gms | German Medical Science

Objective: Therapy of Glioblastoma multiforme (GBM) involves surgery, Temozolomide chemotherapy and γ-irradiation. The latter two act by directly damaging DNA, which causes apoptosis of tumor cells. It has been shown that the methylation status of the MGMT gene correlates with the sensitivity of tumors to treatment with the DNA-alkylating drug Temozolomide. The repair enzyme MGMT removes alkyl-adducts from the DNA and thereby ensures survival of affected cells. Several such repair and control entities can be found in eukaryotic cells. They are activated by DNA-damage or blockage of DNA-replication, delay mitosis and facilitate DNA-repair.

Here we analyzed expression and function of Mad2B in GBM. It has been assumed that Mad2B is involved in regulating the mitotic cell cycle checkpoint. This checkpoint controls fidelity of spindle attachment to the chromatids. However, evidence accumulates that Mad2B might also be involved in DNA-damage control.

Methods: Expression of Mad2B was analyzed in a panel of 15 astrocytomas WHO grade 2, 15 GBM and 3 normal brains by Western blotting. The role of Mad2B in DNA-damage control was evaluated in cisplatin treated HeLa cells with and without Mad2B knock-down by siRNA. Mitotic and nuclear defects were documented by immunofluorescence microscopy.

Results: Mad2B expression was significantly decreased with increasing WHO grading of gliomas. Depletion of Mad2B had no effect on the normal cell cycle. However, DNA alkylation by cisplatin resulted in mitotic spindle defects, abnormalities during chromosome segregation and cytokinesis defects in Mad2B depleted cells. In DNA-damaged cells Mad2B interacted with Cdc20, a regulatory protein of the mitotic spindle checkpoint.

Conclusions: If DNA damage occurs, Mad2B interacts with Cdc20. This interaction inhibits the anaphase-promoting complex/cyclosome (APC/C) and delays chromosome segregation, thereby allowing DNA-repair. Therefore, Mad2B links DNA-damage to mitotic cell cycle control. Reduction of Mad2B expression resulted in mitotic and nuclear defects, such as cells with multiple nuclei and multiple centromeres, and contributes to development of aneuploidy, as it is seen in many tumors including GBM. Especially high grade gliomas show significant reduction in Mad2B expression.

Detection of such repair and cell cycle control factors, which are altered during development of high grade gliomas, could allow development of new therapeutic strategies by intensifying the effect of chemotherapeutics and γ-irradiation.