Inhibition of Glyoxalase-I Leads to Reduced Proliferation, Migration and Colony Formation, and Enhanced Susceptibility to Sorafenib in Hepatocellular Carcinoma.

Maurice Michel,Cristina Ripoll,Alexander Zipprich,Marcus Hollenbach,Sabine Pohl

doi:10.3389/fonc.2019.00785

Abstract

Background: Glyoxalase-I (Glo-I) is essential for detoxification of methylglyoxal (MGO), a byproduct of glycolysis. Overexpression of Glo-I has been linked to multi-drug resistance in cancer therapy. The aim of this study was to analyze Glo-I in hepatocellular carcinoma (HCC) and the effect of the multi-tyrosine kinase inhibitor sorafenib on Glo-I.Methods: Expression and specific activity of Glo-I was measured in human HCC samples, HCC-cell lines (HepG2, Huh7) and a hepatocyte cell line (AML 12). Cells were either treated with Glo-I inhibitors, ethyl pyruvate (EP, 1–20 mM) and BrBzGSHCp2 (1–10 μM), or sorafenib (2.5–10 μM) and protein expression (Western Blot), proliferation (WST-assay), migration (scratch assay), and colony formation (clonogenic assay) were assessed.Results: High expression of Glo-I was detected in human HCC tissue samples. Huh7 showed highest expression and activity of Glo-I and revealed highest proliferation compared to AML 12 and HepG2. Targeting Glo-I by EP or BrBzGSHCp2 led to significantly reduced proliferation (20 mM EP 24 h: 57 ± 12%), migration and colony formation. Glo-I inhibition by 20 mM EP resulted in reduced expression of PDGFR-β (18 ± 10%), VEGFR2 (46 ± 11%), VEGF (61 ± 10%), pERK/ERK (62 ± 6%), NF-κB (44 ± 12%) as well as stimulation of Nrf2 (243 ± 36%). Similar results were seen with BrBzGSHCp2. Sorafenib treatment revealed elevation of Glo-I (10 μM: 209 ± 25%) and MGO. Co-treatment of EP and sorafenib led to an additional reduction of proliferation compared to sorafenib alone.Conclusion: Glo-I is positively correlated with HCC proliferation. Inhibition of Glo-I reduced proliferation, migration, and colony formation. In turn, sorafenib increases Glo-I. Co-treatment using Glo-I inhibitors could enhance susceptibility of HCC to sorafenib.

Highlights

Hepatocellular carcinoma (HCC) is the most frequent primary liver cancer and is ranked as the sixth most common neoplasm and the third leading cause of cancer-related death worldwide [1]
Glo-I Is Highly Expressed in Human HCC Tissue
Proliferation was significantly reduced upon incubation of Huh7 cells with BrBzGSHCp2 for 6 and 24 h, respectively (24 h 10 μM: 32 ± 2%, control: 100 ± 10%, p < 0.001, Figure 4B)

Summary

Introduction

Hepatocellular carcinoma (HCC) is the most frequent primary liver cancer and is ranked as the sixth most common neoplasm and the third leading cause of cancer-related death worldwide [1]. The most frequent used drug, the multi-tyrosine kinase inhibitor sorafenib, has shown beneficial effects in clinical trials of advanced-stage HCC with a median increase in survival of 2.8 months [3]. High glycolytic activity yields toxic by-products, such as the dicarbonyl compound methylglyoxal (MGO). AGEs bind to their receptor, RAGE, and activate intracellular signaling pathways [8]. This so-called “dicarbonyl stress” results in oxidative stress and was shown to be implicated in carcinogenesis [9, 10]. The aim of this study was to analyze Glo-I in hepatocellular carcinoma (HCC) and the effect of the multi-tyrosine kinase inhibitor sorafenib on Glo-I

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Methods

Results

Conclusion