data show the crucial influence of treatment sequence about the cytotoxic effect of the drugs in vitro. Pilot studies demonstrated that hedgehog pathway inhibitor seeding 2,000 endothelial cells per well or 2,000 tumor cells per well for 72 hours allows for evaluation of the result of the medicines while cells were still in linear phase of proliferation.. Together, these demonstrate that TW 37 is more cytotoxic on an equimolar basis than cisplatin in endothelial cells and head and neck cancer cells in vitro. Combination of TW 37 and cisplatin showed enhanced cytotoxic effects for endothelial cells and head and neck cancer cells as compared with single drug therapy TW 37 was discovered using composition based database screening for molecules that interacted with Bcl 2 with high affinity and prevented its interaction with proteins of the Bcl 2 family, such as Bax, Bim, Bad, and Bid. Consequently, it’s not expected that TW 37 could influence Bcl 2 expression levels. Nevertheless, the effect of mixture TW 37 and cisplatin on Bcl 2 expression in endothelial cells and in head Metastatic carcinoma and neck cancer cells is not known. . Here, we observed that concentrations of TW 37 and/or cisplatin that inhibit cell growth don’t influence the expression of Bcl 2 in the endothelial cells or within the head and neck tumor cells. For mix therapy studies in endothelial cells, we selected three concentrations of TW 37 and three concentrations of cisplatin. Combination treatment had significantly higher cytotoxic effect than exposure to single drug in the three cancer cell lines examined here. The CI developments for both drugs in these cell lines were similar to the CI for endothelial cells. The CI was below 0. When higher Lenalidomide ic50 concentrations of TW 37 were found in combination with cisplatin in OSCC3 and UM 9 SCC 74A cells, indicating synergism between drugs. On the other hand, the CI was between 0. 9 and 1. 1 at the focus of TW 37, which illustrates chemical effects of the drug combination. The CI was 1. 1 with most conditions using the lowest concentration of TW 37, which displays lack of additive or synergistic effect of the combination. On the impact of the mix cisplatin and TW 37 treatment sequence features a major impact on the cytotoxicity of TW 37 and cisplatin in vitro Next, we investigated the impact of therapy sequence. We both started treatment simultaneously with both drugs, or finished the experimental period with both drugs together pre-treated with one drug for twenty four hours and then. The concentration of drugs was fixed at the 72 hour IC50 for equally endothelial cells and head and neck cancer cells. When we began treatment with both drugs in the same time the highest effect of combination treatment was observed. Particularly, pre-treatment with either drug essentially eliminated the main benefit of combination therapy, when compared with single drug therapy.