背景:人类嗜T淋巴细胞病毒1(HTLV-1)与人类多种病理和慢性感染的发展有关。可用治疗的低效率和开发保护性疫苗的挑战突出了生产有效的免疫治疗工具的需要。HTLV-1碱性亮氨酸拉链(bZIP)因子(HBZ)在HTLV-1持久性中起着重要作用,通过减少HTLV-1蛋白表达赋予感染细胞生存优势,允许被感染的细胞逃避免疫监视,和增强细胞增殖导致增加的前病毒负荷。
方法:我们已经产生了重组修饰的安卡拉病毒牛痘(MVA-HBZ)和表达基于HBZ肽的多表位蛋白的质粒DNA(pcDNA3.1(+)-HBZ),以研究该病毒衍生蛋白在BALB/c小鼠模型中的免疫原性潜力。以初免-加强异源方案免疫小鼠,并通过流式细胞术对其脾细胞(TCD4和TCD8)进行免疫表型分析,并使用原核载体中产生的HBZ蛋白作为抗原通过ELISA评估体液应答。
结果:HBZ肽(HBZ42-50和HBZ157-176)刺激的TCD4+和TCD8+淋巴细胞对TNF-α/IFN-γ表现出多功能双阳性反应,和TNF-α/IL-2。此外,TCD8+细胞在产生颗粒酶B(CD44+高/CD62L-低)的效应记忆细胞的激活中呈现趋势,通过效应记忆T细胞产生颗粒酶B和CD8T细胞表达CD107a来推断免疫小鼠中细胞毒性T淋巴细胞(CTL)的激活和细胞毒性反应。总体数据与指令和效应器召回响应一致,它可能能够在消除HTLV-1感染的细胞中积极运作,因此,减少前病毒负荷。来自免疫小鼠的血清,与对照动物不同,通过ELISA显示IgG-抗-HBZ产生。
结论:我们的结果强调了从质粒DNA和痘病毒载体表达的HBZ多表位蛋白作为治疗性疫苗候选物的潜力。
Human T-lymphotropic virus 1 (HTLV-1) is associated with the development of several pathologies and chronic infection in humans. The inefficiency of the available treatments and the challenge in developing a protective vaccine highlight the need to produce effective immunotherapeutic tools. The HTLV-1 basic leucine zipper (bZIP) factor (
HBZ) plays an important role in the HTLV-1 persistence, conferring a survival advantage to infected cells by reducing the HTLV-1 proteins expression, allowing infected cells to evade immune surveillance, and enhancing cell proliferation leading to increased proviral load.
We have generated a recombinant Modified Virus Vaccinia Ankara (MVA-
HBZ) and a plasmid DNA (pcDNA3.1(+)-
HBZ) expressing a multiepitope protein based on peptides of
HBZ to study the immunogenic potential of this viral-derived protein in BALB/c mice model. Mice were immunized in a prime-boost heterologous protocol and their splenocytes (T CD4+ and T CD8+) were immunophenotyped by flow cytometry and the humoral response was evaluated by ELISA using
HBZ protein produced in prokaryotic vector as antigen.
T CD4+ and T CD8+ lymphocytes cells stimulated by HBZ-peptides (HBZ42-50 and HBZ157-176) showed polyfunctional double positive responses for TNF-α/IFN-γ, and TNF-α/IL-2. Moreover, T CD8+ cells presented a tendency in the activation of effector memory cells producing granzyme B (CD44+High/CD62L-Low), and the activation of Cytotoxic T Lymphocytes (CTLs) and cytotoxic responses in immunized mice were inferred through the production of granzyme B by effector memory T cells and the expression of CD107a by CD8+ T cells. The overall data is consistent with a directive and effector recall response, which may be able to operate actively in the elimination of HTLV-1-infected cells and, consequently, in the reduction of the proviral load. Sera from immunized mice, differently from those of control animals, showed IgG-anti-
HBZ production by ELISA.
Our results highlight the potential of the HBZ multiepitope protein expressed from plasmid DNA and a poxviral vector as candidates for therapeutic vaccine.