Metagenomic profiling of oral polymicrobial flora in head and neck cancers

描述（由申请人提供）：当胃中的幽门螺杆菌与胃癌相关联，其根除与回归相关时，一个范式转换的致癌模型被创建。许多细菌种类通常存在于口咽粘膜的不同区域，并可能根据宿主的年龄和饮食消费而波动。有趣的是，头颈部鳞状细胞癌（HNSCC）的发病率已被证明与口腔卫生相关，这可能表明细菌与这种特殊恶性肿瘤的发病机制有关。一些细菌会导致炎症，从而导致甲基化增加。利用基因组技术，我们已经证明了基于宏基因组谱的HNSCC和特定细菌种群之间的可能联系。值得注意的是，我们已经证实了HNSCC中特定细菌亚群的存在与MDR1甲基化状态之间的原理关联。因此，我们假设特定的细菌亚群，正如宏基因组分析所揭示的那样，可以触发靶向甲基化事件，然后启动HNSCC的发生和/或进展。我们计划通过三个主要目标来解决这一假设：(1)利用最新的高通量测序技术来确定口咽黏膜中与HNSCC肿瘤发生、进展和/或预防相关的细菌亚群，并以创新的子目标来探索特定的微生物群谱是否属于HNSCC中所见的场效应的一部分；(2)探讨影响HNSCC炎症的特定微生物“联合体”，并由此导致靶肿瘤抑制基因的表观遗传改变；(3)从功能上验证MAD2和/或MDR1参与HNSCC肿瘤抑制。宏基因组图谱所反映的微生物种群将被询问与暴露、临床信息和靶向甲基化的相关性。最后，我们将从功能上验证靶向甲基化是否与HNSCC肿瘤发生有关，还是与炎症反应中的随机甲基化效应有关。为了解决这个问题，我们将在HNSCC细胞系中过表达或敲低MDR1，并检查其对生长、凋亡、细胞形态或分化的影响。这项研究有望揭示特定头颈部细菌菌群与癌症发生和发展的相关性。未来有趣的方向包括临床筛查特定细菌指标的存在，以便提供基于益生菌或抗生素的早期癌症或癌前检测和预防。由于代谢组学图谱能够根据既往放射暴露或化疗区分样本，我们将对难治性疾病患者实施靶向筛选和/或去甲基化治疗。这类数据还将指出卫生工作者和护理人员在早期发现、意识教育和预防方面的更大作用。

DESCRIPTION (provided by applicant): When H. pylori in the stomach was linked with gastric cancer, and its eradication associated with regression, a paradigm-shifting model of carcinogenesis was created. Many bacterial species normally reside in various regions of the oropharyngeal mucosa, and may fluctuate depending on the host's age and dietary consumption. Interestingly, the incidence of head and neck squamous cell carcinoma (HNSCC) has been shown to correlate with dental hygiene, which may suggest the relevance of bacteria in the pathogenesis of this particular malignancy. Some bacteria can result in inflammation, which can cause increased methylation. Using genomic technology, we have demonstrated a possible link between HNSCC and specific bacterial populations based on their metagenomic profiles. Notably, we have proof-of-principle association between the presence of particular bacterial subpopulations per metabiomic profiling and MDR1 methylation status in HNSCC. We therefore hypothesize that specific bacterial subpopulations, as revealed by metagenomic profiling, can trigger targeted methylation events which then initiate HNSCC genesis and/or progression. We plan to address this hypothesis via 3 broad aims: (1) To utilize most recent high throughput sequencing technologies to identify the bacterial subpopulations in the oropharyngeal mucosa associated with HNSCC tumor initiation, progression, and/or prevention, with the innovative sub-aim to explore if specific microbiomic profiles are part of the field effect seen in HNSCC; (2) To interrogate the specific microbial "consortia" influencing inflammation with consequent epigenetic alterations of target tumor suppressor genes in HNSCC; (3) To functionally validate the involvement of MAD2 and/or MDR1 in HNSCC tumor suppression. The resulting microbial populations as reflected by metagenomic profiles will be interrogated for correlation with exposure and clinical information and targeted methylation. Finally, we will functionally validate whether the targeted methylation is relevant in HNSCC tumorigenesis or a random methylation effect in response to inflammation. To address this, we will overexpress or knock-down MDR1 in HNSCC cell lines and examine any effect on growth, apoptosis, cell morphology, or differentiation. This study promises to reveal the relevance of specific head and neck bacterial flora to cancer initiation and progression. Interesting future directions include clinical screening for the presence of specific bacterial indicators in order to provide early cancer or pre-cancer detection and perhaps prevention, based on probiotics or antibiotics. With metabiomic profiles capable of distinguishing samples based on prior radiation exposure or chemotherapy, we will implement targeted screening and/or demethylation treatment for patients with refractory disease. This type of data will also point to larger roles for early detection, awareness education, and prevention from hygienists and nursing. PUBLIC HEALTH RELEVANCE: Over 500,000 new cases of head and neck cancers (HNSCC) are diagnosed worldwide annually, with more than 11,000 individuals/year succumbing to this aggressive malignancy in the United States alone; despite huge strides in research achievements in this topic, overall outcome has remained unchanged for over 3 decades for the majority of patients with advanced stages of this disease. Patients who receive aggressive surgery to remove the cancer have significant disfigurement and diminished quality of life, with impaired ability to swallow, smell, and hear. If successful, then our current proposal promises a new understanding of the role of specific subpopulations of bacteria in the mouth leading to inflammation which shuts off cancer-suppressing genes, thus leading to HNSCC. If true, then we can take advantage of our findings for the earliest diagnosis and even prevention with probiotic therapy. Importantly, even for advanced HNSCC or that which has spread, our findings will help point to antibiotics and agents that can turn back on the cancer-suppressing genes to revolutionize treatment.