Comparative analysis of Rho-GTPase pathway dynamics and epigenetic drug responses in subtype-specific pancreatic cancer cell lines

Zhou, Quan (2024): Comparative analysis of Rho-GTPase pathway dynamics and epigenetic drug responses in subtype-specific pancreatic cancer cell lines. Dissertation, LMU München: Medizinische Fakultät

Abstract

Background: Pancreatic ductal adenocarcinoma (PDAC) is commonly detected at advanced stages, with chemotherapy being the main treatment possibility. Large-scale gene expression studies have identified two major PDAC subtypes, a classical and a basal-like subtype, characterized by an epithelial and mesenchymal phenotype, respectively, that are additionally associated with changes in cytoskeleton organization. The specific transcriptional networks of these subtypes are regulated by epigenetic modifications. Considering the reversible nature of cytoskeleton organization and epigenetic modifications, the presented study aims to explore whether human PDAC cell lines can undergo a subtype-specific reprogramming of cytoskeleton organization and epigenetic modifications towards a less aggressive phenotype. Method: Transcriptional profiles from seven human PDAC cell lines were generated by RNA-sequencing (RNA-seq) to classify the cell lines into a classical and basal-like phenotype. Differentially expressed genes, which are important in regulating epithelial-to-mesenchymal transition (EMT), and cytoskeletal organization as well as Rho-GTPase/ROCK pathway activity, were determined by chromatin-immunoprecipitation (ChIP) followed by qPCR for their histone acetylation status. Differences in Rho-GTPase/ROCK pathway activity were assessed in classical and basal-like cell lines by immunoblot analysis. A ROCK-inhibitor as well as histone acetyltransferase (HAT) and histone deacetylase (HDAC) inhibitors were tested as therapeutic options to induce phenotypical changes towards a less aggressive cancer phenotype. Results: The PDAC cell lines were classified into classical and basal-like subtypes using RNA sequencing and bioinformatic approaches. Classical cell lines exhibited higher expression of epithelial markers and showed enriched H3K27ac at their promoters, while basal-like cell lines had reduced overall histone acetylation levels and increased HDAC2 activity. Basal-like cell lines also demonstrated significant cytoskeletal changes, including increased stress fiber formation, regulated by the Rho-GTPase/ROCK pathway, although ROCK inhibition did not show any treatment effects at the used conditions. Treatment with a HAT inhibitor led to a more aggressive tumor phenotype of classical PDAC cells, marked by the loss of epithelial markers, higher EMT scores, and enhanced cell migration. In contrast, HDAC inhibitors had minimal effects on the reprogramming of both subtypes. Conclusion: The study highlights a crucial role of epigenetic regulation in PDAC subtypes and driving EMT characteristics, particularly through the RhoA-GTPase/ROCK pathway, which significantly influences EMT and cytoskeletal dynamics in basal-like PDAC cell lines. The distinct and heterogeneous cellular responses to HAT and HDAC inhibitors between the two subtypes further emphasize the need for subtype-specific epigenetic therapies. Targeting epigenetic modifications could provide a strategic approach to mitigating tumor aggressiveness.

• Dokumententyp: Dissertationen (Dissertation, LMU München)
• Themengebiete: 600 Technik, Medizin, angewandte Wissenschaften ; 600 Technik, Medizin, angewandte Wissenschaften > 610 Medizin und Gesundheit
• Fakultäten: Medizinische Fakultät
• Sprache der Hochschulschrift: Englisch
• Datum der mündlichen Prüfung: 5. Dezember 2024
• 1. Bericht­erstatter:in: Regel, Ivonne