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: 10249235
• 负责人: Shailesh Agarwal
• 金额: $ 21.44万
• 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 2024-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10249235
• 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; 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 Dissections; 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; gene therapy; 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; secondary lymphedema; 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.

项目概要 淋巴水肿是一种病态的肌肉骨骼和皮肤疾病，影响广大患者群体。 在美国，这种情况主要影响因癌症接受淋巴结手术的人 管理；全世界有超过 3 亿人因淋巴感染而受到影响。 变得肿胀，并出现皮肤增厚和纤维脂肪组织沉积，患者会感到疼痛， 肢体功能受损、慢性伤口，并且有继发性肿瘤（淋巴管肉瘤）的风险。 不幸的是，目前治疗淋巴水肿的治疗策略效果有限。 管理包括加压包扎、手动淋巴引流和气动泵，需要严格的管理 手术方法已被证明有效，但需要患者坚持并耗时。 专门的手术设备和技术培训，限制了向 5+ 百万提供这种选择的能力 在美国受影响的个体中，血管内皮生长因子-C (VEGF-C) 是一种已知的亲-血管内皮生长因子。 淋巴管生成因子已被研究作为治疗继发性药物的方法 提供了腺病毒载体、支架和重复注射等递送策略。 VEGF-C 的重要概念验证支持然而，临床转化尚未实现。 在过去的几年里，我们的实验室开发了一种新技术，使表皮干细胞 进行基因编辑以表达感兴趣的生物学特性，这些干细胞在体外培养以产生皮肤。 我们的方法已证明可以有效治疗糖尿病和糖尿病。 通过小鼠模型的全身生物利用度可卡因滥用在本提案中，我们将在之前的基础上进行研究。 研究结果使这项技术更接近临床实用，同时扩大其应用范围。 首先，我们将表皮干细胞编程为表达VEGF-C，并在体外培养这些干细胞，形成 为了使我们更接近临床转化，我们将从人类皮肤中分离出表皮干细胞。 将证明植皮可以产生生长因子以提高局部生物利用度，从而扩大使用范围 我们将在后肢淋巴水肿模型中使用这些皮肤移植物，从而证明这一点。 该技术具有临床应用潜力，可治疗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。 这种疗法的转化潜力，并且对于体内“生物反应器”移植物的转化具有更广泛的用途。 这项研究结束后，我们将解决癌症患者所经历的具有挑战性的发病率 （继发性淋巴水肿），使用新技术（程序化表皮干细胞）将其用途扩展到 包括生长因子 (VEGF-C) 和局部递送。