Sulforaphane Inhibits the Expression of Long Noncoding RNA H19 and Its Target APOBEC3G and Thereby Pancreatic Cancer Progression

SIMPLE SUMMARY: Pancreatic ductal adenocarcinoma (PDAC) is a highly malignant tumor with poor therapeutic responses and short survival. The identification of factors that make PDAC so deadly and their targeting is important. Sulforaphane has shown promise in experimental and epidemiological studies, as well as in initial patient pilot studies. We examined the influence of sulforaphane to the largely unexplored epigenetic regulators “long noncoding RNAs” (lncRNAs). Sulforaphane modulated the expression of the lncRNAs H19, MALAT1, HOTAIR, HOTTIP and PVT1. The downregulation of the tumor promoter H19 and its target gene APOBEC3G was most significant and converged in inhibition of Smad2/TGF-β, which is involved in prevention of PDAC progression. Our data identified APOBEC3G as a new H19 target and a novel therapeutic target in PDAC, which can be inhibited by sulforaphane. The provided gene array accession numbers are important for future exploration of the suggested mechanism. ABSTRACT: Pancreatic ductal adenocarcinoma (PDAC) is extremely malignant and the therapeutic options available usually have little impact on survival. Great hope is placed on new therapeutic targets, including long noncoding RNAs (lncRNAs), and on the development of new drugs, based on e.g., broccoli-derived sulforaphane, which meanwhile has shown promise in pilot studies in patients. We examined whether sulforaphane interferes with lncRNA signaling and analyzed five PDAC and two nonmalignant cell lines, patient tissues (n = 30), and online patient data (n = 350). RT-qPCR, Western blotting, MTT, colony formation, transwell and wound healing assays; gene array analysis; bioinformatics; in situ hybridization; immunohistochemistry and xenotransplantation were used. Sulforaphane regulated the expression of all of five examined lncRNAs, but basal expression, biological function and inhibition of H19 were of highest significance. H19 siRNA prevented colony formation, migration, invasion and Smad2 phosphorylation. We identified 103 common sulforaphane- and H19-related target genes and focused to the virus-induced tumor promoter APOBEC3G. APOBEC3G siRNA mimicked the previously observed H19 and sulforaphane effects. In vivo, sulforaphane- or H19 or APOBEC3G siRNAs led to significantly smaller tumor xenografts with reduced expression of Ki67, APOBEC3G and phospho-Smad2. Together, we identified APOBEC3G as H19 target, and both are inhibited by sulforaphane in prevention of PDAC progression.