Tc-99m-labeled RGD-conjugated alpha-melanocyte stimulating hormone hybrid peptides with reduced renal uptake

The purpose of this study was to examine whether the replacement of the positively-charged Lys or Arg linker with a neutral linker could reduce the renal uptake of Arg-Gly-Asp (RGD)-conjugated alpha-melanocyte stimulating hormone (alpha-MSH) hybrid peptide. The RGD motif {cyclic(Arg-Gly-Asp-DTyr-Asp)} was coupled to [Cys(3,4,10), D-Phe(7), Arg(11)]alpha-MSH3-13 {(Arg(11))CCMSH} through the neutral beta Ala or Ahx {aminohexanoic acid} linker (replacing the Lys or Arg linker) to generate novel RGD-beta Ala-(Arg(11))CCMSH and RGD-Ahx-(Arg(11))CCMSH hybrid peptides. The receptor-binding affinity and cytotoxicity of RGD-beta Ala-(Arg(11))CCMSH and RGD-Ahx-(Arg(11))CCMSH were determined in B16/F1 melanoma cells. The melanoma targeting and imaging properties of Tc-99m-RGD-beta Ala-(Arg(11))CCMSH and Tc-99m-RGD-Ahx-(Arg(11))CCMSH were determined in B16/F1 melanoma-bearing C57 mice. The replacement of the Lys or Arg linker with the beta Ala or Ahx linker retained nanomolar receptor-binding affinities and remarkable cytotoxicity of RGD-beta Ala-(Arg(11))CCMSH and RGD-Ahx-(Arg(11))CCMSH. The receptor-binding affinities of RGD-beta Ala-(Arg(11))CCMSH and RGD-Ahx-(Arg(11))CCMSH were 0.8 +/- 0.05 and 1.3 +/- 0.1 nM. Three-hour incubation with 0.1 A mu M of RGD-beta Ala-(Arg(11))CCMSH and RGD-Ahx-(Arg(11))CCMSH decreased the survival percentages of B16/F1 cells by 71 and 67 % as compared to the untreated control cells 5 days post the treatment. The replacement of the Arg linker with the beta Ala or Ahx linker reduced the non-specific renal uptake of Tc-99m-RGD-beta Ala-(Arg(11))CCMSH and Tc-99m-RGD-Ahx-(Arg(11))CCMSH by 62 and 61 % at 2 h post-injection. Tc-99m-RGD-beta Ala-(Arg(11))CCMSH displayed higher melanoma uptake than Tc-99m-RGD-Ahx-(Arg(11))CCMSH at 0.5, 2, 4, and 24 h post-injection. Enhanced tumor to kidney uptake ratio of Tc-99m-RGD-beta Ala-(Arg(11))CCMSH warranted the further evaluation of Re-188-labeled RGD-beta Ala-(Arg(11))CCMSH as a novel MC1 receptor-targeting therapeutic peptide for melanoma treatment in the future.