Cognitive impairments are a major clinical feature of the common neurogenetic disease neurofibromatosis type 1 (NF1). Previous studies have demonstrated that increased neuronal inhibition underlies the learning deficits in NF1, however, the molecular mechanism underlying this cell-type specificity has remained unknown. Here, we identify an interneuron-specific attenuation of hyperpolarization-activated cyclic nucleotide-gated (HCN) current as the cause for increased inhibition in Nf1 mutants. Mechanistically, we demonstrate that HCN1 is a novel NF1-interacting protein for which loss of NF1 results in a concomitant increase of interneuron excitability. Furthermore, the HCN channel agonist lamotrigine rescued the electrophysiological and cognitive deficits in two independent Nf1 mouse models, thereby establishing the importance of HCN channel dysfunction in NF1. Together, our results provide detailed mechanistic insights into the pathophysiology of NF1-associated cognitive defects, and identify a novel target for clinical drug development.

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ObjectiveTo investigate whether mammalian target of rapamycin inhibitor everolimus can improve intellectual disability, autism, and other neuropsychological deficits in children with tuberous sclerosis complex (TSC).MethodsIn this 12-month, randomized, double-blind, placebo-controlled trial, we attempted to enroll 60 children with TSC and IQ <80, learning disability, special schooling, or autism, aged 4–17 years, without intractable seizures to be assigned to receive everolimus or placebo. Everolimus was titrated to blood trough levels of 5–10 ng/mL. Primary outcome was full-scale IQ; secondary outcomes included autism, neuropsychological functioning, and behavioral problems.ResultsThirty-two children with TSC were randomized. Intention-to-treat analysis showed no benefit of everolimus on full-scale IQ (treatment effect −5.6 IQ points, 95% confidence interval −12.3 to 1.0). No effect was found on secondary outcomes, including autism and neuropsychological functioning, and questionnaires examining behavioral problems, social functioning, communication skills, executive functioning, sleep, quality of life, and sensory processing. All patients had adverse events. Two patients on everolimus and 2 patients on placebo discontinued treatment due to adverse events.ConclusionsEverolimus did not improve cognitive functioning, autism, or neuropsychological deficits in children with TSC. The use of everolimus in children with TSC with the aim of improving cognitive function and behavior should not be encouraged in this age group.Clinicaltrials.gov identifierNCT01730209.Classification of evidenceThis study provides Class I evidence that for children with TSC, everolimus does not improve intellectual disability, autism, behavioral problems, or other neuropsychological deficits.

Neurofibromatosis type 1 was the first RASopathy and is now one of many RASopathies that are caused by germline mutations in genes that encode components of the Ras/mitogen-activated protein kinase (MAPK) pathway. Their common underlying pathogenetic etiology causes significant overlap in phenotypic features which includes craniofacial dysmorphology, cardiac, cutaneous, musculoskeletal, GI and ocular abnormalities, and a predisposition to cancer. The proceedings from the symposium “Recent Developments in Neurofibromatoses and RASopathies: Management, Diagnosis and Current and Future Therapeutic Avenues” chronicle this timely and topical clinical translational research symposium. The overarching goal was to bring together clinicians, basic scientists, physician-scientists, advocate leaders, trainees, students and individuals with Ras pathway syndromes to discuss the most state-of-the-art basic science and clinical issues in an effort to spark collaborations directed towards the best practices and therapies for individuals with RASopathies.

Neurofibromatosis type 1 (NF1) is an autosomal dominantly inherited disease, characterized by various neurocutaneous symptoms, cognitive impairments and problems in fine and gross motor performance. Although cognitive deficits in NF1 have been attributed to increased release of the inhibitory neurotransmitter γ -amino butyric acid (GABA) in the hippocampus, the origin of the motor deficits is unknown. Cerebellar Purkinje cells, the sole output neurons of the cerebellar cortex, are GABAergic neurons and express neurofibromin at high levels, suggesting an important role for the cerebellum in the observed motor deficits in NF1. To test this, we determined the cerebellar contribution to motor problems in Nf1 +/− mice, a validated mouse model for NF1. Using the Rotarod, a non-specific motor performance test, we confirmed that, like NF1 patients, Nf1 +/− mice have motor deficits. Next, to evaluate the role of the cerebellum in these deficits, mice were subjected to cerebellum-specific motor performance and learning tests. Nf1 +/− mice showed no impairment on the Erasmus ladder, as step time and number of missteps were not different. Furthermore, when compensatory eye movements were tested, no performance deficits were found in the optokinetic reflex and vestibulo-ocular reflex in the dark (VOR) or in the light (VVOR). Finally, Nf1 +/− mice successfully completed short-and longterm VOR adaptation paradigms, tests that both depend on cerebellar function. Thus, despite the confirmed presence of motor performance problems in Nf1 +/− mice, we found no indication of a cerebellar component. These results, combined with recent clinical data, suggest that cerebellar function is not overtly affected in NF1 patients.

To assess emotional and behavioral problems in children and adolescents with neurofibromatosis type 1,parents of 183 individuals aged 10.8 ± 3.1 years (range 6-17) completed the Child Behavior Checklist (CBCL). Also, 173 teachers completed the Teacher's Report Form (TRF), and 88 adolescents (children from 11 to 17 years) completed the Youth Self-Report (YSR). According to parental ratings, 32% scored in the clinical range (above the 90th percentile). This percentage was much lower when rated by teachers or adolescents themselves. Scores from all informants on scales for Somatic complaints, Social problems, and Attention problems were significantly different from normative scores. Attentional problems were associated with lower verbal IQ, male gender, younger age, and ADHD-symptoms. Disease-related factors did not predict behavioral problems scores. Substantial emotional and behavioral problems were reported by parents, teachers, and to a lesser extent by adolescents with NF1 themselves. Possibly, a positive illusory bias affects the observation of behavioral problems by adolescents with NF1.

Children with NF1 show distinct deficits in multiple domains. Many outcome measures showed weak test-retest correlations over the 1-year trial period. Cognitive and behavioral outcomes are complementary. This analysis demonstrates the need to include reliable outcome measures on a variety of cognitive and behavioral domains in clinical trials for NF1.

RASopathies, characterized by germline mutations in genes encoding proteins of the RAS-ERK signaling pathway, show overlapping phenotypes, which manifest themselves with a varying severity of intellectual disability. However, it is unclear to what extent they share the same downstream pathophysiology that underlies the cognitive deficits. Costello syndrome (CS) is a rare RASopathy caused by activating mutations in the HRAS gene. Here we investigated the mechanisms underlying the cognitive deficits of HRas G12V/G12V mice. HRas G12V/G12V mice showed robust upregulation of ERK signaling, neuronal hypertrophy, increased brain volume, spatial learning deficits, and impaired mGluR-dependent long-term depression (LTD). In contrast, long-term potentiation (LTP), which is affected in other RASopathy mouse models was unaffected. Treatment with lovastatin, a HMG-CoA-Reductase inhibitor which has been shown to rescue the behavioral phenotypes of mouse models of NF1 and Noonan syndrome, was unable to restore ERK signaling and the cognitive deficits of HRas G12V/G12V mice. Administration of a potent mitogen-activated protein kinase (MEK) inhibitor rescued the ERK upregulation and the mGluR-LTD deficit of HRas G12V/G12V mice, but failed to rescue the cognitive deficits. Taken together, this study indicates that the fundamental molecular and cellular mechanisms underlying the cognitive aspects of different RASopathies are remarkably distinct, and may require disease specific treatments.