Anti-ICAM-1 scFv from Lactobacilli as a Microbicide

描述（由申请人提供）：鉴于HIV-1传播阻断疫苗的短期前景不佳，以及避孕套作为控制HIV-1性传播的实用机制的失败，迫切需要一种有效且可接受的抗HIV-1杀菌剂。有两个特点可大大提高杀微生物剂的可接受性，即对使用者完全透明和不要求立即在性交前使用。尽管无细胞病毒和细胞相关病毒在人类性传播中的相对作用尚不清楚，但我们通过体内SCID小鼠和体外HIV-1性传播transwell模型发现，HIV-1的细胞相关传播效率很高，而无细胞病毒传播效率较低。这些结果与在猕猴模型系统中报道的结果不同，这可能是由于所有报道的猕猴研究都没有努力将环境调整到在人类环境中发生传播的中性pH值。在我们的模型系统中，针对细胞粘附分子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，该抗体通过一种涉及信号转导的机制起作用，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）的乳酸菌可以作为一种传递机制，提供一种持续的、完全透明的，并且在异性传播的情况下，女性控制的阻断细胞相关病毒传播的方法。为了验证这一假设，我们已经与世界领先的实验室建立了合作，研究乳酸杆菌抗体片段的表达。与位于荷兰的实验室合作，我们寻求1)检验ICAM-1以交联或非交联方式参与内皮单层对感染细胞转移的抗性的不同影响2)工程乳酸菌表达细胞壁结合或分泌的单价单价scFv或scAb，并在transwell实验中评估分泌产物的体外功效3)在SCID小鼠模型中评估工程乳酸菌的定植特性和保护功效在老鼠阴道中繁殖。预计这些研究将为使用这种方法开展临床研究提供必要的原理证明。

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.