Anti-ICAM-1 scFv from Lactobacilli as a Microbicide

描述(申请人提供)：鉴于艾滋病毒-1传播阻断疫苗的近期前景不佳，以及避孕套作为控制艾滋病毒-1性传播的实用机制的失败，迫切需要一种有效和可接受的抗艾滋病毒-1杀微生物剂。应显著提高杀菌剂可接受性的两个特点是对使用者完全透明和不要求立即进行性交前使用。虽然无细胞病毒和细胞相关病毒在人类性行为传播中的相对作用尚不清楚，但我们利用体内SCID小鼠和体外Transwell性传播模型发现，细胞相关病毒对HIV-1的传播是高效的，而无细胞病毒的传播很差。这些结果与猕猴模型系统中报道的结果不同，这可能是因为所有报道的猕猴研究都没有努力将环境调整到中性pH值，在人类环境中发生传播。在我们的模型系统中，细胞黏附分子ICAM-1的抗体在阻断细胞相关传播方面非常有效。初步数据表明，抗体的作用机制涉及信号转导，而不是简单地阻断ICAM-1的对接功能。我们现在假设，表达单链抗体片段(scFv或scab)的乳杆菌可以作为一种传递机制，提供一种持续的、完全透明的，在异性传播的情况下，妇女控制的方法来阻断细胞相关病毒的传播。为了评估这一假设，我们与世界领先的实验室建立了合作关系，研究乳杆菌表达抗体片段。我们与位于荷兰的这个实验室合作，试图1)检测ICAM-1在交联型和非交联型ICAM-1作用下对内皮细胞单层迁移的抵抗力的差异效果2)工程乳酸菌表达细胞壁结合或分泌的单价单价单链抗体或结痂，并评估其分泌产物在Transwell实验中的体外效力3)在SCID小鼠模型中，评价用于定植小鼠阴道的工程乳酸菌的定植特性和保护效果。预计这些研究将提供使用这种方法启动临床研究所需的原则证明。

DESCRIPTION (provided by applicant): Given the poor near-term prospects for an HIV-1 transmission-blocking vaccine and the failure of condoms as a practical mechanism for controlling sexual transmission of HIV-1, there is urgent need for an effective and acceptable anti-HIV-1 microbicide. Two features, which should dramatically enhance microbicide acceptability would be complete transparency to the user and the absence of a requirement for immediate pre-coital application. Although the relative roles of cell-free and cell-associated virus in human sexual transmission are unknown, we have found using in vivo SCID mouse and in vitro transwell models of HIV-1 sexual transmission that cell-associated transmission of HIV-1 is highly efficient and cell-free virus is poorly transmitted. These results differ from those reported in the macaque model system, which is likely accounted for by the fact that all of the reported macaque studies make no effort to adjust the environment to the neutral pH at which transmission occurs in the human setting. In our model systems, antibody to the cell adhesion molecule ICAM-1 is remarkably effective in blocking cell-associated transmission. Preliminary data suggest that the antibody is acting by a mechanism that involves signal transduction rather than by simply blocking the docking function of ICAM-1. We are now hypothesizing that lactobacilli expressing single chain antibody fragments (scFv or scAb) can be used as a delivery mechanism for providing a sustained, totally transparent,and, in the case of heterosexual transmission, woman-controlled method for blocking cell-associated virus transmission. To evaluate this hypothesis we have established a collaboration with the leading laboratory in the world studying the expression of antibody fragments by lactobacilli. In collaboration with this laboratory, located in the Netherlands, we seek to 1) Examine differential effects of engagement of ICAM-1 in a cross-linking or a non-cross-linking manner on resistance of endothelial monolayers to transmigration of infected cells 2)Engineer lactobacilli to express cell-wall bound or secreted monovalent ordivalent scFv or scAb and evaluate the in vitro efficacy of the secreted products in the transwell assay 3) Evaluate in the SCID mouse model the colonization characteristics and protective efficacy of engineered lactobacilli used to colonize the mouse vagina. It is anticipated that these studies will provide the proof of principle necessary to initiate clinical studies using this approach.