miRNA- and Cell Line-Specific Constraints on Precursor miRNA Processing of Stably Transfected Pancreatic Cancer and Other Mammalian Cells.

MicroRNAs (miRNAs) regulate approximately one-third of all human genes. The dysregulation of miRNAs has been implicated in the development of numerous human diseases, including cancers. In our investigation focusing on altering specific miRNA expression in human pancreatic cancer cells, we encountered an interesting finding. While two expression vector designs effectively enhanced miR-708 levels, they were unable to elevate mature forms of miR-29b, -1290, -2467, and -6831 in pancreatic cancer cell lines. This finding was also observed in a panel of other non-pancreatic cancer cell lines, suggesting that miRNA processing efficiency was cell line specific. Using a step-by-step approach in each step of miRNA processing, we ruled out alternative strand selection by the RISC complex and transcriptional interference at the primary miRNA (pri-miRNA) level. DROSHA processing and pri-miRNA export from the nucleus also appeared to be occurring normally. We observed precursor (pre-miRNA) accumulation only in cell lines where mature miRNA expression was not achieved, suggesting that the block was occurring at the pre-miRNA stage. To further confirm this, synthetic pre-miRNA mimics that bypass DICER processing were processed into mature miRNAs in all cases. This study has demonstrated the distinct behaviours of different miRNAs with the same vector in the same cell line, the same miRNA between the two vector designs, and with the same miRNA across different cell lines. We identified a stable vector pre-miRNA processing block. Our findings on the structural and sequence differences between successful and non-successful vector designs could help to inform future chimeric miRNA design strategies and act as a guide to other researchers on the intricate processing dynamics that can impact vector efficiency. Our research confirms the potential of miRNA mimics to surmount some of these complexities.

FOLFIRINOX Pharmacodynamic Interactions in 2D and 3D Pancreatic Cancer Cell Cultures.

The multi-drug combination regime, FOLFIRINOX, is a standard of care chemotherapeutic therapy for pancreatic cancer patients. However, systematic evaluation of potential pharmacodynamic interactions among multi-drug therapy has not been reported previously. Here, pharmacodynamic interactions of the FOLFIRINOX agents (5-fluorouracil (5-FU), oxaliplatin (Oxa) and SN-38, the active metabolite of irinotecan) were assessed across a panel of primary and established pancreatic cancer cells. Inhibition of cell proliferation was quantified for each drug, alone and in combination, to obtain quantitative, drug-specific interaction parameters and assess the nature of drug interactions. The experimental data were analysed assuming Bliss independent interactions, and nonlinear regression model fitting was conducted in SAS. Estimates of the drug interaction term, psi (ψ), revealed that the Oxa/SN-38 combination appeared synergistic in PANC-1 (ψ = 0.6, 95% CI = 0.4, 0.9) and modestly synergistic, close to additive, in MIAPaCa-2 (ψ = 0.8, 95% CI = 0.6, 1.0) in 2D assays. The triple combination was strongly synergistic in MIAPaCa-2 (ψ = 0.2, 95% CI = 0.1, 0.3) and modestly synergistic/borderline additive in PANC-1 2D (ψ = 0.8, 95% CI = 0.6, 1.0). The triple combination showed antagonistic interactions in the primary PIN-127 and 3D PANC-1 model (ψ > 1). Quantitative pharmacodynamic interactions have not been described for the FOLFIRINOX regimen; this analysis suggests a complex interplay among the three chemotherapeutic agents. Extension of this pharmacodynamic analysis approach to clinical/translational studies of the FOLFIRINOX combination could reveal additional pharmacodynamic interactions and guide further refinement of this regimen to achieve optimal clinical responses.