Development of highly neutralizing nanobodies against HIV-1

HIV-1和HIV-2是导致人类获得性免疫缺陷综合征（AIDS）的慢病毒。到目前为止，还没有开发出有效的疫苗，根据世界卫生组织的统计，全世界有近4000万人感染了艾滋病毒。由于预防措施和治疗不普及，艾滋病是撒哈拉以南非洲等地区的主要死亡原因。因此，迫切需要开发有效的方法，包括疫苗，以预防易感人群感染艾滋病毒。在过去的十年中，已经测试了几种药物作为局部杀微生物剂来防止艾滋病毒进入，结果好坏参半。然而，这些方法需要通过阴道环、凝胶、薄膜或灌肠剂进行药物递送，这在世界贫困地区并不实用。一个有趣的替代方案是使用胃肠道和生殖道细菌作为抗病毒肽的体内递送载体。例如，乳酸杆菌递送HIV-1进入抑制剂显著降低了恒河猴中SIV感染的发生率。使用类似的策略通过骆驼科动物仅重链抗体阻断艰难梭菌毒素。来自这些抗体的V（D）J结构域，也称为纳米抗体，特别适合于开发杀微生物剂，因为它们容易由肠道细菌分泌，在不同的pH条件下对变性具有抗性，并且可以容易地穿透宫颈阴道粘膜。此外，由于它们的小尺寸，纳米抗体可以针对抗原裂缝（例如HIV聚糖屏蔽的刺突）开发，这对于体积较大的常规抗体来说是具有挑战性的靶标。纳米抗体也可以相对容易地人源化，这是一种在临床试验中降低其免疫原性的策略。该项目提出了开发高度中和的抗HIV纳米抗体，用于潜在的杀微生物剂或体内治疗。此外，我们提出了一种在小鼠中生产纳米抗体的方法，目的是开发针对HIV-1和其他人类病原体的基于纳米抗体的疗法。

HIV-1 and -2 are lentiviruses that cause acquired immunodeficiency syndrome (AIDS) in humans. To date, no effective vaccine has been developed and according to the World Health Organization nearly 40 million people are infected with HIV across the world. Because preventive measures and treatments are not widely available, AIDS is the leading cause of death in areas such as Sub Saharan Africa. There is therefore an urgent need to develop efficient methods, including vaccines, to prevent HIV infection in susceptible populations. With mixed results, several drugs have been tested in the past decade for use as topical microbicides to prevent HIV entry. However, these approaches require drug delivery through vaginal rings, gels, films or enemas, which are not practical treatments in poor areas of the world. An intriguing alternative is the use of gastrointestinal and genital tract bacteria as in vivo delivery vectors for anti-viral peptides. For instance, Lactobacillus delivery of HIV-1 entry inhibitors markedly reduced the incidence of SIV infection in rhesus monkeys. A similar strategy was used to block Clostridium difficile toxins by means of camelid heavy chain only antibodies. V(D)J domains from these antibodies, also known as nanobodies, are particularly suited for the development of microbicides because they are readily secreted by commensal bacteria, are resistant to denaturation under varied pH conditions, and can easily penetrate the cervicovaginal mucosa. In addition, because of their small size, nanobodies can be developed against antigen crevices such as HIV glycan-shielded spikes, which are challenging targets for bulkier conventional antibodies. Nanobodies can also be humanized with relative ease, a strategy that reduces their immunogenicity in clinical trials. This project proposes the development of highly neutralizing anti-HIV nanobodies for potential use as microbicides or in in vivo therapies. Furthermore, we lay out an approach to produce nanobodies in mice with the goal of developing nanobody-based therapies against HIV-1 and other human pathogens.