The Structural Filed Study Of Berberine Inhibiting Human Colon Cancer Growth Via Retinoid X Receptor Alpha And The Identification Of Derivatives Preliminarily

Retinoid X receptor(RXRa)is a member of nuclear receptor superfamily and involved in cell proliferation,differentiation,metabolism,embryonic development and other physiological processes,especially in carcinogenesis.Its ligands are expected to be antitumor agents.Berberine,an isoquinoline alkaloid,is a traditional oriental medicine which is extracted from coptis chinensis and used to treat diarrhea and gastroenteritis.Recently,it’s a popular trend to study the antitumor activity of berberine.However,the intracellular target mediating the effects of berberine remains elusive.Our previous experiments identified medicine berberine as a novel RXRa ligand.And berberine could increase transcriptional activity of RXRa,as well as degradation of intranuclear β-catenin,to inhibit the growth of colon cancer cells by physically binding to RXRa.Here,we provide evidence that berberine directly binds to a unique region comprising residues Gln275 and Arg316 in nuclear receptor RXRa,where berberine concomitantly binding to and synergistically activating RX-Ra with 9-cis-retinoic acid(9-cis-RA,9cRA),a natural RXRa ligand binding to the classical ligand binding pocket.And there is also a synergistic inhibitory effect of berberine and 9-cis-RA on colon cancer cell growth which is subjected to EdU assays.Real-time PCR analysis of FOX03A,a target gene of RXRa and a tumor repressor,shows a significant increasing of mRNA expression when berberine and 9-cis-RA added together in colon cancer cells.The sy’nergistic rather than competitive effect between berberine and 9-cis-RA exhibits that they might bind to different regions in RXRa.Berberine promotes RXRa interaction with nuclear β-catenin which is analyzed by mammalian two-hybrid assay and this effect is enhanced by berberine dose-dependently.These results thus suggest that binding to RXRa is required for berberine to promote RXRa-β-catenin interaction and inhibit Wnt activity and colon cancer cell growth.Berberine elvates the transcriptional activity of RXRa to inhibit the colon cancer cell growth by binding to RXRa.We preliminary understand how berberine binds to RXRa-LBD with the structural mode analysis of berberine and RXRa,which supports a new sight to design the superior RXRa-targeted antitumor medicine.Based on these researches,we try to keep the advantages of berberine’s low toxicity and high selectivity and develop the analogs with the better solubility and anti-cancer effect.We have designed and synthesized 21 kinds of berberine derivatives and identified these compounds by dual luciferase reporter gene assays and MTT tests to see their transcriptional activity and other biological abilities.We choose a special cyclical pattern combined transformation and indentification to analyze the structure-activity relationship.Now,there are two berberine derivatives 122 and 125,which have a better solubility,activation of RXRa transcription and stronger inhibition in human colon cancer compared with berberine and other derivatives for further drug metabolic analysis and transformation.