Tumor-targeting Salmonella typhimurium, a natural tool for activation of prodrug 6MePdR and their combination therapy in murine melanoma model

Abstract

The PNP/6-methylpurine 2'-deoxyriboside (6MePdR) system is an efficient gene-directed enzyme prodrug therapy system with significant antitumor activities. In this system, Escherichia coli purine nucleoside phosphorylase (ePNP) activates nontoxic 6MePdR into potent antitumor drug 6-methylpurine (6MeP). The Salmonella typhimurium PNP (sPNP) gene has a 96-% sequence homology in comparison with ePNP and also has the ability to convert 6MePdR to 6MeP. In this study, we used tumor-targeting S. typhimurium VNP20009 expressing endogenous PNP gene constitutively to activate 6MePdR and a combination treatment of bacteria and prodrug in B16F10 melanoma model. The conversion of 6MePdR to 6MeP by S. typhimurium was analyzed by HPLC and the enzyme activity of sPNP was confirmed by in vitro (tetrazolium-based colorimetric assay) MTT cytotoxicity assay. After systemic administration of VNP20009 to mice, the bacteria largely accumulated and specifically delivered endogenous sPNP in the tumor. In comparison with VNP20009 or 6MePdR treatment alone, combined administration of VNP20009 followed by 6MePdR treatment significantly delayed the growth of B16F10 tumor and increased the CD8(+) T-cell infiltration. In summary, our results demonstrated that the combination therapy of S. typhimurium and prodrug 6MePdR is a promising strategy for cancer therapy.

Biocatalytic approaches applied to the synthesis of nucleoside prodrugs

“…Additionally, both 6MeP and F-Ade readily diffuse across cell membranes and kill neighbouring tumour cells that do not express ePNP. As alternative, the antitumour effect of mutants hPNP (Afshar et al, 2009) and Salmonella typhimurium PNP (Chen et al, 2013) combined with 6MePdR was tested.…”

Potent and Tumor specific: Arming Bacteria With Therapeutic Proteins

“…In E. coli and Salmonella , the enzyme purine nucleoside phosphorylase converts nontoxic 6-methylpurine 2’-deoxyriboside (6MePdR) into the potent antitumor drug 6-methyl-purine (6MeP) [32]. In mice with B16F10 melanoma tumors, combined administration of Salmonella -purine nucleoside phosphorylase and 6MePdR significantly decreased tumor progression [32]. …”

“…Genetically engineered bacteria can express and release cytotoxic proteins (Figure 1), such as bacterial toxins [19–22], immunoregulatory proteins [23–26] and apoptosis-inducing factors [27,28]. Bacteria can also be engineered to carry enzymes for the conversion of nontoxic prodrugs into cytotoxic drugs [29–32]. Tumor-specific accumulation allows for higher therapeutic doses without toxic side effects.…”

Draft genome sequences of Salmonella Oslo isolated from seafood and its laboratory generated auxotrophic mutant

“…However, its therapeutic potential as an anti-cancer agent remain unexplored. Among more than 2500 NTS serovars, S. Typhimurium VNP20009 [2][3][4], an auxotrophic mutant strain of S. Typhimurium A1-R [5,6], has been successfully studied and was even tested in phase I clinical trial for the treatment of solid tumors [7]. Many features of Salmonella namely the ability to thrive in hypoxic environment of the tumor, to induce innate immune response against tumor or its bactofection, and ability to release anticancer genes within tumor were found to be favorable for tumor regression in animal models [8][9][10].…”