Center for Biologics Evaluation and Research, United States Food and Drug Administration, Bethesda, MD 20892, United States.
Vaccine. 2009 Dec 11;28(2):317-22. doi: 10.1016/j.vaccine.2009.10.047. Epub 2009 Oct 29.
New vaccines and novel immunization strategies are needed to improve the control of the global tuberculosis epidemic. To facilitate vaccine development, we have been creating in vitro mycobacterial intra-macrophage growth inhibition assays. Here we describe the development of an in vitro assay designed for BSL-2 laboratories which measures the capacity of vaccine-induced immune splenocytes to control the growth of isoniazid-resistant Mycobacterium bovis BCG (INH(r) BCG). The use of the INH(r) BCG as the infecting organism allows the discrimination of BCG bacilli used in murine vaccinations from BCG used in the in vitro assay. In this study, we showed that protective immune responses evoked by four different types of Mycobacterium tuberculosis vaccines [BCG, an ESAT6/Antigen 85B fusion protein formulated in DDA/MPL adjuvant, a DNA vaccine expressing the same fusion protein, and a TB Modified Vaccinia Ankara construct expressing four TB antigens (MVA-4TB)] were detected. Importantly, the levels of vaccine-induced protective immunity seen in the in vitro assay correlated with the results from in vivo protection studies in the mouse model of pulmonary tuberculosis. Furthermore, the growth inhibition data for the INH(r) BCG assay was similar to the previously reported results for a M. tuberculosis infection assay. The cytokine expression profiles at day 7 of the INH(r) BCG growth inhibition studies were also similar but not identical to the cytokine patterns detected in earlier M. tuberculosis co-culture assays. Overall, we have shown that a BSL-2 compatible in vitro growth inhibition assay using INH(r) BCG as the intra-macrophage target organism should be useful in developing and evaluating new TB immunization strategies.
需要新的疫苗和新的免疫策略来改善全球结核病流行的控制。为了促进疫苗的开发，我们一直在创建体外分枝杆菌巨噬细胞内生长抑制试验。在这里，我们描述了一种在 BSL-2 实验室中设计的体外检测方法，该方法用于测量疫苗诱导的免疫脾细胞控制异烟肼耐药牛分枝杆菌卡介苗（INH(r)BCG）生长的能力。使用 INH(r)BCG 作为感染生物体可以区分用于小鼠接种的 BCG 杆菌与用于体外检测的 BCG。在这项研究中，我们表明，四种不同类型的结核分枝杆菌疫苗[BCG、用 DDA/MPL 佐剂配制的 ESAT6/抗原 85B 融合蛋白、表达相同融合蛋白的 DNA 疫苗和表达四个结核抗原的 TB 改良痘苗安卡拉构建体（MVA-4TB）]引起的保护性免疫反应都可以被检测到。重要的是，体外检测中观察到的疫苗诱导的保护性免疫水平与肺结核小鼠模型体内保护研究的结果相关。此外，INH(r)BCG 检测的生长抑制数据与先前报道的结核分枝杆菌感染检测结果相似。INH(r)BCG 生长抑制研究第 7 天的细胞因子表达谱与之前在结核分枝杆菌共培养检测中检测到的细胞因子模式也相似但不完全相同。总体而言，我们已经表明，使用 INH(r)BCG 作为巨噬细胞内靶标生物体的 BSL-2 兼容体外生长抑制检测方法应该有助于开发和评估新的结核病免疫策略。
