Anti-ICAM-1 scFv from Lactobacilli as a Microbicide

描述（由申请人提供）：鉴于HIV-1传播阻断疫苗的近期前景不佳，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杀微生物剂。有两个特点将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在性交前立即使用。尽管无细胞和细胞相关病毒在人类性传播中的相对作用尚不清楚，但我们已经发现使用体内SCID小鼠和体外transwell HIV-1性传播模型，HIV-1的细胞相关传播是高效的，而无细胞病毒传播很差。这些结果与猕猴模型系统中报告的结果不同，这可能是由于所有报告的猕猴研究都没有努力将环境调整到中性pH值，在该pH值下，人类环境中发生传播。在我们的模型系统中，细胞粘附分子ICAM-1的抗体在阻断细胞相关传输中非常有效。初步数据表明，该抗体是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）的乳酸杆菌可用作提供持续的、完全透明的递送机制，并且在异性传播的情况下，可用作阻断细胞相关病毒传播的女性控制的方法。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，研究乳酸杆菌表达抗体片段。与位于荷兰的这个实验室合作，我们寻求1）检测ICAM-1以交联或非交联方式参与对内皮单层抵抗感染细胞迁移的不同作用2）工程化乳酸杆菌以表达细胞壁结合或分泌的单价或二价scFv或scAb，并在transwell测定中评价分泌产物的体外功效3）在SCID小鼠模型中评估用于定殖小鼠阴道的工程乳酸杆菌的定殖特性和保护功效。预计这些研究将提供使用该方法启动临床研究所需的原理证明。

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.