Targeting epigenetic abnormalities to inhibit cutaneous squamous cell carcinoma

针对表观遗传异常抑制皮肤鳞状细胞癌

• 批准号: 10705718
• 负责人: Masaoki Kawasumi
• 金额: $ 39.57万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-15 至 2027-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10705718
• 关键词: Acetylation; Affect; Affinity; Automobile Driving; Binding; Binding Sites; CDKN2A gene; CRISPR/Cas technology; Carcinogens; Cell Cycle Proteins; Cell Line; Cells; Cessation of life; Chromatin; Chronic; Communities; Cyclin-Dependent Kinase Inhibitor 2A; DNA; DNA Methylation; DNA Methylation Inhibition; DNA Modification Process; DNA Sequence Alteration; Data; Development; Drug Combinations; EP300 gene; Enhancers; Environmental Risk Factor; Enzymes; Epigenetic Process; Genes; Genetic Induction; Genomic Segment; Genomics; Goals; Guide RNA; Histone Acetylation; Histones; Human; Hypermethylation; Inhibition of Cell Proliferation; Invaded; Laboratories; Large Cell Carcinoma; MYC gene; MYC-Family Oncogene; Malignant Neoplasms; Measures; Methylation; Modification; Neoplasm Metastasis; Oncogenes; Oncogenic; Patients; Phenotype; Play; Positioning Attribute; Pre-Clinical Model; Proliferating; RNA Polymerase II; Risk; Role; Sampling; Signal Transduction; Skin; Skin Cancer; Specimen; Testing; Therapeutic; Tumor Promotion; Tumor Suppressor Proteins; UV Radiation Exposure; UV induced; Ultraviolet Rays; United States; Untranslated RNA; Visualization; demethylation; detection assay; driver mutation; epigenome editing; epigenomics; genome editing; genomic locus; histone modification; in vivo; inhibitor; innovative technologies; malignant phenotype; migration; new technology; novel strategies; novel therapeutic intervention; overexpression; p14ARF Protein; patient derived xenograft model; promoter; rational design; recruit; skin organogenesis; skin squamous cell carcinoma; small molecule inhibitor; targeted treatment; tool; transcription factor; tumor; tumor growth; ultraviolet

Project Summary/Abstract Chronic ultraviolet (UV) radiation induces not only genetic mutations but also aberrant epigenetic modifications that affect the expression of transcription factors, oncogenes, and tumor suppressors, leading to the development of cutaneous squamous cell carcinoma (cSCC), the second most common cancer in the United States. A better understanding of the mechanisms underlying UV-associated epigenetic abnormalities will provide novel approaches to inhibit skin malignancies. Strikingly, chronic UV exposure induces DNA hypermethylation at the promoter of CDKN2A gene, which encodes tumor suppressors p16INK4A and p14ARF. This epigenetic alteration silences CDKN2A, promoting tumor growth and metastasis. However, it remains unclear whether the reversal of this epigenetic aberration can reactivate CDKN2A and inhibit malignant phenotypes of cSCC. In Aim 1, we will use our CRISPR-Cas9-based epigenome editing tools to specifically demethylate the CDKN2A promoter and investigate the effect of targeted DNA demethylation on cancer phenotypes. UV radiation increases the expression of Myc, which is an oncogenic transcription factor. Myc activation is a cancer hallmark, and the Myc oncogene family is deregulated in many human cancers. When overexpressed, Myc binds to noncanonical (low-affinity) motifs, invading enhancers, and potentially activates super-enhancers (noncoding genomic regions defined by unusually high levels of H3K27 acetylation). Cancer-specific super- enhancers strongly upregulate oncogenes, driving malignancies. However, it remains elusive how aberrant super-enhancers are formed in cSCC. We hypothesize that overexpressed Myc binds to low-affinity Myc binding sites, recruits histone acetyltransferase p300, and generates aberrant super-enhancers that drive malignancy. In Aim 2, we will determine the role of overexpressed Myc in super-enhancer formation in cSCC. A subset of cSCC displays poor differentiation, which correlates with poor patient survival. However, it remains unclear what confers the aggressiveness of poorly differentiated cSCC. We hypothesize that small-molecule inhibitors of DNA methylation and/or Myc suppress poorly differentiated cSCC in vivo by reactivating CDKN2A and/or inhibiting Myc-associated super-enhancers. In Aim 3, we will use poorly differentiated cSCC patient- derived xenografts as preclinical models to assess the effects of these inhibitors on tumor growth and metastasis. Innovative technologies will be used for epigenetic analyses: “PIXUL-ChIP” (high-throughput chromatin shearing for robust ChIP signals) and “EVA” (epigenetic visualization assay that detects specific DNA methylation at the single-cell level). With our epigenome editing tools, the proposed study will elucidate the contribution of epigenetic abnormalities to cSCC aggressiveness and reveal therapeutic potential of modulating epigenetic marks to suppress cSCC.

项目摘要/摘要 慢性紫外线辐射不仅能诱发基因突变，还能诱发异常的表观遗传 影响转录因子、癌基因和肿瘤抑制因子表达的修饰，导致 皮肤鳞状细胞癌(CSCC)是美国第二常见的癌症 各州。更好地理解紫外线相关表观遗传异常的机制将 提供新的方法来抑制皮肤恶性肿瘤。 值得注意的是，慢性紫外线暴露诱导了CDKN2A基因启动子的DNA甲基化，这是 编码肿瘤抑制基因p16INK4A和p14ARF。这种表观遗传改变使CDKN2A沉默，促进肿瘤 生长和转移。然而，目前尚不清楚这种表观遗传异常的逆转是否可以 重新激活CDKN2A，抑制CSCC的恶性表型。在目标1中，我们将使用我们基于CRISPR-Cas9的 表观基因组编辑工具特异性去甲基化CDKN2A启动子并研究靶向作用 DNA去甲基化对癌症表型的影响 紫外线辐射增加了一种致癌转录因子Myc的表达。MYC激活是一种 癌症的标志，Myc癌基因家族在许多人类癌症中被解除调控。当过度表达时， MYC结合非规范(低亲和力)基序，入侵增强子，并可能激活超级增强子 (由异常高水平的H3K27乙酰化定义的非编码基因组区域)。癌症特异性超- 增强剂强烈上调癌基因，推动恶性肿瘤的发生。然而，它仍然难以捉摸有多反常。 在CSCC中形成了超级增强剂。我们假设过度表达的Myc与低亲和力的Myc结合 位点，招募组蛋白乙酰转移酶p300，并产生异常的超级增强子，推动恶性肿瘤的发生。 在目标2中，我们将确定过表达的Myc在CSCC中超级增强子形成中的作用。 CSCC的一个子集表现为分化较差，这与患者存活率较低相关。然而，它仍然 不清楚是什么赋予了低分化CSCC的侵袭性。我们假设小分子 DNA甲基化和/或Myc抑制剂通过重新激活CDKN2A抑制体内低分化CSCC 和/或抑制Myc相关的超级增强子。在目标3中，我们将使用低分化的CSCC患者- 衍生异种移植作为临床前模型，以评估这些抑制物对肿瘤生长和转移的影响。 将使用创新技术进行表观遗传学分析：“PIXUL-CHIP”(高通量染色质 剪切以获得强健的芯片信号)和“EVA”(检测特定DNA的表观遗传可视化分析 单细胞水平的甲基化)。使用我们的表观基因组编辑工具，拟议的研究将阐明 表观遗传异常在宫颈鳞癌侵袭性中的作用及其调控治疗潜力 抑制CSCC的表观遗传标记。