32 / 2021-07-27 09:15:04

Amber acid, a novel inhibitor of human ether-à-go-go-1 potassium channel, exerts anti-hepatocellular carcinoma activity

antitumor,Eag1,Inhibitor,amber acid,hepatocellular carcinoma

Hepatocellular carcinoma (HCC) is one of the most common malignancies and is a serious threat to people's health worldwide. However, there is no effective method for the treatment of hepatocellular carcinoma, and it is urgent to find new targets of anticancer drugs. The potassium channel ether à go-go-1 as a new anticancer target is rarely expressed in normal tissues and highly expressed in more than 70% of tumours. Eag1 is related with various physiological processes such as tumour proliferation, malignant transformation, invasion, metastasis, recurrence and prognosis. The specific blockade of human Eag1 (hEag1) may not only allow the dissection of the role of the channel in distinct physiologic processes, but because of the implication of hEag1 in tumor biology, it may also offer an opportunity for the treatment of HCC. Therefore, identifying potent and specific Eag1 channel inhibitors is crucial.

A perforated whole-cell patch technique was used to record the Eag1 currents. Homology modelling and a docking technique were used to investigate the interaction between amber acid and the Eag1 channel. CCK-8 assay, wound healing assay and transwell assay were used to investigate cell viability, cell migration and cell invasion, respectively. Tumour xenograft experiments were used to study tumour growth inhibition in vivo.

Amber acid directly inhibited intracellular Eag1 in a Ca²⁺- independent, specific and dose-dependent manner (IC₅₀ of 844.68±121.5 nM), and the amino acids Lys⁶⁵⁰, His ⁶⁸⁹, and Arg⁶⁹³ in the Eag1 CNBHD domain were critical for amber acid’s inhibitory effect. Additionally, and impotently, amber acid reduced the proliferation, malignant transformation and invasion of Hep G2 (a kind of hepatocellular carcinoma cells) and CHO cells stably expressing Eag1 by inhibiting the Eag1 channel activity. Finally, amber acid (5 mg/kg/d) inhibited tumour growth in mice, demonstrating a 51.46 % inhibitory rate of Hep G2 cell-transplanted tumours without side effects.

A high affinity binding site of amber acid is suggested on intracellular regions of hEag1 channels, leading to profound hEag1 channel inhibition with no effects on the other three potassium channels. Impotently, amber acid inhibits the growth of hepatocellular carcinoma efficiently and without side effects by inhibiting the activity of Eag1. Consequently, identification of amber acid as a novel inhibitor of Eag1 could open new possibilities for addressing problems of liver cancer.