The effect of HPV on apoptosis and cellular proliferation in laryngeal papillomas

HPV对喉乳头状瘤细胞凋亡和增殖的影响

• 批准号: 8190228
• 负责人: Michael P Underbrink
• 金额: $ 22.13万
• 依托单位: UNIVERSITY OF TEXAS MED BR GALVESTON
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-01 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8190228
• 关键词: Address; Adjuvant; Adult; Affect; Apoptosis; Apoptotic; Area; Binding; Biological Assay; Biometry; Caspase; Cell Communication; Cell Culture Techniques; Cell Cycle; Cell Proliferation; Cell physiology; Cells; Child; Clinical; Confocal Microscopy; Cytochromes; DNA Damage; Data; Debridement; Disease; Epidemiology; Epithelium; Functional disorder; Funding; Genes; Genotoxic Stress; Goals; HIV; HPV-High Risk; Hoarseness; Human; Human Papillomavirus; Human papilloma virus infection; Human papillomavirus 11; Human papillomavirus 6; Immunoblotting; Immunohistochemistry; Impairment; Individual; Infection; K-Series Research Career Programs; Knowledge; Larynx; Lead; Learning; Life Cycle Stages; Low risk HPV; Malignant neoplasm of cervix uteri; Maps; Mediating; Mediator of activation protein; Medical; Mentors; Modeling; Molecular; Molecular Analysis; Molecular Biology; Operative Surgical Procedures; Otolaryngology; Outcome; Papilloma; Papillomavirus Transforming Protein E6; Pathway interactions; Patients; Physicians; Positioning Attribute; Process; Proliferating; Property; Proteins; Recurrence; Research; Research Design; Research Project Grants; Research Training; Residencies; Residual state; Risk; Scientist; Skin; Squamous Papilloma of the Larynx; Supervision; Surgeon; Techniques; Testing; Therapeutic; Therapeutic Agents; Training; Undifferentiated; Upper respiratory tract; Virus; Virus Diseases; Voice; Voice Disorders; Writing; airway obstruction; bak protein; career; clinical efficacy; clinically relevant; cost; effective therapy; experience; high risk; human RCN2 protein; human UBE3A protein; improved; innovation; insight; interest; keratinocyte; meetings; molecular pathology; novel; novel therapeutics; post-doctoral training; prevent; respiratory; response; responsible research conduct; skills; small hairpin RNA; success; treatment effect; tumor; tumor growth; ubiquitin ligase; ubiquitin-protein ligase; virology; vocal cord

DESCRIPTION (provided by applicant): The overall goal of this Mentored Clinical Scientist Research Career Development Award proposal is to assist in establishing the independent research career of the candidate in the study of virus-host cell interactions, specifically of the cellular processes contributing to vocal cord dysfunction and upper airway obstruction caused by human papillomavirus (HPV) infections that lead to recurrent respiratory papillomatosis (RRP). The candidate is a board-certified otolaryngology surgeon with a focus on the management of voice disorders and difficult airways. Besides his clinical residency in otolaryngology, the candidate received postdoctoral training in the molecular biology of genus beta HPVs and the mechanisms by which they induce cell proliferation and block apoptosis after DNA damage - issues similar to those to be studied in the research component of this project, focused on the RPP-related genus alpha HPVs. The training and research components of the project are under the supervision of the co-mentors, Dr. Stephen Tyring and Dr. Thomas Albrecht. In addition to the mentored research component, the training plan calls for didactic coursework in such areas as virology, biostatistics, scientific writing, and several aspects of the responsible conduct of research, as well as attending relevant local and national meetings. The research project focuses on elucidating the mechanisms by which RPP-related HPVs block apoptosis and induce cellular proliferation in human keratinocytes. In high-risk HPV infections, the E6 viral protein targets the pro-apoptotic cellular Bak protein (as well as p53) for degradation, and so prevents the normal process of apoptosis from being induced by genotoxic stress. The candidate's preliminary data show that the RRP-related HPVs also retain the ability to target Bak for degradation. Aim 1 of the research project is to use immunoblotting and shRNA knockdown to determine the ability of HPV6 and HPV11 E6 proteins to blunt the apoptotic response, and also to determine the mechanism for Bak degradation. Aim 2 is to use a combination of protein pull down assays to identify conserved (Bak, E6AP) and novel binding partners of the 6E6 and 11E6 proteins. Finally, Aim 3 is to use immunostaining techniques to determine the effects of treatments targeted towards the induction of the extrinsic and/or intrinsic apototis pathways in both undifferentiated and differentiated E6-expressing cell cultures. Upon successful completion of this project, there will be a much better understanding of the molecular biology and pathogenic mechanisms of the "low- risk," RPP-related HPVs; this information will be invaluable in developing effective, function-sparing treatments for this presently incurable disease process. Further, the candidate will have obtained the preliminary data needed to compete successfully for R01 funding, and the skill set necessary for him to achieve his long-range goal - success as an established, extramurally funded surgeon-scientist focused on effective treatments for RPP and other laryngeal-related diseases. PUBLIC HEALTH RELEVANCE: Recurrent respiratory papillomatosis (RRP) is characterized by the growth of tumors in the upper respiratory tract, caused by infection with the human papilloma virus (HPV). Understanding the effects of HPV6 and HPV11 infection in the airway cells (keratinocytes) will clarify how RRP tumors proliferate in the upper respiratory tract and should lead to the discovery of novel, targeted therapies that can be used for the safe, voice-preserving treatment of this disease. Additionally, principles learned through the molecular study of RRP-associated HPV-host interactions will undoubtedly impact the study and treatment of other virus-induced diseases.

描述（由申请人提供）：本指导临床科学家研究职业发展奖提案的总体目标是协助候选人在病毒与宿主细胞相互作用的研究中建立独立的研究生涯，特别是有助于发声的细胞过程人乳头瘤病毒 (HPV) 感染引起的脊髓功能障碍和上呼吸道阻塞，导致复发性呼吸道乳头状瘤病 (RRP)。候选人是经过委员会认证的耳鼻喉外科医生，专注于声音障碍和困难气道的治疗。除了耳鼻喉科临床住院医师实习之外，该候选人还接受了 β HPV 属分子生物学及其在 DNA 损伤后诱导细胞增殖和阻止细胞凋亡的机制方面的博士后培训 - 与本项目研究部分要研究的问题类似，重点关注 RPP 相关的 α HPV 属。该项目的培训和研究部分由共同导师 Stephen Tyring 博士和 Thomas Albrecht 博士监督。除了指导研究部分外，培训计划还要求在病毒学、生物统计学、科学写作和负责任的研究行为的几个方面进行教学课程，以及参加相关的地方和国家会议。该研究项目的重点是阐明 RPP 相关 HPV 阻止人类角质形成细胞凋亡并诱导细胞增殖的机制。在高危 HPV 感染中，E6 病毒蛋白以促凋亡细胞 Bak 蛋白（以及 p53）为目标进行降解，从而阻止基因毒性应激诱导正常的细胞凋亡过程。该候选者的初步数据表明，RRP 相关 HPV 还保留了靶向 Bak 进行降解的能力。该研究项目的目标 1 是利用免疫印迹和 shRNA 敲除来确定 HPV6 和 HPV11 E6 蛋白减弱细胞凋亡反应的能力，并确定 Bak 降解的机制。目标 2 是结合使用蛋白质下拉测定来鉴定 6E6 和 11E6 蛋白质的保守（Bak、E6AP）和新型结合伴侣。最后，目标 3 是使用免疫染色技术来确定针对在未分化和分化的 E6 表达细胞培养物中诱导外在和/或内在凋亡途径的治疗效果。该项目成功完成后，将对“低风险”RPP 相关 HPV 的分子生物学和致病机制有更好的了解；这些信息对于开发有效的、保留功能的治疗方法来治疗这种目前无法治愈的疾病过程非常宝贵。此外，候选人将获得成功竞争 R01 资金所需的初步数据，以及实现其长期目标所需的技能——作为一名成功的、获得外部资助的外科医生科学家，专注于 RPP 和 RPP 的有效治疗。其他喉部相关疾病。 公共卫生相关性：复发性呼吸道乳头状瘤病 (RRP) 的特点是上呼吸道肿瘤生长，由人乳头瘤病毒 (HPV) 感染引起。了解 HPV6 和 HPV11 感染对气道细胞（角质形成细胞）的影响将阐明 RRP 肿瘤如何在上呼吸道增殖，并有助于发现新的靶向疗法，用于安全、保留声音的治疗这种病。此外，通过 RRP 相关 HPV-宿主相互作用的分子研究学到的原理无疑将影响其他病毒引起的疾病的研究和治疗。