Novel Enediyne-Based Antibody-Drug Conjugates for Cancers

两种FDA批准的抗体-药物缀合物（ADC）（Adcetris®和Kadcyla®）和几乎所有的ADC都在FDA批准的药物组合物中使用。 临床试验是通过将药物随机偶联到抗体中的赖氨酸或半胱氨酸残基上进行的， 提供非均相ADC混合物。位点特异性缀合是ADC的一个主要的近期改进 开发，产生具有极大改善的治疗指数的均质ADC。在60+ ADC中 目前在临床开发中，其中近50种使用澳瑞他汀和美登素，这两种有效载荷用于 分别使用Adcetris®或Kadcyla®，其余药物仅使用五种其他药物之一。ADC领域处于关键阶段， 需要在许多肿瘤类型中具有活性的新的、高效的和快速作用的细胞毒性有效载荷。我们 在本申请中提出开发用于癌症治疗的新的基于烯二炔的位点特异性ADC。具体地说， 我们将重点介绍我们最近发现的烯二炔家族的新成员--天环霉素 天然产品。我们的假设是：（i）TNM是优秀的ADC有效载荷候选者，因为它们 精致的效力和验证的作用模式，（ii）TNM生物合成的遗传操作提供了 生产最有前途的TNM类似物以开发接头化学的绝佳机会， （iii）TNM与工程化的thiomab的位点特异性缀合， 硒单抗将产生具有确定的药物与抗体比率（DAR）的均质ADC，以及（iv） HER 2和ROR 1的互补靶向将有助于评估新的基于烯二炔的ADC， 目前的基准和改善乳腺癌患者的治疗结果。具体目标是 获得的授权是：（i）操纵链霉菌CB 03234中的TNM生物合成，以产生最有希望的 用于位点特异性缀合的TNM，（ii）抗HER 2和抗RORl thiomab的产生和纯化，以及 具有2个工程化半胱氨酸（Cys）和2个或1个工程化硒代半胱氨酸（Sec）残基的硒单抗， （iii）开发用于位点特异性缀合的接头化学和递送一组抗- HER 2和抗ROR 1硫代单抗-TNM和硒代单抗-TNM缀合物，和（iv）评价选择性和 抗HER 2和抗ROR 1硫代单抗-TNM和硒代单抗-TNM缀合物针对HER 2+和 体外和体内模型中的HER 2 −/ROR 1+乳腺癌。本申请的结果包括（i） 对ADC的基本贡献和（ii）一组抗HER 2和抗ROR 1硫代单抗-TNM和硒代单抗-TNM。 TNM缀合物作为癌症的下一代ADC治疗剂。我们研究的长期目标是 发现新的微生物天然产物，并利用其精致的细胞毒性作为ADC有效载荷， 抗癌药物的发现

Both the two FDA-approved antibody-drug conjugates (ADCs) (Adcetris® and Kadcyla®) and nearly all ADCs in clinical trials are prepared by randomly conjugating drugs to lysine or cysteine residues in the antibody, affording a heterogeneous ADC mixture. Site-specific conjugation is a major recent improvement to ADC development, yielding homogeneous ADCs with greatly improved therapeutic index. Among the 60+ ADCs currently in clinical development, nearly 50 of them use auristatin and maytansine, the two payloads used in Adcetris® or Kadcyla®, respectively, with the rest using one of only five other drugs. The ADC field is in critical need of new, highly potent, and rapidly acting cytotoxic payloads that are active in many tumor types. We propose in this application to develop new enediyne-based site-specific ADCs for cancer therapy. Specifically, we will focus on tiancimycins (TNMs), which we recently discovered as novel members of the enediyne family of natural products. Our hypotheses are: (i) TNMs are outstanding ADC payload candidates owing to their exquisite potency and validated mode of action, (ii) genetic manipulation of TNM biosynthesis provides outstanding opportunities to produce the most promising TNM analogues to develop the linker chemistry and facilitate site-specific conjugation, (iii) site-specific conjugation of TNMs to engineered thiomabs and selenomabs will generate homogeneous ADCs with defined drug-to-antibody ratios (DARs), and (iv) complementary targeting of HER2 and ROR1 will help evaluate the novel enediyne-based ADCs against current benchmarks and improve therapeutic outcomes for breast cancer patients. The specific aims for this grant are: (i) manipulation of TNM biosynthesis in Streptomyces sp. CB03234 to produce the most promising TNMs for site-specific conjugation, (ii) production and purification of anti-HER2 and anti-ROR1 thiomabs and selenomabs with 2 engineered cysteine (Cys) and 2 or 1 engineered selenocysteine (Sec) residues, respectively, (iii) development of linker chemistry for site-specific conjugation and delivery of a panel of anti- HER2 and anti-ROR1 thiomab-TNM and selenomab-TNM conjugates, and (iv) evaluation of selectivity and potency of the anti-HER2 and anti-ROR1 thiomab-TNM and selenomab-TNM conjugates against HER2+ and HER2−/ROR1+ breast cancers in both in vitro and in vivo models. The outcomes of this application include (i) fundamental contributions to ADCs and (ii) a panel of anti-HER2 and anti-ROR1 thiomab-TNM and selenomab- TNM conjugates as next-generation ADC therapeutics for cancers. The long-term goal of our research is to discover novel microbial natural products and harness their exquisite cytotoxicity as ADC payloads for anticancer drug discovery.