Authors: Mühlebner A., Bongaarts A., Sarnat H.B., Scholl T., Aronica E.

Journal of Anatomy, 2019 March 22, doi: 10.1111/joa.12956

Abstract

In recent years the role of the mammalian target of rapamycin (mTOR) pathway has emerged as crucial for normal cortical development. Therefore, it is not surprising that aberrant activation of mTOR is associated with developmental malformations and epileptogenesis. A broad spectrum of malformations  of cortical development, such as focal cortical dysplasia (FCD) and tuberous sclerosis complex (TSC), have been linked to either germline or somatic mutations in mTOR pathway-related genes, commonly summarised under the umbrella-term 'mTORopathies'. However, there are still a number of unanswered questions regarding the involvement of mTOR in the pathophysiology of these abnormalities. Therefore, a monogentic disease, such as TSC, can be more easily applied as a model to study the mechanisms of epileptogenesis and identify potential new targets of therapy. Developmental neuropathology and genetics demonstrate that FCD IIb and hemimegalencephaly are the same diseases. Constitutive activation of mTOR signalling representa a shared pathogeneic mechanism in a group of developmental malformations that have histopathological and clinical features in common, such as epilepsy, autism and other comorbidities. We seek to understand the effect of mTOR dysregulation in a developing cortex with the propensity to generate seizures as well as the aftermath of the surrounding environment, including the white matter. 

Authors: Drion C.M., Kooijman L., Aronica E., van Vliet E.A., Wadman W.J., Chameau P., Gorter J.A.

Epilepsia 2019 Apr;60(4):605-614.

 

Abstract

Objective: Inhibition of the mammalian target of rapamycin (mTOR) pathway could be antiepileptogenic in temporal lobe epilepsy (TLE), possibly via anti-inflammatory actions. We studied effects of the mTOR inhibitor rapamycin and the anti-inflammatory compound curcumin - also reported to inhibit the mTOR pathway - on epileptogenesis and inflammation in an in vitro organotypic hippocampal-entorhinal cortex slice culture model. 

Methods: Brain slices containing hippocampus and entorhinal cortex were obtained from 6-day old rat pups and maintained in culture for up to 3 weeks. Rapamycin or curcumin was added to the culture medium from day 2 in vitro onward. Electrophysiological recordings revealed epileptiformlike activity that developed over 3 weeks.

Results: In week 3, spontaneous seizurelike events (SLEs) could be detected using whole cell recordings from CA1 principal neurons. The percentage of recorded CA1 neurons displaying SLEs was lower in curcumin-treated slice cultures compared to vehicle-treated slices (25.8% vs 72.5%), whereas rapamycin did not reduce SLE occurrence significantly (52%). Western blot for phosphorylated-S6 (pS6) and phosphorylated S6K confirmed that rapamycin inhibited the mTOR pathway, whereas curcumin only lowered pS6 expression at one phosphorylation site. Real-time quantitative polymerase chain reaction results indicated a trend toward lower expression of inflammatory markers IL-1B and IL-6 and transforming growth factor beta after 3 weeks of treatment with rapamycin and curcumin compared to vehicle. 

Significance: Our results show that curcumin suppresses SLEs in the combined hippocampal-entorhinal cortex slice culture model and suggest that its antiepileptogenic effects should be further investigated in experimental models of TLE. 

Authors: Garcia-Rincon D., Diaz-Alonso J., Paraiso-Luna J., Ortega Z., Aguareles J., de Salas-Quiroga A., Jou C., de Prada I., Martinez-Cerdeno V., Aronica E., Guzman M., Perez-Jimenes M.A., Galve-Roperh I. 

Frontiers in Pharmacology 9:1508. doi:10.3389/fphar.2018.01508

Abstract

Alterations of the PI3K/Akt/mammalian target of rapamycin complex 1 (mTORC1) signaling pathway are causally involved in a subset of malformations of cortical development (MCDs) ranging from cortical dysplasia (FCD) to hemimegalencephaly and megalencephaly. These MCDs represent a frequent cause of refractory pediatric epilepsy. The endocannabinoid system - especially cannabinoid CB1 receptor - exerts a neurodevelopmental regulatory role at least in part via activation of mTORC1 signaling. Therefore, we sought to characterize the possible contribution of the endocannabinoid system signaling to FCD. Confocal microscopy characterization of the CB1 receptor expression and mTORC1 activation was conducted in FCD Type II resection samples. FCD samples were subjected to single nucleotide polymorphism screening for endocannabinoid system elements, as well as CB1 receptor gene sequencing. Cannabinoid CB1 receptor levels were increased in FCD with overactive mTORC1 signaling. CB1 receptors were enriched in phospho-S6-positive cells including balloon cells (BCs) that co-express aberrant markers of undifferentiated cells and dysplastic neurons. Pharmacological regulation of CB1 receptors and the mTORC1 pathway was performed in fresh FCD-derived organotypic cultures. HU-210-evoked activation of CB1 receptors was unable to further activate mTORC1 signaling, whereas CB1 receptor blockade with rimonabant attenuated mTORC1 overactivation. Alterations of the endocannabinoid system may thus contribute to FCD pathological features, and the blockade of cannabinoid signaling might be a new therapeutic intervention in FCD. 

Authors: Urbanska M., Kazmierska-Grebowska P., Kowalczyk T., Caban B., Nader K., Pijet B., Kalita K., Gozdz A., Devijver H., Lechat B., Jaworski T., Grajkowska W., Sadowski K., Jozwiak S., Kotulska K., Konopacki J., Van Leuven F., van Vliet E.A., Aronica E., Jaworski J.

Ebiomedicine 39 (2019), 377-387

Abstract

Background: Glycogen synthase kinase-3beta (GSK3B) is a key regulator of cellular homeostasis. In neurons, GSK3B contributes to the control of neuronal transmission and plasticity, but its role in epilepsy remains to be defined. 

Methods: Biochemical and electrophysiological methods were used to assess the role of GSK3B in regulating neuronal transmission and epileptogenesis. GSK3B activity was increased genetically in GSK3B(S9A) mice. Its effects on neuronal transmission and epileptogenesis induced by kainic acid were assessed by field potential recordings in mice brain slices and video electroencephalography in vivo. The ion channel expression was measured in brain samples from mice and followed by analysis in samples from patients with temporal lobe epilepsy or focal cortical dysplasia in correlation to GSK3B phosphorylation. 

Findings: Higher GSK3B activity decreased the progression of kainic acid induced epileptogenesis. At the biochemical level, higher GSK3B activity increased the expression of hyperpolarization-activated cyclic nucleotide-gated (HCN) channel 4 under basal conditions and in the epileptic mouse brain and decreased phosphorylation of the glutamate alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor subunit GluA1 at Serine 831 under basal conditions. Moreover, we found a significant correlation between higher inhibitory GSK3B phorphorylation at Serine 9 and higher activitating GluA1 phosphorylation at Serine 845 in brain samples from epileptic patients.

Interpretation: Our data imply GSK3B activity in the protection of neuronal networks from hyper-activation in response to epileptogenic stimuli and indicate that the anti-epileptogenic function of GSK3B involves modulation of HCN4 level and the synaptic AMPA receptors pool. 

Authors: Krueger D.A., Capal J.K., Curatolo P., Devinsky O., Ess K., Tzadok M., Koenig M.K., Narayanan V., Ramos F., Jozwiak S., de Vries P., Jansen A.C., Wong M., Mowat D., Lawson J., Bruns S., Franz D.N., TSCure Research Group

Eur J Paediatr Neurol. 2018 Jul 4. pii:S1090-3798(17)31969-4. doi: 10.1016/j/ejpn.2018.06.007.

Abstract

OBJECTIVE: To evaluate the safety of mTOR inhibitors (sirolimus or everolimus) in infants and very young children with tuberoussclerosis complex (TSC) under two years of age.

METHODS: 

Study design was retrospective to capture medical record data from 52 international TSC Centres who initiated treatment with sirolimus or everolimus in TSC children before the age of two years. Data collection included demographic and clinical information including reason(s) for initiating treatment with mTOR inhibitors, treatment duration, dosing, and corresponding serum trough levels, response to treatment, and adverse events (AE).

RESULTS

19 of 52 (37%) TSC Centres reported treatment of at least one child with TSC under the age of two years with everolimus or sirolimus. Treatment-related data were provided for 45 patients meeting inclusion criteria. Everolimus was utilised 87% of the time, compared to 24% for sirolimus (5 subjects, 11%, were treated separately with both). Refractory epilepsy (45%) was the most common primary reason for initiating treatment and treatment was initiated on average at 11.6 ± 7.6 months of age. At least one AE, suspected or definitely treatment-related, occurred in 35 of 45 (78%) treated subjects. Most AEs were mild (Grade 1) or moderate (Grade 2) in severity and most commonly related to infections. Severe AE (Grade 3) was reported in 7 subjects (20%) and no life-threatening AE (Grade 4) or death/disability (Grade 5) was reported. Treatment was discontinued due to an AE in 9 of 45 (20%).

CONCLUSIONS

Everolimus, and to a lesser extent sirolimus, are increasingly being used to treat TSC infants and very young childrenfor multiple TSC-associated clinical indications. While AEs were common, most were not severe and did not prevent continued treatment in the majority of this younger population.