Dual inhibition of FACT and RRM2 suppresses the progression of pancreatic ductal adenocarcinoma driven by the oncohistone H2BG53D

Pancreatic Ductal Adenocarcinoma (PDAC) is a highly lethal cancer with frequent genetic mutations. We previously reported the identification of the histone H2BG53-to-D mutation in PDAC patients and revealed that the mutation destabilizes nucleosomes and promotes cell migration. However, how H2BG53D impacts chromatin and the significance in PDAC development remain largely unknown. Using the genetically engineered mouse models, we demonstrate that H2BG53D accelerates PDAC initiation and metastasis. Moreover, H2BG53D mutation elevates the interaction between nucleosomes and the Facilitates Chromatin Transcription (FACT) complex. FACT depletion reverses transcriptional changes and oncogenic effects induced by H2BG53D mutation. In search of a novel therapeutic strategy for H2BG53D-PDAC, our genome-wide CRISPR/Cas9 screen identified Ribonucleotide Reductase Regulatory Subunit M2 (RRM2) as a druggable target. Remarkably, dual inhibition of RRM2 and FACT exhibits potent cytotoxic effects on PDAC cells in vitro and significantly prolongs the survival of H2BG53D-PDAC mice. This study deciphers the roles and mechanisms of H2BG53D in PDAC progression and proposes a promising therapeutic strategy for this aggressive cancer. Overall design: mouse RNA-seq: KPC and G53D-KPC mice were generated to study the roles of H2BG53D mutation in PDAC development. Primary PDAC tumors and metastases from KPC and G53D-KPC mice were used for this experiment. Gene expression was profiled by RNA-seq. KI cell line RNA-seq: FLAG tagged Wildtype (WT) H2B and H2BG53D knockin PDAC cells were used to study the mechanisms of H2BG53D on transcriptional alteration and oncogenic phenotype. RNA-seq were conducted in in two WT and two H2BG53D cells with or without SPT16 (one subunit of FACT) depletion. KI cell line CUT&RUN seq: FLAG tagged WildtypeH2B (H2B_WT-FLAG)and H2BG53D-FLAG knockin PDAC cells were used to study the mechanisms of H2BG53D on transcriptional alteration and oncogenic phenotype. CUT&RUN seq (targeting FLAG and H2A) were conducted in parental, two WT and two H2BG53D cells with or without SPT16 (one subunit of FACT) depletion. KI cell line CRISPR screen: FLAG tagged WildtypeH2B (H2B_WT-FLAG) and H2BG53D-FLAG knockin PDAC cells were used to identify the synthetic lethal partner for H2BG53D mutation in PDAC.