EACR25-0984

Inhibition of the extracellular signal-regulated kinase 5 (ERK5) increases EGFR expression and its nuclear localization in HCC, generating new targeting opportunities

A. Menconi¹, I. Tusa¹, F. Marra², E. Rovida¹

¹University of Florence, Departiment of Experimental and Clinical Biomedical Sciences “Mario Serio”, Florence, Italy
²University of Florence, Department of Experimental and Clinical Medicine, Florence, Italy

Introduction:

Hepatocellular Carcinoma (HCC) is the most common form of liver cancer and accounts for 90% of cases. Nonetheless, available molecular-targeted therapies are still unsatisfactory, so additional therapeutic targets must be identified. The extracellular signal-regulated kinase 5 (ERK5) is a member of the Mitogen-Activated Protein Kinases (MAPK) family, and is highly expressed in hepatocytes, and its gene has been reported to be amplified in HCC. Moreover, we recently reported that ERK5 regulates the development and growth of HCC, so that ERK5 targeting appears as a promising approach for the treatment of this type of cancer.

Material and method:

The aim of this study was to investigate the interplay between ERK5 and EGFR using two hepatocellular carcinoma cell lines, Huh7 and HepG2. Gene silencing was performed with short harpin RNA for ERK5. For the pharmacological treatments, ERK5 (JWG-071), EGFR (Gefitinib) α/β importin (Ivermectin) inhibitors effects were evaluated after 72h of treatment in term of cell viability using MTT assay. The mRNA expression was analyzed using quantitative RT-PCR. Protein expression levels and intracellular localization were detected by Western blot and immunofluorescence in confocal microscopy.

Result and discussion:

We found that ERK5 knock down (KD) increases EGFR mRNA and protein levels, and downstream PI3K/AKT activation. Furthermore, the amount of EGFR protein was increased in cells treated with the ERK5 inhibitor, JWG-071, compared to control cells. The combination of JWG-071 and the EGFR inhibitor Gefitinib resulted to be more effective than single treatments in reducing cell viability and 3D spheroid growth in both cell lines. In addition, nuclear EGFR was higher in ERK5-KD cells compared to controls. Since EGFR shuttles to the nucleus via the α/β importin system, we performed combined treatment of JWG-071 and the α/β importin inhibitor, Ivermectin. This cotreatment showed a higher antiproliferative effect than single drugs on HCC cell lines.

Conclusion:

In conclusion, these results revealed new potential therapeutic strategies to explore for the treatment of HCC.