Role of Nitric Oxide in the Progression of Murine Cancer

体内、外多种恶性肿瘤组织中氧自由基、一氧化氮(NO)含量均升高。一氧化氮合酶(NOS)的三种同工酶负责在生物体内从精氨酸合成NO：构成神经型NOS(nNOS或NOS1)，主要存在于神经元中；诱导型NOS(iNOS或NOS2)，存在于巨噬细胞和其他细胞中；以及构成内皮细胞NOS(eNOS，ecNOS或NOS3)，主要存在于内皮细胞中。在许多形式的癌症中，NO水平的升高与肿瘤侵袭性的增加有关。在大多数被检查的乳腺癌中发现NO水平升高，并与血管生成加速、细胞凋亡减少和肿瘤侵袭有关。在目前的R03应用中，我们建议阐明NO生成在乳腺癌细胞侵袭特性中的作用。我们假设，NO水平的增加(由NOS活性增加产生)确实与肿瘤侵袭性的增强有关。为了直接研究一氧化氮合酶在细胞中的作用，我们将研究致癌物7-12-二甲基苯并(A)菲(DMBA)在iNOS和eNOS基因敲除小鼠中诱导的小鼠乳腺癌。这些模型将使我们能够确定与肿瘤侵袭性最相关的一氧化氮合酶类型。我们还将通过使用一氧化氮合酶抑制剂改变细胞环境中一氧化氮的量来检验我们的假设。如果NO在这些过程中是一个重要的积极因素，则NG-硝基-L-精氨酸甲酯(L-NAME)或NG-甲基-L-精氨酸(NMMA)对一氧化氮合酶活性的抑制应该可以减少肿瘤的生长和转移。最后，我们预计添加不需要一氧化氮合酶来产生的外源性NO(例如，给予硝普钠或NOC-12[1-羟基-2-氧代-3-(N-乙基-2-氨基乙基)-3-乙基-L-三嗪]将增加肿瘤的侵袭性。我们预计，这些初步研究的结果将为我们提供足够的数据，为随后的R01应用奠定基础，探索NO和NOS在乳腺癌中的作用机制，最终转化为人类疾病，可能在生物标志物和它们的治疗干预方面。

The free oxygen radical, nitric oxide (NO), is elevated in a variety of malignant tumors in vitro and in vivo. Three isoenzymes of nitric oxide synthase (NOS) are responsible for the synthesis of NO from arginine in the organism: constitutive neuronal NOS (nNOS or NOS 1), identified mainly in neurons; inducible NOS (iNOS or NOS 2), found in macrophages and other cells; and constitutive endothelial cell NOS (eNOS, ecNOS, or NOS 3), found predominantly in endothelial cells. Elevated levels of NO have been associated with increased tumor aggressiveness in many forms of cancer. Increased NO levels have been found in most breast cancers examined and are associated with accelerarion of angiogenesis, abrogation of apoptosis, and tumor invasion. In the present R03 application, we propose to elucidate the role of NO generation in the invasive characteristics of breast cancer cells. We hypothesize that increased levels of NO (produced by increased NOS activity) are, indeed, associated with enhanced tumor aggressiveness. To directly examine the role of NOS in cells, we will study murine mammary cancers induced by the carcinogen 7-12- dimethybenz(a)anthracene (DMBA) in iNOS and eNOS knockout mice. These models will enable us to identify the type of NOS that is most associated with tumor aggressiveness. We also will test our hypothesis by chemicall altering the amount of NO in the cellular environment using NOS inhibitors. Inhibition of NOS activity by NG-nitro-L- arginine methyl ester (L-NAME) or by NG-methyl-L-arginine (NMMA) should reduce tumor growth and metastases if NO is a significant positive factor in these processes. Finally, we anticipate that addition of exogenous NO that does not require NOS for its production (e.g., administration of sodium nitroprusside or NOC-12 [1hydroxy-2-oxo-3- (N-ethyl-2-aminoethyl)-3-ethyl-l-triazine] will increase tumor invasiveness. We anticipate that the results obtained from these preliminary studies will provide us with sufficient data to lay the foundation for a subsequent R01 application exploring the mechanisms involved in the actions of NO and NOS in breast cancer, with ultimate translation to human disease, potentially in terms of both biological markers and them therapeutic interventions.