Concentration and timing dependence of lethality enhancement between topotecan, a topoisomerase I inhibitor, and ionizing radiation

Purpose : Topotecan (TPT) is a water-soluble Topoisomerase I (Topo I) inhibitor with reported antineoplastic activity against a variety of solid tumors (including nonsmall cell lung, small cell lung, ovarian, breast, esophageal, and head and neck primaries) and leukemias. We sought to determine : (a) if TPT enhanced the lethal effects of ionizing radiation ; and (b) the biological and biochemical characteristics of the enhancement. Methods and Materials : Quiescent human radioresistant melanoma (Ul-Mel) cells were x-irradiated (1-12 Gy) and exposed to various TPT concentrations (0.1-300 μM) either before (for 4 h), during, or after (for 4 h) radiation. Survival was determined via colony forming assays and normalized to correct for drug cytotoxicity. The effects of TPT on radiation-related potential lethal damage repair (PLDR) and sublethal damage repair (SLDR) were measured. A modification of the SDS-KCI assay was used to quantify DNA-Topo I complexes. Results : Enhanced radiation lethality by TPT was observed using quiescent U1-Mel cells. The sensitizer enhancement ratio (SER) after a 4 h postirradiation exposure of 4 μM TPT was 1.6 at 10% survival. The effect was : (a) dependent on drug concentration, with lethality enhancement and minimal drug lethality alone in the 2-10 μM range for a 4 h posttreatment ; (b) dependent on timing, with enhancement observed only when drug was present at the time of, or shortly after, radiation ; and (c) irreversible, with inhibition of PLDR and SLDR. Exposure to TPT during or after radiation substantially elevated DNA-Topo I complexes (four- to tenfold) over control levels and complex formation correlated to some degree with loss of survival. Conclusions : TPT enhanced radiation lethality in vitro at low drug concentrations that are clinically feasible. The rationale and design of an ongoing Phase I trial which utilizes concurrent TPT and radiation is discussed.