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Fig. 6 | Journal of Experimental & Clinical Cancer Research

Fig. 6

From: Synergistic antitumor interaction of valproic acid and simvastatin sensitizes prostate cancer to docetaxel by targeting CSCs compartment via YAP inhibition

Fig. 6

Valproic acid/simvastatin combination reverts docetaxel-resistance in prostate cancer models. a Dose response curves of 22Rv1 R_39 cells treated for 96 h with increasing concentrations of DTX alone or with VPA/SIM combination and compared with 22Rv1_wt treated with DTX alone. Cell growth is expressed as percentage of control and was assessed by sulforhodamine B colorimetric assay (see Methods). b Combination Index (CI) and Dose Reduction Index (DRI) values (means ±S.D.) of VPA/SIM or VPA/SIM/DTX combinations (50:50 cytotoxic ratio) at 96 h in 22Rv1 R_39 cells from at least three separate experiments performed in quadruplicate computed at 50% of cell kill (CI50) by CalcuSyn software (Biosoft,Cam- bridge, UK). aCIs: < 0.8 strong synergism; CIs < 0.9 sinergysm; between 0.9 and 1.2 additivity; (or > 1.2 antagonism. bDRI values represent the order of magnitude (fold) of dose reduction obtained for IC50 (DRI50) in combination setting compared with each drug alone. c NANOg mRNA expression evaluated by RT-PCR at basal level in both 22Rv1 wt and 22Rv1 R_39 in adhesion conditions and in 1st generation spheres. β-actin was used as housekeeping control gene to normalize RT-PCR reactions. d Surface marker expression (CD44 and CD113) was determined by flow cytometry on 22Rv1 R_39 cells treated for 24 h with VPA and/ SIM at the IC5096h doses. e Western blot analysis of basal pYAP, and YAP, in 22Rv1 (wt), 22Rv1 R_39 and YAP5SA-tansfected 22Rv1 R_39 cells. Densitometric analysis was performed by ImageJ image software; ponceau red serves as loading control. f 22Rv1_wt, 22Rv1 R_39 and YAP5SA –transfected 22Rv1 R_39 2nd generation spheres (sphereD). Cells (40,000/mL) were seeded in sphere medium in low attachment 96 multiwell untreated or treated with VPA and/or SIM and/or DTX at the IC5096h doses relative to 22Rv1 wt cells, then disaggregated and plated again for 72 h without additional treatment. Spheroids viability was assessed by ATP luminescence assay. The values are the means ± S.D. from at least three independent experiments. Statistically significant results are reported (*** indicates P < 0.0005, ** indicates P < 0.005 and * indicates P < 0.05)

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