The first polymeric opioid conjugate vaccine

1.摘要： 阿片成瘾是一种全国性的医疗危机，由于阿片类药物的社会经济后果，预计这种危机将会恶化 SARS-CoV-2大流行。阿片疫苗是一种很有前途的预防阿片类药物过量的治疗策略，并可能 在与现有模式相结合时加强药物治疗。接种疫苗的目的是产生特定的阿片类药物 抗体(-阿片)S与药物结合并将其隔离在血清中，阻止阿片类药物进入大脑和 其他器官，从而防止服药过量和使用者寻求的感觉。而单体阿片类半抗原结合 由阿片单体结合载体蛋白和实验佐剂组成的疫苗是有效的 在啮齿动物中；目前单体阿片疫苗的设计是基于可卡因和尼古丁的单体疫苗， 在人类身上产生相对较弱的、短暂的抗体反应。这项提议的目标是开发一种聚合物 阿片结合疫苗，在没有毒性的情况下诱导强大、持久的-阿片反应。持久抗体 反应是由长寿的抗体分泌细胞(LLASCs)维持的，这些细胞是在B细胞激活时产生的 B细胞受体(BCR)的存在，识别病原体产生的免疫刺激信号，以及 辅助性T细胞共刺激信号的连接。为了获得阿片类药物特异性B细胞激活所需的信号， 我们针对芬太尼的聚合物阿片疫苗--大多数阿片类药物过量死亡的原因--是由一种水溶性的 靶向并激活芬太尼特异性B细胞的共聚物，称为p(Fent-co-TLR7)，与免疫原性偶联 载体蛋白破伤风毒素(TT)通过自焚连接，当被切割时，释放未修饰的TT。我们会 由芬太尼修饰的单体和第二单体合成了无规共聚物p(Fent-co-TLR7)，第二单体 通过Toll样受体7(TLR7)激活B细胞。因此，多价TT-p(Fent-co-TLR7)偶联物被设计成靶向 和交联型阿片特异性B细胞受体(BCR)，引起B细胞活化和BCR介导的内化。一次 内化的TT-p(Fent-co-TLR7)将激活内体TLR7，放大B细胞的激活，释放TT。一次 从p(Fent-co-TLR7)聚合物中释放出来的TT可以被有效地加工，其多肽被呈现在B的表面 细胞转化为TT特异性T细胞，从而产生最佳的T细胞激活和信号的产生，从而驱动B细胞 分化为LLASC。在这项建议中，我们将(1)优化提法并论证机制 负责TT-p(Fent-co-TLR7)的疗效，(2)比较免疫的免疫原性和持久性 TT-p(Fent-co-TLR7)对单体芬太尼-TT结合疫苗的应答，以及(3)证明 TT-p(Fent-co-TLR7)对芬太尼行为效应和致死性的抑制作用。完成这项提案将 验证临床可行的芬太尼疫苗的临床前疗效，并提供一种新的疫苗平台，可以 经修改后可用于治疗其他药物的非法使用以及传染病。

1. Abstract: Opioid addiction is a national medical crisis that is expected to worsen due to the socioeconomic fallout caused by the SARS-CoV-2 pandemic. Opioid vaccines are a promising treatment strategy for preventing opioid overdose and could enhance drug treatment when combined with existing modalities. The aim of vaccination is to generate opioid-specific antibodies (-opioid)s that bind to the drugs and sequester them in the serum, stopping opioids from entering the brain and other organs, and thus preventing overdose and the sensation sought by the user. While monomeric opioid hapten conjugate vaccines composed of opioid monomers conjugated to carrier proteins admixed with experimental adjuvants are efficacious in rodents; the design of current monomeric opioid vaccines is based on monomeric vaccines for cocaine and nicotine that generated relatively weak, short-lived antibody responses in humans. The goal of this proposal is to develop a polymeric opioid conjugate vaccine that induces robust, long-lasting -opioid responses in the absence of toxicity. Durable antibody responses are maintained by long-lived antibody-secreting cells (LLASCs), which are generated upon B cell activation in the presence of B cell receptor (BCR) crosslinking, recognition of pathogen derived immunostimulatory signals, and ligation of co-stimulatory signals from helper T cells. To elicit the signals necessary for opioid-specific B cell activation, our polymeric opioid vaccine against fentanyl—the cause of most opioid overdose deaths—is composed of a water-soluble copolymer that targets and activates fentanyl-specific B cells, termed p(Fent-co-TLR7), conjugated to the immunogenic carrier protein tetanus toxin (TT) via a self-immolative linkage that, when cleaved, releases unmodified TT. We will synthesize p(Fent-co-TLR7) as a random copolymer from a monomer decorated with fentanyl and a second monomer that activates B cells via toll-like receptor 7 (TLR7). Thus, multivalent TT-p(Fent-co-TLR7) conjugates are designed to target and crosslink opioid-specific B cell receptors (BCRs), causing B cell activation and BCR-mediated internalization. Once internalized, TT-p(Fent-co-TLR7) will activate endosomal TLR7, amplifying B cell activation, and release TT. Once released from p(Fent-co-TLR7) polymers, TT can be efficiently processed and its peptides presented on the surface of B cells to TT-specific T cells, resulting in optimal T cell activation and the production of signals that drive B cells to differentiate into LLASCs. In this proposal we will (1) optimize the formulation and demonstrate the mechanisms responsible for the efficacy of TT-p(Fent-co-TLR7), (2) compare the immunogenicity and durability of the immune response generated by TT-p(Fent-co-TLR7) to that of monomeric fentanyl-TT conjugate vaccines, and (3) demonstrate the ability of TT-p(Fent-co-TLR7) to inhibit the behavior effects and lethality of fentanyl. Completion of this proposal will validate the preclinical efficacy of a clinically-viable fentanyl vaccine and deliver a novel vaccine platform that can be modified to treat the illicit use of other drugs as well as infectious diseases.