Novel non-viral reprogramming strategies to treat Discogenic back pain via engineered extracellular vesicles

通过工程细胞外囊泡治疗椎间盘源性背痛的新型非病毒重编程策略

• 批准号: 10606527
• 负责人: Natalia Higuita-Castro
• 金额: $ 51.73万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-04-10 至 2027-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10606527
• 关键词: Address; Age; Animal Model; Animals; Award; Back Pain; Biological; Blood Vessels; Brachyury protein; Cell Survival; Cell Therapy; Cells; Chronic low back pain; DNA; Data; Development; Disease; Drug Delivery Systems; Economic Burden; Engineering; Extracellular Matrix; Extracellular Matrix Proteins; FOXF1 gene; Functional disorder; Gene Expression; Goals; Growth; Health; Human; Hydration status; In Situ; In Vitro; Inflammation; Inflammatory; Injury; Intervertebral disc structure; Ligands; Low Back Pain; Mechanics; Membrane; Mission; Modeling; Mohawk; Mus; Nerve; Operative Surgical Procedures; Pain; Pathologic; Pathology; Patients; Phenotype; Proteomics; Publishing; Quality of life; Research; Role; Specificity; Structure; Therapeutic; Therapeutic Intervention; Tissues; United States National Institutes of Health; Vertebral column; Viral; Vision; Work; age effect; cell type; clinically relevant; comparative; comparison control; discogenic pain; effective therapy; efficacious treatment; extracellular vesicles; gene delivery system; gene therapy; improved; in vivo; in vivo Model; innovation; intervertebral disk degeneration; minimally invasive; mouse model; neurotrophic factor; non-viral gene delivery; novel; nucleus pulposus; opioid epidemic; opioid use; pain behavior; pain reduction; pain symptom; phenotypic biomarker; programs; protein expression; scleraxis; sex; socioeconomics; success; symptom treatment; targeted delivery; targeted treatment; tool; transcription factor; treatment strategy; uptake

Project Summary/Abstract Chronic low back pain is a debilitating disorder of significant socio-economic importance and a major gateway to opioid use. Therefore, minimally invasive non-addictive treatments that aim to address pain by targeting the underlying disease pathology are critical to improve human health and limit the growing opioid crisis. Intervertebral disc (IVD) degeneration is strongly associated with the pathophysiology of chronic low back pain; specifically extracellular matrix (ECM) breakdown, inflammation, and aberrant nerve/vascular ingrowth, all of which are significantly correlated with “Discogenic back pain” (DBP). Therefore, the overall objective of this proposal is to develop novel non-viral reprogramming-based therapies to convert degenerate nucleus pulposus (NP) and annulus fibrosus (AF) IVD cells associated with DBP, into a healthy pro-anabolic, anti-nerve/vascular phenotype, using engineered extracellular vesicles (eEVs). Our central hypothesis is that non-viral reprogramming will restore IVD structure/mechanical function, and limit nerve/vascular ingrowth associated with pain, by converting the patient’s own degenerate IVD cells into a pro-anabolic phenotype in situ. To date, clinicians do not have access to the necessary biological tools to treat IVD degeneration in patients with DBP. A critical barrier to the success of current biologic strategies involves significant logistical and regulatory challenges such as a lack of sustained drug delivery systems, poor long-term cell viability or viral reprogramming that permanently integrates with the host DNA. Our non-addictive strategy focuses on addressing these limitations. Through our recent R61 award and published work, we have shown that human NP cells can be reprogrammed towards a healthier phenotype using developmental transcription factors Brachyury or FOXF1, showing increased ECM accumulation and decreased catabolic/ inflammatory/ neurotrophic factors - all key features of a healthy IVD. Furthermore, in our mouse DBP in vivo model we have also demonstrated significant improvement in NP tissue hydration and dampened pain behaviors in animals treated with eEVs loaded with FOXF1 for up-to 12 weeks, highlighting the potential of our proposed strategy to restore structure/function of the IVD while reducing pain. However, critical gaps remain such as i) understanding the synergistic reprogramming potential of multiple developmental transcription factors in NP and AF cells, ii) how sex and age influence our strategy, and iii) evaluating therapeutic efficiency using more clinically relevant in vivo animal models that simulate the human condition and functionalized eEVs to deliver transcription factors to specific cell types within the IVD (for example, NP or AF cells). Our first aim investigates the synergistic effects of eEVs loaded with multiple transcription factors using in vitro and in vivo clinically relevant models of DBP, and quantifying efficacy of our strategy via assessment of IVD structure/function and pain markers. The second aim involves functionalizing the eEVs with ligands specific for NP or AF cells to selectively deliver cargo to degenerate NP or AF cells of the IVD in vitro and in vivo.

项目摘要/摘要 慢性下腰痛是一种具有重要社会经济重要性的衰弱障碍，并且是一个主要的门户 使用绿o。因此，旨在通过针对的，旨在解决疼痛的最小侵入性的非依恋治疗方法 潜在的疾病病理学对于改善人类健康和限制不断增长的阿片类药物危机至关重要。 椎间盘（IVD）变性与慢性下背部疼痛的病理生理密切相关。 特别是细胞外基质（ECM）分解，感染和异常神经/血管内部 这与“盘状背痛”（DBP）显着相关。因此，总体目标 建议是开发新型的非病毒重编程疗法来转化退化核 果肉（NP）和环纤维（AF）IVD细胞与DBP相关的细胞，成为健康的促胺， 使用工程细胞外蔬菜（EEV），抗神经/血管表型。我们的中心假设是 该非病毒重新编程将恢复IVD结构/机械功能，并限制神经/血管内部 与疼痛相关，通过将患者自己的退化IVD细胞转化为原位的促胺表型。 迄今为止，临床医生无法使用患者治疗IVD变性的必要生物学工具 与DBP。当前生物学策略成功的关键障碍涉及大量后勤和 监管挑战，例如缺乏持续的药物输送系统，长期细胞活力或病毒 重新编程，该编程与宿主DNA永久整合。我们的非依恋战略重点是 解决这些限制。通过我们最近的R61奖和发表的作品，我们已经证明了人类 使用发育转录因子可以将NP细胞重新编程为更健康的表型 Brachyury或FOXF1，显示出ECM的积累增加，分解代谢/炎症/ 神经营养因素 - 健康IVD的所有关键特征。此外，在我们的鼠标DBP体内模型中，我们有 还显示出动物的NP组织水合和该死的疼痛行为的显着改善 用装有FOXF1的EEV处理最多12周，强调了我们提出的策略的潜力 恢复IVD的结构/功能，同时减轻疼痛。但是，仍然存在关键差距，例如我）理解 NP和AF细胞中多个发育转录因子的协同重编程潜力，II） 性别和年龄如何影响我们的策略，以及iii）使用更临床相关的评估治疗效率 体内动物模型，模拟人类状况和功能化EEV以传递转录因子 对于IVD中的特定细胞类型（例如，NP或AF细胞）。 我们的第一个目标研究了使用多个转录因子的EEV的协同作用 DBP的体外和体内临床相关模型，并通过评估IVD来量化我们策略的效率 结构/功能和疼痛标记。第二个目的涉及用特定于配体的EEV功能功能 NP或AF细胞在体外和体内有选择地输送IVD的NP或AF细胞的货物。