Studies on the effect of Pt^IV56MeSS on molecular pathways in A549 lung cancer cells

Abstract

Pt^IV56MeSS, an unconventional anticancer platinum complex with hydroxido axial ligands, exhibits significant cytotoxicity, which is often expressed as IC50 in in vitro cancer models. The intracellular reduction of platinum(IV) to its active platinum(II) precursor relies on the reducing agents that comprise the redox system of the cell. In cancer cells, the intracellular homeostatic balance controlled by these agents is susceptible to instability, which leads to oxidative stress that disturbs cellular processes regulated by redox-sensitive sensors. This thesis explores the principles of 56MeSS cytotoxicity in KRAS mutated cancer cell line models. A fluorometric probe and endpoint biochemical assay were applied to detect oxidative stress and measure the drug's cytotoxicity. Subsequently, quantitative proteomic and bioinformatic methods were used to deconvolute the complex relationship between disturbed molecular processes and the drug's cytotoxicity. Here, we detail the nuances of each methodology and its limitations. The IC50 values of Pt^IV56MeSS in the LoVo colorectal cell line exhibited micromolar values regardless of the in vitro model. A spheroid model of A549 was insufficient to corroborate the nanomolar values of the monolayer model. Label-free protein quantification in the A549 monolayer model quantified over 1,300 proteins involved in translational and vacuole trafficking processes. The number of dysregulated proteins by Pt^IV56MeSS exceeded its reduced platinum(II) form and increased with prolonged treatments. The highest fold change variance was detected in pro-inflammatory cytokine IL18, translational regulator GSPT1, and molecular chaperone for glycoproteins LMAN1. The significant proteins indicate early dysregulation of RNA and protein metabolism and upregulation of proteins involved in ER and Golgi vesicular trafficking systems. This is accompanied by impeded Rho GTPase cycle signalling and mitotic disruption. The results suggest a complex relationship between molecular pathways regulated by redox-sensitive proteins that collectively contribute to Pt^IV56MeSS cytotoxicity.

Date of Award	2022
Original language	English
Awarding Institution	Western Sydney University
Supervisor	Janice Aldrich-Wright (Supervisor)