Abstract
BackgroundTo study whether and how c-MYC expression determines response to radio- and chemotherapy in childhood medulloblastoma (MB).MethodsWe used DAOY and UW228 human MB cells engineered to stably express different levels of c-MYC, and tested whether c-MYC expression has an effect on radio- and chemosensitivity using the colorimetric 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium inner salt (MTS) assay, clonogenic survival, apoptosis assays, cell cycle analysis, and western blot assessment. In an effort to validate our results, we analyzed c-MYC mRNA expression in formalin-fixed paraffin-embedded tumor samples from well-documented patients with postoperative residual tumor and compared c-MYC mRNA expression with response to radio- and chemotherapy as examined by neuroradiological imaging.ResultsIn DAOY - and to a lesser extent in UW228 - cells expressing high levels of c-MYC, the cytotoxicity of cisplatin, and etoposide was significantly higher when compared with DAOY/UW228 cells expressing low levels of c-MYC. Irradiation- and chemotherapy-induced apoptotic cell death was enhanced in DAOY cells expressing high levels of c-MYC. The response of 62 of 66 residual tumors was evaluable and response to postoperative radio- (14 responders (CR, PR) vs. 5 non-responders (SD, PD)) or chemotherapy (23 CR/PR vs. 20 SD/PD) was assessed. c-MYC mRNA expression was similar in primary MB samples of responders and non-responders (Mann-Whitney U test, p = 0.50, ratio 0.49, 95% CI 0.008-30.0 and p = 0.67, ratio 1.8, 95% CI 0.14-23.5, respectively).Conclusionsc-MYC sensitizes MB cells to some anti-cancer treatments in vitro. As we failed to show evidence for such an effect on postoperative residual tumors when analyzed by imaging, additional investigations in xenografts and larger MB cohorts may help to define the exact function of c-MYC in modulating response to treatment.
Highlights
To study whether and how c-MYC expression determines response to radio- and chemotherapy in childhood medulloblastoma (MB)
Susceptibility of DAOY and UW228 cells expressing different levels of c-MYC to IR, etoposide, cisplatin, carboplatin, doxorubicin, methotrexate, and vincristine To investigate whether the sensitivity of MB cells to radioand chemotherapy depend on the c-MYC expression level, we assessed cell viability of DAOY and UW228 human MB cells expressing different levels of c-MYC 72 h following irradiation with 0, 2, 5, or 10 Gy (Figure 1), or 72 h after treatment with different concentrations of etoposide, cisplatin, carboplatin, doxorubicin, methotrexate, or vincristine (Figure 2)
The clonogenic survival of DAOY M2 cells was more reduced after IR, cisplatin, etoposide, and doxorubicin when compared with control cells
Summary
To study whether and how c-MYC expression determines response to radio- and chemotherapy in childhood medulloblastoma (MB). In a recent study we have demonstrated, using small interfering RNA (siRNA) to inhibit c-MYC expression in D341, D425 and DAOY MB cells transiently, that c-MYC down-regulation may reduce sensitivity to radiotherapy, cisplatin, and etoposide treatment [8]. To validate these results and to better understand the effect of c-MYC on MB treatment, we analyzed the response of DAOY and UW228 MB cells engineered to stably express different levels of c-MYC to irradiation and to a panel of chemotherapeutic drugs. We analyzed c-MYC mRNA expression in formalin-fixed paraffin-embedded tumor samples (FFPE) from well-documented patients with postoperative residual tumor mass treated within the prospective multi-center studies HIT’91 and HIT 2000 and compared c-MYC mRNA expression with response to radio- and chemotherapy, as determined by neuroradiological imaging
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