PDE4 as a possible pharmacological target for the treatment of hepatocellular carcinoma

Most human tumors show a decreased level of cyclic nucleotides and an overexpression of phosphodiesterases. Cyclic AMP-specific type 4 phosphodiesterases (PDE4) constitute the largest family with over 20 isoforms encoded by four genes, namely PDE4A, B, C and D, which give rise to many isoforms through alternative mRNA splicing. Their role in liver cancer has not been studied extensively. The goal of this study was to analyze the involvement of PDE4 in the hepatic tumor phenotype using different cell lines that show different degrees of aggressiveness. Hepatocarcinoma cells (HepG2, Hep3B and Huh7) have lower levels of cAMP and higher PDE activity, when compared to the non-tumorigenic, differentiated cells HepaRG. Hepatocarcinoma cells (HCC) also show overexpression of PDE4A and PDE4D proteins by western blot. In agreement, tumor tissues also display stronger staining for both isoforms when compared to normal hepatic tissues. To better analyze the involvement of these isoforms in the carcinogenic phenotype, PDE4D gene silencing experiments were conducted. Silencing of PDE4D reduced cell proliferation and induced apoptosis by regulating key cell cycle molecules. In light of a possible pharmacological application, the different cell lines were also treated with a specific inhibitor of isoform D, Gebr-7b. Also in this case, promising results have been obtained in the inhibition of growth for all the HCC lines under investigation. In addition, treatment of Huh7 cells with Gebr-7b caused inhibition of migration as shown in wound healing assay. In conclusion our study establishes a crucial role of PDE4D in the hepatic tumor phenotype. In addition to being an effective biomarker for diagnosis, PDE4D could represent a specific therapeutic target for the treatment of hepatocellular carcinoma, in particular for those cases that are refractory to existing therapies. Further studies are still needed to identify the specific role of individual PDE4D isoforms produced by alternative mRNA splicing.