Develop a human liver system to study SLC25A13 mutations in citrin deficiency

项目总结 Citrin缺乏症(CD)是一种由SLC25A13基因突变引起的常染色体隐性遗传病，是一种泛种族遗传病 罕见的疾病。目前还没有针对CD的特效药。饮食管理被认为是最有效的 CD的治疗以及对患者临床结果的决定性因素。然而，饮食管理 只是基于小病例报告，患有CD的儿童的饮食管理普遍很差， 尤其是在碳水化合物饮食占主导地位的地区。因此，还需要进一步的研究来 了解疾病的发病机制，促进更具体的治疗发展。 CD表现为年龄相关的表型，可能会有严重的临床结果，包括无法茁壮成长 和突然由柠檬酸缺乏引起的血脂异常(FTTDCD)和瓜氨酸血症2型(CTLN2) 发展为高氨性脑病，以及其他临床表现。然而， SLC25A13功能丧失导致上述疾病表型的机制仍然存在 很大程度上还不清楚。CD的发病年龄和不完全外显是如何表现出来的，目前仍知之甚少 病人是有决心的。 CD研究中一个尚未得到满足的主要需求是缺乏一个可靠的与人类相关的模型来模拟整个CD 这种疾病在人类中的谱系。到目前为止，唯一可用的研究CD的模型是基因敲除(KO)小鼠 (即Slc25a13-KO和SLc25a13和Gpd2-Double KO)。由于物种间显著的基因差异 调节，特别是在糖酵解，这些模型的实用性和临床相关性是有争议的。需要 观察到发病年龄不同且不完整，进一步强调了与人类有关的系统 在怀疑有其他环境和/或遗传触发因素的CTLN2患者中可见外显性。 这项应用的主要研究目标是利用人类诱导的多能干细胞(IPSC)- 我们实验室建立了衍生的多细胞体外肝脏模型，以建立与人类相关的模型 研究了光盘中的SLC25A13。跨越这两个具体目标，我们计划产生含有野生型的肝脏培养物 或SLC25A13 KO细胞，并鉴定与人类CD相关的可能的疾病表型。 用于SLC25A13研究的IPSC来源的可再生和可遗传操作的人体系统的建立 有助于对疾病发病机制的详细剖析。此外，鉴于缺乏合适的人- 相关系统研究光盘，我们的iPSC衍生平台将提供一个迫切需要的新平台来测试 干预措施，促进特定疗法的发展。最后，这一初步支持将使我们能够 执行试行分析和表征，并为更大规模的拨款申请获取关键的初步数据 这将促进这些努力。

PROJECT SUMMARY Citrin deficiency (CD), an autosomal recessive disorder caused by mutations in SLC25A13 gene, is a pan-ethnic rare disease. Currently there is no specific drug for CD. Dietary management is regarded as the most effective treatment for CD, as well as a decisive factor in the clinical outcome of patients. However, dietary management has only been based on small case reports and dietary management for children with CD is generally poor, especially in areas where carbohydrate-based diets are predominant. Thus, further research is needed to understand disease pathogenesis and facilitate more specific therapeutic development. CD manifests as age-dependent phenotypes and can have severe clinical outcome including failure to thrive and dyslipidemia caused by citrin deficiency (FTTDCD) and citrullinemia type 2 (CTLN2) with a sudden development of hyperammonemic encephalopathy, as well as other clinical manifestations. However, the mechanisms by which how SLC25A13 loss-of-function leads to the abovementioned disease phenotypes remain largely unclear. It also remains poorly understood how the age of onset and incomplete penetrance seen in CD patients are determined. A major unmet need in the study of CD is the lack of a reliable human-relevant model that mimics the entire spectrum of this disease in humans. So far, the only available models to study CD are the knockout (KO) mice (i.e., Slc25a13-KO and Slc25a13 and Gpd2-double KO). Owing to marked species-specific differences in gene regulation especially in glycolysis, the utility and clinical relevance of these models are debated. The need for human-relevant system is further underscored by the observations showing variable age of onset and incomplete penetrance seen in CTLN2 patients where additional environmental and/or genetic triggers are suspected. The primary research goal of this application is to leverage a human induced pluripotent stem cell (iPSC)- derived multicellular in vitro liver model developed in our laboratory to create a human-relevant model for studying SLC25A13 in CD. Across the two specific aims, we plan to generate liver cultures harboring wild-type or SLC25A13 KO cells and to characterize the possible disease phenotypes that are relevant to human CD. Developing an iPSC-derived renewable and genetically manipulatable human system for the study of SLC25A13 in CD will facilitate the detailed dissection of disease pathogenesis. Moreover, given the lack of suitable human- relevant systems to study CD, our iPSC-derived platform will provide an urgently needed new platform to test interventions and promote the development of specific therapeutics. Finally, this initial support will allow us to perform pilot analysis and characterization and to acquire critical preliminary data for a bigger grant application that will facilitate these efforts.