| Imatinib mesylate (IM) is first line therapy for patients diagnosed with BCR-ABL+ CML. Unfortunately, drug-resistance has emerged as a serious threat. Minimal residual disease (MRD) persists and many patients eventually relapse. MRD results from innate mechanisms of drug-resistance, notably cell adhesion mediated drug-resistance (CAM-DR). Relapses are most often due to acquired BCR-ABL point mutations including the difficult to treat BCR-ABLT315l mutation. First we asked whether inhibition of cell attachment can overcome innate drug-resistance. Next, we tested the feasibility of targeting acquired drug-resistance by preemptively immunizing against BCR-ABL T315l.;An unbiased genome-wide screen identified the chemokine receptors CXCR3 and CXCR4 as mediators of innate IM drug-resistance. Therefore we evaluated leukemic cell susceptibility to IM and found that RNAi-mediated knockdown of CXCR3 or CXCR4 resulted in a dramatic reduction in leukemia cell viability in the presence of drug. Furthermore, CXCR3 and CXCR4 were both found to activate integrin beta1 as well as cell-to-cell attachment. These results strongly implicate CXCR3 and CXCR4 in triggering CAM-DR in BCR-ABL+ leukemia.;Next, for the prevention of acquired drug-resistance we evaluated an immunotherapeutic approach targeting BCR-ABLT315l Peptides spanning BCR-ABLT315l were analyzed using MHC-binding peptide prediction algorithms, synthesized and evaluated in an in vitro MHC-binding assay. Six candidate T315l H2-Kb binding peptides were identified. Recombinant S. cerevisiae were then engineered to express T315-mutated BCR-ABL (yeastT315l). Administration of yeastT315l significantly extended survival upon challenge with BCR-ABLT315l leukemias. In confirmation of this result, the peripheral circulation of BCR-ABLT315l cells was reduced or eliminated in immunized, but not control, animals. Furthermore, after challenge with a mixed population of BCR-ABLWT and BCR-ABL T315l leukemias, the number of leukemic cells harboring BCR-ABL T315l was significantly reduced relative to cells expressing BCR-ABL WT in mice treated with yeastT315l, but not control yeast.;This work demonstrates the viability of surmounting two distinct mechanisms of drug-resistance. Inhibition of CXCR3 or CXCR4 represents a means of overcoming CAM-DR and sensitizing BCR-ABL+ cells to IM treatment. Likewise, immunotherapy with yeastT315l is a novel approach for the prevention of acquired drug-resistance in vivo. Combined, these two methods may afford an effective means of maintaining IM efficacy. |
