Immuno-gene therapy by secreted gp96-Ig fusion protein

Gp96具有作为肿瘤疫苗产生CD8CTL的特性，不受MHC限制。我们开发了一种分泌型gp96-Ig(gp96-Ig)，方法是删除内质网保留信号KDEL，并用小鼠免疫球蛋白1的Fc部分取代它。与野生型肿瘤相比，高表达gp96-Ig的肿瘤细胞致瘤性较低。对转gp96-Ig(E.G7-gp96-Ig)的小鼠淋巴瘤E.G7(E.G7-gp96-Ig)免疫小鼠，可诱导对E.G7后续攻击的免疫。耗竭研究表明，在整个诱导期和效应期，E.G7-gp96-Ig排斥反应都需要CD8^+细胞，而不需要CD8^+细胞或巨噬细胞。在本研究中，我们检测了gp96-Ig基因转导的肿瘤细胞的免疫治疗效果。在C57BL/6小鼠体内建立E.G7，持续3天。与…相比，从第4天开始每天注射gp96-Ig转导的肿瘤细胞显著抑制了肿瘤的生长更多的是PBS治疗或对照肿瘤治疗，对照肿瘤分泌gp96-Ig。在肿瘤移植后第5天开始接种疫苗，成功的治疗需要每天两次注射E.G7-gp96-LG的疫苗接种计划。在荣格肿瘤模型中。LLC、LLC-gp96-Ig疫苗与从LLC细胞中纯化的gp96的效果相同，与IL-12转导的肿瘤细胞(LLC/1L12)的效果相当。LLC-gp96-Ig和LLC/IL12 1：1混合疫苗没有协同作用，提示这两种疫苗可能通过相似的细胞机制发挥作用。Gp96-Ig转导的肿瘤细胞疫苗不需要CD4^+T细胞。用gp96-Ig转导的肿瘤细胞疫苗诱导CD8~+细胞对野生型肿瘤细胞的杀伤活性。过继转移的卵清蛋白特异性T细胞受体(TCR)转基因CD8^+细胞(OT-1)对EG7-gp96-Ig免疫具有克隆性扩增作用。我们的数据表明，分泌gp96-Ig的肿瘤可能是有效的抗肿瘤疫苗。较少

Gp96 has useful properties as tumor vaccine for the generation of CD8 CTL independent of MHC restriction. We have developed a secreted form of gp96-Ig (gp96-Ig) by deleting the endoplasmic reticulum retention signal, KDEL, and replacing it with the Fc portion of murine IgG_1. Tumor cells transfected with and overexpressing gp96-Ig were less tumorigenic, compared to wild type tumors. Immunization of mice with the murine lymphoma E.G7 transfected with gp96-Ig (E.G7-gp96-Ig), but not with irradiated E.G7, induced immunity to subsequent challenge with E.G7. Depletion studies showed that CD8^+ cells were required for E.G7-gp96-Ig rejection throughout induction phase and effector phase, but not CD4^+ cells or macrophages. In this study, we examined the immunotherapeutical potency by gp96-Ig gene-transfected tumor cells. E.G7 was established in C57BL/6 mice for three days. Daily injections, beginning on day 4, of gp96-Ig-transduced tumor cells suppressed tumor growth remarkably when compared … More to PBS treatment or to treatment with control tumors treatment with control tumors secreting gp96-Ig. Beginning vaccination on day 5 after tumor transplantation, successful therapy required a vaccination schedule of two daily injections of E.G7-gp96-lg. In a Jung tumor model. LLC, LLC-gp96-Ig vaccines were equally effective as gp96 purified from LLC cells and comparable to IL-12-transduced tumor cells (LLC/1L12). There were no synergistic effects by a 1:1 mixture of LLC-gp96-Ig and LLC/IL12, suggesting that the two vaccines may act through similar cellular mechanisms. CD4^+ T cells were not required for vaccines by gp96-Ig-transduced tumor cells. CD8^+ cytotoxic activities specific for wild type tumor cells were induced by vaccines with gp96-Ig-transduced tumor cells. Moreover, adoptively transferred, ovalbumin specific T-cell receptor (TCR) transgenic CD8^+ cells (OT-1) responded with clonal expansion to the immunization with EG7-gp96-Ig. Our data suggest that tumors secreting gp96-Ig may be useful as potent anti tumor vaccines. Less