Targeting RAD51 phosphotyrosine-315 to prevent unfaithful recombination repair in BCR-ABL1 leukemia

Abstract

Chronic myeloid leukemia chronic phase (CML-CP) CD34 ؉ cells contain numerous DNA double-strand breaks whose unfaithful repair may contribute to chromosomal instability and disease progression to blast phase (CML-BP). These phenomena are often associated with the appearance of imatinib-resistant BCR-ABL1 kinase mutants (eg, T315I) and overexpression of BCR-ABL1. Here we show that BCR-ABL1 (nonmutated and T315I mutant) promoted RAD51 recombinase-mediated unfaithful homeologous recombination repair (HomeoRR) in a dosage-dependent manner. BCR-ABL1 SH3 domain interacts with RAD51 proline-rich regions, resulting in direct phosphorylation of RAD51 on Y315 (pY315). RAD51(pY315) facilitates dissociation from the complex with BCR-ABL1 kinase, migrates to the nucleus, and enhances formation of the nuclear foci indicative of recombination sites. HomeoRR and RAD51 nuclear foci were strongly reduced by RAD51(Y315F) phosphorylationless mutant. In addition, peptide aptamer mimicking RAD51(pY315) fragment, but not that with Y315F phosphorylation-less substitution, diminished RAD51 foci formation and inhibited HomeoRR in leukemia cells. In conclusion, we postulate that BCR-ABL1 kinase-mediated RAD51(pY315) promotes unfaithful HomeoRR in leukemia cells, which may contribute to accumulation of secondary chromosomal aberrations responsible for CML relapse and progression. (Blood. 2011;118(4):1062-1068) IntroductionChronic myeloid leukemia in chronic phase (CML-CP) is a myeloproliferative disorder characterized by the presence of the Philadelphia (Ph) chromosome that results from a (9;22)(q34;q11) reciprocal translocation that juxtaposes the c-abl oncogene 1 (ABL1) gene on chromosome 9 with the breakpoint cluster region (BCR) gene on chromosome 22, generating the BCR-ABL1 fusion oncogene. Most CML-CP patients are currently treated with tyrosine kinase inhibitor (TKI) imatinib designed to block the enzymatic action of the ABL1 tyrosine kinase. Approximately 60%-70% of patients achieve and maintain a complete cytogenetic response (CCyR) 5 years after initiating imatinib treatment. Two "second-generation" TKIs, dasatinib and nilotinib, are effective at inducing or restoring CCyR in 40%-50% of patients who appear to have failed primary treatment with imatinib. However approximately 20% of patients presenting with CML-CP fail to respond to both imatinib and a subsequent second-generation TKI; their prognosis is poor because of a higher risk of disease progression. In addition, because TKIs do not eradicate the disease, the patients in CCyR may carry 10 6 to 10 9 leukemia cells, and even BCR-ABL1-negative patients in complete molecular response may have up to 10 6 leukemia cells. 1 These cells may be the "time-ticking bombs" eventually exploding into TKI-resistant clone and/or CML-blast phase (CML-BP) clone on accumulation of additional genetic aberrations. 2 Clinical observations and experimental findings clearly demonstrated that genomic instability in CML is driven, at least in part, by BCR-ABL1 kinase. 2,3 However, TKI-treated CML pat...

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A phosphorylation–deubiquitination cascade regulates the BRCA2–RAD51 axis in homologous recombination

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