Epigenetic-focused CRISPR/Cas9 screen identifies (absent, small, or homeotic)2-like protein (ASH2L) as a regulator of glioblastoma cell survival

Background: Glioblastoma is the most common and aggressive primary brain tumor with extremely poor prognosis, highlighting an urgent need for developing novel treatment options. Identifying epigenetic vulnerabilities of cancer cells can provide excellent therapeutic intervention points for various types of cancers. Method: In this study, we investigated epigenetic regulators of glioblastoma cell survival through CRISPR/Cas9 based genetic ablation screens using a customized sgRNA library EpiDoKOL, which targets critical functional domains of chromatin modifiers. Results: Screens conducted in multiple cell lines revealed ASH2L, a histone lysine methyltransferase complex subunit, as a major regulator of glioblastoma cell viability. ASH2L depletion led to cell cycle arrest and apoptosis. RNA sequencing and greenCUT&RUN together identified a set of cell cycle regulatory genes, such as TRA2B, BARD1, KIF20B, ARID4A and SMARCC1 that were downregulated upon ASH2L depletion. Mass spectrometry analysis revealed the interaction partners of ASH2L in glioblastoma cell lines as SET1/MLL family members including SETD1A, SETD1B, MLL1 and MLL2. We further showed that glioblastoma cells had a differential dependency on expression of SET1/MLL family members for survival. The growth of ASH2L-depleted glioblastoma cells was markedly slower than controls in orthotopic in vivo models. TCGA analysis showed high ASH2L expression in glioblastoma compared to low grade gliomas and immunohistochemical analysis revealed significant ASH2L expression in glioblastoma tissues, attesting to its clinical relevance. Therefore, high throughput, robust and affordable screens with focused libraries, such as EpiDoKOL, holds great promise to enable rapid discovery of novel epigenetic regulators of cancer cell survival, such as ASH2L. Conclusion: Together, we suggest that targeting ASH2L could serve as a new therapeutic opportunity for glioblastoma

Examining the role of chromatin modifying enzymes in medulloblastoma by utilizing a chemical library

Medulloblastoma (MB) is the most common paediatric brain tumor that arises during infancy and childhood and is a major cause of cancer related-morbidity and mortality in children. Recently, medulloblastomas are described as four distinct molecular subgroups (Wnt, sonic hedgehog, Group 3 and Group 4), which have distinct transcriptional, cytogenetic, and mutational spectra. Next-generation studies have revealed that adult medulloblastomas involve remarkably more somatic SNVs and indels than paediatric counterparts, suggesting that epigenetic deregulation might have a foremost role in the initiation and progression of paediatric medulloblastomas

Assessing the function of chromatin modifying enzymes in medulloblastoma

Medulloblastoma is the most common pediatric brain tumor that arises during infancy and childhood and is a major cause of cancer related-morbidity and mortality in children. Recently, medulloblastomas are described as four distinct molecular subgroups (Wnt, sonic hedgehog, Group 3 and Group 4), which have distinct transcriptional, cytogenetic, and mutational spectra

Examining the role of chromatin modifying enzymes in medulloblastoma by utilizing a chemical library

Medulloblastoma (MB) is the most common paediatric brain tumor that arises during infancy and childhood and is a major cause of cancer related-morbidity and mortality in children. Recently, medulloblastomas are described as four distinct molecular subgroups (Wnt, sonic hedgehog, Group 3 and Group 4), which have distinct transcriptional, cytogenetic, and mutational spectra. Next-generation studies have revealed that adult medulloblastomas involve remarkably more somatic SNVs and indels than paediatric counterparts, suggesting that epigenetic deregulation might have a foremost role in the initiation and progression of paediatric medulloblastomas

Investigation of the effects of histone deacetylase inhibitors (HDACi) on apoptotic pathway in glioblastoma multiforme (GBM)

Abstract Not Availabl

Systematic characterization of chromatin modifying enzymes identifies KDM3B as a critical regulator in castration resistant prostate cancer

Androgen deprivation therapy (ADT) is the standard care for prostate cancer (PCa) patients who fail surgery or radiotherapy. While initially effective, the cancer almost always recurs as a more aggressive castration resistant prostate cancer (CRPC). Previous studies have demonstrated that chromatin modifying enzymes can play a critical role in the conversion to CRPC. However, only a handful of these potential pharmacological targets have been tested. Therefore, in this study, we conducted a focused shRNA screen of chromatin modifying enzymes previously shown to be involved in cellular differentiation. We found that altering the balance between histone methylation and demethylation impacted growth and proliferation. Of all genes tested, KDM3B, a histone H3K9 demethylase, was found to have the most antiproliferative effect. These results were phenocopied with a KDM3B CRISPR/Cas9 knockout. When tested in several PCa cell lines, the decrease in proliferation was remarkably specific to androgen-independent cells. Genetic rescue experiments showed that only the enzymatically active KDM3B could recover the phenotype. Surprisingly, despite the decreased proliferation of androgen-independent cell no alterations in the cell cycle distribution were observed following KDM3B knockdown. Whole transcriptome analyses revealed changes in the gene expression profile following loss of KDM3B, including downregulation of metabolic enzymes such as ARG2 and RDH11. Metabolomic analysis of KDM3B knockout showed a decrease in several critical amino acids. Overall, our work reveals, for the first time, the specificity and the dependence of KDM3B in CRPC proliferation