Use of skin grafts programmed to express VEGF-C with biosensor feedback regulation to treat lymphedema

使用编程表达 VEGF-C 并具有生物传感器反馈调节的皮肤移植来治疗淋巴水肿

• 批准号: 10042514
• 负责人: Shailesh Agarwal
• 金额: $ 19.9万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10042514
• 关键词: Address; Adenovirus Vector; Adherence; Affect; Attention; Binding; Biological; Biological Assay; Biological Availability; Biological Products; Bioreactors; Biosensor; Butyrylcholinesterase; CRISPR interference; CRISPR/Cas technology; Cancer Patient; Chronic Disease; Clinical; Cocaine Abuse; Cocaine Dependence; Consumption; Cultured Cells; DNA; DNA Sequence Alteration; Deposition; Dermal; Diabetes Mellitus; Doxycycline; Elements; Etiology; Excision; Feedback; Firefly Luciferases; Gene Delivery; Genes; Genomics; Goals; Growth Factor; Guide RNA; Hindlimb; Histologic; Human; Hypertrophy; Image; Immune response; Impairment; In Vitro; Individual; Infection; Injections; Investigation; Investments; Laboratories; Lead; Limb structure; Lymph; Lymph node excision; Lymphangiogenesis; Lymphatic; Lymphedema; Manual Lymphatic Drainage; Messenger RNA; Microscope; Modeling; Monitor; Morbidity - disease rate; Morphology; Mus; Musculoskeletal; Neoplasm Metastasis; Operative Surgical Procedures; Pain; Patient Dropouts; Patients; Pharmaceutical Preparations; Production; Proteins; Pump; Regulation; Resources; Response Elements; Risk; Series; Skin; Skin graft; Surgical Equipment; Swelling; System; Technology; Testing; Tetanus Helper Peptide; Therapeutic; Time; Tissues; Training; Transgenes; Translations; United States; VEGFC gene; Variant; Vascular Endothelial Growth Factor C; base; cancer surgery; chronic wound; clinical translation; congenital anomaly; design; epidermal stem cell; experience; experimental study; gene product; glucagon-like peptide 1; improved; in vivo; inducible gene expression; instrument; interest; lymph nodes; lymphangiosarcoma; lymphatic drainage; lymphatic vessel; mouse model; new technology; novel; open wound; patient population; prevent; programs; promoter; recurrent infection; relapse risk; scaffold; skills; stem cells; success; transgene expression; translational impact

PROJECT SUMMARY Lymphedema is a morbid musculoskeletal and skin condition affecting a broad population of patients. In the United States, this condition primarily affects individuals who have had lymph node surgery for cancer management; worldwide over 300 million individuals are affected due to lymphatic infection. The affected limb becomes swollen, and develops skin thickening and fibroadipose tissue deposition. Patients experience pain, impaired limb function, chronic wounds, and are at risk for secondary tumors (lymphangiosarcoma). Unfortunately, current therapeutic strategies to treat lymphedema have had limited success. Non-operative management including compression wraps, manual lymphatic drainage, and pneumatic pumps require strict adherence and are time-consuming for patients. Surgical approaches have demonstrated efficacy but require specialized surgical equipment and technical training, limiting the ability to provide this option to the 5+ million affected individuals in the United States. Vascular endothelial growth factor-C (VEGF-C) is a known pro- lymphangiogenic growth factor which has been studied as a drug-based approach to treat secondary lymphedema. Delivery strategies such as adenoviral vectors, scaffolds, and repeat injection have provided important proof-of-concept support for VEGF-C. However, clinical translation has not been realized. Over the past several years, our laboratory has developed a novel technology enabling epidermal stem cells to undergo gene-editing to express biologics of interest. These stem cells are cultured in vitro to produce skin grafts which can be applied to the patient. Our approach has demonstrated efficacy treating diabetes and cocaine abuse through systemic bioavailability in mouse models. In this proposal, we will build on our previous findings to bring this technology closer to clinical utility while broadening its application. First, we will program epidermal stem cells to express VEGF-C and culture these stem cells in vitro, to form skin grafts. To bring us closer to clinical translation, epidermal stem cells will be isolated from human skin. We will demonstrate that skin grafts can produce growth factors for local bioavailability, thereby expanding the use of this technology. We will use these skin grafts in a model of hindlimb lymphedema, thereby demonstrating that this technology has the potential for clinical utility to treat lymphedema isolated to a unilateral limb. Second, we will advance the current skin graft technology by introducing a biosensor mechanism which provides negative feedback regulation of VEGF-C expression. This biosensor will positively impact the translational potential of this therapy, and have broader utility for translation of in vivo “bioreactor” grafts. Upon conclusion of this study, we will have addressed a challenging morbidity experienced by cancer patients (secondary lymphedema), using a novel technology (programmed epidermal stem cells) to expand its utility to include growth factors (VEGF-C) and local delivery.

项目摘要 淋巴水肿是一种病态的肌肉骨骼和皮肤状况，影响广泛的患者。在 美国，这种情况主要影响接受过淋巴结手术的个体 管理;由于淋巴感染，全球超过3亿个人受到影响。受影响的肢体 变得肿胀，形成皮肤增厚和纤维粉蛋白组织的沉积。患者疼痛， 肢体功能受损，慢性伤口，并且有继发性肿瘤的风险（淋巴管肉瘤）。 不幸的是，当前治疗淋巴水肿的治疗策略取得了有限的成功。非手术 管理包括压缩膜，手动淋巴引流和气动泵需要严格 依从性，对患者来说是耗时的。手术方法表明了效率，但需要 专门的手术设备和技术培训，限制了向500万以上提供此选项的能力 在美国影响个人。血管内皮生长因子-C（VEGF-C）是已知的促值 淋巴血管生长因子已被研究为一种基于药物治疗次级的药物 淋巴水肿。提供了腺病毒载体，脚手架和重复注射等递送策略 重要的概念证明支持VEGF-C。但是，尚未实现临床翻译。 在过去的几年中，我们的实验室开发了一种新技术，可以使表皮干细胞能够 要经过基因编辑以表达感兴趣的生物学。这些干细胞在体外培养以产生皮肤 可以应用于患者的移植物。我们的方法表明治疗糖尿病的效率和 可卡因通过鼠标模型中的全身生物利用度滥用。在此提案中，我们将基于以前的 在扩大其应用的同时，使这项技术更接近临床实用性的发现。 首先，我们将对表皮干细胞进行编程以表达VEGF-C并在体外培养这些干细胞，以形成 皮肤移植。为了使我们更接近临床翻译，表皮干细胞将从人的皮肤中分离出来。我们 将证明皮肤移植物可以为局部生物利用度产生增长因素，从而扩大使用 这项技术。我们将在后肢淋巴水肿的模型中使用这些皮肤移植物，从而证明 该技术有可能使临床实用性处理分离到单侧肢体的淋巴水肿。 其次，我们将通过引入生物传感器机制来推进当前的皮肤移植技术 提供VEGF-C表达的负反馈调节。该生物传感器将对 这种疗法的翻译潜力，并具有更广泛的效用，用于翻译体内“生物反应器”移植物。 这项研究结束后，我们将解决癌症患者遇到的挑战发病率 （次要淋巴水肿），使用一种新技术（编程表皮干细胞）将其实用性扩展到 包括生长因子（VEGF-C）和本地分娩。