Choroid plexus carcinomas are rare tumors that typically occur in young children. Prognosis is poor, and very little information is available to optimize treatment protocols. We used a cell culture model to evaluate whether combining chemotherapeutic agents such as methotrexate with histone deacetylase inhibitors (HDACI) such as valproic acid and MS-275 could improve efficacy. Valproic acid increased the cytotoxicity of radiation and of all the chemotherapeutic agents in Z310 and SV11 mouse choroid plexus cell lines, with the exception of methotrexate. Both HDACIs made choroid plexus cells resistant to this folate antagonist. Searching for a molecular explanation, we found that thymidylate synthase was up regulated when the cells were incubated with HDACI. We also confirmed this finding in human choroid plexus carcinoma cells. Methotrexate should not be combined with HDACI in the treatment of choroid plexus carcinoma.