Fig. 5: Identify NR2E3-targeted combinatorial treatments in solid tumors.

A Illustration of treating human uterine endometrial cancer with 11a in explant culture. B Immunoblotting assays showed that one day treatment with 4 µM 11a increased p53 protein level, stimulated p53 acetylation at K319/386, activated p53-targeted genes, and induced cell apoptosis signaling in 2 out of 3 patient samples. C Repurposing screens of FDA-approved anti-cancer drug library (NCI AOD X version) presented Romidepsin as a lead to promote 11a’s cell-killing effect in cervical cancer HeLa cells. Four concentrations (1 nM, 20 nM, 300 nM and 5 µM) of each drug in the library were achieved when mixed with 50 µL of cell suspensions and one concentration of 11a (1.5 µM) were tested in 384-well plates. The cell viability was measured by CellTiter-Glo in two days and the DMSO control in each plate was normalized to 1. The normalized cell viabilities in the presence of 11a (y-axis) were plotted against those in the absence of 11a (x-axis) for each concentration. D ZIP drug synergy score showed the synergy between 11a and Romidepsin. 8 × 9 doses of 11a and Romidepsin were mixed with HeLa cells in 96-well plates for two days. The cell viability of DMSO control was normalized to 100%. The data were evaluated for synergy by ZIP score [39]. E The 11a-Romidepsin combo induced more HeLa cell apoptosis than each individual drug in one day in Annexin V-binding assay. p value was calculated by one-way Anova test and Bonferroni post-hoc test (DMSO vs 11a vs Romidepsin vs Combo). *p < 0.0083; **p < 0.00167; ***p < 0.00017. F The combo had much stronger inhibition of HeLa cell growth than each individual drug in two days. G Repurposing screens of AOD X library as done in (A) identified Bortezomib and Carfilzomib promoting 11a’s cell-killing effects in retinoblastoma Y79 cells. H ZIP drug synergy assay as (D) showed the synergies between 11a and Carfilzomib or Bortezomib in Y79 cells. I The 11a-Bortezomib combo induced more Y79 cell apoptosis than each individual drugs in one day in Annexin V-binding assay. p value was calculated by one-way Anova test and Bonferroni post-hoc test (DMSO vs 11a vs Romidepsin vs Combo). *p < 0.0083; **p < 0.00167; ***p < 0.00017. Mean ± SD shown in all the histograms.