UCSF

Chronic myeloid leukemia (CML), driven by the fusion protein BCR-ABL1, remains highly responsive to treatment with tyrosine kinase inhibitors (TKIs). Although overt resistance and relapse typically occurs through on-target kinase domain mutations, 25-50% of all resistant cases lack such mutations. Second generation BCR-ABL1 inhibitors, such as dasatinib and nilotinib, retain the ability to inhibit a number of kinase domain mutations. Still a number of mutations, notably the T315I gatekeeper mutation, provides continued resistance and only recently with the approval of ponatinib are we able to inhibit all known kinase domain mutations in BCR-ABL1. With ponatinib approval effective inhibitors for all known drug-resistant mutations are available suggesting that poorly understood overt off-target resistance mechanisms should henceforth constitute a greater percentage of overt clinical resistance. Through the use of CML patient samples we sought to define molecular mediators of disease persistence (the occurrence of a hematologic but no cytogenetic response) and overt off-target resistance (a complete loss of cellular response to BCR-ABL1 inhibition), two poorly understood mechanisms of resistance in CML. Chronic phase CML patients exhibiting overexpression of the EVI1 oncogene displayed disease persistence, while the identification of two blast crisis CML patients exhibiting overt off-target resistance were found to contain EVI1 overexpression in conjunction with activated NRAS. Validation of NRAS Q61K, in CML cell lines, exhibited maintenance of cell viability despite TKI treatment and this overt resistance correlated with increased MAPK signaling. Furthermore, expression of MEK-DD, an activated MEK allele, provided a degree of resistance comparable that seen in NRAS Q61K driven off-target resistance. This establishes MAPK signaling as the main culprit downstream of NRAS that mediates the resistant phenotype. This was further substantiated by identifying sensitivity to MEK inhibition both alone and to an even greater extent in combination with BCR-ABL1 inhibitors. In all, we show that EVI1 overexpression can contribute to disease persistence and the presence of activating RAS mutations can lead to overt resistance to BCR-ABL1 tyrosine kinase inhibitors.