Targeting and selectivity of anticancer prodrug: Activation by peptidase-DPP IV

The adverse systemic side effects of chemotherapy are in part attributable to their lack of selectivity. The goal of this project is to develop a prodrug approach to enhance therapeutic efficacy of an anticancer drug. In this research, a prodrug targeting peptidase highly expressed in tumors has been explored to achieve a more selective anticancer action. Based on bioinformatic processing of microarray data and review of the literature, dipeptidyl peptidase IV (DPP IV) was found to demonstrate high levels of expression in several human cancer cell lines. The selectivity of DPP IV in cleaving proline dipeptides at the N-terminus of the substrate, which is less susceptible to nonspecific peptidases and proteases, makes DPP IV a potential candidate for prodrug activation. In this study, we first demonstrated the feasibility of targeting DPP IV by observing a graded activation of glycyl-prolyl-p-nitroanalide, a standard substrate of DPP IV, in a series of cancer cells with high, low and medium expression of DPP IV. A prodrug, glycyl-prolyl-melphalan (GP-Mel), was then synthesized by attaching the gly-pro dipeptide via an amide bond to the primary amine of the anticancer drug melphalan. The activation of prodrug GP-Mel was found to be DPP IV-dependent in the selected cancer cell system. In the cell proliferation assay, the increased selectivity of GP-Mel over melphalan was confirmed with higher DPP IV-expressing cells demonstrating lower IC50, whereas the IC50 of melphalan was stable and unchanged across the seven cell lines. Furthermore, GP-Mel was found to be 3 times less toxic to murine myeloid progenitor cells through evaluation using the colony-forming unit assays. It also appeared to have a promising in vivo anticancer effect in the preliminary B16f10 mouse melanoma model. In summary, cellular DPP IV appears to be a suitable target enzyme for activation of the peptide prodrug GP-Mel. The results of our studies suggest that a prodrug substrate of DPP IV may show improved therapeutic index in tumors that highly express DPP IV.