Role of the chronic inflammatory mediator NO in early mammary tumorigenesis

描述（由申请人提供）：由病毒或细菌感染引起的慢性炎症与大约15%的癌症相关人类死亡有关。转录因子NF-kappaB是这一过程的关键介质，受促炎分子和基因毒性应激的调节。NF-kappaB可能启动肿瘤形成的几种机制之一是通过诱导诱导型一氧化氮合酶（iNOS）导致NO的产生，NO是抑制T细胞免疫和产生DNA损伤活性氮氧化物（RNOS）的重要分子。髓系抑制细胞（MSCs）和肿瘤相关巨噬细胞（tam）是产生NO并介导这些作用的髓系细胞，可浸润实体肿瘤。然而，慢性炎症介质iNOS影响乳腺癌的机制尚未阐明。我们的模型，在C57BI/6小鼠乳腺上皮中表达的多瘤病毒中T癌基因（PyV-mT）被用来证明iNOS促进肿瘤发生，这一发现与人类研究一致，将iNOS表达增加与乳腺癌患者预后不良以及抗炎治疗与乳腺癌发病率降低相关联。具体假设是，慢性炎症期间募集到乳腺的骨髓细胞产生inos诱导的NO，抑制T细胞免疫并导致乳腺上皮细胞DNA损伤，这是癌症发生的重要步骤。针对这一假设的具体目的是：1)通过建立骨髓特异性iNOS基因消融模型，研究骨髓细胞iNOS诱导NO产生对乳腺肿瘤发生的影响；ii)在野生型和iNOS-/- PyV-mT小鼠中表征慢性炎症细胞；iii)进行相互移植实验；iv)骨髓移植实验；2)通过i)髓细胞和上皮细胞共培养和ii)检测p53 DNA损伤途径来评估inos诱导的乳腺上皮细胞DNA损伤。该实验设计将使我们能够确定髓系iNOS诱导NO在乳腺肿瘤发生起始中的作用。我们还希望揭示慢性炎症影响乳腺癌发展机制的新见解，并为使用靶向iNOS/NO途径的治疗方法提供分子基础。

DESCRIPTION (provided by applicant): Chronic inflammation due to viral or bacterial infections has been linked with approximately 15% of cancer-related human deaths. The transcription factor NF-kappaB, a key mediator of this process, is regulated by pro-inflammatory molecules and genotoxic stress. One of several mechanisms by which NF-kappaB may initiate tumor formation is through induction of the inducible nitric oxide synthase (iNOS) leading to production of NO, a molecule important in the suppression of T cell immunity and production of the DNA damaging reactive nitrogen oxide species (RNOS). Myeloid suppressor cells (MSCs) and tumor associated macrophages (TAMs), cells of the myeloid lineage that produce NO and can mediate these effects, infiltrate solid tumors. However mechanisms by which the chronic inflammatory mediator iNOS impacts breast cancer have not been elucidated. Our model, polyomavirus middle T oncogene (PyV-mT) expressed in the mammary epithelium of C57BI/6 mice was used to demonstrate that iNOS promotes tumorigenesis, a finding consistent with human studies associating increased iNOS expression with poor prognosis in breast cancer patients, and anti-inflammatory treatment with reductions in the incidence of breast cancer. The specific hypothesis is that myeloid cells recruited to the mammary gland during chronic inflammation produce iNOS-induced NO that inhibits T cell immunity and causes DNA damage in mammary epithelial cells, important steps in the initiation of cancer. The specific aims to address this hypothesis are: 1) Examine the impact of myeloid cell iNOS induced NO production on mammary tumorigenesis by i) creating a myeloid specific iNOS gene ablation model, ii) characterizing chronic inflammatory cells in wild type and iNOS-/- PyV-mT mice, iii) carrying out reciprocal transplantation experiments, and iv) bone marrow transplantation experiments; 2) Assess iNOS-induced DNA damage to mammary epithelial cells by i) co-culture of myeloid cells and epithelial cells and ii) examination of the p53 DNA damage pathway. This experimental design will enable us to determine the role of myeloid iNOS induced NO in the initiation of mammary tumorigenesis. We also expect to reveal new insights into mechanisms by which chronic inflammation can influence the development of breast cancer and provide a molecular rationale for the use of therapeutics targeting the iNOS/NO pathway.