Bioinformatic analysis of RHO family of GTPases identifies RAC1 pharmacological inhibition as a new therapeutic strategy for hepatocellular carcinoma.

OBJECTIVE: The RHO family of GTPases, particularly RAC1, has been linked with hepatocarcinogenesis, suggesting that their inhibition might be a rational therapeutic approach. We aimed to identify and target deregulated RHO family members in human hepatocellular carcinoma (HCC). DESIGN: We studied expression deregulation, clinical prognosis and transcription programmes relevant to HCC using public datasets. The therapeutic potential of RAC1 inhibitors in HCC was study in vitro and in vivo. RNA-Seq analysis and their correlation with the three different HCC datasets were used to characterise the underlying mechanism on RAC1 inhibition. The therapeutic effect of RAC1 inhibition on liver fibrosis was evaluated. RESULTS: Among the RHO family of GTPases we observed that RAC1 is upregulated, correlates with poor patient survival, and is strongly linked with a prooncogenic transcriptional programme. From a panel of novel RAC1 inhibitors studied, 1D-142 was able to induce apoptosis and cell cycle arrest in HCC cells, displaying a stronger effect in highly proliferative cells. Partial rescue of the RAC1-related oncogenic transcriptional programme was obtained on RAC1 inhibition by 1D-142 in HCC. Most importantly, the RAC1 inhibitor 1D-142 strongly reduce tumour growth and intrahepatic metastasis in HCC mice models. Additionally, 1D-142 decreases hepatic stellate cell activation and exerts an anti-fibrotic effect in vivo. CONCLUSIONS: The bioinformatics analysis of the HCC datasets, allows identifying RAC1 as a new therapeutic target for HCC. The targeted inhibition of RAC1 by 1D-142 resulted in a potent antitumoural effect in highly proliferative HCC established in fibrotic livers.

Development of an Improved Guanidine-Based Rac1 Inhibitor with in†vivo Activity against Non-Small Cell Lung Cancer.

The Rho GTPase Rac1 is involved in the control of cytoskeleton reorganization and other fundamental cellular functions. Aberrant activity of Rac1 and its regulators is common in human cancer. In particular, deregulated expression/activity of Rac GEFs, responsible for Rac1 activation, has been associated to a metastatic phenotype and drug resistance. Thus, the development of novel Rac1-GEF interaction inhibitors is a promising strategy for finding new preclinical candidates. Here, we studied structure-activity relationships within a new family of N,N'-disubstituted guanidine as Rac1 inhibitors. We found that compound 1D-142, presents superior antiproliferative activity in human cancer cell lines and higher potency as Rac1-GEF interaction inhibitor in vitro than parental compounds. In addition, 1D-142 reduces Rac1-mediated TNFα-induced NF-κB nuclear translocation during cell proliferation and migration in NSCLC. Notably, 1D-142 allowed us to show for the first time the application of a Rac1 inhibitor in a lung cancer animal model.

Proapoptotic and antiinvasive activity of Rac1 small molecule inhibitors on malignant glioma cells.

Malignant gliomas are characterized by an intrinsic ability to invade diffusely throughout the normal brain tissue. This feature contributes mainly to the failure of existing therapies. Deregulation of small GTPases signaling, in particular Rac1 activity, plays a key role in the invasive phenotype of gliomas. Here we report the effect of ZINC69391, a specific Rac1 inhibitor developed by our group, on human glioma cell lines LN229 and U-87 MG. ZINC69391 is able to interfere with the interaction of Rac1 with Dock180, a relevant Rac1 activator in glioma invasion, and to reduce Rac1-GTP levels. The kinase Pak1, a downstream effector of Dock180-Rac1 signaling, was also downregulated upon ZINC69391 treatment. ZINC69391 reduced cell proliferation, arrested cells in G1 phase, and triggered apoptosis in glioma cells. Importantly, ZINC69391 dramatically affected cell migration and invasion in vitro, interfering with actin cytoskeleton dynamics. We also evaluated the effect of analog 1A-116, a compound derived from ZINC69391 structure. 1A-116 showed an improved antiproliferative and antiinvasive activity on glioma cells. These findings encourage further preclinical testing in clinically relevant animal models.

Self-assembly of 2,3-dihydroxycholestane steroids into supramolecular organogels as a soft template for the in-situ generation of silicate nanomaterials.

Supramolecular gels are an important and interesting class of soft materials that show great potential for many applications. Most of them have been discovered serendipitously, and understanding the supramolecular self-assembly that leads to the formation of the gel superstructure is the key to the directed design of new organogels. We report herein the organogelating property of four stereoisomers of the simple steroid 2,3-dihydroxycholestane. Only the isomer with the trans-diaxial hydroxy groups had the ability to gelate a broad variety of liquids and, thus, to be a super-organogelator for hydrocarbons. The scope of solvent gelation was analysed with regard to two solvent parameters, namely the Kamlet-Taft and the Hansen solubility parameters. The best correlation was observed with the Hansen approach that revealed the existence of two clear gelation zones. We propose a general model of self-assembly through multiple intermolecular hydrogen bonds between the 1,2-dihydroxy system, which is based on experimental data and computational simulations revealing the importance of the di-axial orientation of the hydroxy groups for the one-dimensional self-assembly. Under controlled conditions, the fibrillar superstructure of the organogel was successfully used as a template for the in-situ sol-gel polymerization of tetraethoxysilane and the further preparation of silica nanotubes. We propose that the driving forces for templating are hydrogen bonding and electrostatic interactions between the anionic silicate intermediate species and the self-assembled fibrillar network.