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.

项目总结 淋巴水肿是一种病态的肌肉骨骼和皮肤疾病，影响着广泛的患者群体。在 在美国，这种情况主要影响因癌症接受过淋巴结手术的个人 管理；全世界有超过3亿人因淋巴感染而受到影响。患肢 变得肿胀，并形成皮肤增厚和纤维脂肪组织沉积。病人会经历痛苦， 肢体功能受损，慢性伤口，并有继发性肿瘤(淋巴管肉瘤)的风险。 不幸的是，目前治疗淋巴水肿的策略收效甚微。非运营 包括压缩包扎、手动淋巴引流和气动泵在内的管理要求严格 依从性对患者来说是很耗时的。手术方法已经证明是有效的，但需要 专门的手术设备和技术培训，将提供这一选择的能力限制在500多万人 在美国受影响的个人。血管内皮生长因子-C(VEGF-C)是一种已知的促血管生成因子。 淋巴管生长因子作为一种药物治疗继发性肝癌的研究进展 淋巴浮肿。已经提供了如腺病毒载体、支架和重复注射等递送策略 对血管内皮生长因子-C的重要概念验证支持然而，临床翻译尚未实现。 在过去的几年里，我们的实验室开发了一种新的技术，使表皮干细胞 进行基因编辑以表达感兴趣的生物制品。这些干细胞在体外培养成皮肤。 可以应用于患者的移植物。我们的方法已经证明了治疗糖尿病和 可卡因滥用通过小鼠模型的全身生物利用度。在这项提案中，我们将在以前的基础上 使这项技术更接近临床实用，同时扩大其应用范围的发现。 首先，我们将编程表皮干细胞表达血管内皮生长因子-C，并在体外培养这些干细胞，以形成 皮肤移植。为了让我们更接近临床移植，我们将从人类皮肤中分离出表皮干细胞。我们 将证明皮肤移植可以产生局部生物利用度的生长因子，从而扩大使用 这项技术。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 这项技术具有治疗孤立于单侧肢体的淋巴水肿的临床实用潜力。 第二，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 提供对血管内皮生长因子-C表达的负反馈调节。这种生物传感器将对 这种疗法的翻译潜力，并对体内“生物反应器”移植物的翻译具有更广泛的实用价值。 在这项研究结束后，我们将解决癌症患者经历的一个具有挑战性的发病率问题。 (继发性淋巴水肿)，使用一种新技术(程序化表皮干细胞)将其用途扩大到 包括生长因子(血管内皮生长因子-C)和局部给药。