Abstract
DNA alkylating drug, temozolomide (TMZ), is a common drug of chemotherapy against brain tumors. However, the therapeutic efficacy of this drug is limited by the development of resistance. It has been reported that the mechanism of this resistance is complex and involves multiple DNA repair pathways. Fanconi anemia (FA) pathway was initially identified by virtue of its inactivation in a rare genetic disorder. In mammalian cells, the FA pathway is frequently activated in response to DNA strand breaks, replication arrest, and other cytotoxic secondary lesions induced by TMZ. The regulation of the FA pathway is highly complex, involving at least 12 proteins (FANCA, B, C, D1, D2, E, F, G, I, J, L, M). In this work, we tested the role of the FA pathway in mediating cellular resistance to TMZ. We used the cultured mouse embryonic fibroblasts; FANCA -/-, FANCC -/-, FANCD2 -/- cells, their parental cells (provided by Fanconi Anemia Cell Repository; Oregon Health and Science Univ., USA) and CHO cells; FANCD1mt, FANCGmt and their parental cells (provided by Dr. Larry H. Thompson; Lawrence Livermore National Laboratory, USA). We examined the cell survival after 3 h TMZ treatment by colony forming assay. The sensitivity of each cell line was assessed from its D50 value, i.e. from the TMZ dose which reduced cell survival to 50%. In order to accurately compare TMZ sensitivities in the repair defective cell lines, the relative D50 values were normalized using the D50 values of the corresponding proficient cell lines. The relative D50 values listed sequentially in the order in which they increase (reflecting decreasing sensitivities to TMZ) are: FANCD1mt cells < FANCGmt cells < FANCD2 -/- cells < FANCA -/- cells < FANCC -/- cells. The most effective target could be FANCD1 in TMZ treatment. Therefore, we are going to apply siRNA of FANCD1 against human glioblastoma cells in TMZ treatment.
