Figure S1: Images for supplementary information on SMM screens; Figure S2: The detail information of compound DSHN; Figure S3: MTT assay in MHCC97H, Huh7 and Hepa1 cells treated with indicated concentration of compound #29 (DSHN) for 72 hours; Figure S4: A-B: Mammalian two hybrid assay; Figure S5: A. Semi-quantitative PCR analysis of mShp mRNA in SHP transgenic mice (stg) liver; Figure S6: The DNA copy number of SHP gene were analyzed from The Cancer Genome Atlas; Figure S7: A. Survival proportions for lung cancer. B. Survival proportions for ovarian cancer stage 4. C. Quantitative RTPCR profiling of the nuclear receptor superfamily in thirty pairs of human lung tumor and corresponding normal epithelium.
ARTICLE ABSTRACT
Small heterodimer partner (SHP, NR0B2) is a nuclear orphan receptor without endogenous ligands. Due to its crucial inhibitory role in liver cancer, it is of importance to identify small molecule agonists of SHP. As such, we initiated a probe discovery effort to identify compounds capable of modulating SHP function. First, we performed binding assays using small molecule microarrays (SMM) and discovered 5-(diethylsulfamoyl)-3-hydroxynaphthalene-2-carboxylic acid (DSHN) as a novel activator of SHP. DSHN transcriptionally activated Shp mRNA, but also stabilized the SHP protein by preventing its ubiquitination and degradation. Second, we identified Ccl2 as a new SHP target gene by RNA-seq. We showed that activation of SHP by DSHN repressed Ccl2 expression and secretion by inhibiting p65 activation of CCL2 promoter activity, as demonstrated in vivo in Shp−/− mice and in vitro in HCC cells with SHP overexpression and knockdown. Third, we elucidated a strong inhibitory effect of SHP and DSHN on HCC cell migration and invasion by antagonizing the effect of CCL2. Lastly, by interrogating a publicly available database to retrieve SHP expression profiles from multiple types of human cancers, we established a negative association of SHP expression with human cancer metastasis and patient survival. In summary, the discovery of a novel small molecule activator of SHP provides a therapeutic perspective for future translational and preclinical studies to inhibit HCC metastasis by blocking Ccl2 signaling. Mol Cancer Ther; 15(10); 2294–301. ©2016 AACR.
