HIV-envelope-specific DARPin-based microbicide strategies

描述（由申请人提供）：HIV包膜特异性DARPin基杀微生物剂策略为了成为可行的候选杀微生物剂，抗HIV策略必须高度有效，阻断相关体外模型系统中的无细胞和细胞相关感染，预防体内感染而不干扰免疫功能或引起任何毒性反应，并且能够以低成本生产。通过最近建立的合作，我们确定了一种新的，高度特异性的方法来抑制艾滋病毒感染使用设计锚蛋白重复蛋白（DARPin）技术。最初的概念验证研究记录了CD 4特异性DARPin的选择，这些DARPin具有高度特异性，并在纳摩尔水平下有效地预防各种HIV分离株以及SIV的体外感染，而不干扰基本的细胞功能。DARPin具有独特的特性，表现出优异的稳定性和特异性，并且生产成本很低。结合这些特征和预防HIV感染的能力，DARPins代表了一种有前途的新策略，可用于探索预防HIV传播。在这项新的合作研究的基础上，我们建议选择能够预防粘膜HIV感染的高活性HIV Env特异性DARPins。我们假设，识别gp 120和gp 41的Env特异性DARPin的混合物将（i）有效地预防在低纳摩尔范围内起作用的许多HIV和SIV分离株的体外无细胞和细胞相关感染，（ii）不损害先天性或适应性免疫功能，和（iii）预防猕猴中SHIV 162 P3的粘膜感染，而对局部免疫功能、植物群、Trkola博士的实验室正在开发这种创新的抗艾滋病毒技术（与Pl <$ckthun博士的小组合作），一旦通过严格的筛选试验选出，将使用Robbiani博士和Trkola博士实验室建立的相关细胞和组织分析在各种体外病毒和免疫系统中对有希望的候选物进行彻底测试。然后通过Robbiani博士的实验室（与Blanchard博士合作）在猕猴中测试先导新候选物的体内活性。利用一种新技术来开发抗HIV Env特异性策略，这种高风险DARPins方法应该在预防HIV感染和传播方面取得关键进展。 项目摘要-相关性：这些研究将提供关于这种新型局部杀微生物剂在体内预防阴道感染的功效的第一批数据。随着概念的证明，这代表了一个有前途的策略，以解决艾滋病毒在粘膜，我们将能够探索这种方法的应用，以性交独立的方式，以及对直肠传播的潜在应用。这些数据将推动这项新技术的设计和未来测试，以改善在女性和男性中测试的杀微生物剂策略。

DESCRIPTION (provided by applicant): HIV-envelope-specific DARPin-based microbicide strategies In order to be a viable candidate microbicide, an anti-HIV strategy must be highly effective, block cell-free and cell-associated infections in relevant in vitro model systems, prevent in vivo infection without perturbing immune function or causing any toxic reactions, and be able to be produced at low costs. Through a recently established collaboration, we identified a novel, highly specific approach to inhibit HIV infection using the Designed Ankyrin Repeat Protein (DARPin) technology. The initial proof of concept studies documented the selection of CD4- specific DARPins that are highly specific and efficiently prevent in vitro infection with a wide range of HIV isolates as well as SIV at nanomolar levels, without interfering with basic cellular functions. DARPins possess unique characteristics exhibiting exceptional stability and specificity, and cost little to produce. Combining these features with the ability to prevent HIV infection, DARPins represent a promising novel strategy to explore for the prevention of HIV spread. Expanding on this new collaborative research, we propose to select highly active HIV Env-specific DARPins that will be able to prevent mucosal HIV infection. We hypothesize that cocktails of Env-specific DARPins, which recognize gp120 and gp41, will (i) efficiently prevent in vitro cell-free and cell-associated infection with numerous HIV and SIV isolates, functioning in the low nanomolar range, (ii) not impair innate or adaptive immune functions, and (iii) prevent mucosal infection with SHIV162P3 in macaques, with no adverse impact on local immune function, flora, and pH. This innovative anti-HIV technology is being developed in Dr. Trkola's laboratory (in collaboration with Dr. Pl¿ckthun's group) and, once selected through rigorous screening assays, promising candidates will be thoroughly tested in a variety of in vitro viral and immune systems using relevant cell and tissue-based assays established in both Drs. Robbiani's and Trkola's laboratories. The lead novel candidate(s) will then be tested for in vivo activity in macaques through Dr. Robbiani's laboratory (in collaboration with Dr. Blanchard). Utilizing a novel technology to develop an anti-HIV Env specific strategy, this high-risk DARPins approach should yield critical advances towards preventing HIV infection and spread. Project Summary-Relevance: These studies will provide the first data on the efficacy of this novel topical microbicide to prevent vaginal infection in vivo. With proof of concept that this represents a promising strategy to tackle HIV at the mucosa, we will be able to explore application of this approach to coitally independent modalities, as well as for the potential application against rectal transmission. These data will advance the design and future testing of this novel technology to improve microbicide strategies to test in women and men.