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1.
Nat Med ; 15(7): 746-9, 2009 Jul.
Article in English | MEDLINE | ID: mdl-19584865

ABSTRACT

Memory T cells promote allograft rejection particularly in co-stimulation blockade-based immunosuppressive regimens. Here we show that the CD2-specific fusion protein alefacept (lymphocyte function-associated antigen-3-Ig; LFA -3-Ig) selectively eliminates memory T cells and, when combined with a co-stimulation blockade-based regimen using cytotoxic T lymphocyte antigen-4 (CTLA-4)-Ig, a CD80- and CD86-specific fusion protein, prevents renal allograft rejection and alloantibody formation in nonhuman primates. These results support the immediate translation of a regimen for the prevention of allograft rejection without the use of calcineurin inhibitors, steroids or pan-T cell depletion.


Subject(s)
Graft Survival/drug effects , Kidney Transplantation , Recombinant Fusion Proteins/pharmacology , Abatacept , Alefacept , Animals , Blood Transfusion , CD2 Antigens/analysis , Immunoconjugates/pharmacology , Immunologic Memory , Macaca mulatta , Sirolimus/pharmacology , T-Lymphocytes/immunology , Transplantation, Homologous
2.
Brain ; 132(Pt 6): 1536-44, 2009 Jun.
Article in English | MEDLINE | ID: mdl-19454532

ABSTRACT

Sporadic inclusion-body myositis (sIBM) is the most common disabling, adult-onset, inflammatory myopathy histologically characterized by intense inflammation and vacuolar degeneration. In spite of T cell-mediated cytotoxicity and persistent, clonally expanded and antigen-driven endomysial T cells, the disease is resistant to immunotherapies. Alemtuzumab is a humanized monoclonal antibody that causes an immediate depletion or severe reduction of peripheral blood lymphocytes, lasting at least 6 months. We designed a proof-of-principle study to examine if one series of Alemtuzumab infusions in sIBM patients depletes not only peripheral blood lymphocytes but also endomysial T cells and alters the natural course of the disease. Thirteen sIBM patients with established 12-month natural history data received 0.3 mg/kg/day Alemtuzumab for 4 days. The study was powered to capture > or =10% increase strength 6 months after treatment. The primary end-point was disease stabilization compared to natural history, assessed by bi-monthly Quantitative Muscle Strength Testing and Medical Research Council strength measurements. Lymphocytes and T cell subsets were monitored concurrently in the blood and the repeated muscle biopsies. Alterations in the mRNA expression of inflammatory, stressor and degeneration-associated molecules were examined in the repeated biopsies. During a 12-month observation period, the patients' total strength had declined by a mean of 14.9% based on Quantitative Muscle Strength Testing. Six months after therapy, the overall decline was only 1.9% (P < 0.002), corresponding to a 13% differential gain. Among those patients, four improved by a mean of 10% and six reported improved performance of daily activities. The benefit was more evident by the Medical Research Council scales, which demonstrated a decline in the total scores by 13.8% during the observation period but an improvement by 11.4% (P < 0.001) after 6 months, reaching the level of strength recorded 12 months earlier. Depletion of peripheral blood lymphocytes, including the naive and memory CD8+ cells, was noted 2 weeks after treatment and persisted up to 6 months. The effector CD45RA(+)CD62L(-) cells, however, started to increase 2 months after therapy and peaked by the 4th month. Repeated muscle biopsies showed reduction of CD3 lymphocytes by a mean of 50% (P < 0.008), most prominent in the improved patients, and reduced mRNA expression of stressor molecules Fas, Mip-1a and alphaB-crystallin; the mRNA of desmin, a regeneration-associated molecule, increased. This proof-of-principle study provides insights into the pathogenesis of inclusion-body myositis and concludes that in sIBM one series of Alemtuzumab infusions can slow down disease progression up to 6 months, improve the strength of some patients, and reduce endomysial inflammation and stressor molecules. These encouraging results, the first in sIBM, warrant a future study with repeated infusions


Subject(s)
Antibodies, Monoclonal/therapeutic use , Antibodies, Neoplasm/therapeutic use , Immunosuppressive Agents/therapeutic use , Myositis, Inclusion Body/drug therapy , Aged , Aged, 80 and over , Alemtuzumab , Antibodies, Monoclonal/adverse effects , Antibodies, Monoclonal, Humanized , Antibodies, Neoplasm/adverse effects , Biopsy , CD4-Positive T-Lymphocytes/drug effects , CD8-Positive T-Lymphocytes/drug effects , Female , Follow-Up Studies , Gene Expression Regulation/drug effects , Humans , Immunosuppressive Agents/adverse effects , Inflammation Mediators/metabolism , Lymphocyte Count , Lymphocyte Depletion/methods , Male , Middle Aged , Muscle Strength/drug effects , Muscle, Skeletal/metabolism , Muscle, Skeletal/pathology , Myositis, Inclusion Body/immunology , Myositis, Inclusion Body/pathology , Myositis, Inclusion Body/physiopathology , RNA, Messenger/genetics , Recovery of Function , Treatment Outcome
3.
Transpl Int ; 22(7): 730-7, 2009 Jul.
Article in English | MEDLINE | ID: mdl-19196448

ABSTRACT

Monocyte accumulation in renal allografts is associated with allograft dysfunction. As monocyte influx occurs acutely following reperfusion, we investigated the effect of ischemia-reperfusion injury (IRI) on monocyte colony stimulating factor (m-CSF), a key cytokine in monocyte recruitment. We hypothesized that renal tubule epithelial cells (RTECs) could produce m-CSF in response to IRI, which could in turn promote monocyte activation. Real time PCR was used to measure levels of intragraft m-CSF transcripts in patients during IRI and clinical rejection. Also, m-CSF production by RTECs following IRI simulation in vitro was measured using ELISA. Monocyte expression of CD40 and CD80 was then analyzed using flow cytometry following co-culture with supernatants of RTECs after IRI. Monocyte expression of CD40, CD80 and HLA-DR was then examined following treatment with rh-m-CSF (10, 36, and 100 ng/ml), as was monocyte size and granularity. We found that intragraft m-CSF transcription was significantly increased postreperfusion (P = 0.002) and during clinical rejection (P = 0.002). We also found that RTECs produced m-CSF in response to IRI in vitro (P = 0.036). Monocytes co-cultured with the supernatants of postischemic RTECs became activated as evidenced by increased expression of CD40 and CD80. Also, monocytes treated with recombinant m-CSF assumed an activated phenotype exhibiting increased size, granularity and expression of CD40, CD80, CD86, and HLA-DR, and demonstrating enhanced phagocytic activity. Taken together, we suggest that renal tubular cell derived m-CSF is a stimulus for monocyte activation and may be an important target for control of IRI-associated immune activation.


Subject(s)
Colony-Stimulating Factors/metabolism , Epithelial Cells/cytology , Kidney Transplantation/methods , Kidney/injuries , Kidney/pathology , Monocytes/metabolism , B7-1 Antigen/biosynthesis , CD40 Antigens/biosynthesis , Enzyme-Linked Immunosorbent Assay , Flow Cytometry/methods , HLA-DR Antigens/metabolism , Humans , Immune System , Ischemia/pathology , Models, Biological , Monocytes/cytology , Reperfusion
4.
Transplantation ; 80(8): 1051-9, 2005 Oct 27.
Article in English | MEDLINE | ID: mdl-16278585

ABSTRACT

BACKGROUND: Perioperative lymphocyte depletion induces allograft tolerance in some animal models, but in humans has only been shown to reduce immunosuppressive requirements. Without maintenance immunosuppression, depleted human renal allograft recipients experience rejection characterized by infiltration of the allograft with monocytes and macrophages. T-cell depletion combined with a brief course of deoxyspergualin (DSG), a drug with inhibitory effects on monocytes and macrophages, induces tolerance in nonhuman primates. We therefore performed a trial to determine if lymphocyte depletion with alemtuzumab combined with DSG would induce tolerance in humans. METHODS: Five recipients of live donor kidneys were treated perioperatively with alemtuzumab and DSG and followed postoperatively without maintenance immunosuppression. Patients were evaluated clinically, by flow cytometry, and by protocol biopsies analyzed immunohistochemically and with real-time polymerase chain reaction. Results were compared to previously studied patients receiving alemtuzumab alone or standard immunosuppression. RESULTS: Despite profound T-cell depletion and therapeutic DSG dosing, all alemtuzumab/DSG patients developed reversible rejection that was similar in timing, histology, and transcriptional profile to that seen in patients treated with alemtuzumab alone. Chemokine expression was marked prior to and during rejections. CONCLUSIONS: We conclude that treatment with alemtuzumab and DSG does not induce tolerance in humans. Chemokine production may not be adequately suppressed using this approach.


Subject(s)
Antibodies, Monoclonal/pharmacology , Antibodies, Neoplasm/pharmacology , Guanidines/pharmacology , Immunosuppressive Agents/pharmacology , Kidney Transplantation/immunology , Lymphocyte Depletion , Transplantation Tolerance/drug effects , Adult , Alemtuzumab , Antibodies, Monoclonal, Humanized , Chemokines/genetics , Chemokines/metabolism , Creatine/blood , Female , Graft Rejection/metabolism , Humans , Kidney/cytology , Kidney/immunology , Kidney/physiology , Lymph Nodes/cytology , Lymphocyte Count , Male , Middle Aged , T-Lymphocytes/drug effects , Transcription, Genetic
5.
J Immunol ; 175(1): 124-30, 2005 Jul 01.
Article in English | MEDLINE | ID: mdl-15972638

ABSTRACT

We examined the role of CD40/CD40L interactions on the development of experimental autoimmune uveoretinitis (EAU), a cell-mediated, Th1-driven autoimmune disease that serves as a model for autoimmune uveitis in humans. EAU-susceptible B10.RIII mice immunized with the retinal autoantigen interphotoreceptor retinoid binding protein in CFA and treated with anti-CD40L Ab (MR1) had reduced incidence and severity of disease. Real-time PCR analysis revealed that the innate and adaptive responses of protected mice were reduced, without an obvious shift toward a Th2 cytokine profile. In contrast to some other reports, no evidence was found for regulatory cells in adoptive transfer experiments. To determine whether CD40L blockade resulted in long-term tolerance, mice protected by treatment with MR1 Ab were rechallenged for uveitis after circulating MR1 Ab levels dropped below the detection limit of ELISA. MR1-treated mice developed severe EAU and strong cellular responses to interphotoreceptor retinoid binding protein, comparable to those of control mice. These responses were higher than in mice that had not received the primary immunization concurrently with anti-CD40L treatment. We conclude that 1) CD40/CD40L interaction is required for EAU and its disruption prevents disease development; 2) CD40L blockade inhibits the innate response to immunization and reduces priming, but does not result in immune deviation; and 3) protection is dependent on persistence of anti-CD40L Abs, and long-term tolerance is not induced. Furthermore, immunological memory develops under cover of CD40L blockade causing enhanced responses upon rechallenge. Taken together, our data suggest that ongoing CD40/CD40L blockade might be required to maintain a therapeutic effect against uveitis.


Subject(s)
Autoimmune Diseases/immunology , CD40 Antigens/metabolism , CD40 Ligand/metabolism , Retinitis/immunology , Uveitis/immunology , Adoptive Transfer , Animals , Antibodies, Monoclonal/pharmacology , Autoimmune Diseases/prevention & control , Autoimmune Diseases/therapy , Disease Models, Animal , Eye Proteins/immunology , Humans , Immune Tolerance , Immunization , Immunologic Memory , Mice , Retinitis/prevention & control , Retinitis/therapy , Retinol-Binding Proteins/immunology , Uveitis/prevention & control , Uveitis/therapy
6.
Am J Transplant ; 5(3): 573-81, 2005 Mar.
Article in English | MEDLINE | ID: mdl-15707413

ABSTRACT

Renal allograft acute cellular rejection (ACR) is a T-cell mediated disease that is diagnosed histologically. However, many normally functioning allografts have T-cell infiltrates and histological ACR, and many nonimmune processes cause allograft dysfunction. Thus, neither histological nor functional criteria are sufficient to establish a significant rejection, and the fundamental features of clinical rejection remain undefined. To differentiate allograft lymphocyte infiltration from clinically significant ACR, we compared renal biopsies from patients with ACR to patients with: sub-clinical rejection (SCR, stable function with histological rejection); no rejection; and nontransplanted kidneys. Biopsies were compared histologically and transcriptionally by RT-PCR for 72 relevant immune function genes. Neither the degree nor the composition of the infiltrate defined ACR. However, transcripts up-regulated during effector T(H)1 T-cell activation, most significantly the transcription factor T-bet, the effector receptor Fas ligand and the costimulation molecule CD152 clearly (p = 0.001) distinguished the patient categories. Transcripts from other genes were equivalently elevated in SCR and ACR, indicating their association with infiltration, not dysfunction. Clinically significant ACR is not defined solely by the magnitude nor composition of the infiltrate, but rather by the transcriptional activity of the infiltrating cells. Quantitative analysis of selected gene transcripts may enhance the clinical assessment of allografts.


Subject(s)
Graft Rejection/diagnosis , Kidney Transplantation , RNA, Messenger , Transcription, Genetic , Adult , Biopsy , Female , Gene Expression Profiling , Graft Rejection/physiopathology , Humans , Kidney/metabolism , Kidney/pathology , Male
7.
Clin Transpl ; : 131-44, 2005.
Article in English | MEDLINE | ID: mdl-17424731

ABSTRACT

The National Institutes of Health has established a clinical transplant research program focusing on translational research in kidney transplantation. The program has been developed with a multidisciplinary approach under a common administrative structure that integrates transplant physicians and surgeons with clinical laboratory and data analysis support personnel. The program has achieved excellent clinical outcomes despite focusing exclusively on investigational methods and serving a diverse and medically complex patient population. Novel approaches toward consenting, computer integration, and tissue acquisition have been layered over interventional and observational studies to serve the scientific mission while delivering quality transplant care.


Subject(s)
Organ Transplantation/statistics & numerical data , Female , Humans , Immunosuppression Therapy , Kidney Transplantation/immunology , Kidney Transplantation/statistics & numerical data , Male , Monitoring, Physiologic , National Institutes of Health (U.S.) , Organ Transplantation/psychology , Patient Selection , Postoperative Period , Racial Groups , Software , Tissue and Organ Procurement/organization & administration , United States
8.
Am J Transplant ; 4(2): 283-5, 2004 Feb.
Article in English | MEDLINE | ID: mdl-14974953

ABSTRACT

Passenger leukocytes have been suggested to be both pro-tolerant and immunogenic. The opportunity to evaluate the role of allogeneic passenger leukocytes in humans was presented by a 47-year-old man who donated bone marrow to his HLA-identical leukemic sister. Eleven years later he developed renal failure. The sister's marrow was noted to be 100% XY karyotype and free of malignancy. She donated a kidney to her brother. Immunosuppression was tapered following transplantation. After 6 months, the recipient was on monotherapy sirolimus, 1 mg every third day. A surveillance biopsy was normal and sirolimus was stopped. Eight weeks later, he presented with severe rejection that reversed with Thymoglobulin. Renal function returned to baseline and has been stable on conventional immunosuppression.


Subject(s)
Kidney Transplantation/immunology , Kidney Transplantation/pathology , Leukocytes/immunology , Antilymphocyte Serum/therapeutic use , Biopsy/methods , Cytokines/genetics , Female , Histocompatibility Testing , Humans , Immunosuppression Therapy/methods , Living Donors , Male , Middle Aged , Polymerase Chain Reaction , Siblings , Sirolimus/therapeutic use , Transplantation, Homologous/immunology , Transplantation, Homologous/pathology
9.
Transplantation ; 76(1): 120-9, 2003 Jul 15.
Article in English | MEDLINE | ID: mdl-12865797

ABSTRACT

BACKGROUND: Profound T-cell depletion before allotransplantation with gradual posttransplant T-cell repopulation induces a state of donor-specific immune hyporesponsiveness or tolerance in some animal models. Alemtuzumab (Campath-1H, Millennium Pharmaceuticals, Cambridge, MA) is a humanized CD52-specific monoclonal antibody that produces profound T-cell depletion in humans and reduces the need for maintenance immunosuppression after renal transplantation. We therefore performed a study to determine if pretransplant T-cell depletion with alemtuzumab would induce tolerance in human renal allografts and to evaluate the nature of the alloimmune response in the setting of T-cell depletion. METHODS: Seven nonsensitized recipients of living-donor kidneys were treated perioperatively with alemtuzumab and followed postoperatively without maintenance immunosuppression. Patients were evaluated clinically by peripheral flow cytometry, protocol biopsies evaluated immunohistochemically, and real-time polymerase chain reaction-based transcriptional analysis. RESULTS: Lymphocyte depletion was profound in the periphery and secondary lymphoid tissues. All patients developed reversible rejection episodes within the first month that were characterized by predominantly monocytic (not lymphocytic) infiltrates with only rare T cells in the peripheral blood or allograft. These episodes were responsive to treatment with steroids or sirolimus or both. After therapy, patients remained rejection-free on reduced immunosuppression, generally monotherapy sirolimus, despite the recovery of lymphocytes to normal levels. CONCLUSIONS: T-cell depletion alone does not induce tolerance in humans. These data underscore a prominent role for early responding monocytes in human allograft rejection.


Subject(s)
Antibodies, Monoclonal/therapeutic use , Antibodies, Neoplasm/therapeutic use , Graft Survival/immunology , Immunosuppressive Agents/therapeutic use , Kidney Transplantation/immunology , Adult , Alemtuzumab , Antibodies, Monoclonal, Humanized , Black People , Female , Follow-Up Studies , Graft Rejection/epidemiology , Graft Survival/drug effects , Humans , Lymph Nodes/immunology , Lymphocyte Depletion , Male , Middle Aged , T-Lymphocytes/immunology , Time Factors , Transplantation, Homologous , United States , White People
10.
Transplantation ; 74(10): 1365-9, 2002 Nov 27.
Article in English | MEDLINE | ID: mdl-12451232

ABSTRACT

BACKGROUND: Antibodies and fusion proteins specific for CD80, CD86, and CD154 have shown promise as agents capable of inducing donor-specific tolerance in rodents. These agents have also been shown to be synergistic with one another in many settings of counter-adaptive immunity. In the nonhuman primate, monoclonal antibodies specific for CD80 and CD86 have prolonged the time to rejection of renal allografts but have not resulted in tolerance. A monoclonal antibody specific for CD154 has resulted in markedly prolonged survival of kidney, islet, cardiac, and skin allografts, but again most animals have eventually developed rejection after prolonged periods of rejection-free survival off therapy. METHODS: A combination of monoclonal antibodies specific for CD80, CD86, and CD154 were used in a mismatched nonhuman primate renal-allograft model. Doses used were based on optimized treatment protocols for each agent individually. RESULTS: Treatment of four rhesus macaques with this combination yielded a mean rejection-free survival of 565 days (311-911 days), significantly greater than untreated controls (mean survival=7.0 days, P=0.001) and animals treated with only a combination of anti-CD80 and CD86 (mean survival=191 days, P=0.01). The survival of animals treated with this combination of monoclonal antibodies was not significantly greater than those treated with anti-CD154 alone, but the production of alloantibody was delayed compared with monotherapy anti-CD154. CONCLUSION: These data suggest that a synergy exists between these agents, particularly with regard to T-dependent B-cell responses, but that they fail to induce durable tolerance in nonhuman primates.


Subject(s)
Antibodies, Monoclonal/administration & dosage , Antigens, CD/immunology , B7-1 Antigen/immunology , CD40 Ligand/immunology , Kidney Transplantation/immunology , Membrane Glycoproteins/immunology , Animals , Antibodies, Monoclonal/adverse effects , Antibody Formation , B7-1 Antigen/physiology , B7-2 Antigen , Graft Rejection , Graft Survival , Humans , Macaca mulatta , Mice , Transplantation, Homologous
11.
Transplantation ; 74(7): 916-23, 2002 Oct 15.
Article in English | MEDLINE | ID: mdl-12394831

ABSTRACT

BACKGROUND Following allotransplantation, renal ischemia-reperfusion (I/R) injury initiates a series of events that provokes counter-adaptive immunity. Though T cells clearly mediate allospecific immunity, the manner in which reperfusion events augment their activation has not been established. In addition, comprehensive analysis of I/R injury in humans has been limited. METHODS To evaluate the earliest events occurring following allograft reperfusion and gain insight into those factors linking reperfusion to alloimmunity, we examined human renal allografts 30 to 60 minutes postreperfusion (n=10) and compared them with allografts with normal function that had resolved their I/R injury insult (>1 month posttransplant, n=6) and to normal kidneys (living donor kidneys before procurement, n=8). Biopsies were processed both for immunohistochemical analysis as well as for transcript analysis by real-time quantitative polymerase chain reaction (RT-PCR). RESULTS Reperfusion injury was characterized by increased levels of gene transcripts known to be involved in cellular adhesion, chemotaxis, apoptosis, and monocyte recruitment and activation. T-cell-associated transcripts were generally absent. However, recovered allografts exhibited increased levels of T-cell and costimulation-related gene transcripts despite normal allograft function. Consistent with these findings, the immediate postreperfusion state was characterized histologically by tubular injury and monocyte infiltration, while the stable posttransplant state was notable for T-cell infiltration. CONCLUSIONS These data suggest that monocytes and transcripts related to their recruitment dominate the immediate postreperfusion state. This gives way to a T-cell dominant milieu even in grafts selected for their stable function and absence of rejection. These data have implications for understanding the fundamental link between I/R injury and alloimmunity.


Subject(s)
Ischemia/physiopathology , Kidney Transplantation , Renal Circulation , Reperfusion Injury/physiopathology , Biopsy , Cell Line , Humans , Immunohistochemistry , Kidney/metabolism , Kidney/pathology , Macrophages/pathology , Monocytes/pathology , RNA/metabolism , Reperfusion Injury/genetics , Reperfusion Injury/metabolism , Reperfusion Injury/pathology , T-Lymphocytes/pathology , Transplantation, Homologous
12.
Transplantation ; 74(7): 940-3, 2002 Oct 15.
Article in English | MEDLINE | ID: mdl-12394834

ABSTRACT

The anti-CD154 antibody hu5C8 prevents acute allograft rejection and prolongs allograft survival after withdrawal of therapy in nonhuman primates. This study describes the use of hu5C8 as a rescue agent for rejection developing after the withdrawal of hu5C8. Twelve rhesus monkeys that had received renal allografts under hu5C8 induction and subsequently rejected were studied. Rescue with hu5C8 was analyzed based on the histological character of the rejection (acute versus chronic) and whether conventional therapy was received at the time of rescue or induction. The diagnosis of rejection and response to therapy was based on allograft function and histology. Four monkeys that had acute rejection associated with conventional immunosuppression and hu5C8 were not reversed by hu5C8 rescue. Four animals with isolated chronic rejection following prolonged rejection-free survival after the withdrawal of hu5C8 did not respond to hu5C8 rescue therapy. Hu5C8 rescue therapy effectively reversed acute rejection occurring in two monkeys after hu5C8 withdrawal. One of two animals with combined acute on chronic rejection responded to hu5C8 rescue therapy. Hu5C8 effectively reverses acute but not chronic allograft rejection and appears to have no synergistic effect with conventional rescue agents.


Subject(s)
Antibodies, Monoclonal/immunology , Antibodies, Monoclonal/therapeutic use , CD40 Ligand/immunology , Graft Rejection/drug therapy , Kidney Transplantation , Salvage Therapy , Acute Disease , Animals , Antibodies, Monoclonal/administration & dosage , Antibodies, Monoclonal, Humanized , Biopsy , Chronic Disease , Drug Administration Schedule , Graft Rejection/pathology , Humans , Immunosuppression Therapy , Kidney/pathology , Macaca mulatta , Transplantation, Homologous
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