The purpose of this notice of funding opportunity (NOFO) is to solicit applications to participate in the Immunobiology of Xenotransplantation Cooperative Research Program (IXCRP), a multi-center program dedicated to resolving immunologic and physiologic barriers to safe and efficacious xenotransplantation using preclinical pig to nonhuman primate (NHP) or human decedent models of pancreatic islet, kidney, heart, lung, or liver xenotransplantation. Transplantation is often the preferred or only therapy for end-stage organ disease. In 2023, 46,630 organ transplants were performed in the United States. In addition, transplantation of pancreatic islets offers a potential therapy for individuals with type 1 diabetes whose disease is not effectively managed with exogenous insulin. Unfortunately, with over 103,500 adults and children on the United Network for Organ Sharing (UNOS) waiting list, those in need of a transplant greatly exceed the number of available organs. It is estimated that 20 people on average die each day waiting for a transplant. Xenotransplantation offers a potential interim or definitive solution to the severe shortage of human organs and pancreatic islets. Pigs are the primary species of interest as xenograft donors due to their favorable reproductive capacity and anatomical and physiological similarities to humans. However, there are multiple barriers to successful clinical xenotransplantation, including immunologic rejection of non-human organs and tissues by the human immune system, physiological differences between non-human and human molecules that prevent proper functioning of various biochemical pathways, and potential transmission of zoonoses.

To address these challenges, the IXCRP was established by the National Institute of Allergy and Infectious Diseases (NIAID) in 2005 with a co-fund for type 1 diabetes from the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) (RFA-AI-04-042). Subsequently, in 2010, the program was renewed solely by NIAID (RFA-AI-09-035), in 2015 (RFA-AI-14-047 and RFA-AI-14-048), and in 2020 (RFA-AI-19-042 and RFA-AI-19-043).. IXCRP investigators and other researchers in the field have made significant advances over the past two decades, and NIAID is committed to support this challenging area of research.

Historically, the most significant hurdle to successful xenotransplantation was hyperacute rejection caused by preformed, xenoreactive naturally-occurring antibodies (XNA) that destroy the xenograft within hours post-transplant. The primary target of XNA is a carbohydrate epitope, galactose-alpha-(1,3)-galactose (Gal), which is not present in humans and Old World NHPs. To overcome this hurdle, two decades ago, the enzyme responsible for terminally linking Gal onto oligosaccharide chains, alpha-1,3 glycosyltransferase (GT), was knocked out in genetically modified pigs. Xenografts from GT knockout (GTKO) pigs elicit substantially less severe hyperacute rejection in NHPs. Cytidine monophosphate-N-acetylneuraminic acid hydroxylase (CMAH) gene knockouts and mutations to beta-1,4-N-acetyl-galactosaminyltransferase 2 (B4GALNT2) were similarly engineered to reduce reactivity to other notable XNA to pig carbohydrate antigens, namely N-glycolylneuraminic acid-modified glycans and SDa swine blood group antigen, respectively.

Over the last decade, application of CRISPR-Cas 9 technology combined with somatic cell nuclear transfer cloning has significantly accelerated the pace of multi-gene modification and donor pig production. Additional genetic modifications, most on the GTKO background, were developed to address key species-to-species incompatibilities and create more human-like organs. These include the insertion of human complement regulatory proteins to minimize the deleterious effects of the complement cascade in antibody-mediated rejection, human thrombomodulin and/or tissue factor pathway inhibitor to overcome coagulation pathway dysfunction, and human anti-inflammatory and/or immune suppressive genes to reduce immune activation contributing to graft rejection. These strategies have dramatically reduced the frequency and severity of hyperacute rejection and have prolonged survival in pig-to-NHP xenotransplantation models for up to 4 years.

Success in prolonging xenograft survival in the pig-to-NHP model allows more in-depth investigation of the remaining immunologic and physiologic issues that must be addressed in order to achieve safe and efficacious clinical xenotransplantation. These include physiologic differences that influence xenograft function and long-term survival, and risks associated with zoonoses. Transmission of pathogenic zoonoses to a human recipient and, potentially, the general population is a concern. To reduce this risk, animals used for xenotransplantation are bred in specific-pathogen-free conditions, weaned early or caesarian-derived, and routinely screened to eliminate most, if not all, known zoonotic agents.

Porcine Cytomegalovirus (PCMV) is a swine pathogen known to have deleterious effects on xenograft survival. In the first human patient to receive a cardiac xenotransplant, conventional testing failed to detect latent PCMV in the donor pig and the virus reactivated post-transplant. The extent to which PCMV reactivation contributed to the patient’s death is unknown; however, this event underscores the need for sensitive and reliable assays to detect both latent and active infection with PCMV. Porcine endogenous retroviruses (PERV), another potential zoonotic threat, were successfully inactivated in a line of pigs through a combination of CRISPR-Cas9 gene-editing and somatic cell nuclear transfer, further highlighting the potential of these technologies to both protect against immunologic attack and reduce the risk of zoonoses.

The field of xenotransplantation has recently witnessed an expansion in research models beyond NHP recipients to include an evaluation of safety, feasibility, and short-term outcomes (2 – 60 days) in humans declared to have irreversible loss of brain function (individuals with brain death, also referred to as human decedents) maintained on cardiopulmonary support. These experiments, using varying genetically modified pig hearts and kidneys transplanted into human decedents whose organs were declined for allotransplantation based on organ quality, have demonstrated early hemodynamic stability, an absence of hyperacute rejection, and basic organ functionality under immunosuppression. No chimerism or transmission of porcine retroviruses were detected; however, many of these experiments have demonstrated thrombotic microangiopathy and/or antibody-mediated injury.

As of the time of this writing, medical teams that include IXCRP-funded investigators have performed two pig-to-human orthotopic heart transplants under expanded access (compassionate use) authorization from the FDA. The two xenograft recipients expired at 8 and 6 weeks, respectively. These initial clinical xenotransplants demonstrated good early xenograft function but also highlighted fundamental gaps in our knowledge of 1) critical pathways leading to inflammation and graft failure, 2) best practices for zoonotic viral surveillance and treatment, 3) optimal design of the donor pig, and 4) postoperative immunosuppression regimen. These knowledge gaps must be addressed prior to broader clinical translation.

Scope and Research Objectives
The re-issue of the IXCRP will support a research program comprised of two or more research projects centered around preclinical NHP and/or human decedent models of porcine pancreatic islet, kidney, heart, lung, or liver xenotransplantation. The research objectives must address one or more of the remaining key immunologic and physiologic barriers to achieving safe and efficacious xenotransplantation, including issues affecting engraftment, survival, and function of xenografts. Research foci may include 1) development or optimization of the models themselves, including genetic modifications of the pig-donor to address FDA concerns, as well as refinement of surgical xenotransplantation techniques, 2) development or optimization of the immunosuppression (IS) regimen to prevent rejection and minimize toxicity, 3) characterization and resolution of physiological and immunological barriers to long-term xenograft survival, and 4) development or optimization of strategies to screen for and prevent pathogen transmission to recipients.

Examples of research topics may include, but are not limited to the following:

Elucidation of the cellular and molecular mechanisms of and development of strategies to prevent hyperacute, acute, and chronic xenograft rejection;
Characterization of the recipient’s innate and adaptive immune responses to the xenograft;
Evaluation of regimens to induce and maintain immune tolerance to xenografts and/or delineation of cellular and molecular mechanisms promoting xenograft tolerance;
Development and characterization of strategies to prolong xenograft survival in life-supporting xenotransplantation models;
Development of approaches to eliminate or minimize the use of immunosuppressive drugs through genetic modifications of donor organs/tissues/cells, utilization of encapsulation techniques, or other tolerogenic approaches;
Characterization of and application of approaches to address differences in the anatomical, physiological, and/or endocrinological features of donor pig organs, tissues, or cells that limit a xenograft’s survival and function in NHP or human decedent recipients;
Delineation and study of cross-match differences between pigs and NHPs or humans;
Development and testing of tools/approaches to diagnose, monitor, and treat porcine zoonoses in human decedent models;
Development and testing of tools/approaches to diagnose, monitor, and treat xenograft rejection; and
Development and testing of tools/approaches to diagnose, monitor, and treat causes of xenograft dysfunction other than immunologic rejection.
Applications proposing any of the following will be considered non-responsive and will not be reviewed:

Pig-to-NHP xenotransplantation studies of any organs, tissues, or cells other than pancreatic islets, kidney, heart, lung, or liver.
Small animal models of xenotransplantation, such as rodent models, unless the model is proposed only as an in vivo bioassay of large animal immune function (e.g., trans in vivo delayed type hypersensitivity assay);
Clinical trials or clinical/human studies of xenotransplantation; (only preclinical human decedent model research is allowed).
Studies of zoonotic agents or infections, except for those studies designed to prevent transmission of, or improve diagnosis, monitoring, or treatment of zoonotic infections in xenograft recipients.
Studies focused on HIV/AIDS-related research
Applications that do not include annual milestones.
Applications that propose studies in human decedents but do not include a Human Decedent Research Plan.
Structure of the IXCRP
Administrative Core: Each application will include an Administrative Core to manage and coordinate all activities to ensure project timelines and objectives are met. The Administrative Core will include a Program Management Plan that will guide its operations and activities. This Core will be responsible for carrying out activities described in the Data Management and Sharing Plan. The Administrative Core will also be responsible for organizing an annual programmatic meeting for all investigators and NIAID staff.

Scientific Core: Scientific Cores are optional and may be included to provide IXCRP investigators with core resources and/or facilities that are essential for the activities of two or more Research Projects. Scientific Core activities must not overlap with each other or with the activities of a Research Project. The Scientific Core may not conduct research independent of the served Research Project.

Research Projects: Applications will propose at least two synergistic Research Projects that meet the goals of the initiative to resolve the immunologic and physiologic barriers to safe and efficacious xenotransplantation using preclinical pig to nonhuman primate (NHP) or human decedent models of pancreatic islet, kidney, heart, lung, or liver xenotransplantation.

Milestones

The research projects must include explicit, detailed, and quantitative annual milestones. These milestones will be used by NIAID program staff to assess annual progress and support funding decisions.

Steering Committee

Program Directors/Principal Investigators (PD(s)/PI(s)) of awards funded under this program will form a Steering Committee after award. The Steering Committee will serve as the main governing body of the IXCRP. At annual meetings, the Steering Committee will review progress of xenotransplantation projects, provide guidance and recommendations to investigators regarding study implementation and conduct, identify scientific opportunities, emerging needs, and impediments to success, and encourage collaborations among consortium members. The voting members of the Steering Committee will include the PD/PI (contact PI) from each single project U01 award and the PD/PI (contact PI) plus one project leader from each multi-project U19 award. Additional PDs/PIs, Project Leaders, Core Leaders, and the NIH Project Scientist will serve as non-voting Steering Committee members. All IXCRP investigators will be required to accept and implement common guidelines and procedures approved by the Steering Committee.

Applicants are encouraged to consider using the following NIAID-supported programs:

The Immunology Database and Analysis Portal (ImmPort)

The Immunology Database and Analysis Portal (ImmPort) program will provide support for public sharing of research data and experimental protocols of the IXCRP. ImmPort is a NIAID-funded data sharing platform, which has developed templates for data collection, standardization, and sharing from various NIAID-supported research programs. The IXCRP recipients are encouraged to participate with ImmPort in developing data standards for IXCRP-specific data types, where applicable, and be responsible for collecting and submitting data and documents into ImmPort. The IXCRP Steering Committee will provide information, consistent with the goals of the program and NIH policy, regarding research data and experimental protocol sharing within the IXCRP and with the public.

The National Swine Resource and Research Center (NSRRC)

The Office of Research Infrastructure Programs within the Division of Program Coordination, Planning, and Strategic Initiatives in the Office of the NIH Director supports the National Swine Resource and Research Center (NSRRC), which is co-sponsored by NIAID and the National Heart, Lung, and Blood Institute (NHLBI). The NSRRC was established in 2003 to develop the infrastructure needed to ensure that biomedical investigators across a variety of disciplines have access to critically needed swine models of human health and disease. The purpose of the NSRRC is to provide the biomedical research community enhanced access to critically needed swine models and to develop genetically modified swine when required for studies involving human health and diseases, including xenotransplantation. NIAID encourages IXCRP-funded investigators to submit relevant cell lines and animal models developed under this NOFO to the NSRRC, when applicable.

The current U19 NOFO is appropriate for investigators proposing a complex research program involving 2 or more research projects supported cores while the companion U01 NOFO (RFA-AI-24-019) should be used for applicants that are proposing a single research project.

Applicants are strongly encouraged to discuss the proposed research with NIAID staff listed in the Scientific/Research contact well in advance of the application submission deadline.

See Section VIII. Other Information for award authorities and regulations.