cells but not in primary human hepatocytes. The effect of NSC130362/ATO, NSC130362/Myr, and NSC130362/BSO combined treatment in breast carcinoma cells and MDA-MB-435 melanoma cells. The effect of NSC130362/ATO and NSC130362/Myr combined treatment in pancreatic, prostate, and lung carcinoma cells as well as in AML cells from cancer patients. Subconfluent cells in a 96-well plate were pre-incubated for 4 h with ATO, Myr, or BSO followed by treatment with NSC130362 for an additional 24 h. At the end of the treatment, the ratio of dead cells was determined by an ATPLite reagent. , P < 0.05. doi:10.1371/journal.pone.0129566.g008 17 / 26 Discovery of a New Component in the TRAIL Pathway Fig 9. PK studies of NSC130362 in the mouse bloodstream. PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19710468 the synergistic effect of ATO and NSC130362 in MIA PaCa-2 cells. The effect of ATO and NSC130362 on the viability of MIA PaCa-2 cells was determined as described in the legend for Fig 8. the ATO/NSC130362 combined treatment retards growth of MIA PaCa-2 xenografts in PG 490 immunodeficient mice. H&E-stained sections of mouse liver and heart. Magnification, 40x. , P = 0.03. doi:10.1371/journal.pone.0129566.g009 18 / 26 Discovery of a New Component in the TRAIL Pathway studies. The fourth group received combination of ATO and NSC130362. As expected, tumor growth in the second and third group was similar to that in the control group. Namely, in the control group the relative and relative median tumor volume was 109235% and 161% of the starting tumor volume, respectively. In the second and third group, the relative/relative median tumor volume was 107197% /135% PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19713490 and 98 244%/142%, respectively. On the other hand, in the fourth group the relative/relative median tumor volume was only 47227%/94%. Since the samples were small and the data did not follow a Gaussian distribution, we chose the Mann-Whitney U-test to assess the statistical significance of the obtained in vivo data. We chose one-tailed version of the test because we wanted to know 
if the treatment slowed down the tumor growth. The differences between the second and the control groups and the third and the control groups were not significant. In contrast, combination of ATO with NSC130362 in the fourth group caused statistically significant retardation in tumor growth, thus confirming the potency of the ATO/ NSC130362 combined treatment in vivo. At study conclusion tumor, liver and heart tissues from four mice per group were collected, fixed, and H&E stained. Pathological evaluation revealed no evidence of toxicity in normal tissue. Discussion The goal of this work was to find chemical compounds that selectively sensitize TRAIL-resistant tumor cells to the TRAIL-activated extrinsic apoptotic pathway without affecting normal cells. These compounds would provide useful research tools for interrogating mechanisms of TRAIL-resistance. They also could serve as the basis for future drug development programs to create a new generation of non-toxic anti-cancer drugs that restore sensitivity to endogenous pathways used by the immune system for eradicating tumors. Some chemotherapeutic agents can augment TRAIL-mediated apoptosis in cancer cells when co-administered with TRAIL in vitro and in vivo, but these treatments rely on the intrinsic apoptotic pathway and risk failure because of the acquired defects in the cell death machinery of tumors found in relapsed cancer patients. In addition, these cytotoxic agents can kill normal proliferating cells. Thus, with currently
