Exploring mechanisms of therapeutic demethylation effects in HPV-associated head and neck cancer

探索 HPV 相关头颈癌去甲基化治疗作用的机制

• 批准号: 10664847
• 负责人: Karen S. Anderson
• 金额: $ 40.47万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-06-01 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10664847
• 关键词: Acceleration; Aftercare; Apoptosis; Azacitidine; Caspase; Cell Death; Cell Line; Cell Survival; Cells; Cervical; Characteristics; Chemotherapy and/or radiation; Clinical Trials; Cytidine Deaminase; DNA; DNA Damage; DNA Double Strand Break; Data; Deglutition; Dental caries; Detection; Diagnosis; Disease; Dose; Double Strand Break Repair; Down-Regulation; Dysmyelopoietic Syndromes; Excision; FDA approved; Family member; Fibrosis; Functional disorder; Gene Expression; Genes; Genetic Transcription; Genomic Instability; Growth; Guidelines; Head and Neck Cancer; Head and Neck Neoplasms; Head and Neck Squamous Cell Carcinoma; Human; Human Papillomavirus; Human papillomavirus 16; Immune; In Vitro; Infection; Inhibition of Matrix Metalloproteinases Pathway; Interferon Activation; Interferons; Interstitial Collagenase; Invaded; Lead; Location; Lymph Node Dissections; Lymphedema; Lymphocytic Infiltrate; Malignant - descriptor; Malignant Neoplasms; Matrix Metalloproteinases; Methods; Methylation; Modeling; Molecular; Molecular Biology; Mus; National Comprehensive Cancer Network; Neoplasm Circulating Cells; Neoplasm Metastasis; Oncogenes; Oncogenic; Operative Surgical Procedures; Oropharyngeal Squamous Cell Carcinoma; Pathologic; Pathway interactions; Patients; Pharmaceutical Preparations; Production; Prognosis; Prognostic Marker; Proliferating; Property; Radiation; Recommendation; Recurrence; Regulation; Retinoblastoma; Role; Site; Skeletal muscle structure of neck; Specimen; Speech; Survival Rate; T-Lymphocyte; TP53 gene; Therapeutic; Tobacco use; Toxic effect; Transcript; Tumor Suppressor Proteins; Tumor-Infiltrating Lymphocytes; United States; Uterus; Xerostomia; Yale Cancer Center; cancer cell; cancer therapy; carcinogenesis; cell transformation; cervical and uterine cancer; chemotherapeutic agent; clinically relevant; cytokine; cytotoxic; demethylation; design; experimental study; falls; human papilloma virus oncogene; immune cell infiltrate; improved; in vitro Assay; molecular modeling; mouse model; multimodality; novel therapeutics; oral HPV-positive head and neck cancers; overexpression; participant enrollment; patient derived xenograft model; pre-clinical; prevent; promoter; public health relevance; response; response biomarker; restoration; side effect; targeted treatment; tumor; tumor growth; tumor xenograft; tumorigenesis

Abstract Oncogenic human papillomaviruses (HPV) are the causative agents of uterine cervical and an increasing portion of head and neck squamous cell carcinomas (HNSCC), but HNSCC is almost exclusively associated with HPV type 16. The oncogenic properties of HPV type 16 are largely attributed to two major HPV oncogenes, E6 and E7, that degrade p53 and retinoblastoma (RB) family members, respectively. Degradation of these tumor suppressors by E6 and E7 results in uncontrolled proliferation, diminished apoptosis and increased genomic instability that predisposes to malignant transformation. The crucial roles of E6/E7 in HPV-related carcinogenesis make them an attractive target for anti-cancer therapy, since methods for decreasing their expression or activities would restore p53 and RB activity in tumors driven by HPV. Our preliminary results indicate that treatment of HPV-positive (HPV+) HNSCC with the demethylating agent, 5-azacytidine (5-aza) at clinically relevant concentrations, resulted in remarkable downregulation of all HPV gene expression, including E6 and E7. 5-aza treatment restored p53 expression and activity in HPV+ head and neck cancer cells, which was partially responsible for the sensitivity of these cells to 5-aza. In addition to restoration of p53, 5-aza also was toxic to HPV+ HNSCC through creation of DNA double strand breaks (DSBs). Mechanistically, 5-aza-induced DNA DSBs in HPV+ HNSCC were dependent on transcription and replication and on overexpression of the cytidine deaminase, APOBEC3B (A3B). Experimental depletion of A3B inhibited 5-aza toxicity and diminished DSB formation, but also indicated that untreated HPV+ HNSCC depend on A3B for clonogenic growth. The observations that untreated HPV+ HNSCC dependent on A3B, but that A3B contributes to 5-aza toxicity and DSBs, suggests an A3B-dependent synthetic lethality upon treatment with 5-aza, and that A3B may serve as a biomarker of response. Treatment of mice bearing HPV+ tumors with 5-aza revealed significant tumor growth inhibition and prevented detection of circulating tumor cells. A window clinical trial in patients with HNSCC using standard dosing for 5 days achieved demethylation (LINE-1) similar to that seen in our in vitro experiments and confirmed that 5-aza treatment: 1) significantly decreased expression of HPV genes; 2) reactivated p53; 3) activated caspases, 4) and inhibited matrix metalloproteinase expression in HPV+ HNSCCs. This proposal is designed to determine molecular mechanisms of demethylation-induced downregulation of HPV oncogenes, elucidate the role of A3B in 5-aza-induced synthetic lethality and DNA DSBs formation, determine effect of demethylation on immune cell infiltration in HPV+ HNSCC, and explore the potential of 5-aza alone or in combination with chemotherapeutic agents to suppress HPV-associated HNSCC metastasis and inhibit growth using patient-derived xenografts. These studies will provide a basis for a new rational targeted therapy for HPV+ HNSCC, which is desperately needed to treat patients with recurrent or metastatic HPV+ HNSCC and to decrease the toxicity associated with current therapy.

抽象的 致癌的人乳头瘤病毒（HPV）是子宫宫颈的致病药物，增加 头部和颈部鳞状细胞癌（HNSCC）的一部分，但HNSCC几乎与 HPV类型16。HPV类型16的致癌特性主要归因于两个主要的HPV癌基因E6 和E7，分别降解p53和视网膜母细胞瘤（RB）家庭成员。这些肿瘤的降解 E6和E7的抑制剂导致不受控制的增殖，细胞凋亡减少和基因组增加 易于恶性转化的不稳定性。 E6/E7在与HPV相关的癌变中的关键作用 使它们成为抗癌疗法的有吸引力的目标，因为用于降低其表达或活动的方法 将恢复由HPV驱动的肿瘤中的P53和RB活性。我们的初步结果表明 HPV阳性（HPV+）HNSCC，具有脱甲基剂，临床相关的5-氮杂丁胺（5-azacytine） 浓度导致所有HPV基因表达（包括E6和E7）的显着下调。 5-aza 治疗恢复了HPV+头颈癌细胞中的p53表达和活性，这部分是 负责这些细胞对5-aza的敏感性。除了恢复p53外，5-aza还有毒 HPV+ HNSCC通过创建DNA双链断裂（DSB）。从机械上讲，5-氮杂诱导的DNA HPV+ HNSCC中的DSB取决于转录和复制以及胞苷的过表达 Deaminase，Apobec3b（A3b）。 A3B的实验耗竭抑制了5-Aza毒性并减少了DSB 形成，但也表明未处理的HPV+ HNSCC依赖于A3B来进行克隆发育。这 观察到未处理的HPV+ HNSCC依赖于A3B，但A3B有助于5-Aza毒性和 DSB提示使用5-Aza治疗时A3B依赖性的合成致死性，并且A3B可以用作一个 反应的生物标志物。用5-aza携带HPV+肿瘤的小鼠显示出明显的肿瘤生长 抑制并阻止检测到循环肿瘤细胞。使用HNSCC患者的窗户临床试验使用 标准剂量5天实现的脱甲基化（线1）类似于我们的体外实验和 确认5-aza治疗：1）显着降低了HPV基因的表达； 2）重新激活p53; 3） 活化的胱天蛋白酶，4）并抑制了HPV+ HNSCC中的基质金属蛋白酶表达。该提议是 旨在确定脱甲基化引起的HPV癌基因下调的分子机制， 阐明A3B在5-Aza诱导的合成致死性和DNA DSB形成中的作用，确定 HPV+ HNSCC中免疫细胞浸润的脱甲基化，并探索单独或单独或在 与化学治疗剂结合抑制与HPV相关的HNSCC转移并抑制生长 使用患者衍生的异种移植物。这些研究将为HPV+进行新的合理靶向疗法提供基础 HNSCC，迫切需要治疗复发或转移性HPV+ HNSCC的患者 减少与当前疗法相关的毒性。

项目成果

Direct Comparison of HPV16 Viral Genomic Integration, Copy Loss, and Structural Variants in Oropharyngeal and Uterine Cervical Cancers Reveal Distinct Relationships to E2 Disruption and Somatic Alteration.

• DOI: 10.3390/cancers14184488
• 发表时间: 2022-09-16
• 期刊: Cancers
• 影响因子: 5.2

Comprehensive Viral Genotyping Reveals Prognostic Viral Phylogenetic Groups in HPV16-Associated Squamous Cell Carcinoma of the Oropharynx.

综合病毒基因分型揭示了 HPV16 相关口咽鳞状细胞癌的预后病毒系统发育群。

• DOI: 10.1158/1541-7786.mcr-21-0443
• 发表时间: 2022
• 期刊: Molecular cancer research : MCR
• 作者: Schrank,TravisP;Landess,Lee;Stepp,WesleyH;Rehmani,Hina;Weir,WilliamH;Lenze,Nicholas;Lal,Asim;Wu,Di;Kothari,Aditi;Hackman,TrevorG;Sheth,Siddharth;Patel,Shetal;Jefferys,StuartR;Issaeva,Natalia;Yarbrough,WendellG
• 通讯作者: Yarbrough,WendellG

Therapeutic Targeting of FGFR Signaling in Head and Neck Cancer.

• DOI: 10.1097/ppo.0000000000000615
• 发表时间: 2022-09-01
• 期刊: Cancer journal (Sudbury, Mass.)