Acute Leukemia in the Elderly: Development of Novel, Targeted Therapeutics

National Institute On Aging

Investigators

Abstract

Despite therapeutic advances, treatment of adult ALL results in long term survival of only 30-40% of patients, with a significantly worse prognosis for patients over the age of 60. We have examined the activity of ABT-888, a potent inhibitor of PARP-1 and -2, in combination with several chemotherapeutic agents against a panel of human ALL and AML cell lines. No enhanced killing was noted when ABT-888 was combined with VP-16 (topoisomerase II inhibitor), trichostatin A (histone deacetylase inhibitor), or cloretazine (alkylating agent). However, synergistic killing of ALL (but not AML) was observed when ABT-888 was combined with the topoisomerase I inhibitor topotecan, and to a greater extent with combination of ABT-888 and temozolomide as determined in a 48h WST-1 screening assay using temozolomide at concentrations from 1-200 microM combined with ABT-888 (5 microM). The pre-B ALL lines Reh, RS4;11, and SUP-B15 were all very resistant to temozolomide alone with IC50s > 200 microM, consistent with the high levels of MGMT expressed by these cell lines. Culture in ABT-888 alone did not inhibit cell proliferation. However, when pre-B ALL cells were incubated for 30 or 60 minutes in clinically achievable concentrations of temozolomide (50-100 microM), washed, and then cultured for 48h in the presence of ABT-888 (5 microM), the IC50s for the combination were 25-50 microM, demonstrating a dramatic synergy for this combination in B-lineage ALL. Interestingly, AML cell lines pulsed in temozolomide and then cultured in ABT-888 as described above showed no inhibition of cell proliferation. Pre-B ALL cells cultured in temozolomide + ABT-888, but not either drug alone, arrested in S-phase and subsequently underwent apoptosis. Increased levels of gamma-H2AX foci were noted in combination-treated cells, and this occurred prior to the onset of apoptosis as determined by PARP cleavage. These data are consistent with a model in which single strand DNA breaks are generated at sites of temozolomide-induced N3-methyladenine adducts due to the actions of DNA glycosylase and APE1, however further repair via the base excision repair (BER) pathway is precluded by PARP inhibition. This leads to double strand DNA breaks during S-phase upon replication fork collapse and subsequent apoptosis. We hypothesize that while AML cells can repair DNA breaks using homologous recombination pathways which resolve dsDNA breaks, pre-B cells are relatively deficient in this pathway and thus exhibit selective sensitivity to this drug combination (synthetic lethal relationship). Experiments are underway to test this hypothesis. The in vitro results predict that the combination of temozolomide + ABT-888 could be an effective new treatment for adult ALL.