New Approaches to Prevent or Treat Breast Cancer Metastases

描述（由申请人提供）： 大约40%的被诊断患有乳腺癌的妇女将进展为转移性疾病。然而，转移瘤不能通过手术或放射去除，并且大多数转移瘤是化学抗性的。因此，专门预防或消除转移的疗法可以带来很大的希望。最近的研究表明，通过疫苗接种增强对乳腺肿瘤的特异性辅助或细胞毒性T淋巴细胞（CTL）反应可能导致特异性消除微转移。然而，疫苗诱导的免疫和临床应答较差，这突出表明需要改进疫苗策略。该研究计划的重点是使用临床前转移性小鼠乳腺肿瘤模型4 T1，通过将疫苗接种与IL-6抑制剂姜黄素相结合来改善针对转移性乳腺癌的疫苗治疗。我们先前证明，在该模型中，Mage-b DNA疫苗接种提供了显著的预防乳腺癌转移的保护，尽管不是完全的。白细胞介素（IL）-6可能导致疫苗功效下降。它在4 T1原发性肿瘤和转移瘤中高度上调，可能通过核因子κ B（NF κ B），并且是体内树突状细胞（DC）分化的有效调节剂。IL-6激活未成熟DC中的转导子和转录激活子（STAT）3的表达，阻止DC成熟和随后向T细胞呈递抗原。这可能导致T细胞无反应性。能够抑制IL-6的药剂可导致增强的疫苗功效。姜黄素可能是这样一种试剂，因为它抑制IL-6的产生。Mage-b和IL-6在人类乳腺癌中经常被检测到，这使得4 T1模型具有临床意义。该提案的长期目标是在4 T1模型中开发Mage-b疫苗接种和姜黄素的新型无毒组合疗法，如果成功，重点是将其转化为人类临床试验。该假说是姜黄素改善了T细胞活化和随后的免疫应答，从而进一步降低了乳腺癌转移的频率。具体目的如下：（1）检测含和不含姜黄素的Mage-b疫苗接种对肿瘤和转移的影响。为此，将用Mage-b DNA疫苗对小鼠进行预防性或治疗性免疫，并用4 T1肿瘤细胞进行攻击。一旦原发性肿瘤可以感觉到姜黄素将每天给药。将确定转移的频率、肿瘤大小和存活时间;（2）测试姜黄素对疫苗和肿瘤诱导的免疫应答以及对肿瘤细胞的增殖和凋亡的直接作用。为此，将分析处理和对照小鼠的脾脏、淋巴结、肿瘤和转移灶中是否存在活化的CD 4和CD 8 T细胞、调节性T细胞和成熟DC。此外，还将分析治疗小鼠和对照小鼠的肿瘤和转移灶的NF κ B和NF κ B调节基因（直接参与免疫应答，如IL-6、TNF 1和STAT 3）的表达，以及增殖和凋亡。公共卫生相关性：目前的癌症治疗对转移无效。在这项研究提案中，将开发一种新型的Mage-b疫苗接种和姜黄素的无毒联合疗法，该疗法可有效对抗乳腺癌转移，使用反映人类转移性乳腺癌的临床前小鼠模型。

DESCRIPTION (provided by applicant): About 40% of the women diagnosed with breast cancer will progress to metastatic disease. However, metastases cannot be removed by surgery or radiation, and most metastases are chemoresistant. Therefore, therapies that specifically prevent or eliminate metastases offer great promise in the outcome. Recent studies indicate that enhancement of specific helper or cytotoxic T lymphocyte (CTL) responses to breast tumors through vaccination could potentially lead to the specific elimination of micro-metastases. However, poor vaccine-induced immune and clinical responses underscore the need for improved vaccine strategies. This research proposal is focused on the improvement of vaccine therapy against metastatic breast cancer by combining vaccination with the IL-6 inhibitor curcumin, using the preclinical metastatic mouse breast tumor model 4T1. We previously demonstrated that Mage-b DNA vaccination provides significant protection against breast cancer metastases in this model, albeit not completely. Interleukin (IL)-6 may contribute to decreased vaccine efficacy. It is highly up regulated in the 4T1 primary tumors and metastases, probably via the Nuclear Factor Kappa B (NFkB), and is a potent regulator of dendritic cells (DC) differentiation in vivo. IL-6 activates the expression of transducer and activator of transcription (STAT)3 in immature DC, preventing the DC from maturation and subsequent presentation of antigens to T cells. This may lead to T cell unresponsiveness. Agents that are able to inhibit IL-6 may lead to enhanced vaccine efficacy. Curcumin could be such an agent, since it inhibits IL-6 production. Both Mage-b and IL-6 are frequently detected in human breast cancer, which makes the 4T1 model clinically relevant. The long-term objective of this proposal is to develop a novel non- toxic combination therapy of Mage-b vaccination and curcumin in the 4T1 model, with a focus on its translation into human clinical trials, if successful. The hypothesis is that curcumin improves T cell activation and subsequent immune responses upon Mage-b DNA vaccination, resulting in further reduction of the frequency of breast cancer metastases. The specific aims are as follows:(1) Testing the effect of Mage-b vaccination with and without curcumin on tumors and metastases. For this purpose, mice will be immunized preventively or therapeutically with Mage-b DNA vaccine and challenged with 4T1 tumor cells. As soon as the primary tumor can be felt curcumin will be administered daily. Frequency of metastases, tumor size, and survival times will be determined; (2) Testing the direct effect(s) of curcumin on vaccine- and tumor-induced immune responses, and on proliferation and apoptosis of tumor cells. For this purpose, spleen, lymph nodes, tumors and metastases of treated and control mice will be analyzed for the presence of activated CD4 and CD8 T cells, regulatory T cells and mature DC. In addition, tumors and metastases of treated and control mice will also be analyzed for the expression of NFkB and NFkB-regulated genes that are directly involved in immune responses such as IL-6, TNF1, and STAT3, and for proliferation and apoptosis. PUBLIC HEALTH RELEVANCE: Current treatment of cancer is ineffective against metastases. In this research proposal a novel non-toxic combination therapy of Mage-b vaccination and curcumin will be developed that is effective against breast cancer metastases, using a preclinical mouse model that reflects metastatic breast cancer in humans.