INFLAMMATORY CHEMOKINE CCL3 IN THE TUMOR MICROENVIRONMENT LEADS TO ENHANCED ANTITUMOR IMMUNE DEVELOPMENT IN THE DRAINING LYMPH NODE

 描述（由申请方提供）：炎性趋化因子CCL 3（MIP-1α）和CCL 4（MIP-1β）在协调免疫淋巴结（LN）中的非随机初始CD 8 + T细胞与树突状细胞接触中非常重要，并且可以增强记忆T细胞的生成。我们推测，将CCL 3和CCL 4整合到肿瘤微环境中可以增强抗肿瘤免疫。使用小鼠结肠肿瘤模型CT 26，我们产生了分泌CCL 3或CCL 4的肿瘤，并将它们植入到幼稚免疫活性受体的足垫中。肿瘤生长动力学测量表明，CCL 3分泌肿瘤导致CD 8 + T细胞依赖性肿瘤进展的减缓或免疫活性小鼠的完全消退，并在用作疫苗时保护随后的致死性肿瘤攻击。接下来，使用荧光激活细胞分选分析（FACS），我们通过肿瘤注射后3天和5天的荧光激活细胞分选分析来分析淋巴细胞募集到引流转移性肿瘤的LN的影响。接种CCL 3分泌性肿瘤的小鼠在第3天导致CD 4+和CD 8 + T细胞的相似增加，范围为WT肿瘤的1.3至1.7倍增加和CCL 4分泌性肿瘤的1.3倍增加。到第5天，分泌CCL 3的肿瘤中的T细胞积累达到与WT和分泌CCL 4的肿瘤相似的水平。此外，动态活体双光子显微镜和组织组织学研究进一步揭示，肿瘤在早期转移期间占据B细胞滤泡，外源性CCL 3和CCL 4梯度可能潜在地改变肿瘤引流淋巴结中的淋巴细胞稳态，导致免疫激活而不是对CT 26的耐受。我们建议询问CCL 3对分泌CCL 3或CCL 4的CT 26肿瘤接种后引流LN中免疫细胞募集、活化、保留和效应子功能递送的生物学效应。这些研究包括树突状细胞和淋巴细胞在暴露于CCL 3后的功能活化、迁移和趋化因子受体表达。此外，我们建议在肿瘤接种后的前5天内研究分泌CCL 3、分泌CCL 4或野生型CT 26肿瘤细胞与肿瘤浸润性LN中的各种免疫细胞之间的动态细胞-细胞相互作用。这些研究将为探索将炎性趋化因子如CCL 3作为抗肿瘤免疫治疗的治疗性佐剂提供科学依据。

 DESCRIPTION (provided by applicant): Inflammatory chemokines CCL3 (MIP-1α) and CCL4 (MIP-1β) are important in orchestrating nonrandom naive CD8+ T cell contacts with dendritic cells in vaccinated lymph nodes (LNs) and can enhance memory T cell generation. We hypothesize that incorporating CCL3 and CCL4 into tumor microenvironment can enhance anti-tumor immunity. Using the murine colon tumor model CT26, we generated tumors that secret CCL3 or CCL4 and inoculate them live into the footpad of naïve immunocompetent recipients. Tumor growth kinetic measurements showed that CCL3-secreting tumors resulted in either a CD8+ T cell dependent slowing of tumor progression or complete regression in immunocompetent mice and protected subsequent lethal tumor challenge when used as a vaccine. Next, using fluorescence-activated cell sorting analysis (FACS) we analyzed the effects of lymphocyte recruitment to the LN draining the metastatic tumor by fluorescence- activated cell sorting analysis 3 and 5 days post-tumor injection. Mice inoculated with CCL3-secreting tumors resulted in similar increases in CD4+ and CD8+ T cells ranging from 1.3 to 1.7-fold increase from WT tumors and 1.3-fold increase from CCL4-secreting tumors on day 3. By day 5 the T cell accumulation in CCL3- secreting tumors reached similar levels as that of WT and CCL4-secreting tumors. Furthermore, dynamic intravital 2-photon microscopy and tissue histology studies further revealed that tumors occupy the B-cell follicle during early metastasis, and exogenous CCL3 and CCL4 gradient may potentially alter lymphocyte homeostasis in tumor draining LNs, resulting in immune activation rather than tolerance against CT26. We propose to interrogate biological effect of CCL3 on immune cell recruitment, activation, retention and effector functional delivery in the draining LNs following CCL3- or CCL4-secreting CT26 tumor inoculation. These studies include functional activation, migration and chemokine receptor expression of dendritic cells and lymphocytes upon exposure to CCL3. Furthermore, we propose to study the dynamic cell-cell interaction between CCL3-secreting, CCL4-secreting, or wild type CT26 tumor cells and various immune cells in the tumor-infiltrating LN within the first 5 days following tumor inoculation. These studies will provide scientific rationale for the exploration of incorporating inflammatory chemokines such as CCL3 as a therapeutic adjuvant to anti-tumor immunotherapy.