Pre-clinical research for diseases caused by IM dysfunction. – IMprove

The IMprove project is novel because it builds on the results on a previous ANR project that generated the concepts and tools that were necessary to build the current translational project. We were able to produce unique models (a knock-in mouse and human neuronal cultures derived from patient's iPS) that are still largely missing in the field and provide us with a strong competitive advantage. We are now able to study pathogenic mechanisms occurring in the whole mouse brain and directly in human Kcnq2-deficient neuronal cultures. Our previous results revealed that neuronal hyperexcitability is transient, suggesting the gradual establishment of a compensation phenomenon whose origin will be studied in the current project. Our models also allow to assess treatment response and to translate our findings to the bedside. The combination of expertise provided by our consortium will allow us to tackle the multiple dimensions of the subject (from natural history in patients to the study of human neuronal cultures, from omics studies to integrated cognitive and motor phenotyping, from electrophysiological characterization to drug testing). We believe that the multidisciplinary nature of our proposal will scale up its translational potential and be a key success factor, each partner having demonstrated its ability to collaborate successfully and to produce high-level publications.

Partner 1 produced new models (KI mice and human neurons derived from iPS). He also produced omics data by performing RNA-seq and proteomic studies. The first treatments of the models have started. Partner 2 showed that mice carrying the p.T274M variant exhibit recorded epileptic seizures (mostly fatal), the occurrence of which decreases during development. These disorders are associated at the cellular level with a reduction in M ??current of approximately 30% and neuronal hyperexcitability. Partner 3 has shown that the T274M variant causes an anomaly of excitability in the interneurons of the central locomotion pacemaker in young animals. Partner 4 produced human neuronal stem cells from pluripotent cells in patients. Several lines have been obtained. Partner 5 created a national KCNQ2 patient registry. This register could be extended to European level in the coming months and will serve to better understand the natural history of the disease and to provide prospective monitoring of the patient population concerned. Communication actions have been carried out for patients.

Our perspective is as set in the intiial project and deadlines will be met.

PLoS Biol 2020;18(11):e3000738.
doi: 10.1371/journal.pbio.3000738.
The M-current works in tandem with the persistent sodium current to set the speed of locomotion.
Verneuil J, Brocard C, Trouplin V, Villard L, Peyronnet-Roux J, Brocard F.

Epilepsia 2020;61(5):868-878.
doi: 10.1111/epi.16494.
A knock-in mouse model for KCNQ2-related epileptic encephalopathy displays spontaneous generalized seizures and cognitive impairment.
Milh M, Roubertoux P, Biba N, Chavany J, Spiga Ghata A, Fulachier C, Collins SC, Wagner C, Roux JC, Yalcin B, Félix MS, Molinari F, Lenck-Santini PP, Villard L.

bioRxiv 2020.05.12.090464; doi: doi.org/10.1101/2020.05.12.090464
Time-limited alterations in cortical activity of a Knock-in mice model of KCNQ2-related Developmental and Epileptic Encephalopathy.
Najoua Biba, Hélène Becq, Marie Kurz, Emilie Pallesi, Laurent Villard, Mathieu Milh, Pierre-Pascal Lenck Santini, Laurent Aniksztejn.

The KCNQ2 gene encodes the Kv7.2 subunit of the potassium Kv7/M channel, known to control neuronal excitability in the brain and spinal chord via the M current (IM). Pathogenic variants in the KCNQ2 gene represent the major cause of early onset epileptic encephalopathy, leading to the concept of KCNQ2-related epileptic encephalopathies (KCNQ2-REE). The patients have a remarkably homogeneous phenotype at the beginning with frequent tonic seizures resulting in abnormal muscle contractions and apnoea in the first days after birth. 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. 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. To date none of the available anti-epileptic drugs has been demonstrated to be effective in KCNQ2-REE.
The IMprove project is novel because it builds on the results on a previous ANR project that generated the concepts and tools that were necessary to build the current translational project. We were able to produce unique models (a knock-in mouse and human neuronal cultures derived from patient's iPS) that are still largely missing in the field and provide us with a strong competitive advantage. We are now able to study pathogenic mechanisms occurring in the whole mouse brain and directly in human Kcnq2-deficient neuronal cultures. Our previous results revealed that neuronal hyperexcitability is transient, suggesting the gradual establishment of a compensation phenomenon whose origin will be studied in the current project. Our models also allow to assess treatment response and to translate our findings to the bedside. The combination of expertise provided by our consortium will allow us to tackle the multiple dimensions of the subject (from natural history in patients to the study of human neuronal cultures, from omics studies to integrated cognitive and motor phenotyping, from electrophysiological characterization to drug testing). We believe that the multidisciplinary nature of our proposal will scale up its translational potential and be a key success factor, each partner having demonstrated its ability to collaborate successfully and to produce high-level publications.
The strongest impact of such a project will be to propose new therapeutic approaches for KCNQ2-REE. Early onset epileptic encephalopathies are very severe diseases, without satisfactory therapeutic options. We believe that one way to move forward is such a pre-clinical project for diseases where clinical research is difficult (small number of patients, unethical placebo, late developmental endpoint).
Our project has integrated an innovative dissemination activity with a specific action to design and organize practical trainings related to genetic epilepsies for relatives of children concerned by this pathology, through a three-day experimental course. IMprove is strongly supported by KCNQ2-France association, which brings together a large proportion of families affected by KCNQ2-REE.