2026-09-23T12:00:29Zhttps://api.repository.cam.ac.uk/server/oai/request

oai:www.repository.cam.ac.uk:1810/3643962024-02-14T01:42:47Zcom_1810_217840com_1810_34581col_1810_218541

Characterisation of HIF2A targets identifies MYC as a key determinant of HIF2A dependency in ccRCC https://doi.org/10.17863/CAM.106089 Dyas, Anna Elizabeth Muñoz-Espín, Daniel Despite advancements in ccRCC treatment, the prognosis for metastatic disease remains poor with a 5-year survival of approximately 10%. VHL loss is an early driver event in the vast majority of ccRCC cases and results in constitutive activation of HIF2A, making HIF2A an important clinical target. While HIF2A inhibitors are showing promising results in clinical trials, many patients do not respond and ultimately most patients progress. To improve our understanding of HIF2A dependency in metastatic ccRCC, I developed and thoroughly validated a comprehensive approach to identify HIF2A transcriptional targets *in vivo*. I demonstrated that HIF2A can be acutely degraded in a xenograft model of ccRCC using AID2 technology, and that combining this with RNA-seq uncouples direct targets from downstream signalling cascades. This unbiased method enabled the shortlisting of clinically relevant candidate HIF2A direct target genes which could then be functionally tested by *in vivo* CRISPR/Cas9 screening and subsequent rescue experiments. CRISPR/Cas9 screening revealed that *MYC* and *CCND1* are key dependencies in ccRCC *in vivo*, in addition to highlighting key differences between *in vitro* and *in vivo* dependencies. By leveraging a doxycycline-inducible HIF2A expression system, combined with exogenous constitutive expression of *MYC* or *CCND1*, I showed that MYC, but not CCND1, is sufficient to rescue tumour maintenance upon HIF2A loss. While both cell cycle-associated genes drive proliferative signalling, I demonstrated that only MYC protects from apoptosis, providing mechanistic insights into the role of HIF2A-dependent *MYC* expression in ccRCC. The results presented here provide a workflow for unbiased, *in vivo* characterisation of oncogenic transcription factors, and novel insights into HIF2A dependency in ccRCC. University of Cambridge 2023-08-29 Thesis Doctoral Doctor of Philosophy (PhD) eng https://www.repository.cam.ac.uk/handle/1810/364396 https://apollo8-f-pro.lib.cam.ac.uk/bitstreams/cd5003a3-e862-4dcc-a1ca-9ce18b887e27/download 87eda9de84448d1f82354d60eee3eb5f https://apollo8-f-pro.lib.cam.ac.uk/bitstreams/9d619b95-45a3-4655-800c-c8f95d243dbb/download 9d3eb8f9a6b02a501c2e6445293df497 https://www.rioxx.net/licenses/all-rights-reserved/ acute protein degradation AID2 Cancer ccRCC CRISPR/Cas9 Genetics HIF2A Oncology Renal cancer Transcriptomics