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：干细胞疗法的承诺尚未完全实现，部分原因是 缺乏适当的临床前模型。由于相似性，猪是人类生物学的绝佳模型 大小，生理和遗传学的 有效地转化为人类临床试验的数据。因此，猪模型更准确地建模人类 需要迫切需要改善再生医学治疗剂的研究结果并最大化安全 翻译到诊所。由于SCID猪模型在异种移植研究中表现出了希望，因此建立了进一步的需求 作为人源化猪作为第二代模型和改进的方法，以常规的后部免疫缺陷猪 用于测量植入治疗细胞的生存，安全性和潜在功效。我们专注于 在临床前再生医学研究中产生这种新知识并证明猪的使用。 我们已经开发了生物培养设施和协议，以增加Artemis（Art） - 突出SCID猪。我们 通过人类诱导的成功异位植入，已经证明了该模型的意义 多能干细胞（IPSC）和人类癌细胞系，以及原位移植的生存 人皮。为了改善模型，我们在男性艺术中突变了白介素2受体伽马基因（IL2RG） 空背景。 ART NULL IL2RG NULL PIG具有预期的T-B-NK-表型，我们已经成功 在这些ART-IL2RG猪中显示出部分人免疫系统的发展。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/功效测试和 人性化仍然是利用这些研究模型的转化医学的先决条件。 本应用程序的具体目标是a）验证SCID猪模型用于细胞的临床前测试和 组织异种移植物，b）使用多种新颖的人性化方法来确定人类的程度 SCID猪中的造血作用，c）建立第二代SCID猪模型管理方案 多个生物内在设施。拟议的研究的理由是，我们的杂种scid猪可能会更多 与小鼠相比，准确反映了拟议的干细胞衍生疗法在人类中的作用。这 项目具有创新性，因为我们将使用一种集成的方法来结合模型异种移植的研究 潜在的研究重点是改善免疫缺陷猪使用的方案，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， 有关异种移植和人性化率的数据以及相关的动物程序，这将是非常需要的 基于SCID的建模，用于研究干细胞疗法的安全性和功效。这些独特的资源 预计将在加速再生医学研究中的翻译中产生重大影响 诊所。

项目成果

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

• DOI: 10.1007/s00335-021-09907-y
• 发表时间: 2022-03
• 期刊: Mammalian genome : official journal of the International Mammalian Genome Society
• 作者: Boettcher AN;Schachtschneider KM;Schook LB;Tuggle CK
• 通讯作者: Tuggle CK

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.

• 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