Variant of the CHEK2 gene as a prognostic marker in glioblastoma multiforme.

OBJECTIVE: Germline mutations of the CHEK2 tumor suppressor gene have been found in families with the Li-Fraumeni syndrome (LFS). Patients with LFS experience a variety of cancers, including malignant astrocytomas. We investigated a potential role for a CHEK2 gene polymorphism in glioblastomas. METHODS: A genetic polymorphism of the CHEK2 gene (CHEK2 SNP rs2017309 A/T) was genotyped in a series of glioblastoma patients (n = 213) and population controls (n = 192). Subsets of tumors were analyzed for loss of heterozygosity 22q(n = 66), loss of heterozygosity CHEK2 (n = 53), CHEK2 expression (n = 21), and CHEK2 coding sequence alterations (n = 18). CHEK2 SNP rs2017309 genotyping findings and traditional clinicopathological parameters were correlated with the patients' prognoses. RESULTS: No association between the CHEK2 SNP and glioblastoma formation was observed. No CHEK2 coding sequence aberrations or tumors completely lacking CHEK2 protein were identified. However, the presence of the CHEK2 rs2017309 A allele was significantly associated with an adverse prognosis (P = 0.034), particularly among patients undergoing postoperative chemotherapy and radiotherapy (n = 28, median survival 10.5 versus 15.5 mo, P = 0.008). We could confirm the patients' age, Karnofsky Performance Scale score, and postoperative radiotherapy and chemotherapy (all P < 0.0001, log-rank test) as decisive prognostic factors. CONCLUSION: Our data suggest that a CHEK2 gene polymorphism might correlate with the prognosis of glioblastoma patients. These findings may point to an as yet unrecognized role for the CHEK2 gene in glioblastomas.

Role of p16 and p14ARF in radio- and chemosensitivity of malignant gliomas.

In addition to cytoreductive surgery, most patients with malignant gliomas also undergo radio- and chemotherapy. An improved understanding of the molecular genetic mechanisms underlying the radio- and chemosensitivity of gliomas may help to identify glioma patients who will benefit from aggressive and, therefore, potentially toxic adjuvant treatment. It may also allow for the development of new therapies aimed at improving the response of these tumors towards chemo- and radiotherapy. The INK4a gene products, p16 and p14ARF, have been suggested as potential regulators of glioma chemo- and radiosensitivity. We have used tetracycline controlled expression of p16 and plasmid-based p14ARF expression to study the chemo- and radiosensitivity of glioma cell lines. Ectopic p16 sensitized U-87MG cells towards treatment with vincristine and possibly also BCNU by approximately 1.5 to 2-fold, and towards ionizing radiation by a factor of 1.5. p14ARF expression was found to render U-87MG cells 2-fold more radioresistant than controls. These findings support a role for p16 and p14ARF as modulators of the radio-and chemosensitivity of gliomas. Further studies of the role of cell cycle regulators in glioma chemo- and radio-sensitivity seem warranted. We would like to point out that such candidate genes which may code for potent growth suppressors (like p16) or even toxic gene products can be successfully investigated using the approach detailed in this manuscript.

Conditional expression of the tumor suppressor p16 in a heterotopic glioblastoma model results in loss of pRB expression.

We have expressed the tumor suppressor p16 under the control of a tetracycline-sensitive promoter in two human glioblastoma cell lines which do not contain endogenous p16. Ectopic p16 expression led to a stable but reversible G1 phase cell cycle arrest, reduced the growth of both cell lines in cell culture, and almost abolished their in vitro tumorigenicity. U-87MG-tTA-p16 glioblastoma cells consistently formed tumors after subcutaneous injection into the flanks of nude mice. p16 expression in these tumors was strictly dependent on the presence or absence of tetracycline in the drinking water. Ectopic p16 reduced the tumor take rate (in vivo tumorigenicity) of U-87MG-tTA-p16 cells from 18/20 (90%) to 5 tumors/12 (42%) tumor cell injections. p16 positive and negative tumors differed with respect to their Ki67 labeling indices (34 +/- 4% vs. 52 +/- 6% , P < 0.001, student's t-test). These data are consistent with an in vitro and in vivo glioma suppressor role for p16. Interestingly, we observed a secondary reduction of pRB expression in tumors (and cell cultures) exposed to p16 for > or = 10 (6) days. pRB is p16's major downstream target. Hence, this finding might explain, why p16 expression neither significantly affected the morphology nor led to a reduction of size or growth rate of the tumors. Loss of pRB following p16 expression might severely limit the potential benefit of p16 gene therapy for glioblastoma.