PHARMACEUTICAL FORMULATIONS OF GHRH MOLECULES
Stabilized solid and liquid pharmaceutical formulations comprising a GHRH molecules as active ingredient, and more particularly GHRH analogs including [trans-3-hexenoyl]hGHRH (1-44) amide, are disclosed. The formulation comprises an anionic surfactant and a non-reducing sugar, and has a pH of about 4.0 to about 7.5. Also disclosed is the use of the formulation for the treatment of various conditions, methods of preparing the formulation, as well as kits containing it.
1 . A solid pharmaceutical formulation comprising:
a GHRH molecule,
an anionic surfactant, and
a non-reducing sugar,
wherein the GHRH molecule is a GHRH analog of formula A:
X-GHRH Peptide (A)
wherein;
the GHRH peptide is a peptide of formula B:
(SEQ ID NO: 1)
A1-A2-Asp-Ala-Ile-Phe-Thr-A8-Ser-Tyr-Arg-Lys-A13-
Leu-A15-Gln-Leu-A18-Ala-Arg-Lys-Leu-Leu-A24-A25-
Ile-A27-A28-Arg-A30-R0 (B)
wherein,
A1 is Tyr or His;
A2 is Val or Ala;
A8 is Asn or Ser;
A13 is Val or Ile;
A15 is Ala or Gly;
A18 is Ser or Tyr;
A24 is Gln or His;
A25 is Asp or Glu;
A27 is Met, Ile or Nle
A28 is Ser or Asn;
A30 is a bond or amino acid sequence of 1 up to 15 residues; and
R0 is NH 2 or NH—(CH 2 )n-CONH 2 , with n=1 to 12; and
X is a hydrophobic tail anchored via an amide bond to the N-terminus of the peptide and the hydrophobic tail defining a backbone of 5 to 7 atoms;
wherein the backbone can be substituted by C 1-6 alkyl, C 3-6 cycloalkyl, or C 6-12 aryl and the backbone comprises at least one rigidifying moiety connected to at least two atoms of the backbone;
said moiety is a double bond, triple bond, saturated or unsaturated C 3-9 cycloalkyl, or C 6-12 aryl.
2 . The formulation of claim 1 , wherein X is:
3 . The formulation of claim 1 , wherein A30 is:
(a) a bond;
(b) an amino acid sequence corresponding to positions 30-44 of a natural GHRH peptide (SEQ ID NO: 6); and
(c) said amino acid sequence of SEQ ID NO: 6, having a 1-14 amino acid deletion from its C-terminus.
4 . The formulation of claim 1 , wherein said GHRH peptide is:
(a) a polypeptide comprising the amino acid sequence of SEQ ID NO: 2 or 3;
(b) a polypeptide comprising the amino acid sequence of SEQ ID NO: 4 or 5; or
(c) said polypeptide of (a) having a 1 to 14 amino acid deletion from its C-terminus.
5 . The formulation of claim 1 , wherein the GHRH analog is (hexenoyl trans-3)hGHRH(1-44)NH 2 (SEQ ID NO: 7).
6 . The formulation of claim 1 , wherein the formulation is lyophilized.
7 . The formulation of claim 1 , wherein the non-reducing sugar is trehalose or sucrose.
8 . The formulation of claim 7 , wherein the non-reducing sugar is trehalose.
9 . The formulation of claim 7 , wherein the non-reducing sugar is sucrose.
10 . The formulation of claim 1 , wherein the anionic surfactant is a polyoxyethylene sorbitan alkyl ester.
11 . The formulation of claim 10 , wherein the anionic surfactant is polysorbate-20.
12 . The formulation of claim 1 , having a pH of about 4.0 to about 7.5 when measured upon suspension in water.
13 . The formulation of claim 1 , wherein the formulation further comprises a bulking agent.
14 . The formulation of claim 13 , wherein the bulking agent is mannitol.
15 . The formulation of claim 1 , wherein the formulation further comprises an anti-oxidant agent.
16 . The formulation of claim 15 , wherein the anti-oxidant agent is methionine.
17 . A liquid pharmaceutical formulation comprising:
a GHRH molecule,
an anionic surfactant, and
a non-reducing sugar,
wherein said formulation has a pH of about 4.0 to about 7.5,
wherein the GHRH molecule is a GHRH analog of formula A:
X-GHRH Peptide (A)
wherein;
the GHRH peptide is a peptide of formula B:
(SEQ ID NO: 1)
A1-A2-Asp-Ala-Ile-Phe-Thr-A8-Ser-Tyr-Arg-Lys-A13-
Leu-A15-Gln-Leu-A18-Ala-Arg-Lys-Leu-Leu-A24-A25-
Ile-A27-A28-Arg-A30-R0 (B)
wherein,
A1 is Tyr or His;
A2 is Val or Ala;
A8 is Asn or Ser;
A13 is Val or Ile;
A15 is Ala or Gly;
A18 is Ser or Tyr;
A24 is Gln or His;
A25 is Asp or Glu;
A27 is Met, Ile or Nle
A28 is Ser or Asn;
A30 is a bond or amino acid sequence of 1 up to 15 residues; and
R0 is NH 2 or NH—(CH 2 )n-CONH 2 , with n=1 to 12; and
X is a hydrophobic tail anchored via an amide bond to the N-terminus of the peptide and the hydrophobic tail defining a backbone of 5 to 7 atoms;
wherein the backbone can be substituted by C 1-6 alkyl, C 3-6 cycloalkyl, or C 6-12 aryl and the backbone comprises at least one rigidifying moiety connected to at least two atoms of the backbone;
said moiety is a double bond, triple bond, saturated or unsaturated C 3-9 cycloalkyl, or C 6-12 aryl.
18 . The formulation of claim 17 , wherein X is:
19 . The formulation of claim 17 , wherein A30 is:
(a) a bond;
(b) an amino acid sequence corresponding to positions 30-44 of a natural GHRH peptide (SEQ ID NO: 6), or
(c) said SEQ ID NO: 6 having a 1-14 amino acid deletion from its C-terminus.
20 . The formulation of claim 17 , wherein said GHRH peptide is:
(a) a polypeptide comprising the amino acid sequence of SEQ ID NO: 2 or 3;
(b) a polypeptide comprising the amino acid sequence of SEQ ID NO: 4 or 5; or
(c) said polypeptide of (a) having a 1 to 14 amino acid deletion from its C-terminus.
21 . The formulation of claim 17 , wherein the GHRH analog is (hexenoyl trans-3)hGHRH(1-44)NH 2 (SEQ ID NO: 7).
22 . The formulation of claim 17 , wherein the non-reducing sugar is trehalose or sucrose.
23 . The formulation of claim 22 , wherein the non-reducing sugar is trehalose.
24 . The formulation of claim 22 , wherein the non-reducing sugar is sucrose.
25 . The formulation of claim 17 , wherein the non-reducing sugar is present in a concentration of about 0.1 to about 5% (w/w).
26 . The formulation of claim 25 , wherein the non-reducing sugar is present in a concentration of about 2% (w/w).
27 . The formulation of claim 17 , wherein the anionic surfactant is a polyoxyethylene sorbitan alkyl ester.
28 . The formulation of claim 27 , wherein the anionic surfactant is polysorbate-20.
29 . The formulation of claim 17 , wherein the surfactant is present in a concentration of about 0.001% (w/w) to about 0.1% (w/w).
30 . The formulation of claim 29 , wherein the surfactant is present at a concentration of about 0.01% (w/w).
31 . The formulation of claim 17 , having a pH of about 5.0 to about 6.0.
32 . The formulation according to claim 31 , having a pH of about 5.0.
33 . The formulation according to claim 31 , having a pH of about 5.5.
34 . The formulation according to claim 31 , having a pH of about 6.0.
35 . The formulation of claim 17 , further comprising a buffer, wherein said buffer is (i) succinate buffer, (ii) histidine buffer, (iii) phosphate buffer or (iv) any combination of (i) to (iii).
36 . The formulation of claim 17 , wherein the formulation further comprises a bulking agent.
37 . The formulation of claim 36 , wherein the bulking agent is mannitol.
38 . The formulation of claim 36 , wherein the bulking agent is present in an amount of about 1 to about 10% (w/w).
39 . The formulation according to claim 38 , wherein the bulking agent is present in an amount of about 4% (w/w).
40 . The formulation of claim 17 , wherein the formulation further comprises an anti-oxidant agent.
41 . The formulation of claim 40 , wherein the anti-oxidant agent is methionine.
42 . The formulation of claim 17 , comprising:
[trans-3-hexenoyl]hGHRH (1-44) amide,
about 0.01% (w/w) of polysorbate-20,
about 2% (w/w) of (i) trehalose, (ii) sucrose or (iii) any combination of (i) and (ii),
about 4% (w/w) of mannitol; and
(i) succinate buffer, (ii) histidine buffer or (iii) any combination of (i) and (ii),
said formulation having a pH of about 5.0 to about 6.0.
43 . The formulation of claim 17 , comprising:
[trans-3-hexenoyl]hGHRH (1-44) amide,
about 0.01% (w/w) of polysorbate-20,
about 2% (w/w) sucrose,
about 4% (w/w) mannitol, and
an histidine buffer,
said formulation having a pH of about 6.0.
44 . The formulation of claim 17 , comprising:
[trans-3-hexenoyl]hGHRH (1-44) amide,
about 0.01% (w/w) polysorbate-20,
about 2% (w/w) sucrose,
about 4% (w/w) mannitol, and
a succinate buffer,
said formulation having a pH of about 5.5.
45 . The formulation of claim 17 , comprising:
[trans-3-hexenoyl]hGHRH (1-44) amide,
about 0.01% (w/w) polysorbate-20,
about 2% (w/w) sucrose,
about 4% (w/w) mannitol, and
a succinate buffer,
said formulation having a pH of about 5.0.
46 . The formulation of claim 17 , comprising:
[trans-3-hexenoyl]hGHRH (1-44) amide,
about 0.01% (w/w) polysorbate-20,
about 2% (w/w) trehalose,
about 4% (w/w) mannitol, and
a succinate buffer,
said formulation having a pH of about 5.5.
47 . A lyophilized pharmaceutical formulation prepared by lyophilizing the liquid formulation of claim 17 .
48 . A lyophilized pharmaceutical formulation prepared by lyophilizing the liquid formulation of claim 21 .
49 . A lyophilized pharmaceutical formulation prepared by lyophilizing the liquid formulation of claim 42 .
50 . A lyophilized pharmaceutical formulation prepared by lyophilizing the liquid formulation of claim 43 .
51 . A lyophilized pharmaceutical formulation prepared by lyophilizing the liquid formulation of claim 44 .
52 . A lyophilized pharmaceutical formulation prepared by lyophilizing the liquid formulation of claim 45 .
53 . A lyophilized pharmaceutical formulation prepared by lyophilizing the liquid formulation of claim 46 .
54 . A method of treating at least one of HIV-associated lipodystrophy, HIV-lipohypertrophy, abdominal obesity, GH deficiency, frailty, mild cognitive impairment, immune deficiency, wasting associated with a chronic disease or long-term disease, or malnutrition associated with a chronic disease or long-term disease in a subject, comprising administering the liquid pharmaceutical formulation of claim 17 to said subject.
55 . A method of treating at least one of HIV-associated lipodystrophy, HIV-lipohypertrophy, abdominal obesity, GH deficiency, frailty, mild cognitive impairment, immune deficiency, wasting associated with a chronic disease or long-term disease, or malnutrition associated with a chronic disease or long-term disease in a subject, comprising administering a liquid pharmaceutical formulation prepared by the suspension of the solid pharmaceutical formulation of claim 1 with a sterile aqueous solution to said subject.
56 . A method of treating at least one of HIV-associated lipodystrophy, HIV-lipohypertrophy, abdominal obesity, GH deficiency, frailty, mild cognitive impairment, immune deficiency, wasting associated with a chronic disease or long-term disease, or malnutrition associated with a chronic disease or long-term disease in a subject, comprising administering a liquid pharmaceutical formulation prepared by the suspension of the lyophilized pharmaceutical formulation of claim 47 with a sterile aqueous solution to said subject.
57 . The method of claim 55 , wherein the administration is via a subcutaneous, intramuscular, intravenous or intraperitoneal route.
58 . The method of claim 56 , wherein the administration is via a subcutaneous, intramuscular, intravenous or intraperitoneal route.
59 . The method of claim 57 , wherein the administration is via a subcutaneous, intramuscular, intravenous or intraperitoneal route.