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 (α-MSH) hybrid peptide. The RGD motif {cyclic(Arg-Gly-Asp-DTyr-Asp)} was coupled to [Cys^3,4,10, D-Phe⁷, Arg¹¹]α-MSH_3-13 {(Arg¹¹)CCMSH} through the neutral βAla or Ahx {aminohexanoic acid} linker (replacing the Lys or Arg linker) to generate novel RGD-βAla-(Arg¹¹)CCMSH and RGD-Ahx-(Arg¹¹)CCMSH hybrid peptides. The receptor-binding affinity and cytotoxicity of RGD-βAla-(Arg¹¹)CCMSH and RGD-Ahx-(Arg¹¹)CCMSH were determined in B16/F1 melanoma cells. The melanoma targeting and imaging properties of ^99mTc-RGD-βAla-(Arg¹¹)CCMSH and ^99mTc-RGD-Ahx-(Arg¹¹)CCMSH were determined in B16/F1 melanoma-bearing C57 mice. The replacement of the Lys or Arg linker with the βAla or Ahx linker retained nanomolar receptor-binding affinities and remarkable cytotoxicity of RGD-βAla-(Arg¹¹)CCMSH and RGD-Ahx-(Arg¹¹)CCMSH. The receptor-binding affinities of RGD-βAla-(Arg¹¹)CCMSH and RGD-Ahx-(Arg¹¹)CCMSH were 0.8∈±∈0.05 and 1.3∈±∈0.1 nM. Three-hour incubation with 0.1 μM of RGD-βAla-(Arg¹¹)CCMSH and RGD-Ahx-(Arg¹¹)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 βAla or Ahx linker reduced the non-specific renal uptake of ^99mTc-RGD-βAla-(Arg¹¹)CCMSH and ^99mTc-RGD-Ahx-(Arg¹¹)CCMSH by 62 and 61 % at 2 h post-injection. ^99mTc-RGD-βAla-(Arg¹¹)CCMSH displayed higher melanoma uptake than ^99mTc-RGD-Ahx-(Arg¹¹)CCMSH at 0.5, 2, 4, and 24 h post-injection. Enhanced tumor to kidney uptake ratio of ^99mTc-RGD-βAla-(Arg¹¹)CCMSH warranted the further evaluation of ¹⁸⁸Re-labeled RGD-βAla-(Arg¹¹)CCMSH as a novel MC1 receptor-targeting therapeutic peptide for melanoma treatment in the future.