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

在人体上的研究是在06-M-0214，NCT00362843协议下进行的。 在2020年的资助期内，我们解决了以下问题：1)用L-1-C-11亮氨酸PET方法测量FXS参与者和清醒状态下的健康志愿者的RCP，2)其他神经发育障碍小鼠模型的蛋白质合成率，3)在一些神经发育障碍中mTORC 1活性作为大脑蛋白质合成的中枢调节因子，4)睡眠和神经发育障碍，5)磷酸二酯酶D抑制剂治疗Fmr1 KO小鼠对行为和RCP的影响。 1)脆性X综合征患者的RCPS测定。采用L-1-C-11亮氨酸PET法测定RCP。我们将努力集中在试图研究不使用镇静剂的脆性X综合征患者。我们使用我们修改的PET方案使一些Fragile X参与者能够清醒地进行研究。我们的初步结果表明，与健康对照组相比，清醒的脆性X参与者的大脑蛋白质合成率降低。我们已经完成了数据收集，正在分析结果。 2)其他几种神经发育障碍的遗传基础表明，翻译控制缺陷可能是核心表型。例如，结节性硬化症(TSC)是由TSC1或TSC2基因的杂合突变引起的，这两个基因都编码对mTOR活性具有负面调节作用的蛋白质。MTOR途径是调节蛋白质合成的节点之一。这种控制的中断被认为是在TSC中观察到的一些表型的基础。我们在TSC的小鼠模型TSC2+/-中测量了RCP，发现RCP在整个大脑中都减少了。我们继续在其他神经发育障碍的小鼠模型上进行RCPS的研究，如Shank3 KO小鼠。我们还在TSC2的条件基因敲除小鼠模型中测量RCP。这些研究正在实验室中进行。 3)据报道，在Fmr1KO小鼠中，mTOR通路过度激活。我们正在研究在Fmr1KO小鼠中mTORC1激活增加的后果。我们用mTORC1抑制剂雷帕霉素对Fmr1KO小鼠进行慢性治疗，以确定雷帕霉素治疗是否可以逆转行为表型。我们发现pS6在Fmr1KO小鼠中上调，并被雷帕霉素治疗后正常化，但除了引起焦虑的作用外，雷帕霉素并不逆转所研究的任何行为表型。此外，雷帕霉素治疗对对照组和Fmr1 KO小鼠的睡眠和社会行为都有不利影响。雷帕霉素治疗的Fmr1KO小鼠的RCPS研究正在进行中。我们正在体内测量RCP，并确定这些小鼠的行为表型。 4)睡眠和神经发育障碍。睡眠异常是被诊断为神经发育综合征的患者中最常见的并发障碍之一。在这些患者中，行为异常的严重程度与睡眠异常的严重程度是相关的。鉴于睡眠在发育可塑性中的重要性，我们继续研究睡眠在神经发育障碍中的作用。我们正在收集许多神经发育障碍小鼠模型的睡眠数据，包括Shank3 KO、催产素受体KO和TSC2+/-小鼠。此外，我们正在用催眠药治疗Fmr1 KO小鼠，以确定对睡眠和其他行为表型的影响。 5)对人类、小鼠和果蝇的研究表明，FXS中FMRP的缺失会影响cAMP水平。我们的研究调查了靶向cAMP水平可能在FXS中具有治疗价值的可能性。我们评估了长期应用磷酸二酯酶-4D负性变构调节剂BPN14770对Fmr1KO小鼠的治疗和神经化学效果。BPN14770治疗对Fmr1KO小鼠的行为表型影响有限。这两种基因都有一些影响，如睡眠时间延长和社会行为增加。对野战行为的影响在WT小鼠中更为显著。BPN14770对WT小鼠在最高剂量时有增加RCP的趋势，但对Fmr1 KO小鼠的作用不明显。结果表明，通过cAMP依赖的途径对翻译的调节存在基因型差异。

The work on humans was conducted under the protocol 06-M-0214, NCT00362843. During the 2020 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 in the awake state, 2) protein synthesis rates in mouse models of other neurodevelopmental disorders, 3) mTORC1 activity as a central regulator of brain protein synthesis in some neurodevelopmental disorders, 4) sleep and neurodevelopmental disorders, 5) 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. 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 PET protocol to enable some fragile X participants to undergo the study awake. Our preliminary results indicate that rates of cerebral protein synthesis in the awake fragile X participants are decreased compared to healthy controls. We have completed data collection and are analyzing results. 2) 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. We are also measuring rCPS in a conditional knockout mouse model of TSC2. These studies are ongoing in the laboratory. 3) 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 and determining behavioral phenotypes in these mice. 4) 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. 5) 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. Effects on open field behavior were more striking in WT mice. BPN14770 treatment in WT mice tended to increase rCPS at the highest dose, but BPN14770 effects were less apparent in Fmr1 KO mice. Results suggest a genotype difference in the regulation of translation via a cAMP-dependent pathway.