DNA methyltransferase 3A (DNMT3A) catalyses the de novo methylation of cytosine at the 5’ position to form 5-methylcytosine. Mutations in DNMT3A occur in 30% of all cytogenetically normal acute myeloid leukaemias (AML) and are associated with poor prognosis. The most common DNMT3A mutation is a substitution at codon 882 that encodes a hypomorphic protein (DNMT3AR882H) which exerts a dominant negative effect over the remaining wild-type protein. DNMT3AR882H has been shown to contribute to leukaemia initiation in vivo, however, whether sustained expression is essential for maintenance of the disease remains unknown.
We have generated a tractable, clinically relevant murine model of AML in which expression of a tetracycline-inducible DNMT3AR882H allele is combined with constitutive expression of two commonly co-occurring oncogenes; NRASG12D and IDHR140Q. Concurrent expression of all 3 constructs in vivo resulted in a fully penetrant leukaemia that was serially transplantable.
Genetic depletion of DNMT3AR882H in established leukaemias in vivo delayed disease progression, and conferred a significant survival advantage. Immunophenotype analysis of leukaemic cells revealed that DNMT3AR882H depletion was associated with loss of the immature marker cKit and a concomitant increase in the mature myeloid marker Cd11b, implicating a role for DNMT3AR882H in blocking differentiation.
Altogether, our results indicate that sustained DNMT3AR882H expression is essential for maintaining the leukaemic phenotype. To further explore DNMT3AR882H dependent signalling pathways we will use RNA and bisulfite sequencing to delineate transcriptional programs and DNA methylation profiles. As DNMT3AR882H cannot be targeted directly by small molecule inhibitors, characterisation of downstream signalling pathways will identify novel protein targets which can be exploited for therapeutic purposes in AML harbouring this mutation.