DISSECTING AND TARGETING THE ROLE OF GALNT14 IN HIGH-RISK OSTEOSARCOMA

剖析和瞄准 GALNT14 在高风险骨肉瘤中的作用

• 批准号: 10761850
• 负责人: Jennifer J Kohler
• 金额: $ 19.37万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-13 至 2024-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10761850
• 关键词: Address; Adjuvant Chemotherapy; Adolescent; Adult; Alternative Therapies; Antigens; Azides; Biological Assay; Biology; Bone neoplasms; Breast; CRISPR/Cas technology; Categories; Cell Communication; Cell Culture Techniques; Cell Line; Cells; Chemicals; Chemoresistance; Chemotherapy-Oncologic Procedure; Childhood Osteosarcoma; Cisplatin; Clinical Trials; Cultured Cells; Databases; Diagnosis; Disease Progression; Disease Resistance; Disease-Free Survival; Dose; Doxorubicin; Drug Kinetics; Enzyme-Linked Immunosorbent Assay; Enzymes; Evaluation; Excision; Exhibits; Family; Future; Genes; Glycobiology; Goals; Immune system; Immunotherapeutic agent; In Vitro; Incidence; Inferior; Informatics; Institution; Knock-out; Libraries; Link; Lung; Malignant Neoplasms; Mass Spectrum Analysis; Measures; Methods; Methotrexate; Mission; Modality; Modeling; Molecular; Molecular Target; Necrosis; Neoadjuvant Therapy; Neoplasm Metastasis; Ontology; Operative Surgical Procedures; Outcome; Patient-Focused Outcomes; Patients; Pediatric Neoplasm; Permeability; Post-Translational Protein Processing; Primary Neoplasm; Prognosis; Prognostic Factor; Prostate; Protein Isoforms; Proteins; Public Health; Rapid screening; Recombinants; Recurrent disease; Refractory; Refractory Disease; Regimen; Relapse; Research; Resected; Resistance; Role; Serine; Signal Transduction; Survival Rate; Testing; Therapeutic; Threonine; Toxic effect; United States National Institutes of Health; chemotherapy; childhood sarcoma; clinical development; effective therapy; experience; functional genomics; gain of function; glycosylation; high risk; high throughput screening; improved; in vitro Model; in vivo; inhibitor; insight; lead candidate; loss of function; new therapeutic target; novel; osteosarcoma; overexpression; patient derived xenograft model; predicting response; protein transport; resistance mechanism; response; sarcoma; small molecule; small molecule inhibitor; standard of care; sugar; synergism; targeted treatment; therapeutic candidate; therapeutic target; therapy resistant; transcriptomics; treatment response; treatment strategy; tumor; tumor growth; tumor progression

Osteosarcoma (OS) is the most common bone tumor in children and adolescents. Regimens of neoadjuvant doxorubicin, cisplatin, and high dose methotrexate have been the standard of care since the 1970s, but no additional chemo or targeted therapies have added significant survival benefits, leaving minimal options for relapsed and resistant disease. These patients have an extremely poor prognosis, with long term outcomes of less than 20%. Therefore, a better understanding of chemoresistance mechanisms is essential towards making our current therapies more effective. Through comprehensive transcriptomic analysis of institutional chemoresistant patient-derived xenograft (PDX) models and the publicly available Therapeutically Applicable Research to Generate Effective Treatments (TARGET) OS database, we identified significant elevation of glycosylation genes in patients with poor response to chemotherapy. Glycosylation is a form of protein post- translational modification that plays a role in protein trafficking, function, and stability. It also provides substrates for cell-cell interactions, antigens for immune system surveillance, and often plays a role in cell signaling. O-type glycosylation, the addition of sugar moieties to serine or threonine residues to proteins, begins with the family of N-acetylgalactosyltransferase enzymes (GALNTs). Our analysis identified overexpression of GALNT14 in poor responding (<90% necrosis) OS tumors and we provide evidence that expression of GALNT14 has significant inverse correlation with overall survival and event-free survival in OS. GALNT14 has been shown to promote chemotherapy resistance and metastasis in adult cancers, such as of the breast, prostate, and lung. Our hypothesis is GALNT14 has a significant impact on disease progression in pediatric OS by promoting chemotherapy resistance and is a novel, viable therapeutic target for small molecule inhibition. We propose two complementary, yet independent aims to address this hypothesis. The proposal will synergize the expertise of Dr. Yustein, in sarcoma biology and modeling, and Dr. Kohler in glycobiology to successfully accomplish the goals. AIM1: Role of GALNT14 in promoting chemoresistance in osteosarcoma models in vitro and in vivo. Using both commercially available and PDX-derived chemoresistant cell lines we are developing both gain and loss-of-function GALNT14 models through overexpression and CRISPR/Cas9 knockout approaches. Using these models, we will evaluate in vitro and in vivo effects on OS sensitivity to chemotherapy and effects on tumor growth and progression. AIM2. Identification and evaluation of small molecule inhibitors of GALNT14. In this aim, we will use high-throughput screening to identify small molecules that inhibit the activity of purified, recombinant GALNT14. Further, we will determine which of these molecules can inhibit GALNT14 in cell culture. GALNT14 inhibitors identified in this aim have the potential to serve as lead candidates for clinical development and may also be used to investigate the mechanistic involvement of GALNT14 in OS. Overall, our studies will lead to the identification of critical candidate therapeutic modalities for the treatment of high-risk OS.

骨肉瘤是儿童和青少年最常见的骨肿瘤。新辅助治疗方案 阿霉素、顺铂和大剂量甲氨蝶呤自20世纪70年代以来一直是治疗的标准，但没有 额外的化疗或靶向治疗增加了显著的生存益处， 复发和耐药疾病。这些患者的预后极差，长期预后为 低于20%。因此，更好地理解化学抗性机制对于使 我们目前的疗法更有效。通过全面的转录组学分析， 化疗耐药患者来源的异种移植物（PDX）模型和公开可用的治疗适用 研究生成有效的治疗（目标）OS数据库，我们确定了显着升高 糖基化基因对化疗反应差的患者。糖基化是一种蛋白质后 在蛋白质运输、功能和稳定性中起作用的翻译修饰。它还提供基质 用于细胞-细胞相互作用，用于免疫系统监视的抗原，并且通常在细胞信号传导中起作用。o型 糖基化，将糖部分添加到蛋白质的丝氨酸或苏氨酸残基，开始于蛋白质的糖基化家族。 N-乙酰半乳糖基转移酶（GALNT）。我们的分析确定了GALNT 14在穷人中的过表达。 我们提供的证据表明，GALNT 14的表达具有显著的生物学效应。 与OS的总生存期和无事件生存期呈负相关。GALNT 14已显示促进 成人癌症（例如乳腺癌、前列腺癌和肺癌）的化疗耐药性和转移。我们 假设GALNT 14通过促进儿童OS的疾病进展， 是一种新的、可行的小分子抑制治疗靶点。我们提出了两 互补，但独立的目的，以解决这一假设。该提案将协同以下方面的专业知识： 博士Yustein，在肉瘤生物学和建模，和科勒博士在糖生物学成功地完成了 目标. AIM 1：GALNT 14在体外骨肉瘤模型中促进化疗耐药性的作用和在骨肉瘤中的作用 vivo.使用市售的和PDX衍生的化学抗性细胞系，我们正在开发两种增益 以及通过过表达和CRISPR/Cas9敲除方法的功能丧失GALNT 14模型。使用 这些模型，我们将评估体外和体内对OS对化疗敏感性的影响以及对肿瘤的影响， 成长和进步。目标2. GALNT 14的小分子抑制剂的鉴定和评价。在 为此，我们将使用高通量筛选来鉴定抑制纯化的， 重组GALNT 14。此外，我们将确定这些分子中的哪一种可以在细胞培养中抑制GALNT 14。 在这一目标中鉴定的GALNT 14抑制剂有可能作为临床开发的主要候选物 也可用于研究GALNT 14参与OS的机制。总之，我们的研究将 导致识别用于治疗高风险OS的关键候选治疗方式。