MONOCLONAL ANTIBODY SEB IMMUNOPROTECTANT

Tulane University Of Louisiana, New Orleans LA

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Abstract

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Staphylococcal enterotoxin B (SEB;Category B agent), a toxin that commonly causes classic food poisoning and can cause a nonmenstrual toxic shock syndrome (TSS), is a potential biological warfare agent. Importantly, SEB is derived from common readily accessible bacteria, is relatively easily produced, and can be delivered in a stable aerosol form. An SEB attack would be devastating to civilian populations as well as on the battlefield during times of war. Currently, there are no preventatives or therapeutics available against SEB. Monoclonal antibodies (Mabs) are a class of FDA-approved therapeutics shown to neutralize toxins. Because of their specificity, stability, high potency, and versatility human Mabs are ideal for biodefense related countermeasures. The goals of this project are to: (1) generate a panel of fully human anti-SEB Mabs;(2) select lead anti-SEB Mabs based upon prophylactic efficacy in mouse intoxication models;(3) compare the protective efficacy of the lead Mabs when expressed in mammalian cell culture with the identical Mab expressed in a rapid, highly scalable manufacturing system. The Long Range Objective is to develop a safe and effective immunoprotectant product for SEB. Two product modalities are envisioned: 1) A preventive product consisting of a human anti-SEB Mab for intramuscular administration prior to potential exposure to weaponized SEB. 2) A therapeutic product consisting of the human anti-SEB Mabs administered in combination with Mab(s) against pro-inflammatory cytokines to provide protection post- exposure. To date we have screened over 33 alpha-SEB antibodies and/or fragments. We have used a number of nonhuman primate-derived peripheral mononuclear blood cells (PMBCs) freshly isolated from blood samples. The PMBCs are derived from blood obtained from Indian and Chinese-origin rhesus macaques (Macaca mulatta), cynomolgus macaques (Macaca fascicularis) and African green monkeys (Chlorocebus athetiops). The in vitro assay we have used for screening purposes is a commercially-available colorimetric proliferation assay that utilizes bromodeoxyuridine (BrdU) (Roche Diagnostics). The BrdU colorimetric assay is a replacement for the tritiated-thymidine radioactive assay. BrdU colorimetric assay incorporates fully into mitogenic activity and provides measurable change indicative of cellular proliferation in the presence of a superantigen or other mitogenic compound. Development and validation of this proliferation assay with this blood source (NHP PMBCs) and SEB was performed for approximately three months prior to testing the alpha-SEB antibodies and fragments. The results of the screenings with the candidate monoclonals thus far have shown minimal modulation of lymphocytic proliferation, indicating that these reagents do not possess the ability to neutralize toxin and minimize the effects from exposure. Concurrent to the proliferation assays, we reestablished the murine models of SEB shock (IP, aerosol, and oral models) in anticipation of the using them for a promising therapeutic antibody that would have been identified in the proliferation assays.