LAURÉAT 2016 & 2019 

Dr. Laurent VILLARD

Le Docteur Laurent VILLARD est Directeur de recherche à l'INSERM UMR-S 1251 - MMG à la Faculté de Médecine de la Timone à Marseille et responsable du diagnostic moléculaire des épilepsies au Département de génétique médicale du Laboratoire de génétique moléculaire, CHU de Marseille - Hôpital de la Timone. 

THÈME DE RECHERCHE :

Integrated omics to understand KCNQ2-Related epileptic encephalopathy. This project is the follow-up of a project funded by the FONDATION JED in 2016.

SUBVENTIONS JED : 65.000 €

OBJECTIF DU PROJET : 

Kcnq2-related epileptic encephalopathy (Kcnq2-REE) is the most severe epilepsy phenotype affecting human neonates. It causes a severe neurological dysfunction. A significant proportion of cases has a genetic origin and the most frequently mutated gene is KCNQ2, encoding Kv7.2, a voltage-dependent potassium channel subunit. The current knowledge on Kcnq2 function and dysfunction has largely been obtained in heterologous setups (e.g. transfected cells, transient transfections in vivo) or mice harbouring genetic variants never found in epileptic encephalopathy patients.

To better study the pathophysiology of Kcnq2-REE and obtain relevant models, we have engineered a knock-in mouse model carrying the p.(Thr274Met) variant identified in a several patients. Heterozygous knock-in animals display generalized spontaneous seizures culminating between P20 and P30. 30% die before 3 months of age and behavioral characterization reveals important cognitive deficits in adult animals but no gross abnormality during early neurosensory development. In addition, we have obtained induced pluripotent stem cells (iPS) cell lines for two patients and are now able to produce glutamatergic and gabaergic human neurons.

We ought to identify relevant transcriptional/proteic changes in order to better understand pathogenic changes during the disease process. We will take advantage of the new models engineered by our group to perform RNA-sequencing and proteomics experiments in mouse and human cells followed by integration of omics data. We believe that such a strategy will allow us to progress toward a better understanding of this devastating and currently intractable epileptic phenotype and also to identify relevant targets amenable to pharmacological intervention.

Pioneer cortical electrical activity emerges from the third trimester of pregnancy and plays an important role in brain development. Early Onset Epileptic Encephalopathies (EOEE) are rare and intractable devastating epileptic syndromes beginning as soon as network activities emerge and characterized by early life seizures associated with a rapid deterioration of motor, cognitive and behavioural skills.

There is a genetic basis for EOEE. We and other laboratories have identified de novo pathogenic variants in the KCNQ2 gene encoding the Kv7.2 subunit of the potassium Kv7/M channel, a channel known to control neuronal excitability in the brain and spinal cord via the M current (IM). Each subunit consists of six transmembrane segments (S) with a voltage-sensor (S1-S4) and a pore domain (S5-P- S6). These subunits are specifically distributed at axon initial segments and nodes of Ranvier where they are co-clustered with sodium (Nav) channels but are also found in the somatodendritic compartment. Pathogenic variants in the KCNQ2 gene represent the major cause of EOEE and the term KCNQ2- related-epileptic encephalopathy (KCNQ2-REE) is now used to define this condition (Weckhuysen et al. 2012; Saitsu et al. 2012; Milh et al. 2013; Milh et al. 2015).

KCNQ2-REE patients have a remarkably homogeneous phenotype at the beginning. Epilepsy emerges during the first week of life, with frequent tonic seizures resulting in abnormal muscle contractions and apnoea. Electro-encephalogram (EEG) often shows a pattern called "suppression-burst" i.e. paroxysmal bursts of activity interspersed with periods of electrical silence. This is a highly recognizable EEG pattern, reflecting the profound and permanent alteration of brain function. This stormy phase of tonic seizures and abnormal EEG pattern lasts 2 to 15 weeks and usually gives place to a calmer period of rare seizures and significant amelioration of the EEG. Despite this apparent positive evolution in terms of seizures, the developmental process is definitively altered and leads to a severe and global neurological impairment. The vast majority of patients have no informative language, autistic behaviour, and significant motor impairment such as tetraplegia, spasticity, ataxia, global hypotonia or dystonia (Milh et al. 2013).

The management of KCNQ2-REE patients is complex because it must take into account multiple disabilities: motor, cognitive and epileptic. Their vulnerability is indeed major and multifactorial: neurological (via tonic seizures that can lead to status epilepticus and death), respiratory, digestive and orthopaedic, due to permanent motor disability.

In recent years, significant progress has been made regarding this condition: the involvement of KCNQ2 was described in 2012, hundreds of mutations have been described, the modes of transmissions have been specified, the initial phenotype, the natural history have also been studied. Electrophysiological studies have documented the impact of KCNQ2 mutations on the IM current in heterologous systems.

Nevertheless, no therapeutic option is available at present: retigabine, an IM agonist that corrects the deleterious effects of most of mutations in vitro, is ineffective in Humans; carbamazepine, which blocks the sodium channels, has a certain effectiveness on seizures but does not alter the cognitive and motor dramatic fate of patients.

The ambition of this proposal is to increase the knowledge of KCNQ2-related epileptic encephalopathies at the molecular level, using integrated omics in new models generated by our laboratory. It also aims to identify targets amenable to pharmacological intervention.