Antisense and drug-resistance gene therapy in a mouse model of chronic myeloid leukemia

Chronic myeloid leukemia (CML) is a disease of the hematopoietic stem cell, characterized by the bcr-abl fusion oncogene and subsequent p210bcr-abl protein expression. The selective p210 kinase inhibitor, STI571, has shown promise in recent clinical trials for treatment of CML. However, STI571-resistant leukemia has emerged in blast-phase CML and acute lymphoid leukemia patients. Autologous marrow transplants of the patient's own marrow for treatment of CML typically result in relapse due to leukemic contamination of the graft or incomplete ablation of leukemia prior to transplant. A drug-resistance and antisense gene therapy strategy has been developed to address the problems of autologous marrow transplant and to provide chemoprotection for improved anti-tumor chemotherapy. In a clinical approach, autologous marrow would be transduced with a retroviral vector expressing a drug-resistant L22Y dihydrofolate reductase ( DHFR) gene which provides hematopoietic protection from antifolate toxicity. The vector also expresses antisense RNA directed against bcr-abl, which is intended to reduce bcr-abl and p210 expression and restore a more normal phenotype to bcr-abl + leukemic cells. Pre-clinical animal testing of the drug-resistant DHFR component in the 32Dp210 mouse model of CML revealed that administration of the antifolate methotrexate exacerbated tumor progression in mice engrafted with marrow expressing the antifolate-resistant DHFR gene. However, administration of an alternate antifolate, trimetrexate, together with a nucleoside transport inhibitor prodrug, NBMPR-P, provided a strong anti-tumor effect, inhibiting growth of the 32Dp210 tumor and completely preventing tumor-associated animal mortality as long as drug administration continued. The antisense component of the gene therapy approach (involving expression of antisense RNA directed against bcr-abl) had previously been found decrease bcr-abl and p210 expression in STI571-sensitive CML cells. Tests of the antisense component in the STI571-resistant Baf/BCR-ABL-r1 murine tumor cell line proved ineffective, as the Baf/BCR-ABL-r1 cells became dependent upon STI571, and exhibited increased apoptosis when STI571 was withdrawn. Further, the STI571-resistant cells were no longer tumorigenic in mice. Recovery from STI571-withdrawal was found to result in partial restoration of tumorigenicity. Alternate STI571-resistant tumor models will be required to further test the efficacy of the bcr-abl antisense for treatment of STI571-resistant leukemia.