Sydney Peters
Pronouns: she/her
Research Mentor(s): Caleb Cheng
Research Mentor School/College/Department: Pathology / Medicine
Program:
Authors: Sydney Peters, Caleb Cheng, Jasmine Wisniewski, Bailey Jackson, Yuanyuan Qiao, Costas Lyssiotis, Arul Chinnaiyan
Session: Session 2: 10:00 am – 10:50 am
Poster: 4
Abstract
Pancreatic Ductal Adenocarcinoma (PDAC) is one of the deadliest cancers, in part due to the lack of effective therapies available. PDAC exists in a harsh microenvironment, which drives the cancer to rely heavily on autophagy to maintain metabolic homeostasis. As such, autophagy is a promising alternative and novel therapeutic target for pancreatic cancer. PIKfyve is a lipid kinase that regulates autophagy. Previous research has shown that pharmacologically inhibiting PIKfyve can suppress autophagy and slow tumor growth in cancers like prostate cancer. Knowing that PDAC has an elevated reliance on autophagy, we aimed to characterize PIKfyve as a target for PDAC therapeutic development. Our preliminary data suggests that pharmacological inhibition of PIKfyve can suppress autophagy and decrease cell growth in both human and mouse pancreatic cancer cell lines; however, to our knowledge, all previous attempts to genetically knock down Pikfyve in murine cells have been unsuccessful. We hypothesized that utilizing CRISPR-i technology would allow us to successfully knock down PIKfyve, further validating the role of PIKfyve in autophagy and pancreatic cancer. To achieve this, we cloned sgRNAs targeting Pikfyve into a pLV hU6-sgRNA hUbC-dCas9-KRAB-T2a-Puro vector and packed this plasmid into a lentivirus. Then, we infected a mouse pancreatic cancer cell line with this virus and performed selection with puromycin. We validated the knockdown of PIKfyve at the RNA and protein levels using qPCR and western blot. Finally, we used a proliferation assay to show that PIKfyve knockdown decreases mouse pancreatic cancer cell growth. This data suggests that Pikfyve knockdown decreased autophagy and suppressed cell growth in mouse pancreatic cancer cell lines. In the context of translational pancreatic cancer research, this project highlights PIKfyve as a therapeutic target for drug development. Furthermore, achieving Pikfyve knockdown in mouse lines is critical to testing its effect in an immunocompetent model. These results also provide molecular support for the new, ongoing Phase II clinical trial utilizing a pharmacologic autophagy inhibitor, ESK981.
