Combined solid-phase/solution synthesis of a 31-residue vasoactive intestinal peptide analog: general method for repetitive coupling of fragments without isolation and purification of intermediates.
A novel analog of vasoactive intestinal peptide (VIP) has been reported which exhibits high potency and enhanced duration of in vivo biological activity. This VIP analog, cyclo-(Lys21-Asp25)Ac[Glu8 Lys12 Nle17 Ala19, Asp25 Leu26,Lys27,28,Gly29,30,Thr31]-VIP, which also has a lactam bridge, has been reported to have relaxant effects that are significantly more potent than other beta-agonists such as salbutamol and salmeterol. Because it has potential use for the treatment of bronchial asthma in humans, various convergent syntheses were evaluated to enable the economic preparation of large quantities of this medium-sized hentriacontapeptide. From these studies we developed a combined solid-phase/solution synthesis which uses four protected fragments (each prepared by solid-phase synthesis with highly acid-labile resins) possessing Nalpha-9-fluorenylmethyloxycarbonyl and side-chain tert-butyl protection. Only equivalent amounts of each fragment were required to achieve near-quantitative coupling reactions using N-[(1H-benzotriazol-1-yl)(dimethylamino)methylene]-N-methylmeth anaminium hexafluorophosphate N-oxide/N-hydroxybenzotriazole. All reagents and side products were removed at each stage by simple extraction procedures. Final deprotection was carried out with 90% trifluoroacetic acid. Under these conditions only low levels of epimerization were observed (<2%). These diastereoisomers and other trace impurities were removed from the product in a single purification by preparative high-performance liquid chromatography. The procedure has been scaled up (10-g scale) and the final product obtained in an overall (nonoptimized) yield of 24%. This procedure for the repetitive coupling of fragments, without isolation of intermediates, may be generally applicable for the economic synthesis of other medium-sized and longer peptides.
Felix
A M
AM
Roche Research Center, Hoffmann-La Roche Inc., Nutley, New Jersey, USA. FelixAssoc@aol.com
Zhao
Z
Z
Lambros
T
T
Ahmad
M
M
Liu
W
W
Daniewski
A
A
Michalewsky
J
J
Heimer
E P
EP
D000595:Amino Acid Sequence; D001671:Biochemistry / Q000379:methods*; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000138:chemical synthesis*; D014660:Vasoactive Intestinal Peptide / Q000031:analogs & derivatives* / Q000138:chemical synthesis*
D010446:Peptide Fragments; D014660:Vasoactive Intestinal Peptide