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: 10438568
• 负责人: Karen S. Anderson
• 金额: $ 40.06万
• 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/10438568
• 关键词: Aftercare; Apoptosis; Azacitidine; Caspase; Cell Death; Cell Line; Cell Survival; Cells; Cervical; Characteristics; Chemotherapy and/or radiation; Clinical Data; 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; Enrollment; 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; Infiltration; Interferon Activation; Interferons; Interstitial Collagenase; Lead; Location; Lymph Node Dissections; Lymphedema; Malignant - descriptor; Malignant Epithelial Cell; 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; Property; Radiation; 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; improved; in vitro Assay; molecular modeling; mouse model; multimodality; novel therapeutics; overexpression; 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.

摘要