Patent ID: 9475837
Filing Date: 2016-10-25
CPC Classification: C07K

Claim Text:
1. A method for synthesizing a therapeutic ghrelin peptide analogue of formula I using stepwise solid-phase fluorenylmethyloxycarbonyl (Fmoc)-chemistry, the method comprising: (a) swelling a Fmoc-Sieber resin comprising an Fmoc group in a first dipolar aprotic solvent; (b) de-protecting the Fmoc group of the Fmoc-Sieber resin with a solution of piperidine in a second dipolar aprotic solvent; (c) washing the resin after Fmoc de-protection with a third dipolar aprotic solvent; (d) activating an Fmoc-amino acid for coupling to the de-protected resin by dissolving the Fmoc-amino acid and one or more coupling reagents in a fourth dipolar aprotic solvent, then adding a base and stirring; (e) contacting the activated Fmoc-amino acid solution and the resin in a reactor; (f) coupling the activated Fmoc-amino acid with the one or more coupling reagents, the one or more coupling reagents comprising: (2-(6-chloro-H-benzotriazole-1-yl)-1,1,3,3-tetramethylaminium hexafluorophosphate) (HCTU) or 2-(1H-benzotriazole-1-yl)-1,1,3,3-tetramethyluronium tetrafluoroborate (TBTU)/1-hydroxybenzotriazole (HOBt); a base; and a fifth dipolar aprotic solvent; (g) washing the resin; (h) repeating steps (b)-(g) until the ghrelin peptide analogue is formed; (i) contacting the resin with a cleavage cocktail to cleave the ghrelin peptide analogue, resulting in cleavage mixture and resin; (j) filtering the cleavage mixture from the resin; and (k) evaporating the cleavage mixture filtrate and precipitating the crude product from the concentrated solution with an organic solvent to yield a partially purified ghrelin peptide analogue; wherein: A 1 is Aib, Apc or Inp; A 2 is D-Bal, D-Bip, D-Bpa, D-Dip, D-1Nal, D-2Nal, D-Ser(Bzl), or D-Trp; A 3 is D-Bal, D-Bip, D-Bpa, D-Dip, D-1Nal, D-2Nal, D-Ser(Bzl), or D-Trp; A 4 is 2Fua, Orn, 2Pal, 3Pal, 4Pal, Pff, Phe, Pim, Taz, 2Thi, 3Thi, Thr(Bzl); A 5 is Apc, Dab, Dap, Lys, Orn, or deleted; R 1 is hydrogen; and R 2 is OH or NH 2 ; provided that when A 5 is Dab, Dap, Lys, or Orn, then: A A A when A 5 is deleted, then: A A