Epigenetic Therapy to Treat Radiation-induced Xerostomia

表观遗传疗法治疗放射引起的口干症

• 批准号: 10646364
• 负责人: Isabelle M.A. Lombaert
• 金额: $ 42.41万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-01 至 2027-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10646364
• 关键词: AQP1 gene; Adverse effects; Affect; CRISPR/Cas technology; Capsid; Cell Culture Techniques; Cells; Chronic; Climacteric; Clinical; Clustered Regularly Interspaced Short Palindromic Repeats; CpG Islands; Cytosine; DNA; DNA Methylation; DNA Sequence; Data; Dental caries; Diagnosis; Disease; Dose; Epigenetic Process; Epithelial Cells; Epithelium; Exclusion; Exposure to; FDA approved; Foundations; Gene Expression; Gene Silencing; Gene Transfer; Genes; Gland; Goals; Grant; Guide RNA; Head and Neck Cancer; Human; Immune; Immune Evasion; Immune response; Immunity; In Vitro; Inflammatory Response; Mediator; Messenger RNA; Methods; Methylation; Miniature Swine; Nature; Non-Viral Vector; Oral cavity; Osmosis; Pain; Patients; Phase; Plasmids; Production; Promoter Regions; Proteins; RNA; Radiation; Radiation Dose Unit; Radiation induced damage; Radiation therapy; Reporting; Saliva; Salivary; Salivary Glands; Speech; Symptoms; System; Techniques; Technology; Testing; Therapeutic; Time; Tissues; Viral; Viral Vector; Water; Water Movements; Xerostomia; bisulfite sequencing; chromatin remodeling; demethylation; drinking water; epigenetic therapy; epigenome editing; expression vector; fitness; gene therapy; head and neck cancer patient; histone modification; in vivo; interest; next generation; non-viral gene therapy; oral infection; overexpression; palliative; pre-clinical; promoter; regenerative; regenerative therapy; side effect; success; therapeutic gene; tool; transcriptome sequencing; treatment planning; ultrasound; water channel; whole genome

ABSTRACT Most head-and-neck cancer patients receiving radiotherapy treatment develop life-long adverse effects. Side effects presented as hyposalivation and chronic xerostomia result from radiation damage to nearby salivary glands. Limited palliative options are currently available for these patients, reinforcing the need for regenerative therapies. Our regenerative efforts, using non-viral gene therapy to overexpress water channel Aquaporin-1 (AQP1) in radiation-surviving salivary gland epithelial cells, showed restorative results by increasing salivary flow into the oral cavity. However, as both viral and non-viral vectors are restricted by the ability to provide long-term expression, a next generation of AQP1 therapy that results in sustained gene expression awaits. Our preliminary data demonstrates that methylation of the endogenous AQP1 promoter in epithelial cells is an important mediator of gene expression. In addition, the methylation status alters after exposure to radiation. Based on these findings, we hypothesize that epigenetic altering of the AQP1 promoter in radiation-surviving epithelial cells facilitates a change towards sustained endogenous AQP1 expression. To test this hypothesis, we adapted a new CRISPR method and aim to (1) demonstrate the negative correlation between the level of methylation and AQP1 expression, and (2) develop in vitro and in vivo methods to optimally alter the methylation level of the AQP1 promoter in irradiated epithelial cells. These studies bring in vivo epigenetic editing to a reality as a means of modulating endogenous gene expression and sustained water movement in damaged salivary glands. As a result, this continuous production of salivary flow can resolve xerostomia in radiated head-and-neck cancer patients in a long-term setting.

摘要 大多数接受放射治疗的头颈部癌症患者会产生终身不良反应。 副作用表现为唾液分泌减少和慢性口干，这是由于辐射损伤了周围组织， 唾液腺目前这些患者的姑息治疗选择有限，加强了对以下方面的需求： 再生疗法我们的再生努力，使用非病毒基因疗法，过度表达水通道 放射存活唾液腺上皮细胞中的水通道蛋白-1（AQP 1），通过 增加唾液流入口腔。然而，由于病毒和非病毒载体都受到限制， 提供长期表达的能力，下一代AQP 1治疗，导致持续的基因表达， 表情等待。 我们的初步数据表明，上皮细胞中内源性AQP 1启动子的甲基化， 基因表达的重要媒介。此外，甲基化状态在暴露于辐射后改变。 基于这些发现，我们假设辐射存活的细胞中AQP 1启动子的表观遗传改变， 上皮细胞促进朝向持续内源性AQP 1表达的变化。为了验证这个假设， 我们采用了一种新的CRISPR方法，目的是（1）证明蛋白质水平之间的负相关性。 甲基化和AQP 1表达，和（2）开发体外和体内方法，以最佳地改变AQP 1的表达。 AQP 1基因启动子甲基化水平。 这些研究使体内表观遗传编辑成为现实，作为调节内源性基因的手段， 表达和持续的水运动受损的唾液腺。因此，这种连续生产 唾液流量的增加可以在长期环境中解决放射性头颈癌患者的口干症。