CRISPR/Cas9 Screen in Gastric Cancer Patient‐Derived Organoids Reveals KDM1A‐NDRG1 Axis as a Targetable Vulnerability

Abstract

Viability CRISPR screens have proven indispensable in parsing genome function. However, their application in new, more physiologically relevant culturing systems like patient-derived organoids (PDOs) has been much slower. To probe epigenetic contribution to gastric cancer (GC), the third leading cause of cancer-related deaths worldwide, the first negative selection CRISPR screen in GC PDOs that faithfully preserve primary tumor characteristics is performed. Extensive quality control measurements showing feasibility of CRISPR screens in primary organoid culture are provided. The screen reveals the histone lysine demethylase-1A (KDM1A) to constitute a GC vulnerability. Both genetic and pharmacological inhibition of KDM1A cause organoid growth retardation. Further, it is shown that most of KDM1A cancer-supporting functions center on repression of N-myc downstream regulates gene-1 (NDRG1). De-repression of NDRG1 by KDM1A inhibitors (KDM1Ai) causes inhibition of Wnt signaling and a strong G1 cell cycle arrest. Finally, by profiling 20 GC PDOs, it is shown that NDRG1 upregulation predicts KDM1Ai response with 100% sensitivity and 82% specificity in the tested cohort. Thus, this work pioneers the use of negative selection CRISPR screens in patient-derived organoids, identifies a marker of KDM1Ai response, and accordingly a cohort of patients who may benefit from such therapy.

Human patient derived organoids: an emerging precision medicine model for gastrointestinal cancer research

“…In CRC, driver genes such as DACH1 ( Hu et al, 2020 ), LARGE2 ( Dietinger et al, 2020 ), and KIT ( Küçükköse et al, 2022 ) were knocked out by using CRIPSR/CAS9 gene editing technology to investigate the promoting effects of intestinal stemness, cell-matrix adhesion, and phenotype switch in carcinomatosis, respectively. In order to explore the epigenetic contribution in GC, the histone lysine demethylase-1A (KDM1A) gene was knocked out in PDOs, which in turn induced inhibition of Wnt signaling and a strong cell cycle arrest ( Mircetic et al, 2023 ). For the drug susceptibility test, Verissimo CS et al evaluated the effect of mutant KRAS on drug response and found that dual inhibition of the EGFR-MEK-ERK pathway in RAS mutant organoids induced a transient cell-cycle arrest rather than cell death ( Verissimo et al, 2016 ).…”

“…TFEB-PGD was considered another signaling axis activated by ATP13A2 to promote tumor growth ( Zhang et al, 2023b ). In GC, signaling axis of KDM1A-NDRG1 and NF-κB-PIEZO1-YAP1-CTGF might serve as potential therapeutic options to block tumorigenesis ( Chen et al, 2023 ; Mircetic et al, 2023 ). Besides, Chromosome eight open reading frame 76 (C8orf76) was for the first time identified as a novel therapeutic target for GC that directly binds to oncogenic lncRNA DUSP5P1 to induce its expression and activate MAPK signaling ( Wang et al, 2019 ).…”

“…In CRC, driver genes such as DACH1 (Hu et al, 2020), LARGE2 (Dietinger et al, 2020), and KIT (Küçükköse et al, 2022) were knocked out by using CRIPSR/CAS9 gene editing technology to investigate the promoting effects of intestinal stemness, cell-matrix adhesion, and phenotype switch in carcinomatosis, respectively. In order to explore the epigenetic contribution in GC, the histone lysine demethylase-1A (KDM1A) gene was knocked out in PDOs, which in turn induced inhibition of Wnt signaling and a strong cell cycle arrest (Mircetic et al, 2023).…”