Article

Functional characterization and in vitro pharmacological rescue of KCNQ2 pore mutations associated with epileptic encephalopathy

Gui-mei Yang1,2, Fu-yun Tian2,3, Yan-wen Shen4,5, Chuan-yan Yang2,6, Hui Yuan1,2, Ping Li1,2,3,7, Zhao-bing Gao1,2,3,7
1 School of Pharmacy, Zunyi Medical University, Zunyi 563000, China
2 Zhongshan Institute for Drug Discovery, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Zhongshan 528400, China
3 Center for Neurological and Psychiatric Research and Drug Discovery, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China
4 Department of Pediatrics, The First Medical Center of PLA General Hospital, Beijing 100853, China
5 Department of Pediatric neurology, Children’s Hospital of Fudan university at Xiamen, Xiamen 361006, China
6 School of Basic Medicine and Clinical Pharmacy, China Pharmaceutical University, Nanjing 211198, China
7 School of Pharmaceutical Sciences, Southern Medical University, Guangzhou 510515, China
Correspondence to: Fu-yun Tian: tianfuyun@zidd.ac.cn, Ping Li: lipingt@simm.ac.cn, Zhao-bing Gao: zbgao@simm.ac.cn,
DOI: 10.1038/s41401-023-01073-y
Received: 10 November 2022
Accepted: 26 February 2023
Advance online: 17 March 2023

Abstract

Mutations in the KCNQ2 gene encoding KV7.2 subunit that mediates neuronal M-current cause a severe form of developmental and epileptic encephalopathy (DEE). Electrophysiological evaluation of KCNQ2 mutations has been proved clinically useful in improving outcome prediction and choosing rational anti-seizure medications (ASMs). In this study we described the clinical characteristics, electrophysiological phenotypes and the in vitro response to KCNQ openers of five KCNQ2 pore mutations (V250A, N258Y, H260P, A265T and G290S) from seven patients diagnosed with KCNQ2-DEE. The KCNQ2 variants were transfected into Chinese hamster ovary (CHO) cells alone, in combination with KCNQ3 (1:1) or with wild-type KCNQ2 (KCNQ2-WT) and KCNQ3 in a ratio of 1:1:2, respectively. Their expression and electrophysiological function were assessed. When transfected alone or in combination with KCNQ3, none of these mutations affected the membrane expression of KCNQ2, but most failed to induce a potassium current except A265T, in which trace currents were observed when co-transfected with KCNQ3. When co-expressed with KCNQ2-WT and KCNQ3 (1:1:2), the currents at 0 mV of these mutations were decreased by 30%-70% compared to the KCNQ2/3 channel, which could be significantly rescued by applying KCNQ openers including the approved antiepileptic drug retigabine (RTG, 10 μM), as well as two candidates subjected to clinical trials, pynegabine (HN37, 1 μM) and XEN1101 (1 μM). These newly identified pathologic variants enrich the KCNQ2-DEE mutation hotspots in the pore-forming domain. This electrophysiological study provides a rational basis for personalized therapy with KCNQ openers in DEE patients carrying loss-of-function (LOF) mutations in KCNQ2.
Keywords: developmental and epileptic encephalopathy (DEE); KCNQ2 pore mutations; retigabine (RTG); pynegabine (HN37); XEN1101; electrophysiology

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