GRAPHICAL ABSTRACT

Highlights

• Porcine islets are seen as a viable alternative to human pancreatic islets for type 1 diabetes.

• Porcine islets survive long-term in non-human primates without zoonosis risks.

• An open-label trial will assess safety and efficacy of porcine islet transplantation in type 1 diabetes.

• Two individuals with poor glycemic control despite intensive insulin will be enrolled.

• The primary goal is porcine islet safety, with a secondary aim to reduce insulin doses and hypoglycemic unawareness.

INTRODUCTION

Diabetes mellitus is a metabolic disorder causing hyperglycemia, resulting from insufficient insulin secretion (type 1 diabetes mellitus) or insulin resistance (type 2 diabetes mellitus) [1]. Patients with type 1 diabetes mellitus facing absolute insulin deficiency, often necessitate insulin treatment, yet some encounter hypoglycemic episodes despite advanced insulin therapy [2]. Islet transplantation, emerging with the Edmonton Protocol in 2000, serves as a viable option for such cases [3]. However, prolonged use of immunosuppressive drugs to prevent rejection and the limited availability of donor pancreases pose significant challenges. To mitigate organ shortages, xenotransplantation, especially using pigs, has been extensively explored for decades [4]. In collaboration with the World Health Organization, The International Xenotransplantation Association (IXA) issued guidelines in 2009 for safe clinical trials of porcine islets, titled “The International Xenotransplantation Association consensus statement on the conditions for undertaking clinical trials of porcine islet products in type 1 diabetes” [5]. A key impediment to the clinical application of porcine islet xenotransplantation is the difficulty in meeting success criteria in non-human primate (NHP) preclinical studies. The 2016 update of the IXA guidelines specifies prerequisites for NHP studies, including maintaining fasting blood glucose levels of <150 mg/dL and non-fasting levels of <200 mg/dL without exogenous insulin or with significantly reduced insulin requirements in at least four out of six consecutive NHPs post-porcine islet transplantation. Follow-up should span a minimum of 6 months in all cases, ideally extending to 12 months in one or two successful cases [6].

Several research groups have reported varying porcine islet graft survival in NHP studies [7,8]. In 2015, our group demonstrated the first proof-of-concept by curing diabetes in five diabetic NHPs through the transplantation of porcine islets, utilizing an immunosuppressive drug (anti-CD154 monoclonal antibody) deemed clinically inapplicable [9]. Subsequently, we presented comparable yet less robust data with a potentially clinically applicable drug, an anti-CD40 monoclonal antibody [10]. Lastly, we reported the maintenance of glycemic control within normal ranges in four out of five diabetic NHPs with significantly reduced exogenous insulin, achieved through a clinically applicable immunosuppressive regimen [11]. Based on these preclinical results in NHPs, we standardized various procedures, including source pig validation, islet product manufacturing, and regulatory frameworks. This groundwork laid the foundation for the development of a relevant clinical trial protocol. Consequently, this study outlines a comprehensive protocol for the inaugural porcine islet xenotransplantation clinical trial. The trial targets patients experiencing labile glycemic control and hypoglycemic unawareness, aligning with IXA guidelines.

METHODS

Overall study design

A clinical trial, titled “A single-center, open-label, sponsor-initiated clinical trial to assess the safety and efficacy of porcine islet transplantation on diabetes patients (Institutional Review Board [IRB] no. GCIRB2023-031),” has received approval. The study flowchart and key considerations at each stage are depicted in Fig. 1. Gachon Hospital will enroll two patients experiencing recurrent hypoglycemic unawareness and brittle diabetes, who will undergo porcine islet xenotransplantation through the portal vein. The administration of commercially available immunosuppressants will follow the prescribed program schedule. Patients will attend regular outpatient clinic visits at Gachon Hospital for postoperative monitoring and blood sampling for a duration of up to 2 years. Monitoring of the patient’s family members, primarily spouses or sexual partners, will also be conducted to investigate the potential occurrence of xenozoonosis. The comprehensive study outline is illustrated in Fig. 2.

Study objectives

The primary aim of this trial will be to evaluate the safety of porcine islet xenotransplantation in patients. This will involve monitoring adverse effects (AEs) and changes in vital signs and laboratory tests throughout the clinical trial period. The secondary objective will be to validate the clinical efficacy of porcine islet xenotransplantation by assessing exogenous insulin injection doses and quantifying the frequency of hypoglycemic unawareness.

Study participants, screening, and informed consent

In this single-center clinical trial, two participants will enroll. The explorative nature of xenotransplantation and the potential for xenozoonosis, such as porcine endogenous retrovirus (PERV) reactivation, has led to a limited number of participants approved during the Investigational New Drug (IND) approval process. The screening period, commencing 6 months before transplantation, aims to identify suitable candidates. Public advertisements and outpatient clinic outreach at the Gachon Intensive Diabetes Care Clinic will be utilized to recruit as many potential diabetic patients as possible. However, only those precisely meeting the inclusion and exclusion criteria will be selected (Table 1). Throughout the screening period, selected patients will undergo a deliberation period. They will be equipped with a continuous glucose monitoring (CGM) system and administered multiple doses of insulin or insulin pump. Comprehensive education will be provided on the objectives of this study, adherence to the program schedule, and the potential benefits and risks of porcine islet xenotransplantation. Notably, patients will have two opportunities to consult with a psychiatrist before providing informed consent, and they retain the freedom to withdraw from the study at any time. In addition to informing patients, their family members will receive information about the purpose of this study, the transplantation procedure, potential benefits and risks of porcine islet xenotransplantation, and their responsibilities. This includes attending continuous monitoring and blood sampling sessions.

Preparation of porcine islets and transplantation

The fundamental surgical, islet isolation, and portal vein transplantation techniques for this study mirror those employed in islet allotransplantation. Preceding transplantation, porcine islets sourced from designated pathogen-free (DPF)-free pigs will undergo isolation, commencing 6 days prior to the procedure, continuing until the total islet count (10,000 islet equivalent/kg of the patient body weight) is achieved. Pancreases will be harvested from DPF-free adult pigs aged over 2 years and transported to the good manufacturing practices (GMP)-compliant islet isolation room at Gachon Hospital. The GMP-compliant collagenase-based Ricord’s method will be employed for the isolation of porcine islets. The isolated porcine islets will undergo scrutiny to ensure compliance with rapid islet product release criteria, including Gram-negative status, mycoplasma-free composition, and a low level of endotoxin (<5 mEU/mL). The collected islets will be resuspended in normal saline and subsequently infused into the portal vein of the patient via percutaneous transhepatic approach by a radiologist. A comprehensive report detailing the islet release criteria will be compiled post-transplantation.

Immunosuppression regimen

In this study, we will administer commercially available immunosuppressive agents, building upon a basic regimen derived from the human islet transplantation setting. To manage robust immune responses in the xenotransplantation context, a subset of agents will be introduced alongside the foundational regimen. The initial patient will undergo transplantation with porcine islets utilizing a conventional immunosuppression approach, featuring anti-thymoglobulin induction and cytokine blockers such as tumor necrosis factor-α (humira) and interleukin 1β (anakinra). Maintenance drugs will include sirolimus and tacrolimus. Subsequently, the second patient will receive porcine islets using the same immunosuppression regimen. However, enhancements will include the incorporation of the B cell-depleting antibody, belimumab, and the substitution of tacrolimus with tofacitinib starting 4 weeks post-transplantation (Fig. 3). The rationale behind the immunosuppressive regimen is to initially expose the first patient to a relatively low-risk immunosuppression, followed by the second patient with an add-on strategy incorporating calcineurin inhibitor-sparing and B-cell depleting effects. These regimens have demonstrated efficacy in previous NHP studies without any observed serious AEs [11].

Infection prophylaxis

In this study, scheduled potent systemic immunosuppression necessitates meticulous infection prophylaxis, akin to protocols routinely employed in human solid organ transplantations like kidney transplantation. Alongside systemic antibiotic prophylaxis, we will administer disease-specific regimens targeting cytomegalovirus infection and pneumocystis pneumonia.

Visits, test, data collection, and data analysis

The patient will undergo continuous monitoring through physical activity assessments, laboratory tests, chest X-rays, and abdominal ultrasound examinations. Discharge from the hospital is anticipated 6 days post-transplantation. Throughout the 2-year follow-up period, the patient will undergo a total of 13 routine visits (Supplementary Tables 1 and 2). To proactively guard against potential xenozoonotic diseases, including PERV, blood samples will be systematically collected from the patient on 13 occasions and from close contact individuals thrice. These samples will be securely archived at Gachon Hospital for a span of 2 years and subsequently transferred to a government institute as mandated by the new law, with archival preservation extending up to 30 years.

Primary endpoint (safety)

To assess the safety of porcine islet xenotransplantation, AEs and significant changes in vital signs and laboratory test results will be systematically monitored and documented for up to 2 years post-transplantation. Case report forms (CRFs) will capture symptoms, initial manifestation and resolution dates, severity, causality of procedures or drugs, treatments, and consequences of all AEs. An adverse drug reaction refers to any deleterious and unintended responses caused by drugs at any dose during the clinical trial, where a causative relationship can be established. An AE will be classified as “serious” (serious adverse event [SAE]) if it leads to outcomes such as death, a life-threatening event, initial hospitalization or prolongation of existing hospitalization, persistent or significant incapacity or disability in daily life functions, a congenital anomaly/birth defect, or clinically significant conditions such as drug dependence/abuse and hematologic disorder. All SAEs will be promptly reported to the IRB in accordance with Gachon Hospital regulations. The severity of AEs will be assessed following the common terminology criteria for AEs version 4.03 by the National Cancer Institute, categorized as grade 1 (mild), grade 2 (moderate), grade 3 (severe), grade 4 (life-threatening consequences), and grade 5 (death). Any AE potentially linked to xenozoonosis will be considered an SAE, regardless of its severity.

Secondary endpoint (efficacy)

To assess the efficacy of porcine islet xenotransplantation, we will monitor and record daily blood glucose levels, exogenous insulin dosage, frequency of hypoglycemic unawareness, and porcine C-peptide in serum on CRFs. Graft rejection will be determined by the absence of porcine C-peptide in serum and a return to the same or higher concentration of glycosylated hemoglobin as observed pre-transplantation.

Statistical analysis

As this study will enroll only two participants, a comprehensive description—instead of detailed statistical analyses—will be provided for the demographic data and evaluation variables related to the safety and efficacy of porcine islet xenotransplantation for each participant.

DISCUSSION

This article presents a clinical trial protocol for treating diabetes in patients through porcine islet xenotransplantation. The protocol has recently gained approval from the Korean Ministry of Food and Drug Safety (MFDS). The approval was driven by the establishment of a robust regulatory framework, successful fulfillment of preclinical criteria outlined by IXA, and the accumulation of valuable prior clinical experience.

In 2020, the Korean National Assembly sanctioned “The act on the safety and support for advanced regenerative medicine and advanced biopharmaceuticals” [12]. This legislative approval sets the stage for advancing the clinical application of porcine islet xenotransplantation in the treatment of diabetes. Approval for the trial was officially granted on December 5, 2022, following an exhaustive 30-month review. Throughout this review, the MFDS meticulously assessed documents across eight key points: (1) ensuring the validity of data obtained from NHPs in a non-good laboratory practices setting; (2) addressing concerns about zoonosis; (3) evaluating the potential for ectopic distribution of transplanted porcine islets beyond the liver; (4) assessing the risk of tumorigenicity; (5) scrutinizing the source pigs; (6) examining the use of immunosuppressants and their dose reduction strategy; (7) refining patient selection and inclusion/exclusion criteria; and (8) investigating the potential transmission of unknown pathogen(s) within the hospital. Of these concerns, confirming the DPF status of the donor pig herd emerged as the most significant and challenging. A comprehensive analysis identified a total of 146 potential pig pathogens. Various analytical methods, including real-time polymerase chain reaction, enzyme-linked immunosorbent assay (ELISA), and whole blood culture, were employed to detect these pathogens (Supplementary Table 3). The donor pig herds underwent testing, confirming the absence of 40 different types of bacteria, six fungi, 27 parasites, and 73 viruses—all yielding negative results. Particularly crucial is the confirmation of the absence of porcine cytomegalovirus (PCMV), porcine lymphotropic herpesvirus types 1 and 2 (PLHV-1 and -2), and PERV. This becomes especially pertinent in light of recent findings suggesting a potential association between PCMV reactivation and early graft failure in pig heart transplantation in human patients [13].

Previous clinical trials involving encapsulated or naked porcine islet transplantation in diabetic patients have mitigated certain risks associated with xenotransplantation and have partially confirmed the safety of porcine islets (Supplementary Table 4) [8].

As porcine islet xenotransplantation has not yet been undertaken in Korea and considering the current uncertainty regarding xenozoonosis, including PERV activation, only two patients will be included in the study. The limited number of participants precludes statistical analyses and definitive scientific assessments of the benefits and risks associated with porcine islet transplantation and the safety of porcine islet xenotransplantation. However, the primary endpoint of the study will be achieved. Furthermore, a 2-year follow-up will provide ample data for in-depth analyses of porcine islet functionality.

The ethical considerations of this clinical trial will undergo thorough evaluation. Recent advancements in glycemic control, facilitated by CGM and automatic injections of various insulin formulations, will enhance the potential for numerous diabetic patients to manage their glycemia within acceptable ranges [14]. This technical progress is anticipated to benefit highly motivated and relatively young patients with diabetes, though it will not prevent various complications [15]. Islet transplantation is poised to offer optimal physiological insulin, counter-regulatory hormone, and glucagon production, thereby mitigating the progression of diabetic complications [16]. Strict benefit and risk analyses, meticulous patient selection, and a comprehensive follow-up schedule will be essential to outweigh the potential risks associated with porcine islet transplantation. In summary, the clinical trial on porcine islet xenotransplantation is expected to make a significant contribution to scientific history and instill hope in many patients with diabetes.

SUPPLEMENTARY MATERIALS

NOTES

CONFLICTS OF INTEREST

The authors from Tascom, Co. Ltd. and GenNBio have no conflicts of interest.

AUTHOR CONTRIBUTIONS

Conception or design: B.J.K., J.S.S., C.G.P., S.J.K., K.W.K.

Acquisition, analysis or interpretation of data: all authors.

Drafting the work or revisiting: B.J.K., J.S.S.

Final approval of the manuscript: B.J.K., J.S.S., K.W.K.

FUNDING

This work was supported by a grant from the Korea Healthcare Technology R&D Project through the Korea Health Industry Development Institute (KHIDI), funded by the Ministry for Health and Welfare, Republic of Korea (Grant No. HI13C0954).

Acknowledgements

We are grateful to the members of the Ethics Committees of IXA and TTS, Drs. Richard N. Pierson, Emanuele Cozzi, Francesco L. Ierino, Wayne Paris, Rita Bottino, Mariachiara Tallacchini, and Ian Dittmer. We extend special thanks to Dr. Richard N. Pierson for his outstanding leadership and hard work. We also thank Drs. Sang Il Kim, Ik Jin Yun, and Myoung-Joon Kim for their valuable participation in the on-site review of the clinical protocols.

Fig. 1.

Overall flowchart of the clinical trial of porcine islet xenotransplantation. Pancreatic islets will be isolated from designated pathogen-free (DPF)-grade Seoul National University (SNU) miniature pigs and transplanted into the portal vein of each patient’s liver. The major points raised by the Korean Ministry of Food and Drug Safety during the review process were broadly categorized into the source of pigs and the definition of DPF status, islet product release criteria, concerns about zoonosis, the use of immunosuppressants, patient selection and inclusion/exclusion criteria, and the sample archive. GMP, good manufacturing practices.

Fig. 2.

Design of the clinical trial of porcine islet xenotransplantation. Patients will be recruited and selected 6 months before the start of the clinical study. During the 6-month-long intensive deliberation period, patients will be monitored using a continuous glucose monitoring system (CGMS) and multiple doses of insulin or insulin pumps. After consulting a psychiatrist, they will have two deliberation opportunities before providing informed consent. F/U, follow-up; W/U, work-up.

Fig. 3.

Immunosuppression regimen. The first patient will be transplanted with porcine islets using a conventional immunosuppression regimen composed of anti-thymoglobulin induction and cytokine blockers such as tumor necrosis factor-α and interleukin 1β, with sirolimus and tacrolimus as maintenance drugs. The second patient will be transplanted using the above immunosuppression regimen plus B cell-depleting antibody, belimumab, and tacrolimus will be replaced with tofacitinib 4 weeks after transplantation. ATG, anti-thymoglobulin; IV, intravenous; PO, per os (by mouth); DO, day 0; BID, bis in die (twice a day); SQ, subcutaneous.

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Kim BJ, Shin JS, Min BH, Kim JM, Park CG, Kang HJ, Hwang ES, Lee WW, Kim JS, Kim HJ, Kwon I, Kim JS, Kim GS, Moon J, Shin DY, Cho B, Yang HM, Kim SJ, Kim KW. Clinical Trial Protocol for Porcine Islet Xenotransplantation in South Korea. Diabetes Metab J. 2024;48(6):1160-1168.

Received: Aug 07, 2023; Accepted: Jan 17, 2024

DOI: https://doi.org/10.4093/dmj.2023.0260.

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