Dysregulation of Protein Synthesis in Fragile X Syndrome and Other Developmental Disorders

National Institute Of Mental Health

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Abstract

The work on humans was conducted under the protocol 06-M-0214, NCT00362843. During the 2021 funding period, we addressed the following: 1) rCPS measured with the L-1-C-11leucine PET method in participants with FXS and healthy volunteers studied under dexmedetomidine sedation, 2) rCPS measured with the L-1-C-11leucine PET method in participants with FXS and healthy volunteers studied in the awake state, 3) protein synthesis rates in mouse models of other neurodevelopmental disorders, 4) mTORC1 activity as a central regulator of brain protein synthesis in some neurodevelopmental disorders, 5) sleep and neurodevelopmental disorders, 6) effects of treatment of Fmr1 KO mice with a phosphodiesterase D inhibitor on behavior and rCPS. 1) Measurement of rCPS in humans with fragile X syndrome studied under sedation. All subjects were males between the ages of 18 and 24 years and free of psychotropic medication. As most fragile X participants were not able to undergo the PET study awake, we used dexmedetomidine for sedation during the imaging studies. We found no differences between rCPS measured during dexmedetomidine-sedation and the awake state in ten healthy controls. In the comparison of rCPS in dexmedetomidine-sedated fragile X participants (n=9) and healthy controls (n=14) we found no statistically significant differences. Our results from in vivo measurements in human brain do not support the hypothesis that rCPS are elevated due to the absence of FMRP. This hypothesis is based on findings in animal models and in vitro measurements in human peripheral cells. The absence of a translation suppressor (FMRP) may produce a more complex response in pathways regulating translation than previously thought. Results of this study were published in Neurobiology of Disease. 2) Measurement of rCPS in humans with fragile X syndrome. We used the L-1-C-11leucine PET method to measure rCPS. We have focused our efforts on trying to study individuals with fragile X syndrome without the use of sedatives. We used our modified and validated PET protocol to enable some fragile X participants to undergo the study awake. We compared rCPS in 10 fragile X participants and 16 age-matched controls all in the awake state with the same procedure. Contrary to the prevailing hypothesis and findings in Fmr1 knockout mice, results indicate that rCPS in awake fragile X subjects are decreased in whole brain and most brain regions by 13-21% compared to healthy controls. 3) The genetic bases of several other neurodevelopmental disorders suggest that defects in translational control may be a core phenotype. For example, tuberous sclerosis complex (TSC) is caused by heterozygous mutations in either the TSC1 or TSC2 gene, both of which encode proteins that are negative regulators of mTOR activity. The mTOR pathway is one of the nodes regulating protein synthesis. Disruption of this control is thought to underlie some of the phenotypes observed in TSC. We have measured rCPS in a mouse model of TSC, TSC2+/-, and found that rCPS are decreased throughout the brain. We continue studies of rCPS in mouse models of other neurodevelopmental disorders such as Shank3 KO mice. SHANK3 is a postsynaptic scaffolding protein that plays a critical role in synaptic development and brain function. Mutations in SHANK3 are implicated in Phelan-McDermid syndrome (PMS), a neurodevelopmental disorder characterized by autistic-like behavior, delayed speech, hypotonia, and intellectual disability (ID). Moreover, mutations in SHANK3 occur in 1-2% of cases of idiopathic autism spectrum disorder (ASD). In fragile X syndrome (FXS), a syndromic form of autism, SHANK3 is one of the 842 targets of fragile X mental retardation protein (FMRP), the protein product of the silenced FMR1 gene. FXS is likely a primary disorder of the regulation of translation, whereas other syndromic forms of ASD/ID, e.g., PMS, appear to be primary disorders of synaptic structure. In this study, we asked if a knockout of the synaptic protein, Shank3, is linked to an effect on translation. Specifically, we measured the effect of Shank3 loss on rates of cerebral protein synthesis (rCPS) in vivo by means of the L-1-14Cleucine quantitative autoradiographic method. We found that Shank3 knockout mice had significantly increased rCPS in every brain region examined. Our results suggest a link in ASD/ID between synaptic structure and regulation of translation. 4) It has been reported that the mTOR pathway is overactivated in Fmr1 KO mice. We are investigating the consequences of increased mTORC1 activation in Fmr1 KO mice. We treated Fmr1 KO mice chronically with an mTORC1 inhibitor, rapamycin, to determine if rapamycin treatment could reverse behavioral phenotypes. We found that pS6 was upregulated in Fmr1 KO mice and normalized by rapamycin treatment, but, except for an anxiogenic effect, rapamycin did not reverse any of the behavioral phenotypes examined. Moreover, rapamycin treatment had an adverse effect on sleep and social behavior in both control and Fmr1 KO mice. Studies of rCPS in rapamycin-treated Fmr1 KO mice are ongoing. We are measuring rCPS in vivo in these mice. 5) Sleep and neurodevelopmental disorders. Sleep abnormalities are one of the most prevalent concurrent disorders in patients diagnosed with neurodevelopmental syndromes. In these patients, the severity of behavioral abnormalities and the severity of sleep abnormalities are correlated. Given the importance of sleep in developmental plasticity, we continue studies on the role of sleep in neurodevelopmental disorders. We are collecting sleep data on numerous mouse models of neurodevelopmental disorders including Shank3 KO, Oxytocin receptor KO, and Tsc2+/- mice. Additionally, we are treating Fmr1 KO mice with hypnotics to determine the effects on sleep and other behavioral phenotypes. 6) Studies in humans, mice and Drosophila indicate that a deficit of FMRP in FXS influences cAMP levels. Our study investigated the possibility that targeting cAMP levels may have therapeutic value in FXS. We assessed the therapeutic and neurochemical effects of chronic administration of the phosphodiesterase-4D negative allosteric modulator, BPN14770, in Fmr1 KO mice. BPN14770 treatment had limited effects on the behavioral phenotype in Fmr1 KO mice. Some effects such as increased sleep duration and increased social behavior occurred in both genotypes. In the open field, the hyperactivity response in Fmr1 KO mice was ameliorated by BPN14770 treatment at low and intermediate doses. BPN14770 treatment tended to increase rCPS in a dose-dependent manner in WT mice, whereas in Fmr1 KO mice effects on rCPS were less apparent. Results indicate BPN14770 treatment improves some behavior in Fmr1 KO mice. Results also suggest a genotype difference in the regulation of translation via a cAMP-dependent pathway. Results of this study were published in Neurobiology of Disease.