Dysregulated Polyamine Metabolism in H. pylori-associated Gastric Inflammation and Disease Progression

胃幽门螺杆菌感染与慢性胃炎、消化性疾病的临床后果 溃疡病，以及疾病进展到胃癌，仍然是退伍军人的主要健康问题。这 病原体感染了世界上一半的人口，而胃癌是导致癌症死亡的第三大原因 全世界。幽门螺杆菌在退伍军人中的流行率很高；部署相关感染是一个问题，特别是 获得与致癌风险较高相关的菌株。三种多胺，腐胺(Put)， 亚精胺(Spd)和精胺(Spm)是普遍存在的具有多种生物效应的分子。看跌期权是综合的 通过限速酶鸟氨酸脱羧酶(ODC)，依次转化为Spd和Spm， 它被精胺氧化酶(SMOX)反向转化为SPD。我们的实验室长期以来一直对 多胺在胃肠道炎症和癌变中的作用，我们有以下与此相关的令人兴奋的发现 续签补助金：1)感染幽门螺杆菌的Smox-/-小鼠胃部炎症、DNA损伤和 与Spd耗竭相关的趋化因子表达；2)这些表型在幽门螺杆菌- 来自Smox-/-小鼠的2D类有机物感染；3)幽门螺杆菌诱导的致癌活性之间的新联系 B-连环蛋白和Smox的研究，包括发现Smox-/-小鼠和用Smox抑制剂治疗的沙土鼠有丢失 活体内b-连环蛋白的激活；4)使用人胃器官培养来证明a 新型高效SMOX抑制剂阻断幽门螺杆菌诱导的SPD相关β-连环蛋白激活 耗竭；5)一种独特的蛋白质翻译形式，即通过脱氧亚硫氨酸的作用而产生的激素化 合成酶(DHPs)，一个与癌症发生有关的过程，似乎被Smox减弱 6)全外显子组测序(WES)显示胃炎和胃炎的体细胞基因组异常。 幽门螺杆菌致癌模型INS-GAS小鼠异型增生组织。我们现在将使有效的 使用重要的分子工具，包括有价值的突变小鼠模型、关键的SMOX抑制剂和高级 使用来自小鼠和VA患者的3D和2D胃有机化合物。我们假设调节失调的多胺 新陈代谢，由于SMOX和相关的SPD和催产素的产生，提供了一个分子 导致胃病进展为癌变风险的途径。我们的具体目标是：1) 直接确定SMOX和Spd在胃癌发生中的作用。我们将测试：a)SMOX在 易患癌症的INS-GAS小鼠，分析幽门螺杆菌诱导的致癌作用，DNA损伤，b-连环蛋白激活， 和GEC在FVB/N Ins-GAS Smox-/-小鼠中的功能；B)一种新的、有效的第二代SMOX 抑制剂SLH150-54在INS-GAS小鼠和沙土鼠中的作用；C)SMOX/Spd对体细胞形成的影响 使用完整外显子组测序的基因组异常。2)确定上皮性DHPs是否介导有害 SMOX/Spd在胃癌发生中的作用我们将分析：a)SMOX在H。 幽门螺杆菌感染在C57BL/6和FVB/N INS-GAS Smox-/-vs WT小鼠中的作用 C57BL/6和FVB/N对胃上皮细胞(GECs)炎症和癌变的影响 GECs中DHPs特异性缺失的INS-GAS小鼠；C)DHPs/亚硫化对体细胞基因组的影响 不稳定。3)利用VA患者组织中的胃有机报告系统建立人 癌症发生的生物标志物。幽门螺杆菌诱导的DNA损伤，b-连环蛋白激活，以及 将研究目标1和目标2，涉及：a)使用CRISPR/Cas9介导的缺失，SMOX的诱导和作用 SMOX和SMOX抑制剂；以及B)使用CRISPR/Cas9介导的缺失DHP的诱导和作用。 DHPS和DHPS抑制剂GC7。这项研究计划将为幽门螺杆菌提供至关重要的新见解-- 在胃癌发生的过程中诱导炎症和疾病进展。翻译成英文 从有价值的动物模型到强大的人体器官系统的发现有望促进这一验证 对VA患者的癌症风险评估、化学预防和治疗的新策略。

Helicobacter pylori infection of the stomach and the resulting clinical consequences of chronic gastritis, peptic ulcer disease, and disease progression to gastric cancer remains a major health concern for Veterans. This pathogen infects half of the world’s population, and gastric cancer is the third leading cause of cancer deaths worldwide. H. pylori prevalence in Veterans is high; deployment-related infection is a problem, especially acquisition of strains associated with higher risk for carcinogenesis. The three polyamines, putrescine (Put), spermidine (Spd), and spermine (Spm), are ubiquitous molecules with many biological effects. Put is synthesized by the rate-limiting enzyme, ornithine decarboxylase (ODC), and is converted sequentially to Spd and Spm, which is back-converted to Spd by spermine oxidase (SMOX). Our Lab has had a long interest in the role of polyamines in GI inflammation and carcinogenesis, and we have the following exciting findings pertinent to this renewal grant: 1) Smox–/– mice infected with H. pylori exhibit decreased gastric inflammation, DNA damage, and chemokine expression, associated with depletion of Spd; 2) These phenotypes are recapitulated in H. pylori- infected 2D organoids from Smox–/– mice; 3) A new link between the H. pylori-induced pro-carcinogenic activation of b-catenin and SMOX, including findings that Smox–/– mice and gerbils treated with a SMOX inhibitor have loss of b-catenin activation in vivo; 4) Use of human gastric organoid cultures to demonstrate the effectiveness of a novel and potent SMOX inhibitor in blocking H. pylori-induced b-catenin activation, associated with Spd depletion; 5) Implication of a unique form of protein translation, hypusination, via the action of deoxyhypusine synthase (DHPS), a process which has been linked to carcinogenesis and appears to be attenuated by Smox deletion; 6) Demonstration of somatic genomic abnormalities by whole exome sequencing (WES) in gastritis and dysplasia tissues of INS-GAS mice, a model of H. pylori-induced carcinogenesis. We will now make effective use of important molecular tools, including valuable mutant mouse models, a key SMOX inhibitor, and advanced use of 3D and 2D gastric organoids from mice and VA patients. We hypothesize that dysregulated polyamine metabolism, due to SMOX and associated generation of Spd and hypusination, provides a molecular pathway leading to risk for gastric disease progression to carcinogenesis. Our specific aims are: 1) To directly determine the role of SMOX and Spd in gastric carcinogenesis. We will test: A) The effect of SMOX in cancer-prone INS-GAS mice, analyzing H. pylori-induced carcinogenesis, DNA damage, b-catenin activation, and GEC function in FVB/N INS-GAS Smox–/– mice +/- Spd; B) A novel, potent, second-generation SMOX inhibitor, SLH150-54, in INS-GAS mice and gerbils; C) The effect of SMOX/Spd on the formation of somatic genomic abnormalities using whole exome sequencing. 2) To determine if epithelial DHPS mediates deleterious effects of SMOX/Spd in gastric carcinogenesis. We will analyze: A) The role of SMOX in hypusination during H. pylori infection in C57BL/6 and FVB/N INS-GAS Smox–/– vs. WT mice +/- H. pylori +/- Spd; B) The effect of hypusination in gastric epithelial cells (GECs) on inflammation and carcinogenesis, using C57BL/6 and FVB/N INS-GAS mice with specific deletion of Dhps in GECs; C) the effect of DHPS/hypusination on somatic genomic instability. 3) To utilize gastric organoid reporter systems in tissues from VA patients to establish human biomarkers for carcinogenesis. H. pylori-induced DNA damage, b-catenin activation, and pathways identified in Aims 1 and 2 will be studied, related to: A) Induction and role of SMOX, using CRISPR/CAS9-mediated deletion of SMOX and SMOX inhibitors; and B) Induction and role of DHPS, using CRISPR/CAS9-mediated deletion of DHPS and the DHPS inhibitor, GC7. This research program will provide crucial new insights into H. pylori- induced inflammation and disease progression along the pathway to gastric carcinogenesis. The translation of findings from valuable animal models into robust human organoid systems is expected to facilitate the validation of new strategies for cancer risk assessment, chemoprevention, and treatment for VA patients.