Drozdowski Brian, Zhou Yuhong, Kline Brad, Spidel Jared, Chan Yin Yin, Albone Earl, Turchin Howard, Chao Qimin, Henry Marianne, Balogach Jacqueline, Routhier Eric, Bavari Sina, Nicolaides Nicholas C, Sass Philip M, Grasso Luigi
Morphotek Inc,, 210 Welsh Pool Road, Exton, PA, USA.
J Immune Based Ther Vaccines. 2010 Dec 21;8:9. doi: 10.1186/1476-8518-8-9.
Staphylococcal enterotoxins are considered potential biowarfare agents that can be spread through ingestion or inhalation. Staphylococcal enterotoxin B (SEB) is a widely studied superantigen that can directly stimulate T-cells to release a massive amount of proinflammatory cytokines by bridging the MHC II molecules on an antigen presenting cell (APC) and the Vβ chains of the T-cell receptor (TCR). This potentially can lead to toxic, debilitating and lethal effects. Currently, there are no preventative measures for SEB exposure, only supportive therapies.
To develop a potential therapeutic candidate to combat SEB exposure, we have generated three human B-cell hybridomas that produce human monoclonal antibodies (HuMAbs) to SEB. These HuMAbs were screened for specificity, affinity and the ability to block SEB activity in vitro as well as its lethal effect in vivo.
The high-affinity HuMAbs, as determined by BiaCore analysis, were specific to SEB with minimal crossreactivity to related toxins by ELISA. In an immunoblotting experiment, our HuMAbs bound SEB mixed in a cell lysate and did not bind any of the lysate proteins. In an in vitro cell-based assay, these HuMAbs could inhibit SEB-induced secretion of the proinflammatory cytokines (INF-γ and TNF-α) by primary human lymphocytes with high potency. In an in vivo LPS-potentiated mouse model, our lead antibody, HuMAb-154, was capable of neutralizing up to 100 μg of SEB challenge equivalent to 500 times over the reported LD50 (0.2 μg) , protecting mice from death. Extended survival was also observed when HuMAb-154 was administered after SEB challenge.
We have generated high-affinity SEB-specific antibodies capable of neutralizing SEB in vitro as well as in vivo in a mouse model. Taken together, these results suggest that our antibodies hold the potential as passive immunotherapies for both prophylactic and therapeutic countermeasures of SEB exposure.
葡萄球菌肠毒素被认为是潜在的生物战剂，可通过摄入或吸入传播。葡萄球菌肠毒素B（SEB）是一种经过广泛研究的超抗原，它可以通过连接抗原呈递细胞（APC）上的MHC II分子和T细胞受体（TCR）的Vβ链，直接刺激T细胞释放大量促炎细胞因子。这可能会导致毒性、使人虚弱和致命的影响。目前，对于SEB暴露没有预防措施，只有支持性疗法。
为了开发一种对抗SEB暴露的潜在治疗候选物，我们制备了三种产生抗SEB人单克隆抗体（HuMAbs）的人B细胞杂交瘤。对这些HuMAbs进行了特异性、亲和力以及在体外阻断SEB活性及其在体内致死效应能力的筛选。
通过BiaCore分析确定，高亲和力HuMAbs对SEB具有特异性，通过ELISA对相关毒素的交叉反应最小。在免疫印迹实验中，我们的HuMAbs与细胞裂解物中混合的SEB结合，而不与任何裂解物蛋白结合。在基于细胞的体外试验中，这些HuMAbs能够高效抑制原代人淋巴细胞中SEB诱导的促炎细胞因子（INF-γ和TNF-α）的分泌。在体内脂多糖增强的小鼠模型中，我们的先导抗体HuMAb-154能够中和高达100μg的SEB攻击，相当于报告的半数致死量（LD50，0.2μg）的500倍，保护小鼠免于死亡。在SEB攻击后给予HuMAb-154时也观察到了生存期延长。
我们制备了高亲和力的SEB特异性抗体，其能够在体外以及小鼠模型体内中和SEB。综上所述，这些结果表明我们的抗体具有作为被动免疫疗法用于SEB暴露的预防和治疗对策的潜力。
