CRISPR-SID: Identifying EZH2 as a druggable target for desmoid tumors via in vivo dependency mapping

Abstract

Cancer precision medicine implies identification of tumor-specific vulnerabilities associated with defined oncogenic pathways. Desmoid tumors are soft-tissue neoplasms strictly driven by Wnt signaling network hyperactivation. Despite this clearly defined genetic etiology and the strict and unique implication of the Wnt/β-catenin pathway, no specific molecular targets for these tumors have been identified. To address this caveat, we developed fast, efficient, and penetrant genetic Xenopus tropicalis desmoid tumor models to identify and characterize drug targets. We used multiplexed CRISPR/Cas9 genome editing in these models to simultaneously target a tumor suppressor gene (apc) and candidate dependency genes. Our methodology CRISPR/Cas9 selection–mediated identification of dependencies (CRISPR-SID) uses calculated deviations between experimentally observed gene editing outcomes and deep-learning–predicted double-strand break repair patterns to identify genes under negative selection during tumorigenesis. This revealed EZH2 and SUZ12, both encoding polycomb repressive complex 2 components, and the transcription factor CREB3L1 as genetic dependencies for desmoid tumors. In vivo EZH2 inhibition by Tazemetostat induced partial regression of established autochthonous tumors. In vitro models of patient desmoid tumor cells revealed a direct effect of Tazemetostat on Wnt pathway activity. CRISPR-SID represents a potent approach for in vivo mapping of tumor vulnerabilities and drug target identification.

Update on Familial Adenomatous Polyposis-Associated Desmoid Tumors

“…49 In recent years, tazemetostat (EZH2 inhibitor) is found to have therapeutic effects on DTs in animal models, and in vitro experiments reveal that tazemetostat could directly inhibit the transcription of Wnt/β-catenin pathway in human sclerofibrosarcoma cells. 50 It is found that DTs lack relevant immune markers and almost all specimen tumor cells are negative for PD-L1, and PD-1 is only partially expressed in lymphocytes surrounding the tumor. 51 This suggests that immune checkpoint inhibitors may not be indicated for the treatment of DTs.…”

“…In recent years, tazemetostat (EZH2 inhibitor) is found to have therapeutic effects on DTs in animal models, and in vitro experiments reveal that tazemetostat could directly inhibit the transcription of Wnt/β-catenin pathway in human sclerofibrosarcoma cells. 50…”

Deep learning in CRISPR-Cas systems: a review of recent studies

“…A substantial contribution to the field of tumorigenesis research was made by Naert et al (2021) , who introduced CRISPR-SID. This approach integrated multiplexed CRISPR-Cas9 genome editing with deep learning, predicting double-strand break repair patterns to identify genes vital for tumorigenesis, suggesting a druggable potential of key genes.…”

Modelling human genetic disorders in Xenopus tropicalis

“…Mimicking missense variants will have a very high impact on the field, since a large proportion of disease variants are missense by nature. Base editing, which changes a single base without cutting the DNA backbone and is, therefore, perfectly suited to missense modelling ( Komor et al, 2016 ), has been shown to work in Xenopus ( Shi et al, 2019 ; Park et al, 2017 ; Naert et al, 2020 and 2021b ). Xenopus are particularly suited for this type of modelling due to the ease of generating many mutant lines.…”