Generation of antibody-drug conjugates by proximity-based sortase-mediated ligation

通过基于邻近的分选酶介导的连接生成抗体-药物缀合物

• 批准号: 10323853
• 负责人: Feifan Yu
• 金额: $ 108.12万
• 依托单位: ALPHATHERA, INC.
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-01 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10323853
• 关键词: Adult; Amino Acid Sequence; Antibodies; Antibody Avidity; Antibody Formation; Antibody Specificity; Antibody-drug conjugates; Binding; Biodistribution; Biological Assay; Blood Circulation; C-terminal; Chemicals; Chemistry; Clinical; Comparative Pathology; Cysteine; Data; Development; Disulfides; Drug Industry; Drug Kinetics; Drug Stability; Effectiveness; Engineering; Enzymes; Exposure to; FDA approved; Formulation; Generations; Goals; Human; Hydrophobicity; IgG Receptors; IgG1; Immunocompetent; Immunoglobulin G; In Vitro; Intravenous; Label; Lead; Ligation; Location; Lysine; Malignant Neoplasms; Malignant neoplasm of pancreas; Measurable; Mediating; Methods; Modality; Modeling; Modification; Molecular; Movement; Mus; Outcome; Pennsylvania; Peptides; Pharmaceutical Preparations; Polysaccharides; Preclinical Testing; Preparation; Property; Proteins; Serum; Site; Solubility; Technology; Testing; Therapeutic; Time; Tissues; Toxic effect; Transglutaminases; Universities; Variant; Vertebral column; antibody engineering; antitumor drug; base; cancer therapy; chemotherapy; clinical translation; cytotoxic; cytotoxicity; effective therapy; enzyme activity; experience; flexibility; formylglycine; glycosylated IgG; interest; lead candidate; neonatal Fc receptor; neoplastic cell; novel strategies; novel therapeutics; pancreatic cancer model; pancreatic cancer patients; selective expression; sortase; targeted treatment; tumor; unnatural amino acids

There has been growing interest in the use of antibody drug conjugates (ADCs) for the treatment of cancer as mounting data suggests an increase in anti-tumor effectiveness and reduced toxicity, compared with the administration of unlabeled antibodies in combination with chemotherapy. Recent evidence has shown that differentially labeled antibodies, i.e. labeled at different locations and with different numbers, can have distinct therapeutic and pharmacokinetic properties and some subpopulations can show little, if any, therapeutic activity yet account for most of the toxicity. Therefore, there has been a movement towards the development of site- specific ADCs, which are precisely labeled with drugs at pre-defined locations. We have recently developed two new approaches for the preparation of highly uniform ADCs, one site-specific bioconjugation approach, Proximity-Based Sortase-mediated protein Ligation (PBS-PL), whereby sortase is used to ligate drugs to a peptide tag that has been introduced into the antibody backbone, and one region- specific bioconjugation approach, Proximity-based Sortase Isopeptide Ligation (PBS-IL), which allows for the labeling of native antibodies with reduced variability compared with current lysine/cysteine residue labeling approaches. Both methods produce ADCs in high yields, are compatible with glycosylated IgG, and offer unlimited flexibility in antibody-drug linker chemistry. Therefore, we believe that these technologies will provide new, favorable approaches for the production of ADCs that will be of interest to the pharmaceutical industry. In this proposal, we have partnered with Integral Molecular and the University of Pennsylvania to develop ADCs against claudin-18 (CLDN 18.2). CLD18.2 is ectopically expressed in several cancers including pancreatic cancer, which is the focus of this proposal. Outcomes for pancreatic cancer patients are poor with a 5-year survival of <8% due to a lack of effective treatment modalities. Therefore, the development of new therapies are a clinical necessity. The highly selective expression of CLDN18.2 in cancer, with no detectable expression on any healthy adult tissues that are accessible to antibodies, make it an attractive option for targeted therapy. We will prepare various ADCs using PBS-PL and PBS-IL and will identify the conjugation approach and ADC formulation that is expected to be most favorable for clinical translation, based on serum stability, pharmacokinetics, and efficacy. Pre-clinical testing will be performed in an orthotopic pancreatic tumor model in syngeneic mice. The specific aims for the proposal are: Aim 1: Produce and characterize anti- CLDN18.2-vcMMAE ADCs using PBS-PL and PBS-IL; Aim 2: Evaluate the binding and efficacy of anti- CLDN18.2-vcMMAE ADCs in vitro; Aim 3. Determine the pharmacokinetics and efficacy of anti-CLDN18.2 ADCs in a murine tumor model

对于使用抗体药物缀合物（ADC）治疗癌症的兴趣日益增加， 越来越多的数据表明，与对照组相比， 施用未标记的抗体与化疗的组合。最近的证据表明， 差异标记的抗体，即在不同位置和用不同数目标记的抗体，可以具有不同的生物学特性。 治疗和药代动力学特性，一些亚群可能显示出很少的治疗活性（如果有的话 但却造成了大部分的毒性因此，有一个运动对网站的发展- 特异性ADC，其在预定义位置精确标记有药物。 我们最近开发了两种新方法来制备高度均匀的ADC，一种是针对特定地点的 生物缀合方法，基于邻近的分选酶介导的蛋白质连接（PBS-PL），其中分选酶是 用于将药物连接到已经引入抗体骨架的肽标签上，并且一个区域- 特异性生物缀合方法，基于邻近的分选酶异肽连接（PBS-IL），其允许将多肽与靶分子结合。 与目前的赖氨酸/半胱氨酸残基标记相比， 接近。两种方法都以高产率产生ADC，与糖基化IgG相容，并提供 抗体-药物接头化学的无限灵活性。因此，我们相信这些技术将提供 新的，有利的方法来生产ADC，这将是制药行业感兴趣的。 在这项提案中，我们与Integral Molecular和宾夕法尼亚大学合作， 针对密蛋白-18的ADC（CLDN 18.2）。CLD18.2在几种癌症中异位表达，包括 胰腺癌，这是这个建议的重点。胰腺癌患者的预后很差， 5-由于缺乏有效的治疗方法，年生存率<8%。因此，新的发展 治疗是临床必需品。CLDN18.2在癌症中的高度选择性表达，没有检测到 在任何健康的成人组织上表达，抗体可以接近，使其成为一个有吸引力的选择， 靶向治疗我们将使用PBS-PL和PBS-IL制备各种ADC，并将鉴定缀合物。 方法和ADC制剂，预计最有利于临床转化，基于血清 稳定性、药代动力学和功效。将在原位胰腺肿瘤中进行临床前试验 在同系小鼠中的模型。该提案的具体目标是： 使用PBS-PL和PBS-IL的CLDN18.2-vcMMAE ADC;目的2：评估抗CLDN18.2-vcMMAE ADC的结合和功效。 体外CLDN18.2-vcMMAE ADC;目的3.确定抗CLDN 18的药代动力学和功效。2 鼠肿瘤模型中的ADC