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US 11,806,387 B1Granted
Process of making abaloparatide
AssigneeRadius Health, Inc.
InventorsMatt Burke, Silvio Campagna
Life of this patent
Estimating the term…
Abstract
Disclosed herein is an improved process for preparing abaloparatide. The process generally utilizes solid phase peptide synthesis employing an Fmoc-protection scheme. Incorporating a systematic recoupling step of a glutamine residue (Gln16) has been found to minimize the formation of an undesirable des-Gln16 abaloparatide impurity, which is often obtained in significant quantities in the conventional process.
Claims28
A process for the preparation of abaloparatide comprising:
- a) providing a peptide bound to a solid resin and having an initial N-terminus, wherein said bound peptide is NH2-Asp(OtBu)-Leu-Arg(Pbf)-Arg20(Pbf)-Arg(Pbf)-Glu(OtBu)-Leu-Leu-Glu25(OtBu)-Lys(Boc)-Leu-Leu-Aib-Lys30(Boc)-Leu-His(Trt)-Thr(tBu)-Ala34-Rink Amide MBHA resin (SEO ID NO:
- 2);
- b) coupling a carboxyl terminus of Fmoc-(Trt)Gln16-OH to the initial N-terminus of the bound peptide, wherein coupling comprises adding to a suspension of the bound peptide in dimethylformamide from about 1.05 to about 2 molar equivalents of Fmoc-(Trt)Gln16-OH, relative to the bound peptide, in the presence of benzotriazol-1-yloxytripyrrolidinophosphonium hexafluorophosphate (PyBOP), 1-Hydroxybenzotriazole (HOBt), N,N′-diisopropylcarbodiimide (DIC), and diisopropylethylamine (DIEA), and allowing the mixture to react for a period of time;
- c) adding to the mixture a further from about 1.05 to about 2 molar equivalents of Fmoc-(Trt)Gln16-OH, relative to the bound peptide, in the presence of additional PyBOP, HOBt, DIC, and DIEA;
- d) selectively cleaving the Fmoc group with a solution comprising an amine base to provide a peptide bound to a solid resin and having a new N-terminus.
2 more elements
A process for the preparation of abaloparatide comprising:
- a) anchoring Fmoc-protected alanine (Fmoc-Ala34-OH) to Rink amide 4-methylbenzhydrylamine (MBHA) resin;
- b) capping the resin obtained in step a) by acetylation;
- c) selectively cleaving the Fmoc group with a solution comprising piperidine to provide an N-terminus;
- d) coupling a carboxyl terminus of Fmoc-Thr(tBu)33-OH to the N-terminus in the presence of a coupling reagent comprising benzotriazol-1-yloxytripyrrolidinophosphonium hexafluorophosphate (PyBOP), 1-Hydroxybenzotriazole (HOBt), N,N′-diisopropylcarbodiimide (DIC), and diisopropylethylamine (DIEA), performing a Kaiser and/or Chloranil test after each step d) to confirm complete incorporation of each Fmoc-protected amino acid, and repeating said step d) if a positive Kaiser or Chloranil test result is obtained;
- e) selectively cleaving the Fmoc group with a solution comprising piperidine to provide an N-terminus;
- f) repeating steps d) and e) sequentially, replacing the Fmoc-Thr(tBu)33-OH with the following series of Fmoc-protected amino acids:
- His(Trt)32-OH, Leu31-OH, (Boc)Lys30-OH, Aib29-OH, Leu28-OH, Leu27-OH, (Boc)Lys26-OH, (OtBu)Glu25-OH, Leu24-OH, Leu23-OH, (OtBu)Glu22-OH, (Pbf)Arg21-OH, (Pbf)-Arg20-OH, (Pbf)Arg19-OH, Leu18-OH, and (OtBu)Asp17-OH, and performing a Kaiser and/or Chloranil test after each step d) before each step e) to confirm complete incorporation of each Fmoc-protected amino acid, and repeating said step d) if a positive Kaiser or Chloranil test result is obtained;
- g) coupling a carboxyl terminus of Fmoc-(Trt)Gln16-OH to the N-terminus, wherein coupling comprises adding to a suspension of the bound peptide in dimethylformamide from about 1.05 to about 2 molar equivalents of Fmoc-(Trt)Gln16-OH, relative to the bound peptide, in the presence of a coupling reagent comprising PyBOP, HOBt, DIC, and DIEA, and allowing the mixture to react for a period of time;
- h) adding to the mixture a further from about 1.05 to about 2 molar equivalents of Fmoc-(Trt)Gln16-OH, relative to the bound peptide, in the presence of additional PyBOP, HOBt, DIC, and DIEA;
- i) selectively cleaving the Fmoc group with a solution comprising piperidine to provide an N-terminus;
- j) coupling a carboxyl terminus of Fmoc-Ile15-OH to the N-terminus in the presence of a coupling reagent comprising PyBOP, HOBt, DIC, and DIEA;
- k) selectively cleaving the Fmoc group with a solution comprising piperidine to provide an N-terminus;
- l) repeating steps j) and k) sequentially, replacing Fmoc-Ile15-OH with the following series of Fmoc-protected amino acids:
- Ser (tBu)14-OH, Lys (Boc)13-OH, Gly21-OH, Lys(Boc)11-OH, Asp(OtBu)10-OH, His(Trt)9-OH, Leu8-OH, Leu7-OH, Gln(Trt)6-OH, His (Trt)5-OH, Glu(OtBu)4-OH, Ser(tBu)3-OH, Val2-OH, and Ala1-OH;
- m) performing a Kaiser and/or Chloranil test after each step j) before each step k) to confirm complete incorporation of each Fmoc-protected amino acid, and repeating said step j) if a positive Kaiser or Chloranil test result is obtained;
- and
- n) treating the resulting anchored, protected abaloparatide with a solution comprising trifluoroacetic acid to provide crude abaloparatide.
14 more elements
Dependencies are inferred from the claim text.