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DOI: 10.1055/s-0036-1587242
Gemcitabine uptake and metabolism of cancer associated fibroblasts in murine pancreatic cancer
Background and Aim: Desmoplasia and hypovascularity were shown to impede drug delivery in pancreatic ductal adenocarcinoma (PDAC). Here, we aim to dissect the differential gemcitabine uptake and metabolism in murine cancer cells and fibroblasts.
Design: We analysed gemcitabine metabolites in primary murine tumour cells, cancer associated fibroblasts (CAFs) and pancreatic stellate cells (PSCs) by LC-MS/MS using a validated protocol. To understand the differential gemcitabine uptake of the various cell types, expression analysis for gemcitabine metabolism pathways was performed in murine cell lines and tissues using qRT-PCR and immunohistochemistry.
Results: In vitro, 3 – 5 fold increased intracellular concentrations of activated dFdCTP were detected in PSCs (n = 2) and CAFs (n = 2) compared to tumour cells (n = 4) (p < 0.02). The inactive gemcitabine metabolite dFdU was detected at significantly lower levels in the supernatant of PSCs and CAFs as compared to tumour cells. Mechanistically, key metabolite enzymes for gemcitabine inactivation such as deoxycytidylate deaminase (Dctd), cytidine deaminase (Cda) and hydrolytic cytosolic 5'-nucleotidases (Ntc1A, Ntc3A) were downregulated in PSCs and CAFs in vitro, and in the LSL-KrasG12D/+;LSL-Trp53R172 H/+;Pdx-1-Cre mouse model in vivo.
Conclusions: Our findings suggest that fibroblast entrap active gemcitabine intracellularly thus making it unavailable for tumour cells. This drug scavenging may contribute to the clinical failure of gemcitabine in stroma rich PDAC.