Investigating the Role of TLR Signaling in Adaptive Rewiring of ARF6-Depleted KRAS-Dependant PDAC

Das, Shubhamay (2023) Investigating the Role of TLR Signaling in Adaptive Rewiring of ARF6-Depleted KRAS-Dependant PDAC. Masters thesis, Indian Institute of Science Education and Research Kolkata.

Text (MS dissertation of Shubhamay Das (18MS034)) Thesis_18MS034.pdf - Submitted Version Restricted to Repository staff only Download (3MB)

Abstract

Pancreatic Ductal Adenocarcinoma (PDAC) is one of the deadliest and ill-prognosed cancers in the world, with less than a 10% five-year survivability rate. Mutations in the KRAS gene are among the most prevalent drivers of PDAC and are present in 90% of tumors. Targeting oncogenic KRAS has proven elusive, with only two drugs approved for clinical trials. Another strategy is to disrupt KRAS signaling by targeting downstream factors. One such axis is given by KRAS-ARF6-ASAP1. ARF6, a small GTPase, together with its effector ASAP1, has been shown to regulate tumor invasion and immune evasion. Previous work in the lab showed that depending on the location of proteins within a signaling cascade, PDAC cells are either able to or unable to find adaptation strategies to continue producing robust phenotypic output. More specifically, time-dependent assays revealed that the knockdown of ARF6, but not ASAP1, allowed the KRAS-dependent PDAC cells to recover invasiveness and proliferation rates. Further investigation using multi-omics time-resolved data pointed towards the involvement of TLR signaling cascades, together with the activation of the NFĸB signaling pathway, enabling this PDAC adaptation process. Here, we perform in-vitro phenotypic characterization assays to evaluate the importance of various TLRs (1-10) in the adaptation process. We show that the knockdown of TLR2 or TLR6, together with the knockdown of ARF6, significantly dampened the ability of KRAS-dependent PDAC cells to regain their mobility and 2D and 3D proliferative ability. Together, our work sheds light on the adaptive landscape of PDAC and paves the way for novel therapeutic strategies.

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