Poster Presentation 35th Lorne Cancer Conference 2023

CRISPR activation screens identify co-drivers of malignant haematopoiesis in a DNMT3a mutant background (#130)

Amali Cooray 1 2 , Erin M Lawrence 1 2 , Maggie A Potts 1 2 , Lin Tai 1 2 , Andrew Kueh 1 2 , Marco Herold 1 2
  1. Walter and Eliza Hall Institute, Parkville, VIC, Australia
  2. Department of Medical Biology, University of Melbourne, Melbourne, VIC, Australia

Genome-wide sequencing studies have shown that the genetic basis of Acute Myeloid Leukaemia (AML) is highly complex. Mutations in de novo methyltransferase 3a (DNMT3a) are overrepresented in patients with AML. However, DNMT3a mutations alone are not sufficient to cause malignancies. Therefore, I aim to identify co-drivers of malignancies in a mutant DNMT3a background. This project utilises a mouse model which has the most frequently found DNMT3a mutation in human AML, in the endogenous mouse DNMT3a locus. Crossing these mice to a recently developed CRISPR activation (CRISPRa) model allows the induction of any gene in a mutant DNMT3a cell. We transduced a CRISPR activation library targeting transcriptional regulators, into DNMT3a mutant/CRISPRa derived haematopoietic stem and progenitor cells (HSPC). These engineered HSPCs were used to reconstitute the haematopoietic systems of lethally irradiated wild type mice. Upon the development of malignancy the mice were euthanised, and next generation sequencing was used to identify the sgRNAs mediating tumour development. Through further validation of hits identified in these experiments, we will be able to detect thus far underappreciated genes or combinations of genes which lead to malignancies in cooperation with mutant DNMT3a.

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  5. Deng, Y., Diepstraten, S.T., Potts, M.A. et al. Generation of a CRISPR activation mouse that enables modelling of aggressive lymphoma and interrogation of venetoclax resistance. Nat Commun 13, 4739 (2022).