Protein Biomarkers of Gastric Cancer

描述(由申请人提供)：幽门螺杆菌(HP)感染了世界上50%的人口的胃粘膜。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感染的流行率很高。这些人群患胃癌的风险也最高。作为感染标志的慢性炎症被认为是胃癌发展的危险因素。感染了携带细胞毒素相关基因A的功能性形式cagA的菌株的人患胃癌的风险增加。因此，幽门螺杆菌是第一种被归类为I类致癌物质的细菌。由于胃癌是第二大致死性癌症，延误诊断会导致预后不良，因此检测生物标记物不仅可以报告感染，而且还可以检测癌症的发展，这显然是重要的。由于十二指肠溃疡疾病和胃癌是相互排斥的，我们推测，比较幽门螺杆菌感染患者和与此感染相关的不同疾病患者的胃上皮细胞蛋白质组图谱将揭示胃癌的重要疾病特异性生物标志物。为了验证这一假说，我们计划解决以下具体目标：目的1.鉴定被诊断为胃癌或十二指肠溃疡的幽门螺杆菌感染患者胃上皮细胞生物标志物的差异。未受感染的捐赠者的蛋白质图谱将被用作基线对照。在这一目标中，胃活检将被用来分离上皮细胞，其蛋白质将被溶解以用于高分辨率双向凝胶电泳法(2-DE)。使用基质辅助激光解吸电离飞行时间质谱仪(MALDI-TOF-MS)的多肽质量指纹图谱将识别在胃癌患者而在十二指肠溃疡患者中表达持续变化的生物标记物。这些蛋白表达的变化将通过特定抗体的免疫印迹和代表性组织的免疫组织化学来证实。目的2.确定幽门螺杆菌(Hp)体外感染胃上皮细胞所诱导的生物标志物。为此，将来自未感染个体的胃上皮细胞感染cagA+的幽门螺杆菌或cagA-菌株作为对照，以确定AIM 1中检测到的哪些生物标记物在感染过程的早期被诱导，从而被认为是细胞转化的早期生物标记物。公共卫生相关性：胃癌是全球第二大致死性癌症，它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，延误诊断导致了与这种癌症相关的高死亡率。该项目的目标是实施高通量技术来识别可能成为候选生物标志物的蛋白质，这些蛋白质将有助于幽门螺杆菌感染和胃癌的早期诊断。

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.