Pegylated arginine deiminase synergistically increases the cytotoxicity of gemcitabine in human pancreatic cancer

Rouzbeh Daylami,Subbulakshmi Virudachalam,Diego J Muilenburg,Richard J Bold

doi:10.1186/preaccept-8245834981349245

Abstract

Pancreatic ductal adenocarcinoma has proven to be one of the most chemo-resistant among all solid organ malignancies. Several mechanisms of resistance have been described, though few reports of strategies to overcome this chemo-resistance have been successful in restoring sensitivity to the primary chemotherapy (gemcitabine) and enter the clinical treatment arena. We examined the ability of cellular arginine depletion through treatment with PEG-ADI to alter in vitro and in vivo cytotoxicity of gemcitabine. The effect on levels of key regulators of gemcitabine efficacy (e.g. RRM2, hENT1, and dCK) were examined. Combination of PEG-ADI and gemcitabine substantially increases growth arrest, leading to increased tumor response in vivo. PEG-ADI is a strong inhibitor of the gemcitabine-induced overexpression of ribonucleotide reductase subunit M2 (RRM2) levels both in vivo and in vitro, which is associated with gemcitabine resistance. This mechanism is through the abrogation of the gemcitabine-mediated inhibitory effect on E2F-1 function, a transcriptional repressor of RRM2. The ability to alter gemcitabine resistance in a targeted manner by inducing metabolic stress holds great promise in the treatment of advanced pancreatic cancer.

Highlights

Pancreatic ductal adenocarcinoma has proven to be one of the most chemo-resistant among all solid organ malignancies
Pegylated arginine deiminase (PEG-ADI) depletes intracellular arginine stores We have previously shown that the L3.3 cell line expresses argininosuccinate synthetase (ASS) and the MIA-PaCa-2 cell line is deficient in ASS [17]
A) Dose-dependent effect of combining gemcitabine with PEG-ADI in MIA-PaCa-2 cells determined by MTT cytotoxicity assay, B) Effect of PEG-ADI, gemcitabine, or the combination on cell death measured by flow cytometry in MIA-PaCa-2, C) Effect of PEG-ADI, gemcitabine, or the combination on cell death measured by flow cytometry in L3.3 with no significant effect of PEG-ADI, either alone or in combination with gemcitabine, D) MIA-PaCa-2 cells treated with PEG-ADI, gemcitabine or the combination with immunoblotting for caspase 3 following gemcitabine, PEG-ADI or the combination with notation of cleaved caspase 3, E) Effect of PEG-ADI, gemcitabine or the combination on Annexin V staining of MIA-PaCa-2 cells

Summary

Introduction

Pancreatic ductal adenocarcinoma has proven to be one of the most chemo-resistant among all solid organ malignancies. There has been a great deal of focus on the mechanisms that confer resistance to apoptosis in pancreatic cancer including aberrations in central mediators of apoptotic cell death. This has led to several investigations that have combined a variety of therapies directed at targets that may increase the sensitivity to gemcitabine [2,3,4]. Gemcitabine is administered as a pro-drug which must first be taken up by cells using the transport protein human equilibrative nucleoside transporter-1 (hENT1) [5] It is converted into an active metabolite by deoxycytidine kinase (dCK), allowing for incorporation into DNA as a nucleoside analog but structurally ending DNA synthesis. Therapies that target the overexpression of any of these proteins altering gemcitabine metabolism may

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Results

Conclusion