基因编辑异种器官移植供体猪的研究进展

doi:10.16174/j.issn.1673-4645.2025.02.001

Abstract

Abstract: Xenotransplantation had attracted much attention as a potential solution to the global organ shortage problem, with gene-edited pigs receiving even more attention as a source of donors. In contrasted to the numerous challenges faced in traditional or gan transplantation, when porcine organs were transplanted into humans, they primarily encounter issued such as immune rejection, coagulation dysfunction, and organ size mismatch. Gene-editing technologies like CRISPR/Cas9 could precisely target relevant gene loci in pigs, enabling targeted improvements in aspects such as the knockout of immunogenic antigens (e.g., knockout of the GGTA1 gene), inhibition of complement activation (e.g., inhibition of the β2M gene), regulation of coagulation function (e.g., regulation of the vWF gene), and control of organ growth (e.g., control of the GHR gene), thereby specifically addressing the problems associated with porcine organ transplantation. Additionally, the introduction of human transgenes(e.g., the hCD47 gene) could further enhance the immune toler ance of porcine organs. Globally, multiple subclinical and clinical studied on the transplantation of genetically edited pig kidneys and livers into humans had been successfully conducted, yielding remarkable progress. However, due to the wide variety of gene-editing tools, there were differences in their mechanisms of action and safety, presenting certain technical and safety thresholds. Therefore, this paper systematically summarized the key genes that need to be edited in donor pigs for xenotransplantation that had entered subclinical and clinical trials and the specific editing methods. It also explored and analyzed the safety and efficiency issues of gene-editing tools, as well as the current opportunities and challenges. On the premise of ensuring the safety and effectiveness of gene editing, this paper aimed to provide some references for optimizing relevant gene-editing methods and promoting subsequent research, so as to advance xenotransplantation technology to a new level.

Key words: pig, gene editing, xenotransplantation, CRISPR/Cas9, animal model

CLC Number:

S828

References

[1] 窦科峰, 张玄, 陶开山. 异种移植存在的问题及国内发展现状[J]. 空军军医大学学报, 2024, 15(1):1-4. DOU KF, ZHANG X, TAO KS. Existing problems and do mestic development of xenotransplantation[J]. Journal of Air Force Medical University, 2024, 15(1):1-4. [2] 张小燕, 王国辉, 韩士超, 等. 国内外异种器官移植的现状及进展[J]. 器官移植, 2024, 15(2):276-281. ZHANG XY, WANG GH, HAN SC, et al. Present situation and progress of xenotransplantation at home and abroad[J]. Organ Transplantation, 2024, 15(2):276-281. [3] 王敏敏. 体外器官支持治疗的进展[J]. 中国血液净化, 2021, 20(8):555-558. WANG MM. The development of extracorporeal organ sup port therapy[J]. Chinese Journal of Blood Purification, 2021, 20(8):555-558. [4] HUANG JF. Expert consensus on clinical trials of human xenotransplantation in China[J]. Health Care Science, 2022, 1(1):7-10. [5] LUNNEY JK, VAN GOOR A, WALKER KE, et al. Importance of the pig as a human biomedical model[J]. Science Transl ational Medicine, 2021, 13(621):eabd5758. [6] 窦科峰, 林智斌, 马先一. 异种移植:从基础到临床的瓶颈与对策[J]. 中国实用外科杂志, 2025, 45(1):5-10. DOU KF, LIN ZB, MA XY. Xenotransplantation: bottlenecks and countermeasures in clinical transformation[J]. Chinese Journal of Practical Surgery, 2025, 45(1):5-10. [7] PORRETT PM, ORANDI BJ, KUMAR V, et al. First clini cal-grade porcine kidney xenotransplant using a human decedent model [J]. American Journal of Transplantation, 2022, 22(4):1037-1053. [8] MONTGOMERY RA, STERN JM, LONZE BE, et al. Results of two cases of pig-to-human kidney xenotransplantation [J]. New England Journal of Medicine, 2022, 386(20):1889-1898. [9] MOAZAMI N, STERN JM, KHALIL K, et al. Pig-to-human heart xenotransplantation in two recently deceased human recipients[J]. Nature Medicine, 2023, 29(8):1989-1997. [10] HEALTH NL. Two-month study of pig kidney xenotrans plantation gives new hope to the future of the organ supply [EB/OL]. https://nyulangone.org/news/two-month-study-pig -kidney-xenotransplantation-gives-new-hope-future-organ-supply.2025-3-31. [11] REGALADO A. A brain-dead man was attached to a gene edited pig liver for three days[EB/OL]. https://www.technol ogyreview.com/2024/01/18/1086791/brain-dead-man-gene-edited-pig-liver.2025-3-31. [12] LOCKE JE, KUMAR V, ANDERSON D, et al. Normal graft function after pig-to-human kidney xenotransplant [J]. JAMA Surgery, 2023, 158(10):1106. [13] MA SJ, QI RC, HAN SC, et al. Plasma exchange and intravenous immunoglobulin prolonged the survival of a porcine kidney xenograft in a sensitized, deceased human recipient [J]. Chinese Medical Journal, 2024:1-15. [14] WANG Y, CHEN G, PAN DK, et al. Pig-to-human kidney xenotransplants using genetically modified minipigs[J]. Cell Reports Medicine, 2024, 5(10):101744. [15] 万恒易. 全球首例！清华长庚董家鸿院士团队合作完成猪—人肝肾联合异种移植 [EB/OL]. https://www.med.tsinghua.edu. cn/info/1405/3275.htm.2025-3-31. WAN HY. World's first! Tsinghua Changgeng Academician Dong Jiahong's team collaborates to complete a combined porcine-human liver and kidney xenotransplantation[EB/OL]. https://www.med.tsinghua.edu.cn/info/1405/3275.htm.2025-3-31. [16] TAO KS, YANG ZX, ZHANG X, et al. Gene-modified pig to-human liver xenotransplantation [J]. Nature, 2025. doi: 10.1038/s41586-025-08799-1. [17] SCHOOL HM. In a first, genetically edited pig kidney is transplanted into human[EB/OL]. https://hms.harvard.edu/news/first-genetically-edited-pig-kidney-transplanted-human. 2025-03-31. [18] HEALTH NL. First-ever combined heart pump & gene-edited pig kidney transplant gives new hope to patient with terminal illness[EB/OL]. https://nyulangone.org/news/node/34872.2025-3-31. [19] 安徽医科大学. 世界首例！我校一附院孙倍成团队成功完成猪肝移植给右叶巨大肝癌病人[EB/OL]. https://www.ahmu.edu. cn/2024/0524/c4325a157504/page.htm.2025-3-31. ANHUI MEDICAL UNIVERSITY. A world first! Sun beicheng's team at the first affiliated hospital of our university successfully completed pig liver transplantation for a patient with giant liver cancer in the right lobe[EB/OL]. https://www.ahmu.edu.cn/2024/0524/c4325a157504/page.htm.2025-3-31. [20] GRIFFITH BP, GOERLICH CE, SINGH AK, et al. Genetically modified porcine-to-human cardiac xenotransplantation[J]. New England Journal of Medicine, 2022, 387(1):35-44. [21] DEBORAH KOTZ. UM medicine faculty-scientists and clinicians perform second historic transplant of pig heart into patient with end-stage cardiovascular disease[EB/OL]. https: //www.medschool.umaryland.edu/news/2023/um-medicine -faculty-scientists-and-clinicians-perform-second-his toric-transplant-of-pig-heart-into-patient-with-end-sta ge-cardiovascular-disease.html.2025-3-31. [22] ANAND RP, LAYER JV, HEJA D, et al. Design and testing of a humanized porcine donor for xenotransplantation[J]. Nature, 2023, 622(7982):393-401. [23] SMOOD B, HARA H, SCHOEL LJ, et al. Genetically-engineered pigs as sources for clinical red blood cell transfusion: what pathobiological barriers need to be overcome?[J]. Blood Reviews, 2019, 35:7-17. [24] DAI YF, VAUGHT TD, BOONE J, et al. Targeted disruption of theα-1, 3-galactosyltransferase gene in cloned pigs[J]. Nature Biotechnology, 2002, 20(3):251-255. [25] LAI LX, KOLBER-SIMONDS D, PARK KW, et al. Production of α-1, 3-galactosyltransferase knockout pigs by nuclear transfer cloning[J]. Science, 2002, 295(5557):1089-1092. [26] CHENG WM, ZHAO H, YU HH, et al. Efficient generation of GGTA1-null Diannan miniature pigs using TALENs combined with somatic cell nuclear transfer[J]. Reproductive Biology and Endocrinology, 2016, 14(1):77. [27] FENG C, LI XR, CUI HT, et al. Highly efficient generation of GGTA1 knockout pigs using a combination of TALEN mRNA and magnetic beads with somatic cell nuclear transfer [J]. Journal of Integrative Agriculture, 2016, 15(7):1540-1549. [28] RYCZEK N, HRYHOROWICZ M, ZEYLAND J, et al. CRISPR/ CAS technology in pig-to-human xenotransplantation research[J]. International Journal of Molecular Sciences, 2021, 22(6):3196. [29] ESTRADA JL, MARTENS G, LI P, et al. Evaluation of human and non-human primate antibody binding to pig cells lacking GGTA1/CMAH/β4GalNT2 genes [J]. Xenotransplantation, 2015, 22(3):194-202. [30] NIU D, WEI HJ, LIN L, et al. Inactivation of porcine endogenous retrovirus in pigs using CRISPR/Cas9 [J]. Science, 2017, 357(6357):1303-1307. [31] YUE YN, XU WH, KAN YN, et al. Extensive germline genome engineering in pigs [J]. Nature Biomedical Engineering, 2021, 5(2):134-143. [32] WANG J, XU K, LIU T, et al. Production and functional verification of 8-gene (ggta1, cmah, β4galnt2, hcd46, hcd55, hcd59, htbm, hcd39)-edited donor pigs for xenotransplan tation [J]. Cell Proliferation, 2025. doi: 10.1111/cpr.70028. Epub ahead of print. [33] HRYHOROWICZ M, LIPI?SKI D, HRYHOROWICZ S, et al. Application of genetically engineered pigs in biomedical research[J]. Genes, 2020, 11(6):670. [34] COOPER DKC, RAZA SS, CHABAN R, et al. Shooting for the moon: genome editing for pig heart xenotransplantation[J]. The Journal of Thoracic and Cardiovascular Surgery, 2023, 166(3):973-980. [35] YUAN YL, CUI YY, ZHAO DY, et al. Complement networks in gene-edited pig xenotransplantation: enhancing transplant success and addressing organ shortage[J]. Journal of Translational Medicine, 2024, 22(1):324. [36] BROOM C, UKNIS ME. Methods of treating antibody-mediated rejection in organ transplant patients with c1-esterase inhibitor[P]. USA: US20150147319, 2015-05-28. [37] HARRIS CL, PETTIGREW DM, LEA SM, et al. Decay-accelerating factor must bind both components of the complement alternative pathway C3 convertase to mediate efficient decay [J]. The Journal of Immunology, 2007, 178(1):352-359. [38] ROOIJAKKERS SHM, VAN STRIJP JAG. Bacterial complement evasion [J]. Molecular Immunology, 2007, 44 (1/2/3): 23-32. [39] JEONG YH, PARK CH, JANG GH, et al. Production of multiple transgenic yucatan miniature pigs expressing human complement regulatory factors, human CD55 CD59 and H-transferase genes[J]. PLoS One, 2013, 8(5):e63241. [40] GOLLACKNER B, GOH SK, QAWI I, et al. Acute vascular rejection of xenografts: roles of natural and elicited xenoreactive antibodies in activation of vascular endothelial cells and in duction of procoagulant activity [J]. Transplantation, 77(11): 1735-1741. [41] WANG Y, DU YN, ZHOU XY, et al. Efficient generation of B2m-null pigs via injection of zygote with TALENs[J]. Scientific Reports, 2016, 6:38854. [42] ZHOU X, LIU Y, TANG C, et al. Generation of genetic modified pigs devoid of GGTA1 and expressing the human leukocyte antigen-G5[J]. Chinese Journal of Biotechnology, 2022, 38(3):1096-1111. [43] IDE K, WANG H, TAHARA H, et al. Role for CD47-SIRPα signaling in xenograft rejection by macrophages [J]. Proceedings of the National Academy of Sciences of the United States of America, 2007, 104(12):5062-5066. [44] LI J, EZZELARAB MB, AYARES D, et al. The potential role of genetically-modified pig mesenchymal stromal cells in xenotransplantation[J]. Stem Cell Reviews and Reports, 2014, 10(1):79-85. [45] PHELPS CJ, BALL SF, VAUGHT TD, et al. Production and characterization of transgenic pigs expressing porcine CTLA4-Ig[J]. Xenotransplantation, 2009, 16(6):477-485. [46] HARA H, WITT W, CROSSLEY T, et al. Human dominant negative class II transactivator transgenic pigs effect on the human anti-pig T-cell immune response and immune status[J]. Immunology, 2013, 140(1):39-46. [47] PUGA YUNG G, BONGONI AK, PRADIER A, et al. Release of pig leukocytes and reduced human NK cell recruitment during ex vivo perfusion of HLA-E/human CD46 double-transgenic pig limbs with human blood[J]. Xenotransplantation, 2018, 25(1). doi: 10.3760/cma.j.issn.1673-4394.2018.01.022. [48] NOWAK-TERPILOWSKA A, LIPINSKI D, HRYHOROWICZM, et al. Production of ULBP1-KO pigs with human CD55 expression using CRISPR technology[J]. Journal of Applied Animal Research, 2020, 48(1):93-101. [49] SHIMIZU A, YAMADA K. Pathology of renal xenograft rejection in pig to non-human primate transplantation[J]. Clinical Transplantation, 2006, 20(s15):46-52. [50] MIWA Y, YAMAMOTO K, ONISHI A, et al. Potential value of human thrombomodulin and DAF expression for coagulation control in pig-to-human xenotransplantation[J]. Xenotransplantation, 2010, 17(1):26-37. [51] CANTU E, BALSARA K, LI B, et al. Prolonged function of macrophage, von willebrand factor-deficient porcine pulmonary xenografts[J]. American Journal of Transplantation, 2007, 7(1):66-75. [52] PARIS LL, CHIHARA RK, REYES LM, et al. ASGR1 expressed by porcine enriched liver sinusoidal endothelial cells mediates human platelet phagocytosis in vitro [J]. Xenotransplantation, 2011, 18(4):245-251. [53] CHOI K, SHIM J, KO N, et al. Production of heterozygous alpha 1, 3-galactosyltransferase (GGTA1) knock-out transgenic miniature pigs expressing human CD39 [J]. Transgenic Research, 2017, 26(2):209-224. [54] WHEELER DG, JOSEPH ME, MAHAMUD SD, et al. Transgenic swine: expression of human CD39 protects against myocardial injury[J]. Journal of Molecular and Cellular Cardiology, 2012, 52(5):958-961. [55] YEOM HJ, KOO OJ, YANG J, et al. Generation and characterization of human heme oxygenase-1 transgenic pigs[J]. PLoS One, 2012, 7(10):e46646. [56] OROPEZA M, PETERSEN B, CARNWATH JW, et al. Trans genic expression of the human A20 gene in cloned pigs provides protection against apoptotic and inflammatory stimuli[J]. Xenotransplantation, 2009, 16(6):522-534. [57] MOHIUDDIN MM, SINGH AK, SCOBIE L, et al. Graft dys function in compassionate use of genetically engineered pig-to-human cardiac xenotransplantation: a case report[J]. The Lancet, 2023, 402(10399):397-410. [58] DENNER J. Porcine endogenous retroviruses and xenotransplantation, 2021[J]. Viruses, 2021, 13(11):2156. [59] MAO HZ, LI JY, LIAO GN, et al. The prevention strategies of swine viruses related to xenotransplantation [J]. Virology Journal, 2023, 20(1):121. [60] YANG HQ, ZHANG J, ZHANG XW, et al. CD163 knockout pigs are fully resistant to highly pathogenic porcine reprod uctive and respiratory syndrome virus[J]. Antiviral Research, 2018, 151:63-70. [61] WANG HY, YANG H, SHIVALILA CS, et al. One-step generation of mice carrying mutations in multiple genes by CRISPR/Cas-mediated genome engineering[J]. Cell, 2013, 153(4):910-918. [62] XUAN YY, PETERSEN B, LIU PT. Human and pig pluripotent stem cells: from cellular products to organogenesis and beyond[J]. Cells, 2023, 12(16):2075. [63] DUAN XY, CHEN CL, DU C, et al. Homozygous editing of multiple genes for accelerated generation of xenotransplantation pigs[J]. Genome Research, 2025. doi: 10.1101/gr.279709.124. [64] TAO JL, BAUER DE, CHIARLE R. Assessing and advancing the safety of CRISPR-Cas tools: from DNA to RNA editing[J]. Nature Communications, 2023, 14:212. [65] ZHANG XH, TEE LY, WANG XG, et al. Off-target effects in CRISPR/Cas9-mediated genome engineering[J]. Molecular Therapy-Nucleic Acids, 2015, 4(11):e264. [66] KARVELIS T, GASIUNAS G, YOUNG J, et al. Rapid characterization of CRISPR/Cas9 protospacer adjacent motif sequence elements[J]. Genome Biology, 2015, 16(1):253. [67] WANG Y, BI DF, QIN GS, et al. Cytosine base editor (hA3A BE3-NG)-mediated multiple gene editing for pyramid breeding in pigs[J]. Frontiers in Genetics, 2020, 11:592623. [68] RYCZEK N, HRYHOROWICZ M, LIPINSKI D, et al. Evaluation of the CRISPR/Cas9 genetic constructs in efficient disruption of porcine genes for xenotransplantation purposes along with an assessment of the off-target mutation formation [J]. Genes, 2020, 11(6):713. [69] FISCHER K, RIEBLINGER B, HEIN R, et al. Viable pigs after simultaneous inactivation of porcine MHC class I and three xenoreactive antigen genes GGTA1 CMAH and B4GALNT2[J]. Xenotransplantation, 2020, 27(1):e12560. [70] ANZALONE AV, KOBLAN LW, LIU DR. Genome editing with CRISPR-Cas nucleases, base editors, transposases and prime editors[J]. Nature Biotechnology, 2020, 38(7):824-844. [71] YUAN HM, YU TT, WANG LY, et al. Efficient base editing by RNA-guided cytidine base editors (CBEs) in pigs[J]. Cellular and Molecular Life Sciences, 2020, 77(4):719-733. [72] FISICARO N, SALVARIS EJ, PHILIP GK, et al. FokI-dCas9 mediates high-fidelity genome editing in pigs[J]. Xenotra nsplantation, 2020, 27(1):e12551.

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