Glycosylation-dependent mechanisms regulating ovarian tumor cell survival

描述（由申请方提供）：表面聚糖谱的改变是肿瘤细胞的一个众所周知的特征;然而，特异性聚糖和相关糖基转移酶调节肿瘤细胞行为的分子机制仍然难以捉摸。我们小组的工作已经确定，ST 6 Gal-I唾液酸转移酶赋予卵巢癌（OC）细胞避免由多种刺激（包括半乳糖凝集素、死亡受体激活剂和化疗药物）诱导的凋亡的能力。在这个建议中，我们首次表明，ST 6 Gal-I蛋白在绝大多数人卵巢肿瘤中上调，包括致命的乳头状浆液性腺癌亚型。ST 6 Gal-I的上调相应地引起所选受体的过度唾液酸化，所述受体包括α 1整联蛋白、Fas和TNFR 1。在范式转换机制研究中，我们发现ST 6 Gal-I介导的Fas和TNFR 1的唾液酸化阻止了受体内化，导致这些受体的信号传导向生存而不是凋亡转变。此外，ST 6 Gal-I活性赋予对凋亡半乳糖凝集素-3和化疗剂顺铂的抗性。该提议的中心假设是ST 6 Gal-I过表达是促成OC致死性的两个关键因素的基础：肿瘤细胞对基于铂的化疗的抗性（Aim 1），以及OC细胞对腹膜肿瘤微环境内的免疫刺激的保护（Aim 2），其富含gal-3以及Fas和TNFR 1的配体。初步研究表明，ST 6 Gal-I促进患者腹膜腹水液中的OC细胞存活;这一发现是重要的，因为OC细胞通过腹膜转运转移。目的1：ST 6 Gal-I介导的受体唾液酸化在化疗耐药性中的作用。我们将定义负责ST 6 Gal-I在顺铂耐药性中的作用的分子机制，并确定受体唾液酸化是否控制完整肿瘤微环境中的化学耐药性。将针对在小鼠中从具有差异ST 6 Gal-I表达的OC细胞建立的原位肿瘤评价顺铂治疗的功效，并且将使用RNAi的体内递送来实施ST 6 Gal-I的治疗性敲低以恢复顺铂敏感性。将在化学敏感性与化学抗性人类肿瘤中定量ST 6 Gal-I以评估其预后潜力。目标2：ST 6 Gal-I对腹膜微环境中肿瘤细胞存活的贡献将阐明腹水中ST 6 Gal-I依赖性OC细胞存活的机制，重点是gal-3/1整合素、FasL/Fas和TNF？TNFR 1相互作用。将检查ST 6 Gal-I介导的OC细胞免受细胞毒性免疫细胞的保护，并筛选人卵巢肿瘤的ST 6 Gal-I和免疫细胞浸润之间的关联。将在免疫活性小鼠中监测原位肿瘤+/-ST 6 Gal-I的生长。这些研究将通过显示ST 6 Gal-I是控制肿瘤/微环境相互作用的主调节分子来产生影响，该主调节分子调节OC细胞的消除与持久性之间的平衡，从而驱动肿瘤复发。创新在于阐明：（1）新型糖基化依赖性肿瘤机制，以及（2）OC发病机制中的新分子参与者和患者化疗耐药性的生物标志物。

DESCRIPTION (provided by applicant): An altered profile of surface glycans is a well-known feature of tumor cells; however the molecular mechanisms by which specific glycans and associated glycosyltransferases regulate tumor cell behavior remain elusive. Work from our group has determined that the ST6Gal-I sialyltransferase endows ovarian cancer (OC) cells with the ability to avoid apoptosis induced by multiple stimuli including galectins, death receptor activators and chemotherapeutic drugs. In this proposal we show for the first time that ST6Gal-I protein is upregulated in the great majority of human ovarian tumors, including the lethal papillary serous adenocarcinoma subtype. Upregulation of ST6Gal-I correspondingly causes hypersialylation of selected receptors including the ¿1 integrin, Fas and TNFR1. In paradigm-shifting mechanistic studies, we find that ST6Gal-I-mediated sialylation of Fas and TNFR1 prevents receptor internalization, causing a shift in the signaling of these receptors toward survival rather than apoptosis. Additionally, ST6Gal-I activity confers resistance to apoptotic galectin-3 and the chemotherapeutic, cisplatin. The central hypothesis of this proposal is that ST6Gal-I overexpression underlies two crucial factors contributing to OC lethality: tumor cell resistance to platinum-based chemotherapy (Aim 1), and OC cell protection against immune stimuli within the peritoneal tumor microenvironment (Aim 2), which is rich in gal-3 as well as ligands for Fas and TNFR1. Preliminary studies show that ST6Gal-I promotes OC cell survival within patient peritoneal ascites fluid; this finding is significant in that OC cells metastasize va peritoneal transit. AIM 1: ST6Gal-I-mediated receptor sialylation in chemoresistance. We will define molecular mechanisms responsible for ST6Gal-I's role in cisplatin resistance, and determine whether receptor sialylation controls chemoresistance within the intact tumor microenvironment. Efficacy of cisplatin treatment will be evaluated for orthotopic tumors established in mice from OC cells with differential ST6Gal-I expression, and in vivo delivery of RNAi will be used to implement therapeutic knockdown of ST6Gal-I to restore cisplatin sensitivity. ST6Gal-I will be quantified in chemosensitive vs. chemoresistant human tumors to assess its prognostic potential. AIM 2: Contribution of ST6Gal-I to tumor cell survival within the peritoneal microenvironment Mechanisms underlying ST6Gal-I-dependent OC cell survival within ascites will be elucidated, with a focus on gal-3/ ¿1 integrin, FasL/Fas, and TNF?/TNFR1 interactions. ST6Gal-I-mediated protection of OC cells from cytotoxic immune cells will be examined, and human ovarian tumors screened for an association between ST6Gal-I and immune cell infiltration. The growth of orthotopic tumors +/- ST6Gal-I will be monitored in immunocompetent mice. The studies will have impact by showing that ST6Gal-I is a master regulatory molecule controlling tumor/ microenvironmental interactions that regulate the balance between elimination vs. persistence of OC cells that drive tumor recurrence. Innovation lies in the elucidation of: (1) novel glycosylation-dependent tumor mechanisms, and (2) a new molecular player in OC pathogenesis and biomarker for patient chemoresistance.