Synthesis and characterization of new radioligands for the mammalian melanin-concentrating hormone (MCH) receptor

Melanin-concentrating hormone (MCH) is a neuropeptide present in the brain of all vertebrates. For the characterization of MCH receptors, a monoiodina ted [Phe(13), Tyr(19)]-MCH radioligand analogue was developed. The high sus ceptibility of [I-125]-[Phe(13), Tyr(19)]-MCH to oxidative damage and its v ery lipophilic nature made it necessary to develop new MCH radioligands. To increase the stability, native methionines were replaced by non-sulphur co ntaining amino acid residues. In one analogue, the L-enantiomer of the phen ylalanine residue at position 13 was substituted by the D-enantiomer, which increased the relative affinity of the ensuing [I-125]-[D-Phe(13), Tyr(19) ]-MCH about 7-fold. The different analogues were iodinated by an enzymatic reaction and used for binding studies with mouse melanoma cells. [I-125]-[M et(O)(4,8), Phe(13), Tyr(19)]-MCH and [I-125]-[Hse(4,8), Phe(13), Tyr(19)]- MCH showed only about 19% of total binding and [I-125]-[Ser(4,8), Phe(13), Tyr(19)]-MCH displayed about 44% of total binding when compared with [I-125 ]-[Phe(13), Tyr(19)]-MCH. Nonspecific binding for all tracers was below 11% of total binding of [I-125]-[Phe(13), Tyr(19)]-MCH binding, [I-125]-[D-Phe (13), Tyr(19)]-MCH was used for saturation binding studies and revealed a K -D of 122.7 +/- 15.3 pmol/l. This radioligand was further characterized by association and dissociation binding studies.