Splicing modulation as novel therapeutic strategy against diffuse malignant peritoneal mesothelioma

Introduction: Therapeutic options for diffuse malignant peritoneal mesothelioma (DMPM) are limited to surgery and locoregional chemotherapy. Despite improvements in survival rates, patients eventually succumb to disease progression. We investigated splicing deregulation both as molecular prognostic factor and potential novel target in DMPM, while we tested modulators of SF3b complex for antitumor activity. Methods: Tissue-microarrays of 64 DMPM specimens were subjected to immunohistochemical assessment of SF3B1 expression and correlation to clinical outcome. Two primary cell cultures were used for gene expression profiling and in vitro screening of SF3b modulators. Drug-induced splicing alterations affecting downstream cellular pathways were detected through RNA sequencing. Ultimately, we established bioluminescent orthotopic mouse models to test the efficacy of splicing modulation in vivo. Results: Spliceosomal genes are differentially upregulated in DMPM cells compared to normal tissues and high expression of SF3B1 correlated with poor clinical outcome in univariate and multivariate analysis. SF3b modulators (Pladienolide-B, E7107, Meayamycin-B) showed potent cytotoxic activity in vitro with IC50 values in the low nanomolar range. Differential splicing analysis of Pladienolide-B-treated cells revealed abundant alterations of transcripts involved in cell cycle, apoptosis and other oncogenic pathways. This was validated by RT-PCR and functional assays. E7107 demonstrated remarkable in vivo antitumor efficacy, with significant improvement of survival rates compared to vehicle-treated controls. Conclusions: SF3B1 emerged as a novel potential prognostic factor in DMPM. Splicing modulators markedly impair cancer cell viability, resulting also in potent antitumor activity in vivo. Our data designate splicing as a promising therapeutic target in DMPM

Tracers to monitor the response to chemotherapy: in vitro screening of four radiopharmaceuticals.

Contains fulltext : 59201.pdf (publisher's version ) (Open Access)OBJECTIVES: It has been postulated that radiopharmaceuticals can be used to predict the therapeutic response to (chemo)therapy, which could lead to individualized treatment regimens. In this study, 18F-deoxyglucose, 99mTc-tetrofosmin, 125I-deoxyuridineribose, and 125I-methyltyrosine were tested for this purpose. METHODS: The uterine sarcoma cell line MES-SA (MDR-) and its multidrug resistant variant, MES-SA/Dx5 (MDR+), were used. The MDR+ cells express high levels of P-glycoprotein, which makes them relatively resistant to various chemotherapeutic agents. Cells were cultured in the presence of escalating concentrations of doxorubicin, and the cellular uptake of the radiopharmaceuticals was determined. RESULTS: Decreasing 18F-deoxyglucose uptake at escalating doxorubicin concentrations reflected the chemosensitivity of the cells: 18F-deoxyglucose uptake in the MDR- cells was reduced to 40% of the baseline level in the presence of 1 microM of doxorubicin, compared to 74% in the MDR+ cells. The 125I-deoxyuridineribose uptake in MDR- cells was reduced to 2% of the baseline level when cultured at a concentration of 1 microM of doxorubicin, while this was 79% in the MDR+ cells. The same trend was observed with 125I-methyltyrosine. The enhanced doxorubicin chemosensitivity of MDR+ cells in the presence of verapamil, a modulator of P-glycoprotein, was reflected by the reduced uptake of 18F-deoxyglucose, 125I-deoxyuridineribose, and 125I-methyltyrosine. Furthermore, baseline 99mTc-tetrofosmin uptake in MDR+ cells was more than six-fold lower than in MDR- cells. CONCLUSION: In the presence of doxorubicin, the uptake of 18F-deoxyglucose, 125I-deoxyuridineribose and, to a lesser extent, 125I-methyltyrosine is more pronouncedly reduced in MDR- cells than in MDR+ cells. The reversal of doxorubicin-resistance of MDR+ cells by verapamil was also reflected by the uptake of 18F-deoxyglucose, 125I-deoxyuridineribose, and 125I-methyltyrosine. 99mTc-tetrofosmin uptake reflected P-glycoprotein expression without exposure to doxorubicin