UCLA

Background: Procollagen-Lysine, 2-Oxoglutarate 5-Dioxygenase 3 (PLOD3) is a gene that encodes a protein crucial for the formation of hydroxylysine in collagens. Overexpression of PLOD3 has been identified in various types of cancer, suggesting its potential role in promoting tumor growth and metastasis. Lipid nanoparticles (LNPs) represent a rapidly evolving method for delivering therapeutic agents and have been the focus of extensive research for enhancing targeted delivery of therapeutic agents to tumors. The goal of my thesis study is to investigate the functional role of PLOD3 and the use of LNPs for delivery of PLOD3 siRNA (siPLOD3-LNPs) in pancreatic cancer.Methods: Analysis of PLOD3 gene expression in pancreatic adenocarcinoma (PAAD) was conducted using the data from the Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA) databases. In addition to these databases, western blot and immunohistochemistry (IHC) were used to assess the expression of the PLOD3 gene in PAAD. In order to discover the role of PLOD3 and evaluate the use of siPLOD3-LNP in pancreatic cancer, CCK-8, migration, Matrigel invasion, and colony-forming assays were carried out on MIA PaCa-2, BxPC-3, and KPC pancreatic cancer cell lines. The synergistic effect of cisplatin and siPLOD3-LNP on pancreatic cancer cell proliferation was also assessed with the CCK-8 assay. Additionally, RNA- sequencing was performed to investigate the changes in global gene expression following siPLOD3-LNP treatment. Results: Based on statistical analysis of the data from the GEO and TCGA databases and the result from IHC, it was observed that PLOD3 gene expression was significantly increased in PAAD tumor tissues compared to normal controls. Moreover, western blot analysis also showed a higher expression level of PLOD3 in pancreatic cancer cells compared to normal pancreatic cells. Western blot analysis demonstrated a significant reduction in PLOD3 protein expression in the pancreatic cancer cells treated with siPLOD3-LNPs compared to the untreated control cells. The administration of siPLOD3-LNPs significantly inhibited the proliferation, migration, and invasion capabilities of MIA PaCa-2, BxPC-3, and KPC cells. Furthermore, when the pancreatic cancer cells were treated concurrently with cisplatin and siPLOD3-LNPs, a more pronounced decline in cell proliferation was observed. RNA-Seq analysis indicated that siPLOD3-LNP treatment significantly altered many signaling pathways, including cytokine-cytokine receptor interaction, chemokine signaling pathway, TNF signaling pathway, toll-like receptor signaling pathway, NF-κB pathway, and HIF-1 pathway in MIA PaCa-2 cells. Conclusion: PLOD3 expression levels in pancreatic cancer tissues/cells were significantly elevated compared to normal pancreatic tissues/cells. Treatment with siPLOD3-LNPs resulted in a significant reduction in PLOD3 protein expression in human pancreatic cancer cells (MIA PaCa-2 and BxPC-3) and mouse KPC cells. Furthermore, the treatment with siPLOD3-LNPs significantly inhibited the proliferation, migration, and invasion of pancreatic cancer cells. A synergistic effect of cisplatin and siPLOD3-LNPs was also observed on the pancreatic cancer cells, as the inhibition of cell proliferation was more pronounced compared to those of individual treatments. It was also found that siPLOD3-LNP treatment significantly altered many KEGG pathways in MIA PaCa-2 cells. Further studies are warranted to understand the molecular mechanism of PLOD3 in PAAD and the in vivo effect of siRNA-LNP treatment on pancreatic cancer. Keywords: LNPs, PLOD3, PAAD, siRNA treatment