Precision Medicine of Cancer

项目1A：微生物组是人类癌症精准医学的关键方面之一。我们正在建立之前发现的与人类肺癌相关的微生物组（Greathouse KL et al., Genome Biol. 19:123-31, 2018）。Actovorax在吸烟者的鳞状细胞癌和TP53突变型癌症中的含量高于腺癌。这些结果产生了我们正在研究的几个假设。首先，Acidovorax是肺癌发生的过客还是贡献者。我们正在人类肺癌动物模型中验证这一假设（Jin C et al., Cell 176:998-1013, 2019）。我们的数据是一致的，因为肺癌在动物模型和体外细胞模型中发生得更早，更大。由于炎症经常与包括肺癌在内的许多类型的人类癌症有关，我们正在研究免疫细胞失调；由酸败引起的。此外，还在研究Acidovorax与乳酸菌、双歧杆菌等抗炎菌的鼻腔联合注射。FACS分析显示产生il - 17和中性粒细胞的γ - δ免疫细胞增加，中性粒细胞可能通过产生自由基和/或其细胞外陷阱起作用（Chaim HR等）。年度Rev Cell Dev Bio， 2020年7月14日)。我们目前正在研究其他假说，例如，Acidovorax，用于在石油泄漏的生物介质中代谢多环芳烃，代谢烟草烟雾中的化学致癌物，以及重新编程肺癌细胞以产生CD47，阻止巨噬细胞吞噬它们。巴雷特食管是一种易患癌症的慢性炎症性疾病，我们也正在确定有或没有巴雷特食管的食管癌患者的微生物组。这项微生物组研究延续了我们悠久的历史，包括24份研究食管癌分子和细胞发病机制的报告。项目1B：我们关注支气管组织学癌前病变转变为1期癌的机制。确定这些过渡机制可能为预防肺癌提供阻断疗法。这些研究使用我们之前研究建立的方法来检查：HOXA9甲基化和表达（Lissa D. et al., Lung Cancer 122:151-59, 2018; Robles et al., J Thoracic Oncology 10:1037-48, 2015）；MALDI成像分析肌酸核糖苷表达（1C项目）；细菌的RNA范围（EUB表达）和Acidovorax使用特异性探针，单细胞RNA测序，空间转录组学，CD8+ T细胞和M1和M2巨噬细胞的定量。这些肿瘤前支气管病变的结果将与我们之前对一期肺癌的研究进行比较。此外，我们目前正在与Roger Reddel（儿童研究基金会研究所）、Ken Aldape （NCI）和Eytan Ruppin （NCI）合作，在一期腺癌和肺鳞状细胞癌的发现和验证队列中研究蛋白质组。本研究还包括项目1D中描述的RNA测序、DNA甲基化、全外显子组基因组测序和代谢组。项目1 c。正如之前在2019年年度报告中详细介绍的那样，我们正在使用尿液、血清和血浆液体活检，通过质谱定量测量肺癌的四种生物标志物（肌酸核糖体（CR）、n -乙酰氨基酸（NANA）、硫酸皮质醇（CS）和27 α -5 - β -胆甾醇-3 α、7 α、12 α、24 α、25 α戊糖醛酸（NCPG）） （Mathe E等人，cancer Res. 74: 359 - 70,014; Haznadar， M等人，cancer Epidemiol）。生物标志物，2016,25:978- 86,2016)。我们正在调查多个队列：国家癌症研究所和马里兰大学医院（NCI-MD）；与Avi Spira、肺癌生物标志物研究网络（LCBRN）合作的军人早期癌症检测（DECAMP1）；范德比尔特医院与皮埃尔·马森合作；以及国家肺筛查试验（NLST），以确定生物标志物的效用：预测经常通过手术治愈的1期和2期肺癌的预后。然而，25%的患者在未来5年内复发，可以从辅助化疗和/或免疫治疗中获益；从癌症中识别良性不确定结节：并优先考虑低剂量CT筛查肺癌候选者。我们还在开发一种基于clia的临床试验，使用液体活检这四种生物标志物。我们和我们的合作者已经合成了重标记肌酸核糖体，目前正在合成重标记NCPG。这些将有助于快速质谱分析在商业临床设置。项目1D：我们发现新的肿瘤来源的生物标志物肌酸核糖苷是尿素循环和戊糖磷酸途径失调的产物，在人类致癌实验室正在研究的主要致死类型的人类癌症中常见，包括肺癌、乳腺癌、前列腺癌、肝癌和食管癌。这是我们与我们实验室的同事Brid Ryan、Perwez Hussain、Stefan Ambs和Xin Wang持续合作的另一个例子。我们有未发表的数据表明，人类间皮瘤产生高水平的肌酸核糖体。我们也有很长的历史来研究石棉的病因和间皮瘤的遗传和分子基础（例如，Gabrielson， EW等，carcinogenes13: 1359- 64,1992； Marrogi a等，Cancer Res 60:3696-700, 2000）。因此，肌酸核苷可能是一种伴随诊断的生物标志物，既可以针对这些癌症，也可以通过精氨酸酶或精氨酸脱氨酶和免疫治疗选择特定的精氨酸消耗治疗。这些疗法正在临床试验中，包括间皮瘤。我们已经开始与Raffit Hassan （NCI）和Peter Zlosarek，英国巴特癌症研究中心合作，研究经常是精氨酸营养不良的间皮瘤。我们将调查以前和正在进行的试验。我们将跟踪这些病人从开始到整个治疗过程。这将提供关于肌酸核苷在四组间皮瘤诊断和治疗中的“原理证明”临床价值的信息。

Project 1A: The microbiome is one of the key facets of the precision medicine of human cancer. We are building on our previous discoveries of the microbiome associated with human lung cancer (Greathouse KL et al., Genome Biol. 19:123-31, 2018). Actovorax was more abundant in squamous cell carcinoma greater than in adenocarcinoma in tobacco smokers and TP53 mutant cancers. These results generated several hypotheses that we are investigating. First, is Acidovorax a passenger or contributor to lung carcinogenesis. We are testing this hypothesis in an animal model of human lung cancer (Jin C et al., Cell 176:998-1013, 2019). Our data are consistent with it being a contributor in that the lung cancers occur earlier and are larger in the animal model and using in vitro cellular models. Because inflammation is frequently associated with many types of human cancer including lung cancer, we are investigating the immune cell dysregulate; on caused by Acidovorax. In addition, the nasal co-injection of Acidovorax and anti-inflammatory bacteria such as Lactobacillus and Bifidobacterium is being investigated. FACS analysis showed an increase in gamma-delta immune cells producing IL17 and neutrophils that may function by producing free radicals and/or by their extracellular traps (Chaim HR et al. Annu Rev Cell Dev Bio July 14, 2020). We are currently investigating other hypotheses e.g., Acidovorax, used to metabolize polycyclic aromatic hydrocarbons in biomediation of oil spills, metabolizing chemical carcinogens found in tobacco smoke and reprogramming lung cancer cells to produce CD47 that prevents macrophages from engulfing them. We are also determining the microbiome of esophageal carcinoma in patients with or without Barrett's esophagus that is a cancer-prone chronic inflammatory disease. This microbiome study continues our long history including 24 reports of investigating the molecular and cellular pathogenesis of esophageal cancer. Project 1B: We are focusing on the mechanism(s) by which histological preneoplastic lesions in the bronchus transition to stage 1 carcinoma. Identifying these transition mechanisms may provide interception therapies to prevent lung cancer. These studies are examining: HOXA9 methylation and expression using the methodology established from our previous studies (Lissa D. et al., Lung Cancer 122:151-59, 2018; Robles et al., J Thoracic Oncology 10:1037-48, 2015); creatine riboside expression (Project 1C) using MALDI imaging analysis; RNA scope of bacteria(EUB expression) and Acidovorax using specific probes, single cell RNA seq, spatial transcriptomics, quantitation of CD8+ T cells and M1 and M2 macrophages. These results of preneoplastic bronchial lesions will be compared to our previous studies of stage one lung cancer. In addition, we are currently investigating the proteome in both discovery and validation cohorts of stage one adenocarcinoma and squamous cell carcinoma of the lung in collaboration with Roger Reddel (Children Research Foundation Institute) and Ken Aldape (NCI) and Eytan Ruppin (NCI). This study also includes the RNA seq, DNA methylation, whole exome genome sequencing and the metabolome descripted in Project 1D. Project 1C. As previously detailed in the 2019 annual report, we are using liquid biopsy of urine, serum and plasma to quantitatively measure four biomarkers (creatine riboside(CR), N-acetylneuminic acid(NANA), cortisol sulfate(CS) , and 27alpha-nor-5beta-cholestane-3aplpha, 7alpha, 12alpha 24alpha, 25alpha Pentol glucuronide(NCPG) of lung cancer by mass spectrometry (Mathe E et al., Cancer Res. 74:3259-70, 014; Haznadar, M et al., Cancer Epidemiol. Biomarker Prev. 25:978-86, 2016). We are investigating multiple cohorts: National Cancer Institute and University of Maryland Hospital (NCI-MD); Detection of Early Cancer among Military Personnel (DECAMP1) in collaboration with Avi Spira, Lung Cancer Biomarker Research Network (LCBRN); Vanderbilt Hospital in collaboration with Pierre Masson; and th National Lung Screening Trial (NLST) to determine the utility of the biomarkers: to predict the prognosis of stage 1 and 2 lung cancer that are frequently cured by surgery. However, 25% recur during the next 5 years and could benefit from adjunct chemotherapy and/or immunotherapy; identify benign indeterminate nodules from cancer: and to prioritize candidates for lung cancer screening by low dose CT. We are also developing a CLIA-based clinical test using liquid biopsy of these four biomarkers. We and our collaborators have synthesized heavy labelled creatine riboside and are currently synthesizing heavy labelled NCPG. These will be useful for rapid mass spectrometry analysis in the commercial clinical setting. Project 1D: We have discovered that the novel tumor-derived biomarker creatine riboside is the product of dysregulated urea cycle and pentose phosphate pathway that is commonly found in major lethal types of human cancer being investigated in the Laboratory of Human Carcinogenesis including lung, breast, prostate, liver and esophageal carcinoma. This is another example of ongoing collaboration with our colleagues Brid Ryan, Perwez Hussain, Stefan Ambs and Xin Wang in our Laboratory. We have unpublished data that human mesothelioma produces high levels of creatine riboside. We also have a long history of investigating the etiology by asbestos and the genetic and molecular basis of mesothelioma (e.g., Gabrielson, EW et al., Carcinogenesis13: 1359-64, 1992; Marrogi A et al, Cancer Res 60:3696-700, 2000). Therefore, creatine riboside may be a companion diagnostic biomarker to both target these cancers and select specific arginine depleting therapy by arginase or arginine deaminase and immunotherapy. These therapies are in clinical trials that including mesothelioma. We have initiated collaborations with Raffit Hassan (NCI) and Peter Zlosarek, Bart Center, Cancer Research, UK to investigate mesothelioma that are frequently arginine auxotrophic. We will investigate both previous and ongoing trials. We will follow these patients from the initiation and throughout their therapy. This will provide information on the "proof of principle" clinical value of creatine riboside in the diagnosis and therapy in four cohorts mesothelioma.