Paloma Riquelme

Cutting Edge: Immunological Consequences and Trafficking of Human Regulatory Macrophages Administered to Renal Transplant Recipients

Regulatory macrophages (M regs) were administered to two living-donor renal transplant recipients. Both patients were minimized to low-dose tacrolimus monotherapy within 24 wk of transplantation and subsequently maintained excellent graft function. After central venous administration, most M regs remained viable and were seen to traffic from the pulmonary vasculature via the blood to liver, spleen, and bone marrow. By 1 y posttransplantation, both patients displayed patterns of peripheral blood gene expression converging upon the IOT-RISET signature. Furthermore, both patients maintained levels of peripheral blood FOXP3 and TOAG-1 mRNA expression within the range consistent with nonrejection. It is concluded that M regs warrant further study as a potential immune-conditioning therapy for use in solid-organ transplantation. The results of this work are being used to inform the design of The ONE Study, a multinational clinical trial of immunomodulatory cell therapy in renal transplantation.

IFN-γ-induced iNOS expression in mouse regulatory macrophages prolongs allograft survival in fully immunocompetent recipients.

Mouse monocytes exposed to macrophage colony-stimulating factor (M-CSF) and interferon-γ (IFN-γ) were driven to a novel suppressor phenotype. These regulatory macrophages (M regs) expressed markers distinguishing them from M0-, M1-, and M2-polarized macrophages and monocyte-derived dendritic cells (DCs). M regs completely suppressed polyclonal T cell proliferation through an inducible nitric oxide synthase (iNOS)-dependent mechanism. Additionally, M regs eliminated cocultured T cells in an allospecific fashion. In a heterotopic heart transplant model, a single intravenous administration of 5 × 10(6) donor-strain M regs before transplantation significantly prolonged allograft survival in fully immunocompetent recipients using both the stringent C3H-to-BALB/c (32.6 ± 4.5 versus 8.7 ± 0.2 days) and B6-to-BALB/c (31.1 ± 12 versus 9.7 ± 0.4 days) strain combinations. Nos2-deficient M regs did not prolong allograft survival, proving that M reg function in vivo is iNOS-dependent and mediated by living cells. M regs were detectable for at least 2 weeks postinfusion in allogeneic recipients. In their origin, development, phenotypic relationship with other in vitro-derived macrophages and functions, there are solid grounds to assert a near-equivalence of mouse and human M regs. It is concluded that mouse M regs represent a novel, phenotypically distinct subset of suppressor macrophages. Clinical applications of M reg therapy as an adjunct immunosuppressive therapy are currently being investigated within The ONE Study.

A cell‐based approach to the minimization of immunosuppression in renal transplantation

Five renal transplant recipients were preoperatively treated with transplant acceptance‐inducing cells (TAICs) in a Phase‐I safety study of TAICs as an adjunct immune‐conditioning therapy in living‐donor kidney transplantation. Initially, patients received anti‐thymocyte globulin induction therapy in combination with tacrolimus and steroid immunosuppression. Over the course of 12 weeks, steroids were withdrawn and tacrolimus therapy was minimized. Three of the five patients were able to tolerate low‐dose tacrolimus monotherapy and one patient was withdrawn from all immunosuppression for over 8 months. No acute or delayed adverse events were associated with the infusion of TAICs. Monitoring of the recipient anti‐donor reactivity of TAIC‐treated patients in mixed lymphocyte cultures demonstrated that, during periods of clinically stable graft function, recipient T‐cell proliferation and cytokine secretion in response to stimulation with donor alloantigen was relatively suppressed. Therefore, although the TAIC‐II trial did not provide conclusive evidence of a beneficial effect of preoperative TAIC treatment, the results were encouraging because they suggest that TAICs promote a state of alloantigen‐specific unresponsiveness, which might allow safe minimization of pharmacological immunosuppression.

Transplant acceptance-inducing cells as an immune-conditioning therapy in renal transplantation.

The transplant acceptance‐inducing cell (TAIC) is a type of immunoregulatory macrophage with the capacity to specifically dampen allogeneic rejection responses to a degree allowing safe minimization of conventional immunosuppressive therapy. In the first part of this report, the production and phenotype of the human TAIC is described. In the second part, an analysis is given of the TAIC‐I clinical trial, in which 12 recipients of renal transplants from deceased donors were treated with donor‐derived TAICs as an adjunct immune‐conditioning therapy. Conventional immunosuppression was gradually withdrawn from 10 of these 12 patients over a period of 8 weeks, starting in the fourth week after transplantation. All but two patients tolerated cessation of steroid therapy, while the remaining eight patients were first weaned from sirolimus and then, in six cases, were also weaned to low‐dose tacrolimus monotherapy. It is concluded that TAIC therapy is both safe and clinically practicable; however, the TAIC‐I trial was unable to provide evidence that postoperative TAIC administration has a beneficial effect.

Human Regulatory Macrophages

Regulatory macrophages (M regs) are a novel type of suppressor macrophage which may be a particularly suitable cell for inducing tolerance of solid organ transplants. In this article, we provide a detailed description of the generation of human M regs from peripheral blood monocytes and methods for the assessment of their phenotype. The uniqueness of the human M reg is best appreciated when the M reg is compared to macrophages in other states of activation; therefore, protocols are provided for generating five comparator macrophage types which have been used as cell type-specificity controls in our work.

Alternative approaches to myeloid suppressor cell therapy in transplantation: comparing regulatory macrophages to tolerogenic DCs and MDSCs

Several types of myeloid suppressor cell are currently being developed as cell-based immunosuppressive agents. Despite detailed knowledge about the molecular and cellular functions of these cell types, expert opinions differ on how to best implement such therapies in solid organ transplantation. Efforts in our laboratory to develop a cell-based medicinal product for promoting tolerance in renal transplant patients have focused on a type of suppressor macrophage, which we call the regulatory macrophage (M reg). Our favoured clinical strategy is to administer donor-derived M regs to recipients one week prior to transplantation. In contrast, many groups working with tolerogenic dendritic cells (DCs) advocate post-transplant administration of recipient-derived cells. A third alternative, using myeloid-derived suppressor cells, presumably demands that cells are given around the time of transplantation, so that they can infiltrate the graft to create a suppressive environment. On present evidence, it is not possible to say which cell type and treatment strategy might be clinically superior. This review seeks to position our basic scientific and early-stage clinical studies of human regulatory macrophages within the broader context of myeloid suppressor cell therapy in transplantation.

Regulatory macrophages as therapeutic targets and therapeutic agents in solid organ transplantation.

PURPOSE OF REVIEW This review aims to provide a basic introduction to human macrophage biology and an appreciation of the diverse roles played by macrophage subsets in allograft damage and repair. Current and future strategies for therapeutically manipulating macrophage behaviour are discussed. RECENT FINDINGS Macrophages are extremely versatile effector cells that exert both immunostimulatory and immunosuppressive effects. This adaptability cannot be explained by differentiation into committed sublineages, but instead reflects the ability of macrophages to rapidly transition between states of functional polarisation. Consequently, categorisation of macrophage subpopulations is not straightforward and this, in turn, creates difficulties in studying their pathophysiology. Nevertheless, particular macrophage subpopulations have been implicated in exacerbating or attenuating ischaemia-reperfusion injury, rejection reactions and allograft fibrosis. Three general strategies for therapeutically targeting macrophages can be envisaged, namely, depletional approaches, in-situ repolarisation towards a regulatory or tissue-reparative phenotype, and ex-vivo generation of regulatory macrophages (M reg) as a cell-based therapy. SUMMARY As critical determinants of the local and systemic immune response to solid organ allografts, macrophage subpopulations represent attractive therapeutic targets. Rapid progress is being made in the implementation of novel macrophage-targeted therapies, particularly in the use of ex-vivo-generated M regs as a cell-based medicinal product.

Preoperative treatment of a presensitized kidney transplant recipient with donor-derived transplant acceptance-inducing cells

This report describes the case of patient FR, a 31‐year‐old recipient of a living‐related kidney transplant from a donor against whom he was presensitized. Seventeen days prior to transplantation, a central venous infusion of transplant acceptance‐inducing cells (TAICs) was administered to the patient. During the 27‐month follow up, the patient experienced no acute rejection episodes under an immunosuppressive regime comprising anti‐thymocyte globulin (ATG) induction, corticosteroids and tacrolimus. In a similar manner to the kidney transplant recipients treated preoperatively with TAICs in a previous study, patient FR achieved a state of donor‐specific hypo‐responsiveness. Most remarkably, the deliberate preoperative exposure of a sensitized patient to the sensitizing alloantigen did not heighten his response; on the contrary, after TAIC treatment and transplantation, HLA‐specific antibodies were no longer detectable. The case of patient FR provides further evidence of the safety of pre‐transplantation treatment with TAICs.

Human regulatory macrophages as a cell-based medicinal product.

Purpose of reviewTo consolidate our basic scientific and technological appreciation of human regulatory macrophages (M reg) as a cell-based medicinal product for use as an adjunct immunosuppressive therapy in organ transplantation. Recent findingsBuilding on the original observation that crude preparations of IFN-&ggr;-stimulated allogeneic macrophages prolong allograft survival in experimental animals, we have arrived at a detailed understanding of the derivation, phenotype and T-cell-suppressive potential of a population of in-vitro-derived human macrophages, which have been designated M regs. This basic scientific knowledge has inspired methodological advances in M reg manufacture, leading to a purer and more homogeneous cell product. In turn, cells produced by these improved protocols have been applied in the clinic, so completing a cycle of technological development. Studying the migration and physiological impact of M reg administration in patients provides a measure of reassurance that the procedure is well tolerated. Cutting-edge strategies to assess the immunological status of solid organ transplant recipients allow the biological effects of M reg treatment to be monitored. SummaryA view of the human M reg as a novel, stringently defined medicinal product is presented, opening exciting possibilities for its future investigation as a therapy in solid organ transplantation and beyond.

Administration of donor‐derived transplant acceptance‐inducing cells to the recipients of renal transplants from deceased donors is technically feasible

Abstract:  Patient KW, a 36‐yr‐old male renal transplant recipient, received transplant acceptance‐inducing cells (TAICs) as an adjunct immunosuppressive therapy. In the weeks post‐transplantation, the patient’s conventional immunosuppressive treatment was gradually minimized, such that, from the 21st wk post‐transplantation onwards, the patient was stably maintained on tacrolimus monotherapy. Treatment with TAICs was without complication, both at the time of administration and in the four‐yr follow‐up period. It is concluded that the production and administration of TAICs to recipients of kidney transplants from deceased donors is technically feasible.