Molecular Etiology of Enchondromatosis

摘要 超过3%的人口患有内生软骨瘤（ECA），这是一种由细胞组成的骨骼良性肿瘤 来自生长板的可引起疼痛、畸形并可导致病理性骨折。 内生软骨瘤可发展为恶性软骨肉瘤（CSA）。异柠檬酸编码基因突变 脱氢酶（IDH 1和2）在大比例的ECA和CSA中被鉴定。在我们之前的工作中，我们 发现IDH突变抑制生长板软骨细胞分化，并且软骨细胞特异性条件性 Idh 1突变小鼠发生ECA。突变IDH独特地产生代谢物2-羟基戊二酸（2-HG），但 我们和其他人发现阻断2-HG β的产生并不改变CSA细胞的生存力。 虽然2-HG具有在肿瘤起始中可能重要的表观遗传效应，但肿瘤维持必须依赖于 其他因素由于IDH在代谢中起着重要作用，因此相关的代谢变化可以驱动 观察到的表型。我们在表达突变IDH的细胞中发现了高水平的糖原。糖原也是 在生长板的增殖和前肥大细胞中发现。在我们以前的工作中，我们发现， 胞内胆固醇的生物合成在IDH突变软骨细胞中被激活，并且它也在 生长板，其活性与糖原水平相关。这就提出了细胞内 胆固醇生物合成，由固醇调节元件结合蛋白（SREBP）激活 转录，也调节糖原。我们的前提是，糖原是一个重要的能量来源， 肥大和肥大生长板软骨细胞和糖原储存是维持 ECA和CSA的肿瘤表型。我们还提出，糖原消耗可以抑制肿瘤细胞的生长， 表型在这个建议中，我们将研究什么调节糖原在生长板，ECA和CSA， 确定糖原在这些生长板和肿瘤软骨细胞中的功能。 为了确定是什么调节生长板、ECA和CSA中的糖原，我们优先考虑了已知的基因， 调节在生长板中和通过IDH突变差异调节的糖原。蛋白 磷酸酶1调节亚基3C（PPP 1 R3 C）是这样一种基因， 在其启动子区含有SREBP结合位点。我们的初步数据表明，SREBP调节 PPP 1 R3 C，然后调节糖原。我们的研究将使用来自人类肿瘤的细胞系， 改良小鼠，发展内生软骨瘤，以确定糖原和PPP 1 R3 C在生长中的功能， 板、ECA和CSA。此外，我们将研究SREBP如何调节PPP 1 R3 C和糖原。糖原 合成酶基因将被删除，或者我们将细胞与药物抑制糖原合成和分解。 这些数据将提供临床前信息，作为ECA和CSA新疗法的基础。

Abstract More than 3% of the population develops an enchondroma (ECA), a benign tumor in bone composed of cells derived from the growth plate that can cause pain, deformity, and can be responsible for pathologic fractures. Enchondromas can progress to malignant chondrosarcoma (CSA). Mutations in genes encoding isocitrate dehydrogenase (IDH1 and 2) were identified in a large proportion of ECAs and CSAs. In our prior work, we found that IDH mutations inhibit growth plate chondrocyte differentiation, and chondrocyte-specific conditional Idh1 mutant mice develop ECAs. Mutant IDH uniquely produces the metabolite 2-hydroxyglutarate (2-HG), but we and others found that blocking the production of 2-HG pharmacologically does not alter CSA cell viability. While 2-HG has epigenetic effects that are likely important in tumor initiation, tumor maintenance must rely on other factors. Since IDH plays an important role in in metabolism, associated metabolic changes could drive the observed phenotype. We found high levels of glycogen in cells expressing a mutant IDH. Glycogen is also found in proliferating and pre-hypertrophic cells of the growth plate. In our previous work, we found that intracellular cholesterol biosynthesis was activated in IDH mutant chondrocytes and that it is also regulated in the growth plate, and its activity corelates with glycogen levels. This raises the possibility that intracellular cholesterol biosynthesis, which is activated by Sterol regulatory-element binding proteins (SREBP) transcription, also regulates glycogen. Our premise is that glycogen is an important energy source for pre- hypertrophic and hypertrophic growth plate chondrocytes and that glycogen stores are required to maintain the neoplastic phenotype in ECA and CSA. We also propose that glycogen depletion can suppress the neoplastic phenotype. In this proposal we will study what regulates glycogen in the growth plate, ECA and CSA, and determine the function of glycogen in these growth plate and neoplastic chondrocytes. To determine what regulates glycogen in the growth plate, ECA, and CSA, we prioritized genes known to regulate glycogen that were differentially regulated in the growth plate and by IDH mutations. Protein phosphatase 1 regulatory subunit 3C (PPP1R3C) is one such gene which is differentially and interestingly, contains SREBP binding sites in its promoter region. Our preliminary data suggest that SREBP regulates PPP1R3C which then regulates glycogen. Our studies will use cell lines from human tumors and genetically modified mice that develop enchondromas to define the function of glycogen and PPP1R3C in the growth plate, ECA, and CSA. In addition, we will study how SREBP regulates PPP1R3C and glycogen. Glycogen synthase will be deleted genetically, or we will cells with drugs that inhibit glycogen synthesis and breakdown. This data will provide pre-clinical information on which to base novel therapies for ECA and CSA.