The connection of innate and adaptive anti-cancer immunity

利用重组鸡痘病毒靶向递送小鼠干扰素(IFN)- γ：自然杀伤（NK）细胞募集到局部淋巴结和启动肿瘤特异性宿主免疫。ifn - γ是一种促炎细胞因子，也是一种有效的免疫调节剂。在这项研究中，我们设计了一种复制缺陷重组禽痘病毒来表达小鼠ifn - γ基因（nf - muifn - γ），其基本原理是将浓缩水平的细胞因子递送到局部组织微环境中。皮下给药rf - muifn - γ导致ifn - γ的产生仅限于注射部位的组织微环境，并且(2)具有生物活性，在给药rf - muifn - γ后，sc生长肿瘤中I类主要组织相容性复合体（MHC）表达水平显著增加。用rf - muifn - γ感染高致瘤性小鼠细胞系MC38，可作为一种有效的肿瘤细胞疫苗，保护小鼠免受原发肿瘤的形成和随后的肿瘤攻击。如果小鼠接种了经rifn - γ预处理的MC38细胞或感染了野生型禽痘病毒（FP-WT），则细胞基疫苗完全无效。对注射rf - muifn - γ肿瘤细胞疫苗部位的局部淋巴结的分析显示，存在肿瘤特异性细胞裂解（CTL），以及针对nk敏感的YAC-1细胞的大量裂解。流式细胞分析结合功能分析证实，注射rf - muifn - γ后，NK1.1（+）细胞在引流淋巴结内持续存在长达5天。在注射rf - muifn - γ感染的MC38肿瘤细胞之前，用NK细胞消耗抗体处理的小鼠没有受到原发肿瘤生长的保护；对nk耗竭小鼠注射部位引流的淋巴结分析显示，肿瘤特异性CTL活性随之丧失。这些发现提供了证据，证明NK细胞，以其对宿主先天免疫的贡献而闻名，也提供了适应性免疫反应发展所需的免疫调节信号，这反过来又保护接种疫苗的小鼠免受肿瘤生长。干扰素- α (IFN-α)是一种多效性细胞因子，具有免疫调节特性，可以提高治疗性癌症疫苗的疗效。该项目的目的是评估重组IFN-&amp；#945；用痘病毒疫苗靶向人癌胚抗原（CEA）在小鼠结肠直肠癌和胰腺腺癌模型中，其中CEA是一种自身抗原。IFN-&#945；在无肿瘤小鼠腹股沟引流淋巴结中进行评价。我们研究了IFN-&amp；#945；给药（局部与远端）抗原特异性免疫应答对痘病毒疫苗。机制研究评估了IFN-a和CEA导向痘病毒疫苗对荷瘤CEA转基因小鼠的疗效。我们确定了IFN-&amp；#945；诱导腹股沟引流淋巴结局部扩张，改善细胞毒性（自然杀伤细胞和CD8+）和抗原呈递。给药IFN-&amp；#945；远端在接种部位的远端IFN-&#945；在CEA+腺癌小鼠中，疫苗抑制肿瘤生长，提高生存率，并引发CEA特异性CTL反应。在患有胰腺肿瘤的小鼠中，IFN- &#945；减缓肿瘤生长，诱导CTL活性，增加CD8+肿瘤浸润淋巴细胞。这些数据表明IFN- &amp；#945；可作为抗原导向痘病毒疫苗的生物反应修饰剂，产生显著的治疗性抗肿瘤免疫反应。这项研究为支持这种免疫治疗策略在cea表达癌患者中的临床试验提供了理论基础和机制见解。

Targeted delivery of murine interferon (IFN)-gamma using a recombinant fowlpox virus: natural killer (NK) cell recruitment to regional lymph nodes and priming of tumor-specific host immunity. IFN-gamma is a proinflammatory cytokine that also acts as a potent immunomodulatory agent. In this study, a replication-deficient recombinant avian (fowlpox) virus was engineered to express the murine IFN-gamma gene (rF-MuIFN-gamma) with the rationale of delivering concentrated levels of the cytokine to a local tissue microenvironment. Subcutaneous (s.c.) rF-MuIFN-gamma administration resulted in IFN-gamma production that was restricted to the tissue microenvironment of the injection site and (2) was biologically active, as evidenced by a significant increase of class I major histocompatibility complex (MHC) expression levels in s.c. growing tumors following rF-MuIFN-gamma administration. Infection of a highly tumorigenic murine cell line, MC38, with rF-MuIFN-gamma functioned as an effective tumor cell-based vaccine by protecting mice from the formation of primary tumors and from subsequent tumor challenge. The cell-based vaccine was completely ineffective if mice were vaccinated with MC38 cells either pretreated with rIFN-gamma or infected with the wild-type fowlpox virus (FP-WT). Analysis of the regional lymph nodes draining the site of injection of the rF-MuIFN-gamma-based tumor cell vaccine revealed the presence of tumor-specific cell lysis (CTL) as well as a significant amount of lysis directed at NK-sensitive YAC-1 cells. Flow cytometric analyses coupled with functional assays confirmed the sustained presence of NK1.1(+) cells within those draining lymph nodes for up to 5 days after rF-MuIFN-gamma injection. Mice treated with NK cell-depleting antibodies prior to the injection of the rF-MuIFN-gamma-infected MC38 tumor cells were not protected from primary tumor growth; analysis of the lymph nodes draining the injection site in NK-depleted mice revealed an accompanying loss of the tumor-specific CTL activity. The findings provide evidence that NK cells, known for their contributions to host innate immunity, also provide immunoregulatory signals required for the development of an adaptive immune response, which, in turn, protected vaccinated mice against tumor growth. Interferon-alpha (IFN-α) is a pleiotropic cytokine possessing immunomodulatory properties that may improve the efficacy of therapeutic cancer vaccines. The aim of this project was to evaluate the effectiveness and compatibility of combining recombinant IFN-α with poxvirus vaccines targeting the human carcinoembryonic antigen (CEA) in murine models of colorectal and pancreatic adenocarcinomas, where CEA is a self-antigen. The phenotypic and functional effects of IFN-α were evaluated in the draining inguinal lymph nodes of tumor-free mice. We studied the effect of the site of IFN-α administration (local versus distal) on antigen-specific immune responses to poxvirus vaccination. Mechanistic studies were conducted to assess the efficacy of IFN-a and CEA-directed poxvirus vaccines in tumor-bearing CEA transgenic mice. We identified a dose and schedule of IFN-α that induced a locoregional expansion of the draining inguinal lymph nodes and improved cellular cytotoxicity (natural killer and CD8+) and antigen presentation. Suppression of the vaccinia virus was avoided by administering IFN-α distal to the site of vaccination. The combination of IFN-α and vaccine inhibited tumor growth, improved survival, and elicited CEA-specific CTL responses in mice with CEA+ adenocarcinomas. In mice with pancreatic tumors, IFN- α slowed tumor growth, induced CTL activity, and increased CD8+ tumor-infiltrating lymphocytes. These data suggest that IFN- α can be used as a biological response modifier with antigen-directed poxvirus vaccines to yield significant therapeutic antitumor immune responses. This study provides the rationale and mechanistic insights to support a clinical trial of this immunotherapeutic strategy in patients with CEA-expressing carcinomas.