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New drug therapies to kill TSC2-deficient cell lines

McCann, Henry 2019. New drug therapies to kill TSC2-deficient cell lines. PhD Thesis, Cardiff University.
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Abstract

Tuberous sclerosis complex (TSC) is a rare autosomal dominant genetic disorder which results in the global formation of hamartomas, along with epilepsy, autism and learning difficulties. TSC is caused by mutations in TSC1 and/or TSC2 genes, which are involved in regulating the mTOR pathway, an essential pathway involved in cell cycling, proliferation, survival and growth. For non-surgically viable tumours, the use of mTOR inhibitors such as rapamycin (and rapalogs) have been shown to reduce tumour size and incidence of seizures and to improve intellectual and social development. However, these tumours will grow back rapidly when treatment ceases Research has shown that mTOR-hyperactive cells have increased basal levels of endoplasmic reticulum (ER) stress compared to wildtype cells which could be seen as a potential drug target. Several combinations containing nelfinavir, a known ER stress enhancer, have been shown to selectively target Tsc2-/- mouse embryonic fibroblasts (MEFs) and mTOR-hyperactive sporadic cancer cells while being well tolerated by wildtype cells. In this thesis, the key findings were the identification of three nelfinavir-based combinations (mefloquine, Bortezomib and cepharanthine). Results showed that optimised combinations caused selective cytotoxicity in Tsc2-/- MEFs and mTOR-hyperactive sporadic cancer cells while being tolerated by wildtype control cells (measured by DRAQ7 staining). All combinations caused cytotoxicity in a 3D environment (using tumour spheroids). The mechanism of action for each combination was investigated via several method including western blot analysis, rescue assays and RNA sequencing. Results show that mefloquine/nelfinavir and cepharanthine/nelfinavir combinations caused cell death via combined energy stress (cell death was rescued by the addition of methyl pyruvate) and potentially prolonged ER stress while the Bortezomib/nelfinavir combination caused cell death via prolonged ER stress and proteasome inhibition. This work highlights critical vulnerabilities in cancer cells with hyperactive mTORC1 activity that lack flexibility in homeostatic pathways.

Item Type: Thesis (PhD)
Status: Unpublished
Schools: Medicine
Date of First Compliant Deposit: 10 July 2019
Last Modified: 10 Jul 2019 10:58
URI: http://orca-mwe.cf.ac.uk/id/eprint/124098

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