Synthesis of biologically active tritiated amylin and salmon calcitonin analogues

Amylin is a hormone belonging to the calcitonin protein family of peptides. To facilitate receptor screening Studies, alternatively radiolabeled and b iologically active amylin and salmon calcitonin analogues were synthesized by reductive methylation. Free amino groups of amylin and salmon calcitonin were methylated by reaction of peptides with formaldehyde and sodium [H-3] borohydride. Radioactively labeled peptides were purified by size exclusion chromatography followed by HPLC. Analysis by MALDI-TOF mass spectrometry o f purified amylin and salmon calcitonin peptides revealed incorporation of both two and four tritiated methyl groups per peptide molecule. Specific ac tivities of 22.6 and 23.2 GBq/mmol were measured for amylin and salmon calc itonin, respectively. Methylation of rat amylin and salmon calcitonin did n ot affect:their biological activities as both retained their potency to inh ibit insulin-stimulated glycogen synthesis in isolated rat soleus muscle. T he synthesis of these tritiated analogues provides an alternative chemicall y stable radiolabeled ligand which may be useful in exploring receptor inte ractions within the calcitonin peptide family. (C) 2000 Academic Press.