Analyzing the Role of PCSK9 in Cholesterol Uptake in Pancreatic Ductal Adenocarcinoma
Board Location: #17
Discipline: Biological Sciences
Subcategory: Cancer Research
Session: 3
Varsha Subramanyam - University of California, Los Angeles
Co-Author(s): Dr. Gilles Rademaker, University of California San Francisco, San FranciscoDr. Rushika Perera, University of California San Francisco, San Francisco
Pancreatic Ductal Adenocarcinoma (PDAC) is a highly aggressive form of cancer known for its tumor heterogeneity and resistance to traditional chemotherapy. It poses a significant threat worldwide, with a 5-year survival rate of approximately 12% following diagnosis, primarily due to its invasive nature and propensity to metastasize to the liver and lungs. Past research has shown that PDAC can be classified into two distinct subtypes termed basal and classical based on their differential gene expression. Basal PDAC cells tend to have higher levels of LDL (low density lipoprotein) uptake and lower levels of cholesterol biosynthesis while classical PDAC cells tend to have lower levels of LDL uptake and thus higher levels of cholesterol biosynthesis. Samples from the same patient tended to have only one of two cell types and interestingly, patients with basal PDAC tended to have a lower life expectancy than those with classical PDAC. In this study, we investigated the metabolic differences between basal and classical PDAC cells. We find that a protein known as proprotein convertase subtilisin/kexin type 9 (PCSK9), an LDL-receptor inhibitor that inhibits LDL-uptake by the cell is highly expressed in classical PDAC and absent in basal PDAC. Thus, we aimed to knock out PCSK9 in a classical PDAC cell line to see if a more basal phenotype could be observed. By employing CRISPR gene editing, we have successfully knocked out the PCSK9 gene in two classical PDAC cell lines. Through immunofluorescence imaging, we were able to observe increased uptake of LDL, reminiscent of a basal-like phenotype in the classical cell lines. Reintroducing the PCSK9 gene in the classical PDAC cells through viral transfection strategy was able to rescue the classical phenotype, characterized by decreased LDL uptake and increased de novo cholesterol biosynthesis. These findings highlight a potentially important role of differential cholesterol uptake and utilization, as a key determinant distinguishing classical and basal PDAC cell types. This insight sheds light on the underlying mechanisms driving PDAC progression in patients and provides potential targets for therapeutic interventions in the future.
Funder Acknowledgement(s): Many thanks to Dr. Rushika Perera and Dr. Gilles Rademaker for mentoring me and giving me the opportunity to work in the Perera Lab. Thank you to the UCSF Student Research Training Program and Amgen for funding my research this summer and providing me with valuable resources and experience for my future career in research.
Faculty Advisor: Dr. Rushika Perera, Rushika.Perera@ucsf.edu
Role: I was able to employ CRISPR to edit the genomes of 2 classical PDAC cell lines to knock out the PCSK9 gene. I synthesized lentivirus containing the PCSK9 gene and introduced this virus to my knock out cell lines as a control experiment. Finally, I used immunofluorescence imaging to visualize the LDL-uptake and cholesterol levels within my engineered classical PDAC cells.