A genome-scale CRISPR/Cas9 knockout screening reveals SH3D21 as a sensitizer for gemcitabine

Gemcitabine, 2′,2′-difluoro-2′-deoxycytidine, is used as a pro-drug in treatment of variety of solid tumour cancers including pancreatic cancer. After intake, gemcitabine is transferred to the cells by the membrane nucleoside transporter proteins. Once inside the cells, it is converted to gemcitabine triphosphate followed by incorporation into DNA chains where it causes inhibition of DNA replication and thereby cell cycle arrest and apoptosis. Currently gemcitabine is the standard drug for treatment of pancreatic cancer and despite its widespread use its effect is moderate. In this study, we performed a genome-scale CRISPR/Cas9 knockout screening on pancreatic cancer cell line Panc1 to explore the genes that are important for gemcitabine efficacy. We found SH3D21 as a novel gemcitabine sensitizer implying it may act as a therapeutic target for improvement of gemcitabine efficacy in treatment of pancreatic cancer.

Factors influencing the intracellular exposure of gemcitabine triphosphate in children with CNS tumors.

e13547 Background: Gemcitabine (dFdC), a cytosine analog, is currently being evaluated as a therapy to treat children and young adults with central nervous systems (CNS) tumors in two clinical trials (NCT01878617 and NCT03434262). Once in the cell, dFdC undergoes complex and highly variable conversion to gemcitabine triphosphate (dFdCTP), which is responsible for its cytotoxic effects. Our objective is to identify the factors influencing dFdC and dFdCTP exposures in children. Methods: As part of treatment, patients with newly diagnosed medulloblastoma and patients with recurrent/refractory medulloblastoma or ependymoma receive 28-day cycles of dFdC, as a 30-minute IV infusion at 750, 1000 or 1250 mg/m2 on days 1 and 15 of each cycle. Patients with recurrent/refractory disease also receive concomitant ribociclib. Pharmacokinetic (PK) studies were performed after one dose of dFdC alone (day 1) and with ribociclib (day 15). dFdC and dFdCTP concentrations were measured in plasma and in peripheral blood mononuclear cells respectively, for up to 4h post-dose, using validated LC-MS/MS methods. The concentrations were analyzed using a population PK model. A preliminary covariate analysis was performed to evaluate the impact of factors (e.g., patient demographics and infusion rate) on dFdC and dFdCTP exposure (area under the curve). Results: dFdC and dFdCTP concentrations were fitted with a three-compartment linear model. dFdC infusion rate significantly influenced the drug exposures with a faster plasma dFdC elimination and higher dFdCTP formation observed at the lower infusion rate. On average, patients receiving dFdC as 25 or 33 mg/m2/min had a dFdCTP exposure of 526 µM·h vs 205 µM·h with a rate of 42 mg/m2/min (p < 0.001). Patient with lower body surface area also showed higher dFdC clearance. Significant intra-patient differences were observed in dFdCTP concentrations measured on days 1 and 15. Conclusions: This preliminary PK analysis shows a saturable formation of dFdCTP in children with CNS tumors, and suggests the need for longer infusion duration with higher dFdC dosages. Future work will evaluate potential drug-drug interaction with ribociclib and the effect of genotypic variants on dFdCTP exposure.