Antitumor Mechanisms of anti-CD40/CpG-activated Macrophages

描述（由申请人提供）：通过连接免疫系统细胞表面CD40分子来激活免疫系统细胞，可以在荷瘤小鼠中诱导T细胞依赖性和非依赖性抗肿瘤作用。我们最近发现，通过抗 CD40 单克隆抗体 (1CD40) 和 CpG 这两种免疫调节剂的组合，CD40 和 Toll 样受体 9 (TLR9​​) 连接可诱导巨噬细胞的协同激活，甚至在缺乏 T、B 和溶细胞 NK 细胞的小鼠中也能产生有效的抗肿瘤反应。由于巨噬细胞对化疗具有相对抵抗性，我们假设用 1CD40 加 CpG 疗法激活体内巨噬细胞将增强荷瘤宿主先前化疗的抗肿瘤效果。最近的初步数据支持了这一假设。我们还假设，使用由与 CCL16 趋化因子连接的肿瘤反应性单克隆抗体组成的融合蛋白将巨噬细胞靶向肿瘤微环境，将进一步增强其对已形成肿瘤的小鼠的抗肿瘤作用。我们还假设，已知的干扰肿瘤诱导的免疫抑制的策略将在 1CD40 和 CpG 治疗后实现更大的巨噬细胞介导的抗肿瘤活性。该项目的目的是表征 1CD40 CpG 激活巨噬细胞的抗肿瘤机制和治疗用途。具体来说，我们将通过以下 4 个目标来实现这一目标，其中包括小鼠巨噬细胞的体外和体内分析及其对小鼠肿瘤（特别是 B16 黑色素瘤）的影响： 1. 利用 1CD40 CpG 克服肿瘤诱导的 MF 抑制。 2. 确定特定化疗药物的预处理如何与 1CD40 CpG 激活的 MF 的抗肿瘤作用产生协同作用。 3. 确定使用 chTNT-3/LEC FP 将 1CD40 CpG-MF 定位在肿瘤中是否会产生更好的抗肿瘤效果。 4. 评估联合 1CD40 CpG 治疗的临床前策略。这项研究将确定如何通过干扰肿瘤诱导的体内免疫抑制来增强 1CD40 和 CpG 激活的巨噬细胞的抗肿瘤反应性。然后，它将确定联合治疗方案，进一步增强已患有肿瘤的小鼠的抗肿瘤作用。由于小鼠和人类巨噬细胞都可以通过 CD40 和 TLR9 激活，这些基础和临床前研究应该为未来的临床试验提供一个框架，利用这种将化疗与巨噬细胞激活和定位方式相结合的新策略。公共健康相关性：巨噬细胞是白细胞的重要衍生物，对于我们防御感染和激活保护性免疫反应至关重要。该提议基于巨噬细胞也可以被激活以介导直接抗癌作用的概念；我们假设，癌症患者巨噬细胞的适当激活具有巨大的治疗潜力，特别是与其他抗癌疗法（包括常规化疗）相结合。该提案的总体目标是表征并进一步开发这种在癌性小鼠中激活巨噬细胞的新策略，其中已经显示出有益的抗癌作用，以确定如何最好地利用它作为治疗人类癌症的未来策略。

DESCRIPTION (provided by applicant): Activation of immune system cells via ligation of their cell surface CD40 molecules can induce both T cell- dependent and independent antitumor effects in tumor-bearing mice. We have recently shown that CD40 and Toll-like receptor-9 (TLR9) ligation by the combination of two immunomodulators, anti-CD40 monoclonal antibody (1CD40) and CpG, induces synergistic activation of macrophages and results in potent antitumor responses even in mice lacking T, B and cytolytic NK cells. As macrophages can be relatively resistant to chemotherapy, we hypothesize that in vivo macrophage activation with 1CD40 plus CpG therapy will augment the antitumor effect of prior chemotherapy in tumor-bearing hosts. Recent preliminary data support this hypothesis. We also hypothesize that targeting macrophages to the tumor microenvironment using a fusion protein consisting of a tumor reactive monoclonal antibody linked to the CCL16 chemokine will further enhance their antitumor effects in mice with established tumors. We also hypothesize that strategies known to interfere with tumor induced immune-suppression will enable even greater macrophage mediated antitumor activity following 1CD40 and CpG treatment. The purpose of this project is to characterize the antitumor mechanisms and therapeutic utility of 1CD40+CpG-activated macrophages. Specifically we will accomplish this objective through the following 4 aims, which include in vitro and in vivo analyses of murine macrophages and their effects on murine tumors, particularly the B16 melanoma: 1. Utilize 1CD40+CpG to overcome tumor-induced suppression of MF. 2. Determine how pretreatment with specific chemotherapy agents synergizes with the antitumor effects of MF activated with 1CD40+CpG. 3. Determine if localizing of 1CD40+CpG-MF in the tumor using chTNT-3/LEC FP will result in better antitumor effects. 4. Evaluate preclinical strategies of combinatory 1CD40+CpG therapy. This research will determine how to augment antitumor reactivity by macrophages activated with 1CD40 and CpG by interfering with tumor induced immune suppression in vivo. It will then identify combined regimens that further enhance these antitumor effects in mice with established tumors. As both murine and human macrophages can be activated through CD40 and TLR9, these basic and preclinical studies should provide a framework for future clinical trials utilizing this novel strategy of combining chemotherapy with macrophage- activating and localizing modalities. PUBLIC HEALH RELEVANCE: Macrophages are important derivatives of white blood cells that are critical in our defense against infections and in activating protective immune responses. This proposal is based on the concept that macrophages can also be activated to mediate direct anticancer effects; we hypothesize that proper activation of macrophages in patients with cancer has great therapeutic potential, especially in combination with other anti-cancer therapies, including conventional chemotherapy. The overall goal of this proposal is to characterize and further develop this novel strategy for macrophage activation in cancerous mice, where beneficial anti-cancer effects have already been shown, in order to determine how best to use this as a future strategy for treating human cancers.