In vitro studies of a FLT3 inhibitor combined with chemotherapy: sequence of administration is important to achieve synergistic cytotoxic effects

Patients with acute myeloid leukemia (AML) harboring internal tandem duplication mutations of the FLT3 receptor (FLT3/ITD mutations) have a poor prognosis compared to patients lacking such mutations. Incorporation of FLT3 inhibitors into existing chemotherapeutic regimens has the potential to improve clinical outcomes in this high-risk group of patients. CEP-701, an indolocarbazole-derived selective FLT3 inhibitor, potently induces apoptosis in FLT3/ITD-expressing cell lines and primary leukemic blasts. We conducted a series of in vitro cytotoxicity experiments combining CEP-701 with chemotherapy using the FLT3/ITD-expressing cell lines MV4-11 and BaF3/ITD as well as a primary blast sample from a patient with AML harboring a FLT3/ITD mutation. CEP-701 induced cytotoxicity in a synergistic fashion with cytarabine, daunorubicin, mitoxantrone, or etoposide if used simultaneously or immediately following exposure to the chemotherapeutic agent. In contrast, the combination of pretreatment with CEP-701 followed by chemotherapy was generally antagonistic, particularly with the more cell cycle-dependent agents such as cytarabine. This effect appears to be due to CEP-701 causing cell cycle arrest. We conclude that in FLT3/ITD-expressing leukemia cells, CEP-701 is synergistic with standard AML chemotherapeutic agents, but only if used simultaneously with or immediately following the chemotherapy. These results should be considered when designing trials combining chemotherapy with each of the FLT3 inhibitors currently in clinical development.