Targeting the Hippo pathway in malignant mesothelioma
AuthorKulkarni, Aishwarya Shrikant
AffiliationSir Peter MacCallum Department of Oncology
Document TypePhD thesis
Access StatusThis item is embargoed and will be available on 2023-06-23.
© 2021 Aishwarya Shrikant Kulkarni
Malignant mesothelioma is an aggressive cancer of the mesothelium for which asbestos-exposure is the predominant etiological factor. The standard treatment regime for mesothelioma is systemic chemotherapy, which exhibits poor efficacy with a median post-diagnosis survival of 12 months. This highlights the need for alternate therapies, such as targeted therapies, to improve patient outcomes. Genomic and cell biological analyses of mesotheliomas indicate that deficiency of key tumour suppressor genes including BAP1 or members of the Hippo tumour suppressor pathway such as NF2, are frequent genetic aberrations in this cancer. Inhibition of cancer-promoting events that occur downstream of these genetic aberrations represents a potential mechanism by which mesothelioma patient outcomes can be improved. Originally identified in Drosophila, the Hippo pathway is highly conserved in mammals and regulates the activity of the oncoproteins and transcriptional coactivators YAP and TAZ and their nuclear interaction partners, the TEAD family of transcription factors. YAP/TAZ-TEAD hyperactivation is an oncogenic driver in Hippo pathway deregulated mesotheliomas, indicating that inhibition of these proteins could represent a new treatment strategy for this disease. An emerging class of small molecule inhibitors of YAP/TAZ-TEAD target the recently identified post-translational modification of TEAD palmitoylation. One such compound, VT107, has shown promising pre-clinical activity in in vitro and in vivo models of Hippo pathway deregulated mesothelioma. Whilst VT107 is a promising therapeutic, the current understanding of its molecular impact is incomplete. Moreover, further identification of therapeutic targets in BAP1 or NF2 deficient mesothelioma cells might enable development of additional targeted therapies to treat candidate tumours. To address these considerations, I aimed to answer three key questions: 1) Which genes are potential therapeutic targets for treatment of NF2 or BAP1 mutant mesothelioma?; 2) How does VT10- 7 modulate the behaviour and transcription regulatory activity of YAP/TAZ-TEAD?; and 3) How does VT107 inhibit the proliferation of Hippo pathway mutant mesothelioma cells? 1) To identify candidate therapeutic targets for treatment of NF2 or BAP1 deficient mesothelioma, I conducted a genome-wide synthetic lethality screen. Due to technical limitations with BAP1 deficient cells, I conducted the screen with only NF2 deficient cells. While I identified a mild synthetic lethal interaction between NF2 and CREB3, I concluded that the screen was unsuccessful in identifying robust synthetic lethal interaction partners of NF2. 2) I assessed the molecular impact of VT107 in a Hippo pathway mutant mesothelioma cell line (NCI-H2052) using single molecule tracking of YAP and TEAD1. With this advanced live microscopy approach, I observed that VT107 increased the nuclear mobility, and reduced the DNA residence-time, of both YAP and TEAD1. Thus, I provided a high resolution molecular understanding of how VT107 influences the ability of YAP and TEAD1 to regulate transcription. 3) I conducted a genome-wide CRISPR screen to investigate the mechanism of action of VT107 in NCI-H2052 cells. I identified that loss of SOCS3, VGLL4, BTAF1 and DR1, among other genes, increased cellular resistance to VT107. I also found that loss of several genes, including TCEB2, could confer VT107 sensitivity. These findings provide unbiased insights into the mechanism by which YAP/TAZ-TEAD inhibition modulates the proliferation of mesothelioma cells. Furthermore, I identified potential drivers of resistance to VT107- information which might guide its optimal application in the clinic for the treatment of Hippo pathway mutant mesothelioma.
KeywordsHippo pathway; Malignant mesothelioma; YAP/TAZ; TEAD; CRISPR screen; Targeted therapy; Single molecule tracking; Synthetic lethality; Therapy resistance; NF2; BAP1; Functional genomics
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