Developing second generation SCID pig models: filling the gaps to improve translation of therapeutics in regenerative medicine

开发第二代 SCID 猪模型：填补空白，改善再生医学疗法的转化

• 批准号: 10559550
• 负责人: Christopher Tuggle
• 金额: $ 73.04万
• 依托单位: IOWA STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-02-15 至 2025-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10559550
• 关键词: Academia; Acceleration; Advisory Committees; Age; Age Months; Animal Model; Animals; B-Lymphocytes; Bone Marrow; Breeding; CD3 Antigens; Cancer cell line; Cardiac Myocytes; Cell Therapy; Cells; Clinic; Clinical Trials; Collaborations; Communication; Communities; Community Medicine; Data; Drug Implants; Engraftment; Family suidae; Fetus; Funding; Future; Generations; Genes; Genetic; Germ; Goals; Hematopoiesis; Hematopoietic stem cells; Human; Human Biology; Immune; Immune system; Immunologic Deficiency Syndromes; Immunophenotyping; In Vitro; Industry; Injections; Interleukin 2 Receptor Gamma; Knowledge; Link; Location; Longevity; Measurement; Methods; Modeling; Mus; Mutate; Natural Killer Cells; Neonatal; Operative Surgical Procedures; Outcomes Research; Phenotype; Physiology; Population; Pre-Clinical Model; Preclinical Testing; Preparation; Procedures; Protocols documentation; R24; Recommendation; Regenerative Medicine; Reporting; Research; Resources; Rodent Model; Safety; Severe Combined Immunodeficiency; Skin; Systems Development; T-Lymphocyte; Techniques; Testing; Therapeutic; Thymus Gland; Tissues; Translating; Translations; Transplantation; Xenograft Model; Xenograft procedure; artemis; biocontainment facility; cancer therapy; conditioning; efficacy testing; fetal; genetic resource; genome editing; human model; improved; in vivo; induced pluripotent stem cell; innovation; interest; intraperitoneal; male; mutant; neonatal human; novel; null mutation; pathogen; porcine model; postnatal; pre-clinical; safety study; safety testing; stem cell therapy; stem cells; tool; translational medicine; translational model; translational therapeutics; transplant model

Project Summary/Abstract: The promise of stem cell therapies has not been fully realized, due in part to a paucity of appropriate pre-clinical models. The pig is an excellent model of human biology, due to similarities of size, physiology, and genetics, and in some cases may be superior to rodent models, which often fail to provide data which effectively translates to human clinical trials. Thus, pig models that more accurately model humans are critically needed to improve research outcomes of regenerative medicine therapeutics and maximize safe translation to the clinic. As SCID pig models show promise in xenograft studies, this establishes further needs for humanized pigs as second-generation models and improved methods to routinely rear immunodeficient pigs for measurement of survival, safety, and potential efficacy of implanted therapeutic cells. We are focused on generating such new knowledge and demonstrating SCID pig use in preclinical regenerative medicine research. We have developed biocontainment facilities and protocols to raise Artemis (ART)-mutant SCID pigs. We have demonstrated this model's significance through successful ectopic engraftment with human induced pluripotent stem cells (iPSC) and human cancer cell lines, as well as survival of orthotopically transplanted human skin. To improve the model, we mutated the Interleukin 2 Receptor gamma gene (IL2RG) in a male ART null background. The ART null IL2RG null pig has the expected T-B-NK- phenotype, and we have successfully demonstrated partial human immune system development in these ART-IL2RG pigs. Thus we have taken pioneering steps to establish a pig SCID model, but improvements in xenograft safety/efficacy testing and humanization remain prerequisites to harness these research models for translational medicine. The specific objectives of this application are to a) validate SCID pig models for preclinical testing of cell and tissue xenografts, b) use multiple novel humanization approaches to determine the extent of human hematopoiesis in SCID pigs, and c) establish second-generation SCID pig model management protocols at multiple biocontainment facilities. The rationale for the proposed research is that our outbred SCID pig may more accurately reflect how proposed stem cell derived therapies will function in humans compared to mice. This project is innovative because we will use an integrated approach to combine research on the model's xenograft potential with research focused on protocols for improving the use of immunodeficient pigs, including humanization methods. We expect that the successful completion of this project will create genetic resources, data on xenograft and humanization rates, and associated animal procedures that will be highly desirable for SCID based modeling for research on the safety and efficacy of stem cell therapeutics. These unique resources are expected to have a significant impact in accelerating the translation of regenerative medicine research into the clinic.

项目摘要/摘要：干细胞疗法的前景尚未完全实现，部分原因是 缺乏适当的临床前模型。由于相似之处，猪是人类生物学的优秀模型 大小、生理学和遗传学方面的差异，在某些情况下可能优于啮齿动物模型，而啮齿动物模型通常无法提供 有效转化为人体临床试验的数据。因此，猪模型可以更准确地模拟人类 迫切需要改善再生医学疗法的研究成果并最大限度地提高安全性 翻译到诊所。随着 SCID 猪模型在异种移植研究中显示出前景，这确立了进一步的需求 作为第二代模型的人源化猪和常规饲养免疫缺陷猪的改进方法 用于测量植入治疗细胞的存活率、安全性和潜在功效。我们专注于 产生此类新知识并展示 SCID 猪在临床前再生医学研究中的应用。 我们开发了生物防护设施和方案来饲养 Artemis (ART) 突变 SCID 猪。我们 通过人类诱导的成功异位植入证明了该模型的重要性 多能干细胞 (iPSC) 和人类癌细胞系，以及原位移植的存活率 人类的皮肤。为了改进模型，我们对男性 ART 中的白细胞介素 2 受体 γ 基因 (IL2RG) 进行了突变 空背景。 ART null IL2RG null 猪具有预期的 T-B-NK- 表型，我们成功地 在这些 ART-IL2RG 猪中证明了部分人类免疫系统的发育。因此我们采取了 建立猪 SCID 模型的开创性步骤，但异种移植安全性/功效测试和 人性化仍然是利用这些研究模型进行转化医学的先决条件。 该应用的具体目标是 a) 验证用于细胞和临床前测试的 SCID 猪模型 组织异种移植，b) 使用多种新颖的人源化方法来确定人源化的程度 SCID 猪的造血功能，以及 c) 建立第二代 SCID 猪模型管理方案 多个生物防护设施。拟议研究的基本原理是，我们的远交 SCID 猪可能更 准确地反映了所提出的干细胞衍生疗法与小鼠相比如何在人类中发挥作用。这 该项目具有创新性，因为我们将使用综合方法来结合模型异种移植的研究 重点研究改善免疫缺陷猪的使用方案的潜力，包括 人性化方法。我们预计该项目的成功完成将创造遗传资源， 有关异种移植和人源化率的数据，以及相关的动物程序，这些数据对于 基于 SCID 的模型，用于研究干细胞疗法的安全性和有效性。这些得天独厚的资源 预计将对加速再生医学研究的转化产生重大影响 诊所。

项目成果

Swine models for translational oncological research: an evolving landscape and regulatory considerations.

用于转化肿瘤学研究的猪模型：不断变化的格局和监管考虑。

• 发表时间: 2022-03
• 作者: Boettcher, Adeline N;Schachtschneider, Kyle M;Schook, Lawrence B;Tuggle, Christopher K
• 通讯作者: Tuggle, Christopher K

Effect of ARTEMIS (DCLRE1C) deficiency and microinjection timing on editing efficiency during somatic cell nuclear transfer and in vitro fertilization using the CRISPR/Cas9 system.

ARTEMIS (DCLRE1C) 缺陷和显微注射时机对使用 CRISPR/Cas9 系统的体细胞核移植和体外受精过程中编辑效率的影响。

• DOI: 10.1016/j.theriogenology.2021.04.003
• 发表时间: 2021-08
• 期刊: Theriogenology
• 影响因子: 2.8
• 作者: Li Y;Adur MK;Wang W;Schultz RB;Hale B;Wierson W;Charley SE;McGrail M;Essner J;Tuggle CK;Ross JW
• 通讯作者: Ross JW