Protein Biomarkers of Gastric Cancer

描述（由申请人提供）：幽门螺杆菌（Hp）感染世界上>50%人口的胃粘膜。感染这种革兰氏阴性细菌会导致严重的胃十二指肠疾病，包括慢性胃炎，溃疡和癌症。在代表性不足的少数民族和低收入人群中，Hp感染的患病率很高。这些群体也有最高的风险发展胃癌。慢性炎症是感染的标志，被认为是胃癌发展的危险因素。感染携带细胞毒素相关基因A（cagA）功能形式的菌株的个体患胃癌的风险增加。因此，Hp是第一个被列为I类致癌物的细菌因子。由于胃癌是第二种最致命的癌症形式，诊断延误会导致预后不良，因此不仅报告感染而且还允许检测癌症发展的生物标志物的检测显然很重要。由于十二指肠溃疡疾病和胃癌是相互排斥的，我们假设比较感染Hp但与此感染相关的不同疾病患者的胃上皮细胞的蛋白质组学特征将揭示胃癌的重要疾病特异性生物标志物。为了验证这一假设，我们计划解决以下具体目标：目标1。描述被诊断为胃癌或十二指肠溃疡的Hp感染患者的胃上皮细胞生物标志物谱的差异。未感染供体的蛋白质谱将用作基线对照。在这一目标中，胃活检将用于分离上皮细胞，其蛋白质将被溶解，用于通过高分辨率二维凝胶电泳（2-DE）进行分析。将使用基质辅助激光解吸电离飞行时间MS（MALDI-TOF- MS）通过肽质量指纹图谱鉴定在胃癌患者中表达持续改变而在十二指肠溃疡患者中表达不一致的生物标志物。这些蛋白质表达的变化将通过使用特异性抗体的蛋白质印迹法进行验证，并通过代表性组织的免疫组织化学进行确认。目标2.确定幽门螺杆菌体外感染胃上皮细胞诱导的生物标志物。在该目的中，来自未感染个体的胃上皮细胞将被Hp的cagA+菌株或cagA-菌株感染作为对照，以确定在Aim 1中检测到的哪些生物标志物在感染过程的早期被诱导，并且因此可以被认为是细胞转化的早期生物标志物。 公共卫生相关性：胃癌是全球第二大致命癌症，与幽门螺杆菌感染有关。由于胃癌在早期阶段不会引起任何症状，因此诊断延误导致与这种癌症相关的高死亡率。该项目的目的是实施高通量技术来鉴定可能成为候选生物标志物特征的蛋白质，这将有助于幽门螺杆菌感染和胃癌发展的早期诊断。

DESCRIPTION (provided by applicant): Helicobacter pylori (Hp) infects the gastric mucosa of >50% of the world's population. Infection with this gram negative bacterium results in significant gastroduodenal diseases that include chronic gastritis, ulcers and cancer. Prevalence rates of Hp infection are high among underrepresented minority and lower income populations. These groups also have the highest risk of developing gastric cancer. The chronic inflammation that is a hallmark of the infection is considered a risk factor for the development of gastric cancer. Individuals infected with strains that carry the functional form of the cytotoxin-associated gene A, cagA, have an increased risk of developing gastric cancer. Hp is thus the first bacterial agent to be classified as a class I carcinogen. Since gastric cancer is the second deadliest form of cancer and delays in diagnosis contribute to poor prognosis, the detection of biomarkers that not only report the infection, but also allow detection of carcinoma development is clearly important. Since duodenal ulcer disease and gastric cancer are mutually exclusive, we hypothesize that a comparison of the proteomic profile of gastric epithelial cells of patients infected with Hp but with different diseases linked to this infection will reveal important disease-specific biomarkers of gastric cancer. To test this hypothesis we plan to address the following specific aims: Aim 1. Characterize the differences in the biomarker profile of gastric epithelial cells from patients infected with Hp who are diagnosed with gastric cancer or duodenal ulcers. The protein profile of uninfected donors will be used as a baseline control. In this aim, gastric biopsies will be used to isolate epithelial cells, whose proteins will be solubilized for profiling by high resolution two-dimensional gel electrophoresis (2-DE). Biomarkers whose expression is consistently altered in gastric cancer patients and not in duodenal ulcer patients will be identified by peptide mass fingerprinting using matrix-assisted laser desorption ionization-time-of-flight MS (MALDI-TOF- MS). Changes in expression of these proteins will be validated by western blotting with specific antibodies and confirmed by immunohistochemistry of representative tissues. Aim 2. Determine what biomarkers are induced by in vitro infection of gastric epithelial cells with Hp. In this aim, gastric epithelial cells from uninfected individuals will be infected with cagA+ strains of Hp or cagA- strains, as controls, to determine what biomarkers detected in Aim 1 are induced early in the course of infection and could thus be considered early biomarkers of cell transformation. PUBLIC HEALTH RELEVANCE: Gastric cancer is the second deadliest cancer worldwide and its linked to infection with the Helicobacter pylori bacterium. Since in its early stages gastric cancer causes no symptoms, delays in diagnosis contribute to the high mortality rate associated with this form of cancer. The objective of this project is to implement high throughput technology to identify proteins that could become candidate biomarker signatures that would aid in early diagnosis of Helicobacter pylori infection and gastric cancer development.