Role of Alcohol as a risk factor in the induction of Pancreatic Carcinogenesis

酒精作为诱发胰腺癌的危险因素的作用

• 批准号: 9891302
• 负责人: Sharmila Shankar
• 金额: --
• 依托单位: SOUTHEAST LOUISIANA VETERANS HEALTH CARE
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-07-01 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9891302
• 关键词: Acetaldehyde; Address; Alcohol consumption; Alcohols; Analysis of Variance; Attenuated; BALB/c Nude Mouse; Binding; Biochemical; Biological; Biological Process; Biology; Blood; CRISPR/Cas technology; CYP2E1 gene; Cancer cell line; Carcinogens; Cell Survival; Cells; ChIP-seq; Characteristics; Chronic; Complex; Data; Development; Disease; Ductal Epithelial Cell; Early Diagnosis; Epithelial Cells; Ethanol; Ethanol toxicity; Etiology; Face; Future; Gene Expression; Genes; Genetic Transcription; Glutathione S-Transferase; Goals; Growth; Growth and Development function; Histologic; Homeobox; Human; IL8 gene; Implant; In Vitro; Inflammation; Inflammatory; Interleukin-1 beta; Interleukin-6; International Agency for Research on Cancer; Knockout Mice; Knowledge; Liver; Malignant - descriptor; Malignant Neoplasms; Malignant neoplasm of pancreas; Measures; Mediating; Military Personnel; Molecular; Monitor; Mus; Neoplasm Metastasis; Oncogenic; PET/CT scan; Pancreas; Pancreatic duct; Pathway interactions; Pattern; Population; Production; Quantitative Reverse Transcriptase PCR; Radiation therapy; Recurrence; Regulation; Risk; Risk Factors; Role; Social Problems; Survival Rate; TNF gene; Techniques; Testing; Time; Tissues; Transgenic Mice; Veterans; Vulnerable Populations; Western Blotting; X-Ray Computed Tomography; alcohol effect; alcohol exposure; alcohol use disorder; aldehyde dehydrogenases; animal imaging; base; cancer initiation; cancer risk; cancer stem cell; cell transformation; chemotherapy; comparison group; cytokine; diagnostic biomarker; glutathione peroxidase; improved; in vivo; inflammatory milieu; knock-down; metaplastic cell transformation; mouse model; notch protein; novel; novel strategies; outcome forecast; overexpression; pancreatic tumorigenesis; pluripotency; problem drinker; self-renewal; stem cells; transcription factor; transcriptome; transcriptome sequencing; tumor; tumor growth; tumorigenic

Pancreatic cancer is one of the most lethal and devastating human malignancies, with a 5-year survival rate of approximately 6%. Early diagnosis of pancreatic cancer (CaP) is challenging due to lack of reliable diagnostic markers and limited understanding of the etiological factors. Heavy alcohol consumption constitutes a significant social problem and has been identified as a risk factor for pancreatic cancer. Alcohol use disorders frequently co-occur in military veterans and civilians; thus, these populations are vulnerable to face a disproportionate burden of increased risk to develop pancreatic cancer. This application will address evaluating the cellular and molecular mechanism by which alcohol increases pancreatic cancer risk. It will be highly significant and beneficial to understand the underlying mechanism of this complex disease association. Therefore, the overarching goal of this application is to examine the biological role of alcohol toxicity in the induction of pancreatic cancer and to identify genes mediating alcohol induced cellular transformation and progression of pancreatic cancer. Pancreatic cancer growth and persistence depend on the cancer stem cells (CSCs) that also induce recurrence and resist current chemo- and radiotherapies. Our preliminary data demonstrate that ethanol induced cellular transformation of normal pancreatic ductal epithelial cells. Chronic ethanol exposure induced malignant transformation of HPNE cells by causing inflammation (upregulated IL-6, IL-8, TNFα, and IL-1β genes, inhibited glutathione S-transferases (GSTs) and glutathione peroxidases (GPx), and induced stem cell markers). Interestingly, we identified that the expression pattern of Engrailed-2 (EN2), a homeobox-containing transcription factor was significantly altered in ethanol-transformed HPNE cells, and the induction of EN2 correlated with the rate of transformation. However, the biological function and molecular mechanism of EN2 in pancreatic cancer have never been examined. We demonstrate that overexpression of EN2 in human pancreatic normal ductal epithelial cells induces malignant transformation by generating CSCs, and regulating Notch-Hes1 pathway; suggesting that the expression of EN2 is oncogenic for pancreatic cancer. Further, we have shown that EN2 regulates transcription of pluripotency maintaining factors, that are required for sustaining the stem cells capacity for self-renewal and that the expression of EN2 is tightly regulated in cancer. EN2 is highly expressed in pancreatic CSCs and cancer cell lines, but not in normal pancreatic cells or normal pancreatic tissues. Further, we have shown that inhibition of EN2 attenuated CSC characteristics, Notch pathway, and tumor growth in mice. Based on our preliminary studies, we hypothesize that alcohol induces inflammatory environment during transformation resulting in induction of EN2, which acts as a key regulator of pluripotency and self-renewal and inhibition of EN2 in ethanol-transformed cells attenuates their tumorigenic potential. To examine the role of alcohol in pancreatic cancer and address the above hypothesis we propose to integrate functional, molecular, biochemical and histological approaches. Gain and loss of EN2 functional studies will be performed in vitro and in vivo. EN2 expression will be knocked down by CRISPR- Cas9 technology in vitro. Novel triple transgenic mice (LSL-KrasG12D; LSL-EN2+/+; Pdx-Cre) will be generated and effects of alcohol on pancreatic carcinogenesis will be compared in the presence or absence of EN2 (KrasG12D/EN2+/+, and KrasG12D/EN2-/- mice). We will examine the involvement of Notch-Hes1 pathway to assess the biological effects of alcohol mediated by EN2. The studies proposed in this application will enhance our understanding of the biology of ethanol-induced cellular transformation of normal pancreatic ductal epithelial cells, which will be significant for the management of pancreatic cancer.

胰腺癌是最致命和最具破坏性的人类恶性肿瘤之一，其 5 年生存率为 由于缺乏可靠的诊断，胰腺癌 (CaP) 的早期诊断具有挑战性。 标志物和对病因的了解有限。 严重的社会问题，并已被确定为胰腺癌的危险因素。 退伍军人和平民经常同时发生；因此，这些人群很容易面临 该应用程序将评估罹患胰腺癌的风险增加的负担。 酒精增加胰腺癌风险的细胞和分子机制。 对于理解这种复杂疾病关联的潜在机制具有重要意义和有益意义。 因此，本申请的首要目标是检查酒精毒性在酒精中毒中的生物学作用。 诱导胰腺癌并鉴定介导酒精诱导细胞转化的基因 胰腺癌的进展和持续性取决于癌症干细胞。 （CSC）也会诱发复发并抵抗当前的化疗和放疗。 证明乙醇诱导正常胰腺导管上皮细胞慢性细胞转化。 乙醇暴露通过引起炎症（上调 IL-6、 IL-8、TNFα 和 IL-1β 基因，抑制谷胱甘肽 S-转移酶 (GST) 和谷胱甘肽过氧化物酶 (GPx)， 暗示，我们确定了 Engrailed-2 (EN2) 的表达模式， 乙醇转化的 HPNE 细胞中含有同源盒的转录因子发生显着改变，并且 EN2的诱导与转化率相关，但其生物学功能和分子水平有关。 我们从未研究过 EN2 在胰腺癌中的过度表达。 人胰腺正常导管上皮细胞中的EN2通过生成CSC诱导恶性转化， 并调节Notch-Hes1通路；表明EN2的表达对胰腺癌具有致癌作用。 此外，我们已经证明 EN2 调节多能性维持因子的转录，这是必需的 维持干细胞自我更新的能力，并且 EN2 的表达在 EN2 在胰腺 CSC 和癌细胞系中高表达，但在正常胰腺细胞或癌细胞中不表达。 此外，我们还发现 EN2 的抑制会减弱 CSC 的特征， 根据我们的初步研究，我们服用了酒精。 在转化过程中诱导炎症环境，从而诱导 EN2，这是一个关键 乙醇转化细胞中多能性和自我更新的调节剂和 EN2 的抑制会减弱其 研究酒精在胰腺癌中的作用并解决上述假设。 我们建议整合功能、分子、生化和组织学方法。 功能研究将在体外进行，EN2 表达将被 CRISPR 敲低。 体外Cas9技术将产生新型三重转基因小鼠（LSL-KrasG12D；LSL-EN2+/+；Pdx-Cre）。 并且将在存在或不存在 EN2 的情况下比较酒精对胰腺癌发生的影响 （KrasG12D/EN2+/+ 和 KrasG12D/EN2-/- 小鼠）我们将检查 Notch-Hes1 通路对小鼠的影响。 评估 EN2 介导的酒精的生物效应将增强本申请中提出的研究。 我们对乙醇诱导的正常胰腺导管细胞转化生物学的理解 上皮细胞，这对于胰腺癌的治疗具有重要意义。