Compound (XI) was prepared by solid phase peptide synthesis on a Rink amide 4-methylbenzhydrylamine (MBHA) resin. After removal of Fmoc protecting group from the resin with 20% piperidine in N-methyl-2-pyrrolidinone (NMP), a coupling cycle was carried out with Fmoc-Thr(O-t-Bu)OH (I) using bromotripyrrolidino phosphonium hexafluorophosphate (PyBroP) and diisopropylethylamine (DIEA) in NMP, followed by Fmoc removal with 20% piperidine in NMP. To the Thr-bound resin (II) they were stepwise coupled in subsequent coupling and deprotecting cycles the following amino acids: Fmoc- (N-allyloxycarbonylpropyl)PheOH (IV), Fmoc-ValCl (VI), Fmoc-Lys(Boc)OH (VIII), and Fmoc-D-TrpOH (X) to yield the corresponding peptide-bound resins (V), (VII), (IX) and (XI), respectively.
Compound (XVI) was prepared by solid phase peptide synthesis on a Rink amide 4-methylbenzhydrylamine (MBHA) resin. To the Thr-bound resin (XI) they were stepwise coupled in subsequent coupling and deprotecting cycles the following amino acids: Fmoc-Tyr(t-Bu)OH (XII), and Fmoc- (N-allyloxycarbonylaminoethyl)PheOH (XIV) to yield the corresponding peptide-bound resins (XIII) and (XV), respectively. Then, allyl and allyoxycarbonyl protecting groups were both removed from the linear peptide-bound resin (XV) by treatment with tetrakis(triphenylphosphine)palladium, acetic acid, and N-methylmorpholine (NMM) in chloroform to yield compound (XVI).
The cyclization between free amino and acid groups of compound (XVI) was effected by reaction with benzotriazol-1-yloxy-tripyrrolidinophosphonium hexafluorophosphate (PyBOP) to give the cyclic peptide (XVII). Finally, treatment with moist trifluoroacetic acid (TFA) containing a trace of ethanedithiol (EDT) and triisopropylsilane (TIS) at 0 C removed all protecting groups and liberated from the resin the peptide amide, which was finally purified by reversed-phase preparative HPLC.