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Acknowledgements
The researcher would like to thank Shawndra Fordham, the Education and Outreach Program Developer at the University of Colorado Cancer Center, for her help as a mentor in answering questions about the methods of data collection surrounding the 96-well plate trials and providing the contact information for my acquisition of the RIN-m cells. They would also like to thank Dr. David Wagner and Dr. Gisella Vaitaitis, the Associate Professor / Head of the Immunology Section of the Department of Medicine and Webb-Waring Center at the University of Colorado Anschutz Medical Campus and his Senior Professional Research Assistant respectively, for their donation of the RIN-m cell frozen stock, 500 mL of 10% FBS RPMI-1640 media, 15 mL of 10% DMSO FBS freeze media, and advice about the definition of a data point versus trial in my experiment. They would like to thank Ian Fleming for his advice on RIN-m cell culture in the RCHS laboratory. Additionally, they would like to thank Susanne Petri and Nikki Dobos, their PI of the RCHS Laboratory and their AP Research Instructor, respectively, for their instruction and expertise on lab equipment and experimental design. They would like to thank Azzota Scientific for waiving the shipping and providing a 15% discount for the 75 cm2 cell culture flasks used in the experiment. They would also like to thank Rocky Mountain Biologicals, LLC, for their donation of 50 mL of Iron Fortified Calf Bovine Serum for this experiment. Lastly, they would like to thank Rock Canyon High School and Douglas County School District for the funding and lab space that was provided for this research project.
Abstract
Targeted cancer therapies exploit genetic deficiencies in cancer cells. One therapy, Poly-adenosine-diphosphate-ribose polymerase (PARP) inhibition, attacks homologous recombination repair (HRR) deficient cancer by inducing DNA damage. Novel research suggests coumarin compounds might enhance the efficacy of PARP inhibitors by inhibiting a secondary HRR pathway. This study examined the efficacy of Olaparib (PARP inhibitor), Novobiocin (coumarin compound), and dual treatment on RIN-m insulinoma cell viability. It was hypothesized dual treatment would decrease cell viability the most in comparison to a dimethyl sulfoxide (DMSO) negative control due to increased HRR inhibition. A 230 µM Olaparib solution and 300 µM Novobiocin solution were tested alone and in combination against a 0.5% DMSO negative control. A 96-well plate was seeded with 50 µL of 10% FBS RPMI-1640 media containing 1.5 x 104 cells and supplemented with 100 µL of treatment solution. A Trypan-blue exclusion assay viability assessment occurred at 168 hours. Olaparib and Novobiocin treatments were equally effective (6800 and 5650 viable cells/well, respectively) with a 0.084 p-value. The combined treatment (3950 cells/well) was most effective, with p-values of 0.0013 (Olaparib) and 0.0074 (Novobiocin) respectively. The Novobiocin and combined treatments were statistically significant compared to the control (8500 cells/well) with p-values of 0.0014 and 1.08 x 10-5, respectively. It was concluded that the combination of Olaparib and Novobiocin resulted in the most successful RIN-m cell viability reduction. Future studies should test non-tumorigenic beta cell viability under the same conditions to confirm selective tumorigenic toxicity.
The embedded link corresponds with my AP Research final submission, which is the most comprehensive and digestible version of the research article that currently exists. All major, unknown acronyms and enzymes are defined in the Appendix at the bottom of the paper. All visuals that are not cited were taken by the researcher. Blacked out sections of text correspond to personal, redacted information.
Final Presentation
The embedded video is a recording of my official AP Research presentation. Recording was done by my AP Research teacher, Mrs. Dobos.
Grammer 4-12-22.movSources
All factual information in this post not directly obtained through my own experiment was taken directly from my AP Research Paper, which can be accessed here.
Credit to Mrs. Dobos for the embedded video.
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