OXIDIZED DNA BASES AS BIOMARKERS OF TUMOR PROGRESSION

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 早期的致癌理论认为，DNA受到环境因素的破坏，从而导致DNA密码的变化。这些变化被称为突变。一旦积累了足够多的突变，细胞就不能再正常运作，癌症就会出现。吸烟会导致肺癌，阳光暴晒会导致皮肤癌，这两个事实都支持这一理论。然而，这种直接的环境联系在人类乳腺癌中并不明显。一项更新的理论表明，氧气的新陈代谢会产生自由基，从而导致致癌的DNA损伤。为了验证后一种理论，有必要测量细胞内自由基损伤的产物。活性氧对细胞造成的损害称为氧化。在一些乳腺癌中，氧化DNA损伤水平的增加与肿瘤的进展有关。然而，最常用的DNA氧化标记8-oxoG已知在化学上不稳定，很难准确测量。重要的是，8-oxoG本身很容易氧化形成几个二次氧化产物。我们建议开发新技术，允许在大鼠和人类乳腺癌细胞中检测这些产品。这种化验方法可以作为诊断的基础，以便在未来的人类临床和流行病学研究中使用。这些生物标志物的另一个好处是它们最终将应用于动物和人类的化学预防。 作为一种高通量测量方法，AMS是一种理想的技术，因为它具有敏感度和稳健性。相比之下，较少定量的MS方法，如FT-MS，在常规操作中缺乏AMS的精密度，特别是在每个细胞几个DNA加合物的预期水平上。这两种技术的结合将使蛋白质多肽的分离和检测达到或低于attomole水平，这对于小的法医样本是必要的。这个项目是对AMS RR赠款项目3的补充，特别是关于使用AMS进一步了解基于DNA的标记物的癌症病因。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. An early theory of carcinogenesis proposes that DNA is damaged by environmental factors, which lead to changes in the DNA code. These changes are called mutations. Once enough mutations accumulate, cells can no longer function properly, and cancer arises. The facts that smoking causes lung cancer, and that sunlight exposure causes skin cancer support this theory. However, such a direct environmental link is not evident for breast cancer in humans. A more recent theory suggests that the metabolism of oxygen produces free radicals that cause carcinogenic DNA damage. In order to validate the latter theory, it is necessary to measure the products of free radical damage in cells. Damage to cells caused by reactive oxygen is called oxidation. In some breast cancers, increased levels of oxidative DNA damage have been associated with tumor progression. However, the most commonly used marker of DNA oxidation, called 8-oxoG, is known to be chemically unstable, and is difficult to measure accurately. Importantly, 8-oxoG is itself easily oxidized to form several secondary oxidation products. We propose to develop new technology that will allow detection of the products in rat and human breast cancer cells. Such an assay can be used as a foundation for making diagnostics for use in future human clinical and epidemiological studies. An additional benefit of these biomarkers will be their eventual application to chemoprevention in animals and humans. AMS is a desirable technique due to its sensitivity and robustness as a method for high-throughput measurement. In comparison, less quantitative MS methods such as FT-MS lack the precision of AMS for routine operations, especially at the expected level of a few DNA adducts per cell. The combination of these two techniques will allow for the fractionation and detection of protein polypeptides at or below attomole levels, which is necessary for small forensic samples. This project is complementary aims to those of Project 3 of the AMS RR grant, particularly with regard to the use of AMS to further understand cancer etiology using DNA-based markers.