1--34)NH2; [Aibl6' 34]hPTH(1--34)NH2;
[Aibl9~ 34]hPTH(l-34)NH2; [Cha7~ 11, Nle8~ 13 Aib16, 19 Tyr34]hpTH(l-34)NH2; tCha7~ 11, Nle8, 18, 31 Aib16, 19 Tyr34]hPTH(1-34)NH2; [Cha7, Aibl6]hPTH(1-34)NH2; [Chal1, o Aibl6]hPTH(1-34)2; [Cha7, Aib34]hPTH(1-34)NH2; [Chall, Aib34]hPTH(1-34)NH2; [Cha27, Aibl6]hPTH(1-34)NH2; [Cha27, Aib34]hPTH(1-34)NH2; [Cha23, Aibl6]hPTH(1-34)NH2; [Cha28, Aib34]hPTH(1-34)NH2; [Nle31]hPTH(1-34)NH2; [hArg27]hPTH(l-34)NH ; [Dap1 Nle8~ 18, Tyr34]hPTH(1~34)NH2;
[Nle31]bPTH(1-34)NH2; [Nle3l]rPTH(1-34)NH2; [hArg27]bPTH(l-34)NH2; [hArg27]rPTH(1-34)NH2; [Cha7~ 11, Aib19, Lys30]hPTH(1-34)NH2; [Aibl2]hPTH(1-34)NH2; [Cha24~ 28~ 31, Lys30]hPTH(1-34)NH2; [Cha28~ 31]hPTH(1-34)NH2; [Cha7~ 11, Nle8~ 18, Aib34]hPTH(1-34)NH2; [Aib3]hPTH(1-34)NH2;
[Cha8]hPTH(1-34)NH2; [Chal5]hPTH(1-34)NH2; [Cha7~ 11, Aibl9]hPTH(1-34)NH2; [Cha7~ 1l, Aibl6 ]hPTH(1-34)NH2;
[Aib17]hPTH(1-34)NH2; [Cha5]hPTH(1-34)NH2;[Cha7 15]hPTH(1-34)NH2; [Cha7~ 11, Nle8~ 18, Aibl9, Tyr ]hPTH(1-34)NH2; [Cha7~ l1, Nle8~ 13 Aibl9 L 30 25 Tyr34]hPTH(1-34)NH2; [Cha7~ 15]hPTH(1-34)NH2;
[Aibl7]hPTH(1-34)NH2; [Cha7~ 11, Leu27]hPTH(1-34) NH2;
[Cha7~ 1l, 15, Leu27]hPTH(1-34)NH2; [Cha7~ ll, 27] hPTH(1-Ch 7, 11, 15, 27]hPTH (1-34)NH2; [Trp ]
34)NH2; [Nall5]hPTH(1-34) NH2; [Trpl5, Cha23]hPTH(1-34)NH2;
[Chal5~ 23]hPTH(1-34)NH2; [Phe7~ 11]hPTH(1-34)NH2; [Nal7 ]hPTH(1-34)NH2; [Trp7~ ll]hPTH (1-34)NH2; [Phe7 5]hPTH(1-34)NH2; [Nal7~ l5]hPTH(1-34)NH2; [Trp7 ]hPTH(1-34)NH2; and [Tyr7~ l5]hpTH(l-34)NH
In another aspect, this invention relates to 35 peptides covered by the following formula:
W O 97/02834 PCTrUS96/11292 Al - val - A3 - Glu - A5 - Gln - A7 - A8 - His - Alo - All - Al2 - Lys A14 A15 A16--A17-A18-Alg~Arg~Arg~A22~A23~A24~A25~A26~A27~A28 wherein A1 is Ala, Ser, or Dap;
A3 is Ser or Aib;
A5 is His, Ile, or Cha;
A7 is Leu, Cha, Nle, ~-Nal, Trp, Pal, Phe, or p-X-Phe in which X is OH, a halogen, or CH3;
A8 is Leu, Met, or Cha;
A1o is Asp or Asn;
Al1 is Lys, Leu, Cha, Phe, or ~-Nal;
A12 is Gly or Aib;
A14 is Ser or His;
A15 is Ile, or Cha;
A16 is Gln or Aib; .
A17 is Asp or Aib;
A18 is Leu, Aib, or Cha;
Alg is Arg or Aib;
A22 is Phe, Glu, Aib, or Cha;
A23 is Phe, Leu, Lys, or Cha;
A24 is Leu, Lys, or Cha;
A25 is His, Aib, or Glu;
A26 is His, Aib, or Lys;
A27 is Leu, Lys, or Cha;
A28 is Ile, Leu, Lys, or Cha;
A29 is Ala, Glu, or Aib;
A30 is Glu, Cha, Aib, or Lys;
A31 is Ile, Leu, Cha, Lys, or deleted;
A32 is His or deleted;
A33 is Thr or deletedj CA 02226l77 l997-l2-30 WO 97/02834 PCTrUS96/11292 A34 is Ala or deleted;
each of R1 and R2 is, independently, H, Cl_l2 alkanyl, C7_20 phenylalkyl, C11_20 napthyalkyl, Cl_l2, hydroxyalkyl, C2_12 hydroxyalkenyl, C7_20 hydroxyphenylalkyl, or Cll_20 hydroxynapthylalkyl; or one and only one of Rl and R2 is COEl in which El is Cl_l2 alkyl, C2 12 alkyl, C2_12 alkenyl~ C7_20 phenYlalkYl' 11-20 napthylalkyl, Cl_l2 hydroxyalkyl, C2_12 hydroxyalkenyl, C7_ 20 hydroxyphenylalkyl, or Cll_20 hydroxynapthylalkyl; and lo R3 is OH, NH2, Cl_l2 alkoxy, or NH-Y-CH2-Z in which Y is a C1_12 hydrocarbon moiety and Z is H, OH, C02H or CONH2;
provided that (i) at least one of A5, A7, A8, All, Als~ A18~ A22~ A23~ A24, A27, A28, A30, or A31 is Cha, or at 15 least one of A3, A12, A16, A17, A18, Als~ A22~ A2s~ A26~
A29, A30, or A34 is Aib; or that (ii) at least one of A23, A24, A27, A28, or A31 is Lys; or a pharmaceutically acceptable salt thereof. In one embodiment, at least one of A7 and All is Cha. In another embodiment, at least one 20 of A16 or Alg is Aib. Specific examples of peptides of the just-recited formula include, but are not limited to, [Cha7]hPTHrP(1-34)NH2; [Cha11]hPTHrP(1 -34)NH2; [Cha7 ]hPTHrP(1-34)NH2; [Aib16, Tyr34hPTHrP(1-34)NH2;
[Aibl9]hPTHrP(1-34)NH2; [Aibl6~ 19]hPTHrP(1-34)NH2; [Cha7~
25 11, Aibl6hPTHrP(1-34)NH2; [Cha7~ 1l, Aibl9]hPTHrP(1-34)NH2;
[cha22 LeU23, 28, 31, Glu25, 29, Lys26~ 3o]hpTHrp(l-34)NH
[Glu ~ , 29, LeU23, 28, 31, LyS26, 27, 30]hpTH
[Cha 2~ 23, GlU25~ 29, Leu28, 31 LyS26~ 30]hPTHrP(1 34)NH
[GlU22, 25 LeU23, 28, 31, Aib29, Ly526~ 3o]hpTHrp(l-34)NH
[GlU22, 25, 29 Lys23, 26, 30, Leu28~ 31] hPTHrP(1-34)NH2;
[GlU22, 25, 29 Leu23, 28, 31, LyS26, Cha30]hPTHrP(l-34)NH2;
22, 25, 29 Leu23, 28, 31, LyS26, Aib3o]hpTHrp(l-34)NH2;
[GlU22, 25, 29 Leu23, 31, LyS26, 28~ 30]hPTHrP(1-34) NH2;
[cha22, 23, 24, 27, 28, 31, Glu25, 29, Lys26~ 30]hPTHrP(1-35 34)NH2; [Glu22~ 25~ 29, Cha23, 24, 28, 31 LyS26, 27, CA 02226l77 l997-l2-30 W O 97/02834 PCT~US96/11292 30]hPTHrP(1-34)NH2; tGlU22~ 25, 29, Cha23, 24, 27, 31 LyS26, 28~ 30]hpTHrp(l-34)NH2; [Glu22~ 25, 29 Lys23, 26, 30 Cha24, 27~ 28~ 31]hPTHrP(1-34)NH2; tCha22, Leu23~ 28~ 31, Glu25~ 29, Lys26~ 27~ 30] hPTHrP(1-34)NH2; [Cha22, Leu23- 31, Glu25~ 29, 5 Lys26~ 28~ 30]hPTHrP(1-34)NH2; [Cha22, Lys23~ 26~ 30, Glu25 29, Leu28~ 31]hPTHrP(1-34)NH2; [Cha22 Leu23~ 28~ 31 Glu25 Lys26~ 30, Aib29] hPTHrP(1-34)NH2; [Cha22, Leu23~ 28~ 31, Glu25~ 29 Lys26 Aib30]hPTHrP(1-34)NH2; [Glu22~ 25, Leu23 28~ 31, Lys26~ 27~ 30, Aib29]hPTHrP(1-34)NH2; [Glu22~ 25, 23 26, 30 Leu 28, 31 Aib29] hPTHrP(1-34)NH2; [Glu Leu23, 31 Lys 26, 28, 30 Aib29]hpTHrp(l-34)NH2; [Cha7' 1 GlU22, 25, 29 Leu23, 28, 31, LyS26~ 30] hPTHrP(1-34)NH2;
[Cha ' , 22, LeU23, 28, 31, Glu25' 29 Lys26' 30]h 34)NH2; [Cha7~ 11, Glu22, 25, 29, Leu23~ 28, 31 LyS26, 27, 30]
15 hPTHrP(1-34)NH2; [Cha7~ 11, 22, 23 GlU25, 29 LeU28, 31 Lys26~ 3~]hPTHrP(1-34)NH2; [Cha7~ 11, GlU22, 25, 29 LyS23, 26~ 30, Leu28~ 31]hPTHrP(1-34)NH2; [Cha7~ 11, Glu22~ 25~ 29, LeU23, 31 LyS26, 28, 30] hPTHrP(1-34)NH2; [Cha7~ 11, Glu22' 25 Leu23' 28, 31 Aib29, Lys26~ 3~]hPTHrP(1-34)NH2; [Cha7' , Glu , 25, 29, Leu23~ 28, 31 Lys26 Aib30]hPTH P( 34)NH2; [Chal5, G1U22~ 25, 29, Leu23~ 28, 31 LyS26, 3~]hPTHrP(1-34) NH2; [Chal5~ 22, Leu23, 28, 31 GlU25, 29 Lys26~ 3~]hPTHrP(1-34)NH2; [chal5, Glu22, 25, 29 LeU23, 28, 31, Lys26~ 27~ 30]hPTHrP(1-34)NH2; [Chal5~ 22~ 23, Glu25~ 29, 25 LeU28~ 31 Lys26~ 3~]hPTHrP(1-34) NH2; [Chal5~ Glu22~ 25 LeU23, 28, 31 Aib29, Ly526~ 3~]hpTHrP(l-34)NH2; [Chal5, Glu22, 25, 29 Lys23~ 26, 30, Leu25' 31]hPTHrP(1-34) NH2;
Ch 15 Glu22~ 25~ 29 Leu23~ 28, 31, LyS26, Aib30] hPTHrp 34)NH2; [Chal5, Glu22, 28, 29, Leu23, 31 LyS26, 28, 30 3~]hPTHrP(1-34)NH2; [chal5~ 30, GlU22, 25, 29 Leu23, 28, 31 Lys26]hPTHrP(1-34)NH2; [Cha7~ 8~ 22, Leu23~ 25, 31 Glu25~ 29 Lys26~ 30]hPTHrP(1-34)NH2; [Cha7~ 8, Glu22~ 25~ 29 Leu23~ 25 31, Lys26~ 27~ 30]hPTHrP(1-34)NH2; [Cha7~ 8~ 22~ 23, Glu25~
29 LeU28, 31 LyS26~30] hPTHrP (1-34)NH2; [cha7~ 8~ Glu22' 35 25, 29 LeU23, 28, 31 Lys26,30] hpTHrp(l-34)NH2; [cha7~ 8, W O 97/02834 PCTrUS96/11292 GlU22, 25 LeU23, 28, 31, Aib29 , LyS26, 30]hPTHrP(1-34) NH2;
tCha7~ 8 GlU22, 25, 29 LyS23, 26~ 30, LeU2s, 31]hPTHrP(1-34)NH2; [Cha7~ 8, Glu22~ 25~ 29, LeU23, 28, 31 LyS26 Aib30]
hPTHrP(1-34)NH2; [Cha7~ 8, Glu22~ 25, 29, Leu23~ 31 LyS26, 28~ 3o]hpTHrp(l-34)NH2; [Cha7~ 8, 30 GlU22, 25, 29 LeU23, 28, 31, Lys26]hPTHrP(1-34)NH2; [Serl, Ile5, Cha7~ 11, 22, Met8, Asn , Hisl4, Leu23, 28, 31, Glu25~ 29 LyS26, 30]hpT
NH2; [Ser , Ile5, Cha7~ 11, Met8 Asn10 Hi514 Gl 22 25 29 LeU23, 28, 31 Ly526~ 27~ 3~]hpTHrp(1-34)NH2; [Ser1, lo Ile5, Cha7~ 11, Met8, Asn10, Hi514, GlU22, 25, 29 LeU23, 31 Lys26~ 28~ 3~]hPTHrP(1-34)NH2; Serl, Ile5, Cha7~ 11, Met8, Asn , His14, G1U22~ 25, 29 LYS23, 26, 30 L 28 ( 34)NH2; [Ser1, Ile5, Cha7~ 11 Metg A 1o Hisl4 GlU22, 25 LeU23, 28, 31, Aib29, Lys26~ 3~]hPTHrP(1-34) 15 NH2; [Ser , Ile5, Cha7~ 11, Met8 Asn1o His14 Gl 22 25 29 LeU23, 28, 31 Ly526~ Aib30] PTHrP(1-34)NH2; [Ser1, Iles Cha7~ 11, 22, 23 Met8, A5nlo~ Hisl4, Glu25~ 29, Leu23 31, Lys26~ 30]hPTHrP(1-34) NH2; [Serl, Ile5, Cha7~ 11, 15, Met3, Asn10, Hisl4]hPTHrP(1-34)NH2; [Serl, Ile5, Met3, 20 Asn10, Leull, Hisl4, Aibl6]hPTHrP (1-i4)NH2; [Serl~ Ile5, M t8 A5n1~ LeU11, 28, 31, His14, Cha22~ 23, Glu ~ , LyS26, 30]hPTHrP (1-34)NH2; [Ser1, Ile5, Cha7~ 11 Met8 Asnl~, Hisl4, GlU22~ 25~ 29, Leu23, 28, 31 LyS26, 30]hpTH p (1-34)NH2; [Serl, Ile5, Met8, Asnl~ Hisl4 Chal5 Gl 22 25 25, 29 LeU23, 28, 31, LyS26, 3o]hpTHrp (1-34)NH2; [Ser Ile5, Cha7~ 8, Asn10, Hi5l4, Glu22, 25, 29 LeU23, 28, 31 Lys26~ 30]hPTHrP (l-34)NH2;[Glu22~ 25, 29 LeU23, 28, 31 Lys24~ 26~ 30]hPTHrP(1-34)NH2; [Aib22, Leu23~ 28~ 31, Glu25 29, Lys26~ 30]hPTHrP(1-34)NH2; [Glu22~ 29, Leu23~ 28~ 31, 30 Aib25, Lys26~ 30]hPTHrP(1-34)NH2; [Glu22~ 25~ 29, LeU23~ 28~
31, Aib26, Lys30]hPTHrP (1-34)NH2; [Glu22, 25, 29 LeU23, 28 - Lys 26~ 30, 31] hPTHrP(1-34)NH2; [Ser1, Ile5, Met8, Asn10, Leul1, 23, 28, 31, Hisl4, Cha22, Glu25' 29, Lys26~ 30]
~ hPTHrP(1-34)NH2; [Ser1, Ile5, Met8, Asn10, Leul1~ 28~ 31, 35 HiS14 GlU22, 25, 29, Lys23, 26, 30]pTHrp(l-34)NH2; [Ser1~
CA 02226l77 l997-l2-30 W 097r02834 PCT~US96/11292 Il 5 Met3 Asn10 Leull~ 23~ 28~ 31, Hisl4, Glu2 ~ ~ , Lys26~ 27~ 30] hPTHrP(1-34)NH2; [Serl, Ile5, Met5, Asnl0, Leu1l~ 23~ 31, Hi514, Glu22, 25, 29 LyS26, 28, 30] hPTH P(1 34)NH2; tSer1, Ile5, Met5, A5nlo~ Leull, 23, 28, 31 HiS14 5 GlU22, 25 Aib29, LyS26~ 30] hpTHrp(l-34)NH2; [Serl, Ile5, Met8 Asnl~ LeUll, 23, 28, 31, His14, Glu22~ 25~ 29, Lys Aib30] hPTHrP(1-34)NH2; or [Ser1, Ile5, Met3]hPTHrP(1-34)NH2.
With the exception of the N-terminal amino acid, lo all abbreviations (e.g. Ala or A1) of amino acids in this disclosure stand for the structure of -NH-CH(R)-C0-, wherein R is a side chain of an amino acid (e.g., CH3 for Ala). For the N-terminal amino acid, the abbreviation stands for the structure of =N-CH(R)-C0-, wherein R is a 15 side chain of an amino acid. ~-Nal, Nle, Dap, Cha, Nva, Amp, Pal, and Aib are the abbreviations of the following ~-amino acids: ~-(2-naphthyl)alanine, norleucine, ~,~-diaminopropionic acid, cyclohexylalanine, norvaline, 4-amino-phenylalanine, 3-pyridinylalanine, and ~-20 aminoisobutyric acid, respectively. In the aboveformula, hydroxyalkyl, hydroxyphenyl-alkyl, and hydroxynaphthylalkyl may contain 1-4 hydroxy substituents. Also, COE1 stands for -C=O E1. Examples of -C=O E1 include, but are not limited to, acetyl and 25 phenylpropionyl.
A peptide of this invention is also denoted herein by another format, e.g., [Cha7~ ll]hPTH(1-34)NH2, with the substituted amino acids from the natural sequence placed between the second set of brackets (e.g., Cha7 for Leu7, 30 and Chal1 for Leu11 in hPTH). The abbreviation hPTH
stands for human PTH, hPTHrP for human PTHrP, rPTH for rat PTH, and bPTH for bovine PTH. The numbers between the parentheses refer to the number of amino acids present in the peptide (e.g., hPTH(1-34) is amino acids 1 35 through 34 of the peptide sequence for human PTH). The CA 02226l77 l997-l2-30 W O 97/02834 PCT~US96/11292 sequences for hPTH(1-34), hPTHrP(1-34), bPTH(1-34), and rPTH(1-34) are listed in Nissenson, et al., Receptor, 3:193 (1993). The designation "NH2" in PTH(1-34)NH2 indicates that the C-terminus of the peptide is amidated.
~ 5 PTH(1-34), on the other hand, has a free acid C-terminus.
Each of the peptides of the invention is capable of stimulating the growth of bone in a subject (i.e., a mammal such as a human patient). Thus, it is useful in the treatment of osteoporosis and bone fractures when lo administered alone or concurrently with antiresorptive therapy, e.g., bisphosphonates and calcitonin.
The peptides of this invention can be provided in the form of pharmaceutically acceptable salts. Examples of such salts include, but are not limited to, those formed with organic acids (e.g., acetic, lactic, maleic, citric, malic, ascorbic, succinic, benzoic, methanesulfonic, toluenesulfonic, or pamoic acid), inorganic acids (e.g., hydrochloric acid, sulfuric acid, or phosphoric acid), and polymeric acids (e.g., tannic 20 acid, carboxymethyl cellulose, polylactic, polyglycolic, or copolymers of polylactic-glycolic acids).
A therapeutically effective amount of a peptide of this invention and a pharmaceutically acceptable carrier substance (e.g., magnesium carbonate, lactose, or a 25 phospholipid with which the therapeutic compound can form a micelle) together form a therapeutic composition (e.g., a pill, tablet, capsule, or liquid) for administration (e.g., orally, intravenously, transdermally, pulmonarily, vaginally, subcutaneously, nasally, iontophoretically, or 30 by intratracheally) to a subject. The pill, tablet, or capsule that is to be administered orally can be coated ~ with a substance for protecting the active composition from the gastric acid or intestinal enzymes in the ~ stomach for a period of time sufficient to allow it to pass undigested into the small intestine. The W 097/02834 PCT~US96/11292 therapeutic composition can also be in the form of a biodegradable or nonbiodegradable sustained release formulation for subcutaneous or intramuscular a~r;n;~tration. See, e.g., U.S. Patents 3,773,919 and 5 4,767,628 and PCT Application No. WO 94/15587.
Continuous a~m; n; ~tration can also be achieved using an implantable or external pump (e.g., INFUSAID~ pump). The administration can also be conducted intermittently, e.g., single daily injection, or continuously at a low lo dose, e.g., sustained release formulation.
The dose of a peptide of the present invention for treating the above-mentioned diseases or disorders varies depending upon the manner of administration, the age and the body weight of the subject, and the condition of the subject to be treated, and ultimately will be decided by the attending physician or veterinarian.
Also contemplated within the scope of this invention is a peptide covered by the above generic formula for use in treating diseases or disorders 20 associated with deficiency in bone growth or the like, e.g., osteoporosis or fractures.
Other features and advantages of the present invention will be apparent from the detailed description and from the claims.
Detailed Description of the Invention Based on the description herein, the present invention can be utilized to its fullest extent. The following specific examples are to be construed as merely illustrative, and not limitative of the remainder of the 30 disclosure in any way whatsoever. Further, all publications cited herein are incorporated by reference.
Structure PTH(1-34) has been reported to have two amphophilic alpha helical dom~;n~ See, e.g., Barden, et W O 97/02834 PCTrUS96/11292 al., Biochem., 32:7126 (1992). The first ~-helix is formed between amino acid residues 4 through 13, while the second ~-helix is formed between amino acid residues 21 through 29. Some peptides of this invention contain - 5 the substitution of Cha for one or more residues within or near these two regions of PTH(1-34), e.g., Cha7 and Cha11 within the first ~-helix or Cha27 and Cha23 within the second ~-helix.
Also covered by this invention are variants of lo PTH(1-34) with the substitution of Aib for a residue adjacent to the ~-helixes, e.g., Aib16, Aib19, and Aib34;
hArg27 and Nle31, or the substitution of Dpa for the N-terminal residue.
Svnthesis The peptides of the invention can be prepared by standard solid phase synthesis. See, e.g., Stewart, J.M., et al., Solid Phase Synthesis (Pierce Chemical Co., 2d ed. 1984). The following is a description of how [Aib34]hPTH(1-34)NH2 was prepared. Other peptides of the 20 invention can be prepared in an analogous manner by a person of ordinary skill in the art.
The peptide was synthesized on an Applied Biosystems (Foster City, CA) model 430A peptide synthesizer which was modified to do accelerated Boc-25 chemistry solid phase peptide synthesis. See Schnoize,et al., Int. J. Peptide Protein Res., 90:180 (1992). 4-Methylbenz-hydrylamine (MBHA) resin (Peninsula, Belmont, CA) with the substitution of 0.93 mmol/g was used. The Boc amino acids (Bachem, CA, Torrance, CA; Nova Biochem., 30 LaJolla, CA) were used with the following side chain protection: Boc-Arg(Tos)-OH, Boc-Asp(OcHxl)-OH, Boc-Asn(Xan)-OH, Boc-Glu(OcHxl)-OH, Boc-His(DNP)-OH, Boc-Asn-GH, Boc-Val-OH, Boc-Leu-OH, Boc-Ser-OH, Boc-Gly-OH, Boc-Met-OH, Boc-Gln-OH, Boc-Ile-OH, Boc-Lys(ZClZ)-OH, Boc-CA 02226l77 l997-l2-30 W 097/02834 PCTrUS96/11292 Ser(Bzl)-OH, and Boc-Trp(Fm)-OH. The synthesis was carried out on a 0.14 mmol scale. The Boc groups were removed by treatment with 100% TFA for 2 x 1 min. Boc amino acids (2.5 mmol) were pre-activated with HBTU (2.0 mmol) and DI~A (l.o mL) in 4 mL of DMF and were coupled without prior neutralization o~ the peptide--resin TFA
salt. Coupling times were 5 min except for the Boc-Aib-OH and the following residue, Boc-Asn(Xan)-OH, wherein the coupling times were 20 min.
At the end of the assembly of the peptide chain, the resin was treated with a solution of 20%
mercaptoethanol/10% DIEA in DMF for 2 x 30 min. to remove the DNP group on the His side chain. The N-terminal Boc group was then removed by treatment with 100% TFA for 2 x 15 Z min. After neutralization of the peptide-resin with 10% DIEA in DMF (1 x 1 min.), the formyl group on the side chain of Trp was removed by treatment with a solution of 15% ethanolamine/15% water/70% DMF for 2 x 30 min. The partially-deprotected peptide-resin was washed 20 with DMF and DCM and dried under reduced pressure. The final cleavage was done by stirring the peptide-resin in 10 mL of HF containing 1 mL of anisole at 0~C for 75 min.
HF was removed by a flow of nitrogen. The residue was washed with ether (6 x 10 mL) and extracted with 4N HOAc (6 x 10 mL).
The peptide mixture in the aqueous extract was purified on a reversed-phase preparative high pressure liquid chromatography (HPLC) using a reversed phase Vydac~ C18 column (Nest Group, Southborough, MA). The 30 column was eluted with a linear gradient (10% to 45% of solution B over 130 min.) at a flow rate of 10 mL/min (Solution A = 0.1% agueous TFA; Solution B = acetonitile contA;n;ng 0.1% of TFA). Fractions were collected and checked on analytical HPLC. Those contA; n; ng pure 35 product were combined and lyophilized to dryness. 62.3 W O 97/02834 PCTrUS96/11292 mg of a white solid was obtained. Purity was >99~ based on analytical HPLC analysis. Electro-spray mass spectrometer analysis gave the molecular weight at 4054.7 (in agreement with the calculated molecular weight of 5 4054-7)-- The synthesis and purification of [Cha7~11]hPTH
(1-34)NH2 was carried out in the same manner as the above synthesis of [Aib34]hPTH(1-34)NH2. The protected amino acid Boc-Cha-OH was purchased from Bachem, CA. The o purity of the final product was >98%, and the electron-spray mass spectrometer gave the molecular weight at 4197.0 (calculated molecular weight is 4196.9).
The full names for the abbreviations used above are as follows: Boc for t-butyloxycarbonyl, HF for hydrogen fluoride, Fm for formyl, Xan for xanthyl, Bzl for benzyl, Tos for tosyl, DNP for 2,4-dinitrophenyl, DMF
for dimethylformamide, DCM for dichloromethane, HBTU for 2-(lH-Benzotriazol-1-yl)-1,1,3,3-tetramethyl uronium hexafluorophosphate, DIEA for diisopropylethylamine, HOAc for acetic acid, TFA for trifluoroacetic acid, 2ClZ for 2-chlorobenzyloxycarbonyl and OcHxl for O-cyclohexyl.
The substituents R1 and R2 ~f the above generic formula may be attached to the free amine of the N-terminal amino acid by standard methods known in the art.
For example, alkyl groups, e.g., C1_12 alkyl, may be attached using reductive alkylation. Hydroxyalkyl groups, e.g., C1_12 hydroxyalkyl, may also be attached using reductive alkylation wherein the free hydroxy group is protected 30 with a t-butyl ester. Acyl groups, e.g., COEl, may be attached by coupling the free acid, e.g., E1COOH, to the free amine of the N-terminal amino acid by m i~;ng the - completed resin with 3 molar equivalents of both the free acid and diisopropylcarbodiimide in methylene chloride r 35 for one hour and cycling the resulting resin through steps (a) to (f) in the above wash program. If the free acid contains a free hydroxy group, e.g., p-CA 02226l77 l997-l2-30 hydroxyphenylpropionic acid, then the coupling should be performed with an additional 3 molar equivalents of HOBT.
Other peptides of this invention can be prepared in an analogous manner by a person of ordinary skill in 5 the art.
Functional Assays A. Binding to PTH Receptor The peptides of the invention were tested for their ability to bind to the PTH receptor present on lo SaOS-2 (human osteosarcoma cells). SaOS-2 cells (American Type Culture Collection, Rockville, MD; ATCC
#HTB 85) were maintained in RPMI 1640 medium (Sigma, St.
Louis, MO) supplemented with 10~ fetal bovine serum (FBS) and 2 mM glutamine at 37~C in a humidified atmosphere of 15 5% CO2 in air. The medium was changed every three or four days, and the cells were subcultured every week by trypsinization.
SaOS-2 cells were maintained for four days until they had reached confluence. The medium was replaced 20 with 5~ FBS in RPMI 1640 medium and incubated for 2 hrs at room temperature with 10 x 104 cpm mono-l25I-[Nle8~l8, Tyr34(3-125I)] bPTH(1-34)NH2 in the presence of a competing peptides of the invention at various concentrations between 10-1lM to 10-4 M. The cells were 25 washed four times with ice-cold PBS and lysed with 0.1 M
NaOH, and the radioactivity associated with the cells was counted in a scintillation counter. Synthesis of mono-3 18 T 34(3-l25I)] bPTH(1-34)NH2 was c as described in Goldman, M.E., et al., Endocrinol., 30 123:1468 (1988).
The binding assay was conducted with various peptides of the invention, and the IC50 value, (half maximal inhibition of binding of mono-l25I-[Nle Tyr34(3-l25I)]bPTH(1-34)NH2, for each peptide was 35 calculated.
W O 97/02834 PCTrUS96/11292 As shown in Table I, all of the tested peptides had a high binding affinity for the PTH receptor on the SaOS-2 cell.
B. Stimulation of Adenylate Cyclase Activity s The ability of the peptides of the invention to - induce a biological response in SaOS-2 cells were measured. More specifically, any stimulation of the adenylate cyclase was determined by measuring the level of synthesis of cAMP (adenosine 3',5'-monophosphate) as 10 described previously in Rodan, et al., J. Clin. Invest.
72: 1511 (1983) and Goldman, et al., Endocrinol., 123:1468 (1988). Confluent SAOS-2 cells in 24 wells plates were incubated with 0.5 ~Ci [3H]adenine (26.9 Ci/mmol, New England Nuclear, Boston, MA) in fresh medium 15 at 37~C for 2 hrs, and washed twice with Hank's balanced salt solution (Gibco, Gaithersburg, MD). The cells were treated with 1 mM IBMX [isobutylmethyl-xanthine, Sigma, St. Louis, MO] in fresh medium for 15 min, and the peptides of the invention were added to the medium to 20 incubate for 5 min. The reaction was stopped by the addition of 1.2 M trichloroacetic acid (TCA) (Sigma, St.
Louis, MO) followed by sample neutralization with 4 N
KOH. cAMP was isolated by the two-column chromatographic method (Salmon, et al., 1974, Anal. Biochem. 58, 541).
25 The radioactivity was counted in a scintillation counter (Liquid Scintillation Counter 2200CA, PACKARD, Downers Grove, IL).
The respective EC50 values (half maximal stimulation of adenylate cyclase) for the tested peptides 30 were calculated and shown in Table I. All tested peptides were found to be potent stimulators of adenylate cyclase activity, which is a biochemical pathway ~ indicative as a proximal signal for osteoblast proliferation (e.g., bone growth).
W O 97/02834 PCTrUS96/11292 TABLE I
PEPTIDE Kd (~LM) ECSo (nM) Cha7~ 11]hPTH(1-34)NH2 0.01 0.6 lCha231hPTH(1-34)NH2 0.2 20 ICha24]hPTH(1-34)NH2 Ø1 10 Nlc8~ 18, Cha27]hPTH(1-34)NH2; 0.05 2 lCha28]hPTH(1-34)NH2 0 05 25 ¦Cha31]hPTH(1-34)NH2 Q 03 4 IAibl61hPTH(1-34)NH2; O.OW 0.7 10 lAibl9]hPTH(1-34)NH2; O.OOS 0.6 [Aib34]hPTH(1-34)NH2; 0.007 3 [Nle31]hPTH(1-34)NH2; O.OW 0.7 [hArg27]hPTH(1-34)NH2 0 007 [Dap, Nle8~ 18. Tyr34]hPTH(1-34)NH2 0.1S0 10 15 [cha24, 28, 31, Lys30]hPTH(1-34)NH2; 05 7 [Cha ~ , Nle ~ , Tyr ]hPTH(1-34)NH2 0.006 0.6 [Cha7~ 11 Nle8~ 18 Ajbl6~ 19, Tyr34]hPTH (1-34)NH2 ~.~~S 15 [Cha7~ 11 Nle8, 18, 31 Ajbl6, 19, Tyr34]hPTH(1-34)NH2 O.W 4 lChall]hPTH(1-34)NH2 0.005 2 20 [Cha28~ 31]hPTH(1-34)NH2 0.06 7 [Cha7~ 11,Nle8~ 18, Aib34]hPTH(1-34)NH2 0.03 15 [ChalS]hPT11(1-34)NH2 Q oo5 1.3 [Cha7~11, Aibl9]hPTH(1-34)NH2 0.007 05 [Cha7~11, Aibl6]hPTH(1-34)NH2 O.OW 1.1 2 5 [Aib1fi~ 19]hPTH(1-34)NH2 0.004 0.6 [Aib12]hPTH(1-34)NH2 O.OOS 2 [Aib3]hPTH(1-34)NH2 O.OW 1.1 [Cha7~11, Aibl9, Lys30]hPTH(1-34)NH2 0.004 2 [Cha7]hPTH(1-34)NH2 0.02 2.3 30 [Cha24~28~ 31]hPTH(1-34)NH2 1.0 30 [Aibl7]hPTH(1-34) 0.05 3 [Cha7~ 15]hPTH(1-34) 0.01 lA
W O 97/02834 PCTrUS96111292 Other Embodiments It is to be understood that while the invention has been described in conjunction with the detailed description thereof, that the foregoing description is intended to illustrate and not limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages, and modifications are within the claims.
Backqround of the Invention n Parathyroid hormone ("PTH") is a polypeptide 5 produced by the parathyroid glands. The mature circulating form of the hormone is comprised of 84 amino acid residues. The biological action of PTH can be reproduced by a peptide fragment of its N-terminus (e.g.
amino acid residues 1 through 34). Parathyroid hormone-10 related protein ("PTHrP") is a 139 to 173 aminoacid-protein with N-terminal homology to PTH. PTHrP
shares many of the biological effects of PTH including binding to a common PTH/PTHrP receptor. Tregear, et al., Endocrinol., 93:1349 (1983). PTH peptides from many 15 different sources, e.g., human, bovine, rat, chicken, have been characterized. Nissenson, et al., Receptor, 3:193 (1993).
PTH has been shown to both improve bone mass and quality. Dempster, et al., Endocrine Rev., 14:690 (1993); and Riggs, Amer. J. Med., 91 (Suppl. 5B):37S
(1991). The anabolic effect of intermittently a~m;n;~tered PTH has been observed in osteoporotic men and women either with or without concurrent antiresorptive therapy. Slovik, et al., J. Bone Miner.
25 Res., 1:377 (1986); Reeve, et al., Br. Med. J., 301:314 (1990); and Hesch, R-D., et al., Calcif. Tissue Int'l, 44:176 (1989).
Summary of the Invention In one aspect, the invention relates to peptide 30 variants of PTH(1-34) of the following generic formula:
CA 02226l77 l997-l2-30 W O 97/02834 PCTrUS96/11292 R
1~
V 1 A Glu-A5-Gln-A7-A8-His-Asn-Al1 A12 Ly 15 A16 A17 Al8 - Al9 - Arg - A2l - Glu - A23 - A24 - Arg - Lys - A27 - A
wherein A1 is Ser, Ala, or Dap;
o A3 is Ser, Thr, or Aib;
A5 is Leu, Nle, Ile, Cha, ~-Nal, Trp, Pal, Phe or p-X-Phe, in which X is OH, a halogen, or CH3;
A7 is Leu, Nle, Ile, Cha, ~-Nal, Trp, Pal, Phe, or p-X-Phe in which X is OH, a halogen, or CH3;
A8 is Met, Nva, Leu, Val, Ile, Cha, or Nle;
All is Leu, Nle, Ile, Cha, ~-Nal, Trp, Pal, Phe or p-X-Phe in which X is OH, a halogen, or CH3;
A12 is Gly or Aib;
A15 is Leu, Nle, Ile, Cha, ~-Nal, Trp, Pal, Phe, 20 or p-X-Phe in which X is OH, a halogen, or CH3;
A16 is Ser, Asn, Ala, or Aib;
A17 is Ser, Thr, or Aib;
A18 is Met, Nva, Leu, Val, Ile, Nle, Cha, or Aib;
Alg is Glu or Aib;
25 A21 is Val, Cha, or Met;
A23 is Trp or Cha;
A24 is Leu or Cha;
A27 is Lys, Aib, Leu, hArg, Gln, or Cha;
A28 is Leu or Cha;
30 A30 is Asp or Lys;
A31 is Val, Nle, Cha, or deleted;
A32 is His or deleted;
A33 is Asn or deleted;
A34 is Phe, Tyr, Amp, Aib, or deleted;
W O 97/02834 PCT~US96!11292 each of R1 and R2 is, independently, H, Cl_l2 alkyl, C2_12 alkenyl, C7_20 phenylalkyl, Cll_20 napthylalkyl, Cl_l2 hydroxyalkyl, C2_12 hydroxyalkenyl, C7_ 20 hydroxyphenylalkyl, or Cll_20 hydroxynapthylalkyl; or one and only one of Rl and R2 is COE1 in which El is Cl_l2 alkyl, C2_12 alkenyl, C7_20 phenylalkyl, Cll_20 napthylalkyl, C1_12 hydroxyalkyl, C2_12 hydroxyalkenyl, C7_ 20 hydroxy-phenylalkyl, or Cll_20 hydroxynapthylalkyl; and R3 is OH, NH2, Cl_l2 alkoxy, or NH-Y-CH2-Z in which o Y is a C1_12 hydrocarbon moiety and Z is H, OH, C02H, or CoNH2;
provided that (i) at least one of A5, A7, A8, A11, Al5~ A18, A21, A23, A24, A27, A28, and A31 is Cha, or at least one of A3, A12, Al6, Al7, A18, Alg, and A34 is Aib;
lS or that (ii) at least Al is Dap, A7 is ~-Nal, Trp, Pal, Phe, or p-X-Phe, A15 is ~-Nal, Trp, Pal, Phe, or p-X-Phe, A27 is hArg, or A31 is Nle; or a pharmaceutically acceptable salt thereof.
A subset of the compounds covered by the above 20 formula are those in which at least one of A5, A7, All, Als~ Al8, A2l, A23, A24, A27, A28, and A31 is Cha. For example, A3 is Ser; A5 is Ile; A7 is Leu or Cha; A8 is Met, Nva, Leu, Val, Ile, or Nle; A11 is Leu or Cha; A12 is Gly; A15 is Leu or Cha; A16 is Asn or Aib; A17 is Ser; A18 2s is Met or Nle; A21 is Val; A27 is Lys, hArg, or Cha; A32 is His; A31 is Val, Nle, or Cha; A33 is Asn; A34 is Phe, Tyr, Amp, or Aib; Rl is H; R2 is H; and R3 is NH2;
provided that at least one of A5, A7, A8, All, Al5, A18, A2l, A23, A24, A27, A28, and A31 is Cha, or at least one of 30 A3~ A12, A16, A17, Al8, Alg, and A34 is Aib. If desired, at least one of A7 and A11 can be Cha; or at least one of A1s~ A23, A24, A27, A28, and A31 is Cha.
In another subset, at least one of A3, A12, A16, A17, A18, Alg, and A34 is Aib. For example, A3 is Ser or 3s Aib; A5 is Ile; A7 is Leu or Cha; A8 is Met, Nva, Leu, CA 02226l77 l997-l2-30 W O 97/02834 PCT~US96/11292 Val, Ile, or Nle; All is Leu or Cha; A15 is Leu or Cha, A16 is Asn or Aib; A18 is Met, Aib, or Nle; A21 is Val; A27 is Lys, Aib, Leu, hArg, or Cha; A31 is Val, Nle, or Cha;
A32 is His; A33 is Asn; A34 is Phe, Tyr, Amp, or Aib; R1 5 is H; R2 is H; and R3 is NH2; provided that at least one A5~ A7~ A8, A11~ A15, A18, A21, A23~ A24~ A27, A25, and A31 is Cha, or at least one of A3, A12, A16~ A17, A18, A19, and A34 is Aib. If desired, at least one of A7 and A11 can be Cha; or at least one of A1s, A23, A24~ A27~ A28~ and o A31 is Cha.
In a still further subset, at least one of A5, A7, A8, Al1, A15, A18~ A21~ A23, A24, A27, A28, and A31 is Cha, or at least one of A3, A12, A16, A17, A18~ A1s~ and A34 is Aib. For example, A3 is Ser or Aib; A5 is Ile; A7 is Leu 15 or Cha; A8 is Met, Nva, Leu, Val, Ile, or Nle; All is Leu or Cha; A15 is Leu or Cha; A16 is Asn or Aib; A18 is Met, Aib, or Nle; A21 is Val; A27 is Lys, Aib, Leu, hArg, or Cha; A31 is Val, Nle, or Cha; A32 is His; A33 is Asn; A34 is Phe, Tye, Amp, or Aib; R1 is H; R2 is H; and R3 is NH2.
20 If desired, at least one of A7 and All is Cha and at least one of A16, Alg, and A34 is Aib; or at least one of A24, A28, and A31 is Cha and at least one of A16 and A17 is Aib.
In yet another subset, at least one of A1 is Dap, A7 is ~-Nal, Trp, Pal, Phe or p-X-Phe, A13 is ~-Nal, Trp, 25 Pal, Phe, or p-X-Phe. For example, Al is Ser, Gly, or Dap; A3 is Ser or Aib; A8 is Met, Nva, Leu, Val, Ile, or Nle; A16 is Asn or Aib; A18 is Met, Aib, or Nle; A21 is Val; A27 is Lys, Aib, Leu, hArg, or Cha; A31 is Val, Nle, or Cha; A32 is His; A33 is Asn; A34 is Phe, Tyr, Amp, or 30 Aib; Rl is H; R2 is H; and R3 is NH2 The following are examples of the peptide of this invention as covered by the above formula: [Cha7]hPTH(1-34)NH2; [Chall]hPTH(1-34)NH2; [Chal5]hPTH(1-34)NH2; [cha7 ( 34)NH2i [Cha7~ 11, Nle8, 18 Tyr34]h~5 [Cha23]hPTH(1-34)NH2; [Cha24]hPTH(1-34)NH2; [Nle3~ 18, WO 97/02834 PCTrUS96/11292 Cha27]hPTH(1--34)NH2; [Cha28]hPTH(1--34)NH2; [Cha31]hPTH(l--34)NH2; [Cha27]hPTH(1--34)NH2; [Cha27~ 29]hPTH(1--34)NH2;
[Cha28]bPTH(1-34)NH2; [Cha28]rPTH(1-34)NH2; [Cha24~ 28~
31]hPTH(1--34)NH2; [Aibl6]hPTH(1--34)NH2; [Aibl9]hPTH(l--34)NH2; [Aib34]hPTH(1--34)NH2; [Aib16~ 19]hPTH(1--34)NH2;
[Aibl6' 19, 34]bPTH(1--34)NH2; [Aibl6' 34]hPTH(1--34)NH2;
[Aibl9~ 34]hPTH(l-34)NH2; [Cha7~ 11, Nle8~ 13 Aib16, 19 Tyr34]hpTH(l-34)NH2; tCha7~ 11, Nle8, 18, 31 Aib16, 19 Tyr34]hPTH(1-34)NH2; [Cha7, Aibl6]hPTH(1-34)NH2; [Chal1, o Aibl6]hPTH(1-34)2; [Cha7, Aib34]hPTH(1-34)NH2; [Chall, Aib34]hPTH(1-34)NH2; [Cha27, Aibl6]hPTH(1-34)NH2; [Cha27, Aib34]hPTH(1-34)NH2; [Cha23, Aibl6]hPTH(1-34)NH2; [Cha28, Aib34]hPTH(1-34)NH2; [Nle31]hPTH(1-34)NH2; [hArg27]hPTH(l-34)NH ; [Dap1 Nle8~ 18, Tyr34]hPTH(1~34)NH2;
[Nle31]bPTH(1-34)NH2; [Nle3l]rPTH(1-34)NH2; [hArg27]bPTH(l-34)NH2; [hArg27]rPTH(1-34)NH2; [Cha7~ 11, Aib19, Lys30]hPTH(1-34)NH2; [Aibl2]hPTH(1-34)NH2; [Cha24~ 28~ 31, Lys30]hPTH(1-34)NH2; [Cha28~ 31]hPTH(1-34)NH2; [Cha7~ 11, Nle8~ 18, Aib34]hPTH(1-34)NH2; [Aib3]hPTH(1-34)NH2;
[Cha8]hPTH(1-34)NH2; [Chal5]hPTH(1-34)NH2; [Cha7~ 11, Aibl9]hPTH(1-34)NH2; [Cha7~ 1l, Aibl6 ]hPTH(1-34)NH2;
[Aib17]hPTH(1-34)NH2; [Cha5]hPTH(1-34)NH2;[Cha7 15]hPTH(1-34)NH2; [Cha7~ 11, Nle8~ 18, Aibl9, Tyr ]hPTH(1-34)NH2; [Cha7~ l1, Nle8~ 13 Aibl9 L 30 25 Tyr34]hPTH(1-34)NH2; [Cha7~ 15]hPTH(1-34)NH2;
[Aibl7]hPTH(1-34)NH2; [Cha7~ 11, Leu27]hPTH(1-34) NH2;
[Cha7~ 1l, 15, Leu27]hPTH(1-34)NH2; [Cha7~ ll, 27] hPTH(1-Ch 7, 11, 15, 27]hPTH (1-34)NH2; [Trp ]
34)NH2; [Nall5]hPTH(1-34) NH2; [Trpl5, Cha23]hPTH(1-34)NH2;
[Chal5~ 23]hPTH(1-34)NH2; [Phe7~ 11]hPTH(1-34)NH2; [Nal7 ]hPTH(1-34)NH2; [Trp7~ ll]hPTH (1-34)NH2; [Phe7 5]hPTH(1-34)NH2; [Nal7~ l5]hPTH(1-34)NH2; [Trp7 ]hPTH(1-34)NH2; and [Tyr7~ l5]hpTH(l-34)NH
In another aspect, this invention relates to 35 peptides covered by the following formula:
W O 97/02834 PCTrUS96/11292 Al - val - A3 - Glu - A5 - Gln - A7 - A8 - His - Alo - All - Al2 - Lys A14 A15 A16--A17-A18-Alg~Arg~Arg~A22~A23~A24~A25~A26~A27~A28 wherein A1 is Ala, Ser, or Dap;
A3 is Ser or Aib;
A5 is His, Ile, or Cha;
A7 is Leu, Cha, Nle, ~-Nal, Trp, Pal, Phe, or p-X-Phe in which X is OH, a halogen, or CH3;
A8 is Leu, Met, or Cha;
A1o is Asp or Asn;
Al1 is Lys, Leu, Cha, Phe, or ~-Nal;
A12 is Gly or Aib;
A14 is Ser or His;
A15 is Ile, or Cha;
A16 is Gln or Aib; .
A17 is Asp or Aib;
A18 is Leu, Aib, or Cha;
Alg is Arg or Aib;
A22 is Phe, Glu, Aib, or Cha;
A23 is Phe, Leu, Lys, or Cha;
A24 is Leu, Lys, or Cha;
A25 is His, Aib, or Glu;
A26 is His, Aib, or Lys;
A27 is Leu, Lys, or Cha;
A28 is Ile, Leu, Lys, or Cha;
A29 is Ala, Glu, or Aib;
A30 is Glu, Cha, Aib, or Lys;
A31 is Ile, Leu, Cha, Lys, or deleted;
A32 is His or deleted;
A33 is Thr or deletedj CA 02226l77 l997-l2-30 WO 97/02834 PCTrUS96/11292 A34 is Ala or deleted;
each of R1 and R2 is, independently, H, Cl_l2 alkanyl, C7_20 phenylalkyl, C11_20 napthyalkyl, Cl_l2, hydroxyalkyl, C2_12 hydroxyalkenyl, C7_20 hydroxyphenylalkyl, or Cll_20 hydroxynapthylalkyl; or one and only one of Rl and R2 is COEl in which El is Cl_l2 alkyl, C2 12 alkyl, C2_12 alkenyl~ C7_20 phenYlalkYl' 11-20 napthylalkyl, Cl_l2 hydroxyalkyl, C2_12 hydroxyalkenyl, C7_ 20 hydroxyphenylalkyl, or Cll_20 hydroxynapthylalkyl; and lo R3 is OH, NH2, Cl_l2 alkoxy, or NH-Y-CH2-Z in which Y is a C1_12 hydrocarbon moiety and Z is H, OH, C02H or CONH2;
provided that (i) at least one of A5, A7, A8, All, Als~ A18~ A22~ A23~ A24, A27, A28, A30, or A31 is Cha, or at 15 least one of A3, A12, A16, A17, A18, Als~ A22~ A2s~ A26~
A29, A30, or A34 is Aib; or that (ii) at least one of A23, A24, A27, A28, or A31 is Lys; or a pharmaceutically acceptable salt thereof. In one embodiment, at least one of A7 and All is Cha. In another embodiment, at least one 20 of A16 or Alg is Aib. Specific examples of peptides of the just-recited formula include, but are not limited to, [Cha7]hPTHrP(1-34)NH2; [Cha11]hPTHrP(1 -34)NH2; [Cha7 ]hPTHrP(1-34)NH2; [Aib16, Tyr34hPTHrP(1-34)NH2;
[Aibl9]hPTHrP(1-34)NH2; [Aibl6~ 19]hPTHrP(1-34)NH2; [Cha7~
25 11, Aibl6hPTHrP(1-34)NH2; [Cha7~ 1l, Aibl9]hPTHrP(1-34)NH2;
[cha22 LeU23, 28, 31, Glu25, 29, Lys26~ 3o]hpTHrp(l-34)NH
[Glu ~ , 29, LeU23, 28, 31, LyS26, 27, 30]hpTH
[Cha 2~ 23, GlU25~ 29, Leu28, 31 LyS26~ 30]hPTHrP(1 34)NH
[GlU22, 25 LeU23, 28, 31, Aib29, Ly526~ 3o]hpTHrp(l-34)NH
[GlU22, 25, 29 Lys23, 26, 30, Leu28~ 31] hPTHrP(1-34)NH2;
[GlU22, 25, 29 Leu23, 28, 31, LyS26, Cha30]hPTHrP(l-34)NH2;
22, 25, 29 Leu23, 28, 31, LyS26, Aib3o]hpTHrp(l-34)NH2;
[GlU22, 25, 29 Leu23, 31, LyS26, 28~ 30]hPTHrP(1-34) NH2;
[cha22, 23, 24, 27, 28, 31, Glu25, 29, Lys26~ 30]hPTHrP(1-35 34)NH2; [Glu22~ 25~ 29, Cha23, 24, 28, 31 LyS26, 27, CA 02226l77 l997-l2-30 W O 97/02834 PCT~US96/11292 30]hPTHrP(1-34)NH2; tGlU22~ 25, 29, Cha23, 24, 27, 31 LyS26, 28~ 30]hpTHrp(l-34)NH2; [Glu22~ 25, 29 Lys23, 26, 30 Cha24, 27~ 28~ 31]hPTHrP(1-34)NH2; tCha22, Leu23~ 28~ 31, Glu25~ 29, Lys26~ 27~ 30] hPTHrP(1-34)NH2; [Cha22, Leu23- 31, Glu25~ 29, 5 Lys26~ 28~ 30]hPTHrP(1-34)NH2; [Cha22, Lys23~ 26~ 30, Glu25 29, Leu28~ 31]hPTHrP(1-34)NH2; [Cha22 Leu23~ 28~ 31 Glu25 Lys26~ 30, Aib29] hPTHrP(1-34)NH2; [Cha22, Leu23~ 28~ 31, Glu25~ 29 Lys26 Aib30]hPTHrP(1-34)NH2; [Glu22~ 25, Leu23 28~ 31, Lys26~ 27~ 30, Aib29]hPTHrP(1-34)NH2; [Glu22~ 25, 23 26, 30 Leu 28, 31 Aib29] hPTHrP(1-34)NH2; [Glu Leu23, 31 Lys 26, 28, 30 Aib29]hpTHrp(l-34)NH2; [Cha7' 1 GlU22, 25, 29 Leu23, 28, 31, LyS26~ 30] hPTHrP(1-34)NH2;
[Cha ' , 22, LeU23, 28, 31, Glu25' 29 Lys26' 30]h 34)NH2; [Cha7~ 11, Glu22, 25, 29, Leu23~ 28, 31 LyS26, 27, 30]
15 hPTHrP(1-34)NH2; [Cha7~ 11, 22, 23 GlU25, 29 LeU28, 31 Lys26~ 3~]hPTHrP(1-34)NH2; [Cha7~ 11, GlU22, 25, 29 LyS23, 26~ 30, Leu28~ 31]hPTHrP(1-34)NH2; [Cha7~ 11, Glu22~ 25~ 29, LeU23, 31 LyS26, 28, 30] hPTHrP(1-34)NH2; [Cha7~ 11, Glu22' 25 Leu23' 28, 31 Aib29, Lys26~ 3~]hPTHrP(1-34)NH2; [Cha7' , Glu , 25, 29, Leu23~ 28, 31 Lys26 Aib30]hPTH P( 34)NH2; [Chal5, G1U22~ 25, 29, Leu23~ 28, 31 LyS26, 3~]hPTHrP(1-34) NH2; [Chal5~ 22, Leu23, 28, 31 GlU25, 29 Lys26~ 3~]hPTHrP(1-34)NH2; [chal5, Glu22, 25, 29 LeU23, 28, 31, Lys26~ 27~ 30]hPTHrP(1-34)NH2; [Chal5~ 22~ 23, Glu25~ 29, 25 LeU28~ 31 Lys26~ 3~]hPTHrP(1-34) NH2; [Chal5~ Glu22~ 25 LeU23, 28, 31 Aib29, Ly526~ 3~]hpTHrP(l-34)NH2; [Chal5, Glu22, 25, 29 Lys23~ 26, 30, Leu25' 31]hPTHrP(1-34) NH2;
Ch 15 Glu22~ 25~ 29 Leu23~ 28, 31, LyS26, Aib30] hPTHrp 34)NH2; [Chal5, Glu22, 28, 29, Leu23, 31 LyS26, 28, 30 3~]hPTHrP(1-34)NH2; [chal5~ 30, GlU22, 25, 29 Leu23, 28, 31 Lys26]hPTHrP(1-34)NH2; [Cha7~ 8~ 22, Leu23~ 25, 31 Glu25~ 29 Lys26~ 30]hPTHrP(1-34)NH2; [Cha7~ 8, Glu22~ 25~ 29 Leu23~ 25 31, Lys26~ 27~ 30]hPTHrP(1-34)NH2; [Cha7~ 8~ 22~ 23, Glu25~
29 LeU28, 31 LyS26~30] hPTHrP (1-34)NH2; [cha7~ 8~ Glu22' 35 25, 29 LeU23, 28, 31 Lys26,30] hpTHrp(l-34)NH2; [cha7~ 8, W O 97/02834 PCTrUS96/11292 GlU22, 25 LeU23, 28, 31, Aib29 , LyS26, 30]hPTHrP(1-34) NH2;
tCha7~ 8 GlU22, 25, 29 LyS23, 26~ 30, LeU2s, 31]hPTHrP(1-34)NH2; [Cha7~ 8, Glu22~ 25~ 29, LeU23, 28, 31 LyS26 Aib30]
hPTHrP(1-34)NH2; [Cha7~ 8, Glu22~ 25, 29, Leu23~ 31 LyS26, 28~ 3o]hpTHrp(l-34)NH2; [Cha7~ 8, 30 GlU22, 25, 29 LeU23, 28, 31, Lys26]hPTHrP(1-34)NH2; [Serl, Ile5, Cha7~ 11, 22, Met8, Asn , Hisl4, Leu23, 28, 31, Glu25~ 29 LyS26, 30]hpT
NH2; [Ser , Ile5, Cha7~ 11, Met8 Asn10 Hi514 Gl 22 25 29 LeU23, 28, 31 Ly526~ 27~ 3~]hpTHrp(1-34)NH2; [Ser1, lo Ile5, Cha7~ 11, Met8, Asn10, Hi514, GlU22, 25, 29 LeU23, 31 Lys26~ 28~ 3~]hPTHrP(1-34)NH2; Serl, Ile5, Cha7~ 11, Met8, Asn , His14, G1U22~ 25, 29 LYS23, 26, 30 L 28 ( 34)NH2; [Ser1, Ile5, Cha7~ 11 Metg A 1o Hisl4 GlU22, 25 LeU23, 28, 31, Aib29, Lys26~ 3~]hPTHrP(1-34) 15 NH2; [Ser , Ile5, Cha7~ 11, Met8 Asn1o His14 Gl 22 25 29 LeU23, 28, 31 Ly526~ Aib30] PTHrP(1-34)NH2; [Ser1, Iles Cha7~ 11, 22, 23 Met8, A5nlo~ Hisl4, Glu25~ 29, Leu23 31, Lys26~ 30]hPTHrP(1-34) NH2; [Serl, Ile5, Cha7~ 11, 15, Met3, Asn10, Hisl4]hPTHrP(1-34)NH2; [Serl, Ile5, Met3, 20 Asn10, Leull, Hisl4, Aibl6]hPTHrP (1-i4)NH2; [Serl~ Ile5, M t8 A5n1~ LeU11, 28, 31, His14, Cha22~ 23, Glu ~ , LyS26, 30]hPTHrP (1-34)NH2; [Ser1, Ile5, Cha7~ 11 Met8 Asnl~, Hisl4, GlU22~ 25~ 29, Leu23, 28, 31 LyS26, 30]hpTH p (1-34)NH2; [Serl, Ile5, Met8, Asnl~ Hisl4 Chal5 Gl 22 25 25, 29 LeU23, 28, 31, LyS26, 3o]hpTHrp (1-34)NH2; [Ser Ile5, Cha7~ 8, Asn10, Hi5l4, Glu22, 25, 29 LeU23, 28, 31 Lys26~ 30]hPTHrP (l-34)NH2;[Glu22~ 25, 29 LeU23, 28, 31 Lys24~ 26~ 30]hPTHrP(1-34)NH2; [Aib22, Leu23~ 28~ 31, Glu25 29, Lys26~ 30]hPTHrP(1-34)NH2; [Glu22~ 29, Leu23~ 28~ 31, 30 Aib25, Lys26~ 30]hPTHrP(1-34)NH2; [Glu22~ 25~ 29, LeU23~ 28~
31, Aib26, Lys30]hPTHrP (1-34)NH2; [Glu22, 25, 29 LeU23, 28 - Lys 26~ 30, 31] hPTHrP(1-34)NH2; [Ser1, Ile5, Met8, Asn10, Leul1, 23, 28, 31, Hisl4, Cha22, Glu25' 29, Lys26~ 30]
~ hPTHrP(1-34)NH2; [Ser1, Ile5, Met8, Asn10, Leul1~ 28~ 31, 35 HiS14 GlU22, 25, 29, Lys23, 26, 30]pTHrp(l-34)NH2; [Ser1~
CA 02226l77 l997-l2-30 W 097r02834 PCT~US96/11292 Il 5 Met3 Asn10 Leull~ 23~ 28~ 31, Hisl4, Glu2 ~ ~ , Lys26~ 27~ 30] hPTHrP(1-34)NH2; [Serl, Ile5, Met5, Asnl0, Leu1l~ 23~ 31, Hi514, Glu22, 25, 29 LyS26, 28, 30] hPTH P(1 34)NH2; tSer1, Ile5, Met5, A5nlo~ Leull, 23, 28, 31 HiS14 5 GlU22, 25 Aib29, LyS26~ 30] hpTHrp(l-34)NH2; [Serl, Ile5, Met8 Asnl~ LeUll, 23, 28, 31, His14, Glu22~ 25~ 29, Lys Aib30] hPTHrP(1-34)NH2; or [Ser1, Ile5, Met3]hPTHrP(1-34)NH2.
With the exception of the N-terminal amino acid, lo all abbreviations (e.g. Ala or A1) of amino acids in this disclosure stand for the structure of -NH-CH(R)-C0-, wherein R is a side chain of an amino acid (e.g., CH3 for Ala). For the N-terminal amino acid, the abbreviation stands for the structure of =N-CH(R)-C0-, wherein R is a 15 side chain of an amino acid. ~-Nal, Nle, Dap, Cha, Nva, Amp, Pal, and Aib are the abbreviations of the following ~-amino acids: ~-(2-naphthyl)alanine, norleucine, ~,~-diaminopropionic acid, cyclohexylalanine, norvaline, 4-amino-phenylalanine, 3-pyridinylalanine, and ~-20 aminoisobutyric acid, respectively. In the aboveformula, hydroxyalkyl, hydroxyphenyl-alkyl, and hydroxynaphthylalkyl may contain 1-4 hydroxy substituents. Also, COE1 stands for -C=O E1. Examples of -C=O E1 include, but are not limited to, acetyl and 25 phenylpropionyl.
A peptide of this invention is also denoted herein by another format, e.g., [Cha7~ ll]hPTH(1-34)NH2, with the substituted amino acids from the natural sequence placed between the second set of brackets (e.g., Cha7 for Leu7, 30 and Chal1 for Leu11 in hPTH). The abbreviation hPTH
stands for human PTH, hPTHrP for human PTHrP, rPTH for rat PTH, and bPTH for bovine PTH. The numbers between the parentheses refer to the number of amino acids present in the peptide (e.g., hPTH(1-34) is amino acids 1 35 through 34 of the peptide sequence for human PTH). The CA 02226l77 l997-l2-30 W O 97/02834 PCT~US96/11292 sequences for hPTH(1-34), hPTHrP(1-34), bPTH(1-34), and rPTH(1-34) are listed in Nissenson, et al., Receptor, 3:193 (1993). The designation "NH2" in PTH(1-34)NH2 indicates that the C-terminus of the peptide is amidated.
~ 5 PTH(1-34), on the other hand, has a free acid C-terminus.
Each of the peptides of the invention is capable of stimulating the growth of bone in a subject (i.e., a mammal such as a human patient). Thus, it is useful in the treatment of osteoporosis and bone fractures when lo administered alone or concurrently with antiresorptive therapy, e.g., bisphosphonates and calcitonin.
The peptides of this invention can be provided in the form of pharmaceutically acceptable salts. Examples of such salts include, but are not limited to, those formed with organic acids (e.g., acetic, lactic, maleic, citric, malic, ascorbic, succinic, benzoic, methanesulfonic, toluenesulfonic, or pamoic acid), inorganic acids (e.g., hydrochloric acid, sulfuric acid, or phosphoric acid), and polymeric acids (e.g., tannic 20 acid, carboxymethyl cellulose, polylactic, polyglycolic, or copolymers of polylactic-glycolic acids).
A therapeutically effective amount of a peptide of this invention and a pharmaceutically acceptable carrier substance (e.g., magnesium carbonate, lactose, or a 25 phospholipid with which the therapeutic compound can form a micelle) together form a therapeutic composition (e.g., a pill, tablet, capsule, or liquid) for administration (e.g., orally, intravenously, transdermally, pulmonarily, vaginally, subcutaneously, nasally, iontophoretically, or 30 by intratracheally) to a subject. The pill, tablet, or capsule that is to be administered orally can be coated ~ with a substance for protecting the active composition from the gastric acid or intestinal enzymes in the ~ stomach for a period of time sufficient to allow it to pass undigested into the small intestine. The W 097/02834 PCT~US96/11292 therapeutic composition can also be in the form of a biodegradable or nonbiodegradable sustained release formulation for subcutaneous or intramuscular a~r;n;~tration. See, e.g., U.S. Patents 3,773,919 and 5 4,767,628 and PCT Application No. WO 94/15587.
Continuous a~m; n; ~tration can also be achieved using an implantable or external pump (e.g., INFUSAID~ pump). The administration can also be conducted intermittently, e.g., single daily injection, or continuously at a low lo dose, e.g., sustained release formulation.
The dose of a peptide of the present invention for treating the above-mentioned diseases or disorders varies depending upon the manner of administration, the age and the body weight of the subject, and the condition of the subject to be treated, and ultimately will be decided by the attending physician or veterinarian.
Also contemplated within the scope of this invention is a peptide covered by the above generic formula for use in treating diseases or disorders 20 associated with deficiency in bone growth or the like, e.g., osteoporosis or fractures.
Other features and advantages of the present invention will be apparent from the detailed description and from the claims.
Detailed Description of the Invention Based on the description herein, the present invention can be utilized to its fullest extent. The following specific examples are to be construed as merely illustrative, and not limitative of the remainder of the 30 disclosure in any way whatsoever. Further, all publications cited herein are incorporated by reference.
Structure PTH(1-34) has been reported to have two amphophilic alpha helical dom~;n~ See, e.g., Barden, et W O 97/02834 PCTrUS96/11292 al., Biochem., 32:7126 (1992). The first ~-helix is formed between amino acid residues 4 through 13, while the second ~-helix is formed between amino acid residues 21 through 29. Some peptides of this invention contain - 5 the substitution of Cha for one or more residues within or near these two regions of PTH(1-34), e.g., Cha7 and Cha11 within the first ~-helix or Cha27 and Cha23 within the second ~-helix.
Also covered by this invention are variants of lo PTH(1-34) with the substitution of Aib for a residue adjacent to the ~-helixes, e.g., Aib16, Aib19, and Aib34;
hArg27 and Nle31, or the substitution of Dpa for the N-terminal residue.
Svnthesis The peptides of the invention can be prepared by standard solid phase synthesis. See, e.g., Stewart, J.M., et al., Solid Phase Synthesis (Pierce Chemical Co., 2d ed. 1984). The following is a description of how [Aib34]hPTH(1-34)NH2 was prepared. Other peptides of the 20 invention can be prepared in an analogous manner by a person of ordinary skill in the art.
The peptide was synthesized on an Applied Biosystems (Foster City, CA) model 430A peptide synthesizer which was modified to do accelerated Boc-25 chemistry solid phase peptide synthesis. See Schnoize,et al., Int. J. Peptide Protein Res., 90:180 (1992). 4-Methylbenz-hydrylamine (MBHA) resin (Peninsula, Belmont, CA) with the substitution of 0.93 mmol/g was used. The Boc amino acids (Bachem, CA, Torrance, CA; Nova Biochem., 30 LaJolla, CA) were used with the following side chain protection: Boc-Arg(Tos)-OH, Boc-Asp(OcHxl)-OH, Boc-Asn(Xan)-OH, Boc-Glu(OcHxl)-OH, Boc-His(DNP)-OH, Boc-Asn-GH, Boc-Val-OH, Boc-Leu-OH, Boc-Ser-OH, Boc-Gly-OH, Boc-Met-OH, Boc-Gln-OH, Boc-Ile-OH, Boc-Lys(ZClZ)-OH, Boc-CA 02226l77 l997-l2-30 W 097/02834 PCTrUS96/11292 Ser(Bzl)-OH, and Boc-Trp(Fm)-OH. The synthesis was carried out on a 0.14 mmol scale. The Boc groups were removed by treatment with 100% TFA for 2 x 1 min. Boc amino acids (2.5 mmol) were pre-activated with HBTU (2.0 mmol) and DI~A (l.o mL) in 4 mL of DMF and were coupled without prior neutralization o~ the peptide--resin TFA
salt. Coupling times were 5 min except for the Boc-Aib-OH and the following residue, Boc-Asn(Xan)-OH, wherein the coupling times were 20 min.
At the end of the assembly of the peptide chain, the resin was treated with a solution of 20%
mercaptoethanol/10% DIEA in DMF for 2 x 30 min. to remove the DNP group on the His side chain. The N-terminal Boc group was then removed by treatment with 100% TFA for 2 x 15 Z min. After neutralization of the peptide-resin with 10% DIEA in DMF (1 x 1 min.), the formyl group on the side chain of Trp was removed by treatment with a solution of 15% ethanolamine/15% water/70% DMF for 2 x 30 min. The partially-deprotected peptide-resin was washed 20 with DMF and DCM and dried under reduced pressure. The final cleavage was done by stirring the peptide-resin in 10 mL of HF containing 1 mL of anisole at 0~C for 75 min.
HF was removed by a flow of nitrogen. The residue was washed with ether (6 x 10 mL) and extracted with 4N HOAc (6 x 10 mL).
The peptide mixture in the aqueous extract was purified on a reversed-phase preparative high pressure liquid chromatography (HPLC) using a reversed phase Vydac~ C18 column (Nest Group, Southborough, MA). The 30 column was eluted with a linear gradient (10% to 45% of solution B over 130 min.) at a flow rate of 10 mL/min (Solution A = 0.1% agueous TFA; Solution B = acetonitile contA;n;ng 0.1% of TFA). Fractions were collected and checked on analytical HPLC. Those contA; n; ng pure 35 product were combined and lyophilized to dryness. 62.3 W O 97/02834 PCTrUS96/11292 mg of a white solid was obtained. Purity was >99~ based on analytical HPLC analysis. Electro-spray mass spectrometer analysis gave the molecular weight at 4054.7 (in agreement with the calculated molecular weight of 5 4054-7)-- The synthesis and purification of [Cha7~11]hPTH
(1-34)NH2 was carried out in the same manner as the above synthesis of [Aib34]hPTH(1-34)NH2. The protected amino acid Boc-Cha-OH was purchased from Bachem, CA. The o purity of the final product was >98%, and the electron-spray mass spectrometer gave the molecular weight at 4197.0 (calculated molecular weight is 4196.9).
The full names for the abbreviations used above are as follows: Boc for t-butyloxycarbonyl, HF for hydrogen fluoride, Fm for formyl, Xan for xanthyl, Bzl for benzyl, Tos for tosyl, DNP for 2,4-dinitrophenyl, DMF
for dimethylformamide, DCM for dichloromethane, HBTU for 2-(lH-Benzotriazol-1-yl)-1,1,3,3-tetramethyl uronium hexafluorophosphate, DIEA for diisopropylethylamine, HOAc for acetic acid, TFA for trifluoroacetic acid, 2ClZ for 2-chlorobenzyloxycarbonyl and OcHxl for O-cyclohexyl.
The substituents R1 and R2 ~f the above generic formula may be attached to the free amine of the N-terminal amino acid by standard methods known in the art.
For example, alkyl groups, e.g., C1_12 alkyl, may be attached using reductive alkylation. Hydroxyalkyl groups, e.g., C1_12 hydroxyalkyl, may also be attached using reductive alkylation wherein the free hydroxy group is protected 30 with a t-butyl ester. Acyl groups, e.g., COEl, may be attached by coupling the free acid, e.g., E1COOH, to the free amine of the N-terminal amino acid by m i~;ng the - completed resin with 3 molar equivalents of both the free acid and diisopropylcarbodiimide in methylene chloride r 35 for one hour and cycling the resulting resin through steps (a) to (f) in the above wash program. If the free acid contains a free hydroxy group, e.g., p-CA 02226l77 l997-l2-30 hydroxyphenylpropionic acid, then the coupling should be performed with an additional 3 molar equivalents of HOBT.
Other peptides of this invention can be prepared in an analogous manner by a person of ordinary skill in 5 the art.
Functional Assays A. Binding to PTH Receptor The peptides of the invention were tested for their ability to bind to the PTH receptor present on lo SaOS-2 (human osteosarcoma cells). SaOS-2 cells (American Type Culture Collection, Rockville, MD; ATCC
#HTB 85) were maintained in RPMI 1640 medium (Sigma, St.
Louis, MO) supplemented with 10~ fetal bovine serum (FBS) and 2 mM glutamine at 37~C in a humidified atmosphere of 15 5% CO2 in air. The medium was changed every three or four days, and the cells were subcultured every week by trypsinization.
SaOS-2 cells were maintained for four days until they had reached confluence. The medium was replaced 20 with 5~ FBS in RPMI 1640 medium and incubated for 2 hrs at room temperature with 10 x 104 cpm mono-l25I-[Nle8~l8, Tyr34(3-125I)] bPTH(1-34)NH2 in the presence of a competing peptides of the invention at various concentrations between 10-1lM to 10-4 M. The cells were 25 washed four times with ice-cold PBS and lysed with 0.1 M
NaOH, and the radioactivity associated with the cells was counted in a scintillation counter. Synthesis of mono-3 18 T 34(3-l25I)] bPTH(1-34)NH2 was c as described in Goldman, M.E., et al., Endocrinol., 30 123:1468 (1988).
The binding assay was conducted with various peptides of the invention, and the IC50 value, (half maximal inhibition of binding of mono-l25I-[Nle Tyr34(3-l25I)]bPTH(1-34)NH2, for each peptide was 35 calculated.
W O 97/02834 PCTrUS96/11292 As shown in Table I, all of the tested peptides had a high binding affinity for the PTH receptor on the SaOS-2 cell.
B. Stimulation of Adenylate Cyclase Activity s The ability of the peptides of the invention to - induce a biological response in SaOS-2 cells were measured. More specifically, any stimulation of the adenylate cyclase was determined by measuring the level of synthesis of cAMP (adenosine 3',5'-monophosphate) as 10 described previously in Rodan, et al., J. Clin. Invest.
72: 1511 (1983) and Goldman, et al., Endocrinol., 123:1468 (1988). Confluent SAOS-2 cells in 24 wells plates were incubated with 0.5 ~Ci [3H]adenine (26.9 Ci/mmol, New England Nuclear, Boston, MA) in fresh medium 15 at 37~C for 2 hrs, and washed twice with Hank's balanced salt solution (Gibco, Gaithersburg, MD). The cells were treated with 1 mM IBMX [isobutylmethyl-xanthine, Sigma, St. Louis, MO] in fresh medium for 15 min, and the peptides of the invention were added to the medium to 20 incubate for 5 min. The reaction was stopped by the addition of 1.2 M trichloroacetic acid (TCA) (Sigma, St.
Louis, MO) followed by sample neutralization with 4 N
KOH. cAMP was isolated by the two-column chromatographic method (Salmon, et al., 1974, Anal. Biochem. 58, 541).
25 The radioactivity was counted in a scintillation counter (Liquid Scintillation Counter 2200CA, PACKARD, Downers Grove, IL).
The respective EC50 values (half maximal stimulation of adenylate cyclase) for the tested peptides 30 were calculated and shown in Table I. All tested peptides were found to be potent stimulators of adenylate cyclase activity, which is a biochemical pathway ~ indicative as a proximal signal for osteoblast proliferation (e.g., bone growth).
W O 97/02834 PCTrUS96/11292 TABLE I
PEPTIDE Kd (~LM) ECSo (nM) Cha7~ 11]hPTH(1-34)NH2 0.01 0.6 lCha231hPTH(1-34)NH2 0.2 20 ICha24]hPTH(1-34)NH2 Ø1 10 Nlc8~ 18, Cha27]hPTH(1-34)NH2; 0.05 2 lCha28]hPTH(1-34)NH2 0 05 25 ¦Cha31]hPTH(1-34)NH2 Q 03 4 IAibl61hPTH(1-34)NH2; O.OW 0.7 10 lAibl9]hPTH(1-34)NH2; O.OOS 0.6 [Aib34]hPTH(1-34)NH2; 0.007 3 [Nle31]hPTH(1-34)NH2; O.OW 0.7 [hArg27]hPTH(1-34)NH2 0 007 [Dap, Nle8~ 18. Tyr34]hPTH(1-34)NH2 0.1S0 10 15 [cha24, 28, 31, Lys30]hPTH(1-34)NH2; 05 7 [Cha ~ , Nle ~ , Tyr ]hPTH(1-34)NH2 0.006 0.6 [Cha7~ 11 Nle8~ 18 Ajbl6~ 19, Tyr34]hPTH (1-34)NH2 ~.~~S 15 [Cha7~ 11 Nle8, 18, 31 Ajbl6, 19, Tyr34]hPTH(1-34)NH2 O.W 4 lChall]hPTH(1-34)NH2 0.005 2 20 [Cha28~ 31]hPTH(1-34)NH2 0.06 7 [Cha7~ 11,Nle8~ 18, Aib34]hPTH(1-34)NH2 0.03 15 [ChalS]hPT11(1-34)NH2 Q oo5 1.3 [Cha7~11, Aibl9]hPTH(1-34)NH2 0.007 05 [Cha7~11, Aibl6]hPTH(1-34)NH2 O.OW 1.1 2 5 [Aib1fi~ 19]hPTH(1-34)NH2 0.004 0.6 [Aib12]hPTH(1-34)NH2 O.OOS 2 [Aib3]hPTH(1-34)NH2 O.OW 1.1 [Cha7~11, Aibl9, Lys30]hPTH(1-34)NH2 0.004 2 [Cha7]hPTH(1-34)NH2 0.02 2.3 30 [Cha24~28~ 31]hPTH(1-34)NH2 1.0 30 [Aibl7]hPTH(1-34) 0.05 3 [Cha7~ 15]hPTH(1-34) 0.01 lA
W O 97/02834 PCTrUS96111292 Other Embodiments It is to be understood that while the invention has been described in conjunction with the detailed description thereof, that the foregoing description is intended to illustrate and not limit the scope of the invention, which is defined by the scope of the appended claims. Other aspects, advantages, and modifications are within the claims.
Claims (31)
1. A peptide of the formula:
wherein A1 is Ser, Ala, or Dap;
A3 is Ser, Thr, or Aib;
A5 is Leu, Nle, Ile, Cha, .beta.-Nal, Trp, Pal, Phe or p-X-Phe, in which X is OH, a halogen, or CH3;
A7 is Leu, Nle, Ile, Cha, .beta.-Nal, Trp, Pal, Phe, or p-X-Phe in which X is OH, a halogen, or CH3;
A8 is Met, Nva, Leu, Val, Ile, Cha, or Nle;
A11 is Leu, Nle, Ile, Cha, ,.beta.-Nal, Trp, Pal, Phe or p-X-Phe in which X is OH, a halogen, or CH3;
A12 is Gly or Aib;
A15 is Leu, Nle, Ile, Cha, .beta.-Nal, Trp, Pal, Phe, or p-X-Phe in which X is OH, a halogen, or CH3;
A16 is Ser, Asn, Ala, or Aib;
A17 is Ser, Thr, or Aib;
A18 is Met, Nva, Leu, Val, Ile, Nle, Cha, or Aib;
A19 is Glu or Aib;
A21 is Val, Cha, or Met;
A23 is Trp or Cha;
A24 is Leu or Cha;
A27 is Lys, Aib, Leu, hArg, Gln, or Cha;
A28 is Leu or Cha;
A30 is Asp or Lys;
A31 is Val, Nle, Cha, or deleted;
A32 is His or deleted;
A33 is Asn or deleted;
A34 is Phe, Tyr, Amp, Aib, or deleted;
each of R1 and R2 is, independently, H, C1-12 alkyl, C2-12 alkenyl, C7-20 phenylalkyl, C11-20 napthylalkyl, C1-12 hydroxyalkyl, C2-12 hydroxyalkenyl, C7-20 hydroxyphenylalkyl, or C11-20 hydroxynapthylalkyl; or one and only one of R1 and R2 is COE1 in which E1 is C1-12 alkyl, C2-12 alkenyl, C7-20 phenylalkyl, C11-20 napthylalkyl, C1-12 hydroxyalkyl, C2-12 hydroxyalkenyl, C7-20 hydroxy-phenylalkyl, or C11-20 hydroxynapthylalkyl; and R3 is OH, NH2, C1-12 alkoxy, or NH-Y-CH2-Z in which Y is a C1-12 hydrocarbon moiety and Z is H, OH, CO2H, or CONH2;
provided that at least one of A5, A7, A8, A11, A15, A18, A21, A23, A24, A27, A28, and A31 is Cha, or at least one of A3, A12, A16, A17, A18, A19, and A34 is Aib; or a pharmaceutically acceptable salt thereof.
wherein A1 is Ser, Ala, or Dap;
A3 is Ser, Thr, or Aib;
A5 is Leu, Nle, Ile, Cha, .beta.-Nal, Trp, Pal, Phe or p-X-Phe, in which X is OH, a halogen, or CH3;
A7 is Leu, Nle, Ile, Cha, .beta.-Nal, Trp, Pal, Phe, or p-X-Phe in which X is OH, a halogen, or CH3;
A8 is Met, Nva, Leu, Val, Ile, Cha, or Nle;
A11 is Leu, Nle, Ile, Cha, ,.beta.-Nal, Trp, Pal, Phe or p-X-Phe in which X is OH, a halogen, or CH3;
A12 is Gly or Aib;
A15 is Leu, Nle, Ile, Cha, .beta.-Nal, Trp, Pal, Phe, or p-X-Phe in which X is OH, a halogen, or CH3;
A16 is Ser, Asn, Ala, or Aib;
A17 is Ser, Thr, or Aib;
A18 is Met, Nva, Leu, Val, Ile, Nle, Cha, or Aib;
A19 is Glu or Aib;
A21 is Val, Cha, or Met;
A23 is Trp or Cha;
A24 is Leu or Cha;
A27 is Lys, Aib, Leu, hArg, Gln, or Cha;
A28 is Leu or Cha;
A30 is Asp or Lys;
A31 is Val, Nle, Cha, or deleted;
A32 is His or deleted;
A33 is Asn or deleted;
A34 is Phe, Tyr, Amp, Aib, or deleted;
each of R1 and R2 is, independently, H, C1-12 alkyl, C2-12 alkenyl, C7-20 phenylalkyl, C11-20 napthylalkyl, C1-12 hydroxyalkyl, C2-12 hydroxyalkenyl, C7-20 hydroxyphenylalkyl, or C11-20 hydroxynapthylalkyl; or one and only one of R1 and R2 is COE1 in which E1 is C1-12 alkyl, C2-12 alkenyl, C7-20 phenylalkyl, C11-20 napthylalkyl, C1-12 hydroxyalkyl, C2-12 hydroxyalkenyl, C7-20 hydroxy-phenylalkyl, or C11-20 hydroxynapthylalkyl; and R3 is OH, NH2, C1-12 alkoxy, or NH-Y-CH2-Z in which Y is a C1-12 hydrocarbon moiety and Z is H, OH, CO2H, or CONH2;
provided that at least one of A5, A7, A8, A11, A15, A18, A21, A23, A24, A27, A28, and A31 is Cha, or at least one of A3, A12, A16, A17, A18, A19, and A34 is Aib; or a pharmaceutically acceptable salt thereof.
2. A peptide of claim 1, wherein at least one of A7, A11, A15, A23, A24, A27, A28, and A31 is Cha; or a pharmaceutically acceptable salt thereof.
3. A peptide of claim 2, wherein A3 is Ser;
A5 is Ile;
A7 is Leu or Cha;
A8 is Met, Nva, Leu, Val, Ile, or Nle;
A11 is Leu or Cha;
A12 is Gly;
A15 is Leu or Cha;
A16 is Asn or Aib;
A17 is Ser;
A18 is Met or Nle;
A21 is Val;
A27 is Lys, hArg, or Cha;
A32 is His;
A31 is Val, Nle, or Cha;
A33 is Asn;
A34 is Phe, Tyr, Amp, or Aib;
R1 is H;
R2 is H; and R3 is NH2;
or a pharmaceutically acceptable salt thereof.
A5 is Ile;
A7 is Leu or Cha;
A8 is Met, Nva, Leu, Val, Ile, or Nle;
A11 is Leu or Cha;
A12 is Gly;
A15 is Leu or Cha;
A16 is Asn or Aib;
A17 is Ser;
A18 is Met or Nle;
A21 is Val;
A27 is Lys, hArg, or Cha;
A32 is His;
A31 is Val, Nle, or Cha;
A33 is Asn;
A34 is Phe, Tyr, Amp, or Aib;
R1 is H;
R2 is H; and R3 is NH2;
or a pharmaceutically acceptable salt thereof.
4. A peptide of claim 3, wherein at least one of A7 and A11 is Cha; or a pharmaceutically acceptable salt thereof.
5. A peptide of claim 4, wherein said peptide is [Cha7, 11]hPTH(1-34)NH2, [Cha7, 11, Nle8, 18, Tyr34]hPTH(1-34)NH2; [Cha11]hPTH(1-34)NH2; [Cha7,11,15]hPTH(1-34)NH2; or [Cha7]hPTH(1-34)NH2; or a pharmaceutically acceptable salt thereof.
6. A peptide of claim 3, wherein at least one of A15, A23, A24, A27, A28, and A31 is Cha; or a pharmaceutically acceptable salt thereof.
7. A peptide of claim 6, wherein said peptide is [Cha23]hPTH(1-34)NH2, [Cha24]hPTH(1-34)NH2, [Nle8, 18, Cha27]hPTH (1-34)NH2, [Cha23]hPTH(1-34)NH2, [Cha31]hPTH(1-34 NH2, [Cha24, 28, 31]hPTH(1-34)NH2; [Cha24, 28, 31, Lys30]hPTH(1-34)NH2; [Cha28, 31]hPTH(1-34)NH2; or [Cha15]hPTH(1-34)NH2; or a pharmaceutically acceptable salt thereof.
8. A peptide of claim 1, wherein at least one of A3, A12, A16, A17, A18, A19, and A34 is Aib; or a pharmaceutically acceptable salt thereof.
9. A peptide of claim 8, wherein A3 is Ser or Aib;
A5 is Ile;
A7 is Leu or Cha;
A8 is Met, Nva, Leu, Val, Ile, or Nle;
A11 is Leu or Cha;
A15 is Leu or Cha;
A16 is Asn or Aib;
A18 is Met, Aib, or Nle;
A21 is Val;
A27 is Lys, Aib, Leu, hArg, or Cha;
A31 is Val, Nle, or Cha;
A32 is His;
A33 is Asn;
A34 is Phe, Tyr, Amp, or Aib;
R1 is H;
R2 is H; and R3 is NH2;
or a pharmaceutically acceptable salt thereof.
A5 is Ile;
A7 is Leu or Cha;
A8 is Met, Nva, Leu, Val, Ile, or Nle;
A11 is Leu or Cha;
A15 is Leu or Cha;
A16 is Asn or Aib;
A18 is Met, Aib, or Nle;
A21 is Val;
A27 is Lys, Aib, Leu, hArg, or Cha;
A31 is Val, Nle, or Cha;
A32 is His;
A33 is Asn;
A34 is Phe, Tyr, Amp, or Aib;
R1 is H;
R2 is H; and R3 is NH2;
or a pharmaceutically acceptable salt thereof.
10. A peptide of claim 9, wherein at least one of A3, A12, A16, A17, A19, and A34 is Aib; or a pharmaceutically acceptable salt thereof.
11. A peptide of claim 10, wherein said peptide is [Aib16]hPTH(1-34)NH2, [Aib19]hPTH(1-34)NH2, [Aib34]hPTH(1-34)NH2; [Aib16, 19]hPTH(1-34)NH2;
[Aib3]hPTH(1-34)NH2; [Aib17]hPTH(1-34)NH2; or [Aib12]hPTH(1-34)NH2; or a pharmaceutically acceptable salt thereof.
[Aib3]hPTH(1-34)NH2; [Aib17]hPTH(1-34)NH2; or [Aib12]hPTH(1-34)NH2; or a pharmaceutically acceptable salt thereof.
12. A peptide of claim 1 wherein at least one of A7, A11, A15, A23, A24, A27, A28, and A31 is Cha and at east one of A3, A12, A16. A17, A18, A19, and A34 is Aib;
or a pharmaceutically acceptable salt thereof.
or a pharmaceutically acceptable salt thereof.
13. A peptide of claim 12, wherein A3 is Ser or Aib;
A5 is Ile;
A7 is Leu or Cha;
A8 is Met, Nva, Leu, Val, Ile, or Nle;
A11 is Leu or Cha;
A15 is Leu or Cha;
A16 is Asn or Aib;
A18 is Met, Aib, or Nle;
A21 is Val;
A27 is Lys, Aib, Leu, hArg, or Cha;
A31 is Val, Nle, or Cha;
A32 is His;
A33 is Asn;
A34 is Phe, Tye, Amp, or Aib;
R1 is H;
R2 is H; and R3 is NH2;
or a pharmaceutically acceptable salt thereof.
A5 is Ile;
A7 is Leu or Cha;
A8 is Met, Nva, Leu, Val, Ile, or Nle;
A11 is Leu or Cha;
A15 is Leu or Cha;
A16 is Asn or Aib;
A18 is Met, Aib, or Nle;
A21 is Val;
A27 is Lys, Aib, Leu, hArg, or Cha;
A31 is Val, Nle, or Cha;
A32 is His;
A33 is Asn;
A34 is Phe, Tye, Amp, or Aib;
R1 is H;
R2 is H; and R3 is NH2;
or a pharmaceutically acceptable salt thereof.
14. A peptide of claim 13, wherein at least one of A7 and A11 is Cha and at least one of A16, A19, and A34 is Aib; or a pharmaceutically acceptable salt thereof.
15. A peptide of claim 14, wherein said peptide is [Cha7, 11 Nle8, 18, Aib 16, 19, Tyr34]hPTH(1-34)NH2, [Cha7, 11, Nle8, 18, 31, Aib16, 19, Tyr34]hPTH(1-34)NH2;
[Cha7, 11, Aib19]hPTH(1-34)NH2; [Cha7, 11, Aib16]hPTH(1-34)NH ;
[Cha7, 11, Nle8, 18, Aib34]hPTH(1-34)NH2; or [Cha7, 11, Aib19, Lys30]hPTH(1-34)NH2; or a pharmaceutically acceptable salt thereof.
[Cha7, 11, Aib19]hPTH(1-34)NH2; [Cha7, 11, Aib16]hPTH(1-34)NH ;
[Cha7, 11, Nle8, 18, Aib34]hPTH(1-34)NH2; or [Cha7, 11, Aib19, Lys30]hPTH(1-34)NH2; or a pharmaceutically acceptable salt thereof.
16. A peptide of claim 13, wherein at least one of A24, A28, and A31 is Cha and at least one of A16 and A17 is Aib; or a pharmaceutically acceptable salt thereof.
17. A peptide of claim 16, wherein said peptide is [Cha28 Nle8, 18, Aib16, 19, Tyr34]hPTH(1-34)NH2, or [Cha28, Aib16,19] PTH(1-34)NH2; or a pharmaceutically acceptable salt thereof.
18. A peptide of the formula:
wherein A3 is Ser, Thr, or Aib;
A5 is Leu, Nle, Ile, Cha, .beta.-Nal, Trp, Pal, Phe or p-X-Phe, in which X is OH, a halogen, or CH3;
A7 is Leu, Ile, Nle, Cha, .beta.-Nal, Trp, Pal, Phe, or p-X-Phe in which X is H, OH, a halogen, or CH3;
A8 is Met, Nva, Leu, Val, Ile, Cha, or Nle;
A11 is Leu, Nle, Ile, Cha, .beta.-Nal, Trp, Pal, Phe or p-X-Phe in which X is OH, a halogen, or CH3;
A12 is Gly or Aib;
A15 is Leu, Nle, Ile, Cha, .beta.-Nal, Trp, Pal, Phe, or p-X-Phe in which X is OH, a halogen, or CH3;
A16 is Ser, Asn, Ala, or Aib;
A17 is Ser, Thr, or Aib;
A18 is Met, Nva, Leu, Val, Ile, Nle, Cha, or Aib;
A19 is Glu or Aib;
A21 is Val, Cha, or Met;
A23 is Trp or Cha;
A24 is Leu or Cha;
A27 is Lys, Aib, Leu, hArg, Gln, or Cha;
A28 is Leu or Cha;
A30 is Asp or Lys;
A31 is Val, Nle, Cha, or deleted;
A32 is His or deleted;
A33 is Asn or deleted;
A34 is Phe, Tyr, Amp, Aib, or deleted;
each of R1 and R2 is, independently, H, C1-12 alkyl, C2-12 alkenyl, C7-20 phenylalkyl, C11-20 napthylalkyl, C1-12 hydroxyalkyl, C2-12 hydroxyalkenyl, C7-20 hydroxyphenylalkyl, or C11-20 hydroxynapthylalkyl; or one and only one of R1 and R2 is COE1 in which E1 is C1-12 alkyl, C2-12 alkenyl, C7-20 phenylalkyl, C11-20 napthylalkyl, C1-12 hydroxyalkyl, C2-12 hydroxyalkenyl, C7-20 hydroxy-phenylalkyl, or C11-20 hydroxynapthylalkyl;
R3 is OH, NH2, C1-12 alkoxy, or NH-Y-CH2-Z in which Y is a C1-12 hydrocarbon moiety and Z is H, OH, CO2H, or CONH2;
provided that at least A1 is Dap, A7 is .beta.-Nal, Trp, Pal, Phe, or p-X-Phe; A15 is .beta.-Nal, Trp, Pal, Phe, or p-X-Phe, A27 is hArg, or A31 is Nle; or a pharmaceutically acceptable salt thereof.
wherein A3 is Ser, Thr, or Aib;
A5 is Leu, Nle, Ile, Cha, .beta.-Nal, Trp, Pal, Phe or p-X-Phe, in which X is OH, a halogen, or CH3;
A7 is Leu, Ile, Nle, Cha, .beta.-Nal, Trp, Pal, Phe, or p-X-Phe in which X is H, OH, a halogen, or CH3;
A8 is Met, Nva, Leu, Val, Ile, Cha, or Nle;
A11 is Leu, Nle, Ile, Cha, .beta.-Nal, Trp, Pal, Phe or p-X-Phe in which X is OH, a halogen, or CH3;
A12 is Gly or Aib;
A15 is Leu, Nle, Ile, Cha, .beta.-Nal, Trp, Pal, Phe, or p-X-Phe in which X is OH, a halogen, or CH3;
A16 is Ser, Asn, Ala, or Aib;
A17 is Ser, Thr, or Aib;
A18 is Met, Nva, Leu, Val, Ile, Nle, Cha, or Aib;
A19 is Glu or Aib;
A21 is Val, Cha, or Met;
A23 is Trp or Cha;
A24 is Leu or Cha;
A27 is Lys, Aib, Leu, hArg, Gln, or Cha;
A28 is Leu or Cha;
A30 is Asp or Lys;
A31 is Val, Nle, Cha, or deleted;
A32 is His or deleted;
A33 is Asn or deleted;
A34 is Phe, Tyr, Amp, Aib, or deleted;
each of R1 and R2 is, independently, H, C1-12 alkyl, C2-12 alkenyl, C7-20 phenylalkyl, C11-20 napthylalkyl, C1-12 hydroxyalkyl, C2-12 hydroxyalkenyl, C7-20 hydroxyphenylalkyl, or C11-20 hydroxynapthylalkyl; or one and only one of R1 and R2 is COE1 in which E1 is C1-12 alkyl, C2-12 alkenyl, C7-20 phenylalkyl, C11-20 napthylalkyl, C1-12 hydroxyalkyl, C2-12 hydroxyalkenyl, C7-20 hydroxy-phenylalkyl, or C11-20 hydroxynapthylalkyl;
R3 is OH, NH2, C1-12 alkoxy, or NH-Y-CH2-Z in which Y is a C1-12 hydrocarbon moiety and Z is H, OH, CO2H, or CONH2;
provided that at least A1 is Dap, A7 is .beta.-Nal, Trp, Pal, Phe, or p-X-Phe; A15 is .beta.-Nal, Trp, Pal, Phe, or p-X-Phe, A27 is hArg, or A31 is Nle; or a pharmaceutically acceptable salt thereof.
19. A peptide of claim 18, wherein A1 is Ser, Gly, or Dap;
A3 is Ser or Aib;
A8 is Met, Nva, Leu, Val, Ile, or Nle;
A16 is Asn or Aib;
A18 is Met, Aib, or Nle;
A21 is Val;
A27 is Lys, Aib, Leu, hArg, or Cha;
A31 is Val, Nle, or Cha;
A32 is His;
A33 is Asn;
A34 is Phe, Tyr, Amp, or Aib;
R1 is H;
R2 is H; and R3 is NH2;
or a pharmaceutically acceptable salt thereof.
A3 is Ser or Aib;
A8 is Met, Nva, Leu, Val, Ile, or Nle;
A16 is Asn or Aib;
A18 is Met, Aib, or Nle;
A21 is Val;
A27 is Lys, Aib, Leu, hArg, or Cha;
A31 is Val, Nle, or Cha;
A32 is His;
A33 is Asn;
A34 is Phe, Tyr, Amp, or Aib;
R1 is H;
R2 is H; and R3 is NH2;
or a pharmaceutically acceptable salt thereof.
20. A peptide of claim 19, wherein said peptide is [Nle31]hPTH(1-34)NH2, [hArg27 ]hPTH(1-34)NH2, or [Dap1, Nle8, 18, Tyr34]hPTH(1-34)NH2; or a pharmaceutically acceptable salt thereof.
21. A peptide of the formula:
wherein A1 is Ala, Ser, or Dap;
A3 is Ser or Aib;
A5 is His, Ile, or Cha;
A7 is Leu, Cha, Nle, .beta.-Nal, Trp, Pal, Phe, or p-X-Phe in which X is OH, a halogen, or CH3;
A8 is Leu, Met, or Cha;
A10 is Asp or Asn;
A11 is Lys, Leu, Cha, Phe, or .beta.-Nal;
A12 is Gly or Aib;
A14 is Ser or His;
A15 is Ile, or Cha;
A16 is Gln or Aib;
A17 is Asp or Aib;
A18 is Leu, Aib, or Cha;
A19 is Arg or Aib;
A22 is Phe, Glu, Aib, or Cha;
A23 is Phe, Leu, Lys, or Cha;
A24 is Leu, Lys, or Cha;
A25 is His, Aib, or Glu;
A26 is His, Aib, or Lys;
A27 is Leu, Lys, or Cha;
A28 is Ile, Leu, Lys, or Cha;
A29 is Ala, Glu, or Aib;
A30 is Glu, Cha, Aib, or Lys;
A31 is Ile, Leu, Cha, Lys, or deleted;
A32 is His or deleted;
A33 is Thr or deleted;
A34 is Ala or deleted;
each of R1 and R2 is, independently, H, C1-12 alkanyl, C7-20 phenylalkyl, C11-20 napthyalkyl, C1-12 hydroxyalkyl, C2-12 hydroxyalkenyl, C7-20 hydroxyphenylalkyl, or C11-20 hydroxynapthylalkyl; or one and only one of R1 and R2 is COE1 in which E1 is C1-12 alkyl, C2-12 alkyl, C2-12 alkenyl, C7-20 phenylalkyl, C11-20 napthylalkyl, C1-12 hydroxyalkyl, C2-12 hydroxyalkenyl, C7-20 hydroxyphenylalkyl, or C11-20 hydroxynapthylalkyl; and R3 is OH, NH2, C1-12 alkoxy, or NH-Y-CH2-Z in which Y is a C1-12 hydrocarbon moiety and Z is H, OH, CO2H or CONH2;
provided that at least one of A5, A7, A8, A11, A15, A18, A22, A23, A24, A27, A28, A30, or A31 is Cha, or at least one of A3, A12, A16, A17, A18, A19, A22. A25, A26, A29, A30, or A34 is Aib; or a pharmaceutically acceptable salt thereof.
wherein A1 is Ala, Ser, or Dap;
A3 is Ser or Aib;
A5 is His, Ile, or Cha;
A7 is Leu, Cha, Nle, .beta.-Nal, Trp, Pal, Phe, or p-X-Phe in which X is OH, a halogen, or CH3;
A8 is Leu, Met, or Cha;
A10 is Asp or Asn;
A11 is Lys, Leu, Cha, Phe, or .beta.-Nal;
A12 is Gly or Aib;
A14 is Ser or His;
A15 is Ile, or Cha;
A16 is Gln or Aib;
A17 is Asp or Aib;
A18 is Leu, Aib, or Cha;
A19 is Arg or Aib;
A22 is Phe, Glu, Aib, or Cha;
A23 is Phe, Leu, Lys, or Cha;
A24 is Leu, Lys, or Cha;
A25 is His, Aib, or Glu;
A26 is His, Aib, or Lys;
A27 is Leu, Lys, or Cha;
A28 is Ile, Leu, Lys, or Cha;
A29 is Ala, Glu, or Aib;
A30 is Glu, Cha, Aib, or Lys;
A31 is Ile, Leu, Cha, Lys, or deleted;
A32 is His or deleted;
A33 is Thr or deleted;
A34 is Ala or deleted;
each of R1 and R2 is, independently, H, C1-12 alkanyl, C7-20 phenylalkyl, C11-20 napthyalkyl, C1-12 hydroxyalkyl, C2-12 hydroxyalkenyl, C7-20 hydroxyphenylalkyl, or C11-20 hydroxynapthylalkyl; or one and only one of R1 and R2 is COE1 in which E1 is C1-12 alkyl, C2-12 alkyl, C2-12 alkenyl, C7-20 phenylalkyl, C11-20 napthylalkyl, C1-12 hydroxyalkyl, C2-12 hydroxyalkenyl, C7-20 hydroxyphenylalkyl, or C11-20 hydroxynapthylalkyl; and R3 is OH, NH2, C1-12 alkoxy, or NH-Y-CH2-Z in which Y is a C1-12 hydrocarbon moiety and Z is H, OH, CO2H or CONH2;
provided that at least one of A5, A7, A8, A11, A15, A18, A22, A23, A24, A27, A28, A30, or A31 is Cha, or at least one of A3, A12, A16, A17, A18, A19, A22. A25, A26, A29, A30, or A34 is Aib; or a pharmaceutically acceptable salt thereof.
22. A peptide of claim 21, wherein at A22 is Phe or Cha; A23 is Phe or Cha; A25 is His; A26 is His; A27 is Leu or Cha; A28 is Ile or Cha; A29 is Ala; A30 is Glu or Lys; A31 is Ile or Cha; A32 is His; A33 is Thr; and A34 is Ala; or a pharmaceutically acceptable salt thereof.
23. A peptide of claim 22, wherein at least one of A7 and A11 is Cha; or a pharmaceutically acceptable salt thereof.
24. A peptide of claim 22, wherein at least one of A16 or A19 is Aib; or a pharmaceutically acceptable salt thereof.
25. A peptide of claim 21, wherein A22 is Glu, Aib, or Cha; A23 is Leu, Lys, or Cha; A25 is Aib or Glu;
A26 is Aib or Lys; A28 is Leu, Lys, or Cha; A29 is Glu or Aib; A30 is Cha, Aib, or Lys; A31 is Leu, Cha, or Lys; A32 is His; A33 is Thr; and A34 is Ala; or a pharmaceutically acceptable salt thereof.
A26 is Aib or Lys; A28 is Leu, Lys, or Cha; A29 is Glu or Aib; A30 is Cha, Aib, or Lys; A31 is Leu, Cha, or Lys; A32 is His; A33 is Thr; and A34 is Ala; or a pharmaceutically acceptable salt thereof.
26. A peptide of claim 25, wherein at least one of A7 and All is Cha; or a pharmaceutically acceptable salt thereof.
27. A peptide of claim 25, wherein at least one of A16 or A19 is Aib; or a pharmaceutically acceptable salt thereof.
28. A peptide of the formula:
wherein A1 is Ala, Ser, or Dap;
A3 is Ser or Aib;
A5 is His, Ile, or Cha;
A7 is Leu, Cha, Nle, .beta.-Nal, Trp, Pal, Phe, or p-X-Phe in which X is OH, a halogen, or CH3;
A8 is Leu, Met, or Cha;
A10 is Asp or Asn;
A11 is Lys, Leu, Cha, Phe, or .beta.-Nal;
A12 is Gly or Aib;
A14 is Ser or His;
A15 is Ile, or Cha;
A16 is Gln or Aib;
A17 is Asp or Aib;
A18 is Leu, Aib, or Cha;
A19 is Arg or Aib;
A22 is Phe, Glu, Aib, or Cha;
A23 is Phe, Leu, Lys, or Cha;
A24 is Leu, Lys, or Cha;
A25 is His, Aib, or Glu;
A26 is His, Aib, or Lys;
A27 is Leu, Lys, or Cha;
A28 is Ile, Leu, Lys, or Cha;
A29 is Ala, Glu, or Aib;
A30 is Glu, Cha, Aib, or Lys;
A31 is Ile, Leu, Cha, Lys, or deleted;
A32 is His or deleted;
A33 is Thr or deleted;
A34 is Ala or deleted;
each of R1 and R2 is, independently, H, C1-12 alkanyl, C7-20 phenylalkyl, C11-20 napthyalkyl, C1-12, hydroxyalkyl, C2- 12 hydroxyalkenyl, C7-20 hydroxyphenylalkyl, or C11-20 hydroxynapthylalkyl; or one and only one of R1 and R2 is COE1 in which E1 is C1-12 alkyl, C2 12 alkyl, C2-12 alkenyl, C7-20 phenylalkyl, C11-20 napthylalkyl, C1-12 hydroxyalkyl, C2-12 hydroxyalkenyl, C7-20 hydroxyphenylalkyl, or C11-20 hydroxynapthylalkyl; and R3 is OH, NH2, C1-12 alkoxy, or NH-Y-CH2-Z in which Y is a C1-12 hydrocarbon moiety and Z is H, OH, CO2H or CONH2;
provided that at least one of A23, A24, A27, A28, or A31 is Lys; or a pharmaceutically acceptable salt thereof.
wherein A1 is Ala, Ser, or Dap;
A3 is Ser or Aib;
A5 is His, Ile, or Cha;
A7 is Leu, Cha, Nle, .beta.-Nal, Trp, Pal, Phe, or p-X-Phe in which X is OH, a halogen, or CH3;
A8 is Leu, Met, or Cha;
A10 is Asp or Asn;
A11 is Lys, Leu, Cha, Phe, or .beta.-Nal;
A12 is Gly or Aib;
A14 is Ser or His;
A15 is Ile, or Cha;
A16 is Gln or Aib;
A17 is Asp or Aib;
A18 is Leu, Aib, or Cha;
A19 is Arg or Aib;
A22 is Phe, Glu, Aib, or Cha;
A23 is Phe, Leu, Lys, or Cha;
A24 is Leu, Lys, or Cha;
A25 is His, Aib, or Glu;
A26 is His, Aib, or Lys;
A27 is Leu, Lys, or Cha;
A28 is Ile, Leu, Lys, or Cha;
A29 is Ala, Glu, or Aib;
A30 is Glu, Cha, Aib, or Lys;
A31 is Ile, Leu, Cha, Lys, or deleted;
A32 is His or deleted;
A33 is Thr or deleted;
A34 is Ala or deleted;
each of R1 and R2 is, independently, H, C1-12 alkanyl, C7-20 phenylalkyl, C11-20 napthyalkyl, C1-12, hydroxyalkyl, C2- 12 hydroxyalkenyl, C7-20 hydroxyphenylalkyl, or C11-20 hydroxynapthylalkyl; or one and only one of R1 and R2 is COE1 in which E1 is C1-12 alkyl, C2 12 alkyl, C2-12 alkenyl, C7-20 phenylalkyl, C11-20 napthylalkyl, C1-12 hydroxyalkyl, C2-12 hydroxyalkenyl, C7-20 hydroxyphenylalkyl, or C11-20 hydroxynapthylalkyl; and R3 is OH, NH2, C1-12 alkoxy, or NH-Y-CH2-Z in which Y is a C1-12 hydrocarbon moiety and Z is H, OH, CO2H or CONH2;
provided that at least one of A23, A24, A27, A28, or A31 is Lys; or a pharmaceutically acceptable salt thereof.
29. A peptide of claim 28, wherein A22 is Glu, Aib, or Cha; A23 is Leu, Lys, or Cha; A25 is Aib or Glu;
A26 is Aib or Lys; A28 is Leu, Lys, or Cha; A29 is Glu or Aib; A30 is Cha, Aib, or Lys; A31 is Leu, Cha, or Lys; A32 is His; A33 is Thr; and A34 is Ala; or a pharmaceutically acceptable salt thereof.
A26 is Aib or Lys; A28 is Leu, Lys, or Cha; A29 is Glu or Aib; A30 is Cha, Aib, or Lys; A31 is Leu, Cha, or Lys; A32 is His; A33 is Thr; and A34 is Ala; or a pharmaceutically acceptable salt thereof.
30. A peptide of claim 29, wherein at least one of A7 and A11 is Cha; or a pharmaceutically acceptable salt thereof.
31. A peptide of claim 29, wherein at least one of A16 or A19 is Aib; or a pharmaceutically acceptable salt thereof.
Applications Claiming Priority (7)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| US110595P | 1995-07-13 | 1995-07-13 | |
| US60/001,105 | 1995-07-13 | ||
| US330595P | 1995-09-06 | 1995-09-06 | |
| US60/003,305 | 1995-09-06 | ||
| US08/626,186 | 1996-03-29 | ||
| US08/626,186 US5723577A (en) | 1995-07-13 | 1996-03-29 | Analogs of parathyroid hormone |
| PCT/US1996/011292 WO1997002834A1 (en) | 1995-07-13 | 1996-07-03 | Analogs of parathyroid hormone |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CA2226177A1 CA2226177A1 (en) | 1997-01-30 |
| CA2226177C true CA2226177C (en) | 2010-09-07 |
Family
ID=42727572
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA2226177A Expired - Lifetime CA2226177C (en) | 1995-07-13 | 1996-07-03 | Analogs of parathyroid hormone |
Country Status (1)
| Country | Link |
|---|---|
| CA (1) | CA2226177C (en) |
-
1996
- 1996-07-03 CA CA2226177A patent/CA2226177C/en not_active Expired - Lifetime
Also Published As
| Publication number | Publication date |
|---|---|
| CA2226177A1 (en) | 1997-01-30 |
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Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| EEER | Examination request | ||
| MKEX | Expiry |
Effective date: 20160704 |