Identification of synthetic lethal interactions with the KRAS oncogene for targeted cancer treatment
AuthorMorgan, Kimberly Jane
Document TypePhD thesis
Access StatusThis item is embargoed and will be available on 2023-08-25.
© 2021 Kimberly Jane Morgan
Cancer is a major public health issue globally, ranking as the second most common cause of death. Molecularly targeted therapies, focused on exploiting tumour cell dependency on certain oncogenic driver mutations for growth and survival, have greatly improved patient outcomes. However, despite these advances, some of the most frequent oncogenic mutations in cancer, such as those found in KRAS, are extremely challenging to target directly. One promising strategy to expand the range of actionable targets for cancer drug development is the exploitation of synthetic lethal interactions. Synthetic lethality is the term used to describe the death of cells in response to the co-existing disruption of two genes, neither of which is lethal alone. In this setting, targeting a gene that is synthetic lethal with a cancer-relevant mutation has the potential to induce the death of vulnerable cancer cells while leaving healthy cells unaffected. With this background in mind, my lab participated in a focused ENU mutagenesis screen in zebrafish with the aim of identifying genes that are essential for high rates of cell proliferation during endodermal organ development but not required by quiescent tissues. This yielded mutants that exhibited either ‘cell death’ or ‘growth arrest’ phenotypes in the liver, intestine and pancreas. I investigated two of the underlying mutant genes, ahctf1 and rnpc3, for their capacity to engage in synthetic lethal interactions with the kras oncogene. In Chapter 3, I investigated the impact of ahctf1 heterozygosity on the growth and survival of KrasG12V-expressing hepatocytes in a zebrafish model of hepatocellular carcinoma (HCC), TO(krasG12V). ahctf1 encodes Elys, a multifunctional nucleoporin with essential roles in nuclear pore assembly and mitosis. I found that ahctf1 heterozygosity impairs nuclear pore formation, mitotic spindle assembly and chromosome segregation, leading to DNA damage and activation of Tp53-dependent and Tp53-independent cell death pathways which reduced tumour burden. Importantly, ahctf1 heterozygosity did not impact normal liver development, advancing ELYS as an attractive target for cancer therapy with a viable therapeutic window. In Chapter 4, I examined if rnpc3 heterozygosity also reduced tumour burden in the TO(krasG12V) model. rnpc3 encodes 65K, a unique protein component of the U12-dependent spliceosome, a specialised splicing machinery required for the correct splicing of a very small percentage (3.7%) of genes. In hepatocytes expressing krasG12V, rnpc3 heterozygosity reduced the number of cells in S phase of the cell cycle and increased cell death, together reducing tumour burden, without affecting normal tissue. In Chapter 5, I demonstrated that the zebrafish model of HCC is a powerful platform for testing novel therapeutics. I evaluated the efficacy of PRMT5 and KAT6A/B inhibitors early in their development, and showed that they were effective in reducing tumour growth and worthy of future investigation. In conclusion, my studies revealed two promising new targets for cancer treatment. I also demonstrated that the zebrafish HCC model is highly amenable to pharmacological inhibition and provides a valuable system for the pre-clinical examination of drug treatments in vivo.
KeywordsKras; ahctf1; elys; nucleoporin; synthetic lethality; hepatocellular carcinoma; zebrafish; cancer
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