Median Graft Survival Research Articles

Intralipid® attenuates acute cardiac allograft rejection in relation to promoting CD4+CD25+Foxp3+ regulatory T-cells and inhibiting toll-like receptor 4 expression

Clinically available agents that promote the suppression of inflammatory processes with minimal adverse effects are highly desired. An FDA-approved nutrition supplement, Intralipid®, has shown its immunomodulatory potential. However, its influence on organ transplant rejection has not been tested. Here, we have investigated whether Intralipid® affect allograft outcome in a fully major histocompatibility complex (MHC)-mismatched rat model of heterotopic working-heart transplantation. Recipients were treated intravenously with Intralipid® (1.5g/kg, clinical dosage) or saline (untreated control) on post-transplantation days (PODs) 1, 3, and 6. Treatment with Intralipid® results in a significant prolongation of median graft survival from 7 days to 12 days. On POD 7, while the allografts from our untreated (i.e., control) group exhibit severe rejection, the allografts from Intralipid®-treated group show milder rejection demonstrating preserved myocardium integrity and decreasing infiltration of inflammatory cells (ED1+ and CD3+ cells). Flow cytometry analysis reveals a marked increase of CD4+CD25+Foxp3+ regulatory T-cells (Tregs) in the peripheral blood of Intralipid®-treated recipients beginning on POD 3 (7.0% vs. 14.4%, untreated vs. Intralipid®-treated). Intralipid® treatment also promotes Tregs in the spleens and allograft hearts. Toll-like receptor 4 expression is markedly suppressed in the Intralipid®-treated allografts. Our results suggest a new use of Intralipid® to modulate immune responses in the setting of heart transplantation. This study could stimulate further mechanistic as well as treatment studies leading to the clinical application of Intralipid® as an adjunctive therapy with current immunosuppression agents to manage organ rejection.

Renal Allograft Survival in Nonhuman Primates Infused With Donor Antigen-Pulsed Autologous Regulatory Dendritic Cells.

Systemic administration of autologous regulatory dendritic cells (DCreg; unpulsed or pulsed with donor antigen [Ag]), prolongs allograft survival and promotes transplant tolerance in rodents. Here, we demonstrate that nonhuman primate (NHP) monocyte-derived DCreg preloaded with cell membrane vesicles from allogeneic peripheral blood mononuclear cells induce T cell hyporesponsiveness to donor alloantigen (alloAg) in vitro. These donor alloAg-pulsed autologous DCreg (1.4-3.6 × 106 /kg) were administered intravenously, 1 day before MHC-mismatched renal transplantation to rhesus monkeys treated with costimulation blockade (cytotoxic T lymphocyte Ag 4 immunoglobulin [CTLA4] Ig) and tapered rapamycin. Prolongation of graft median survival time from 39.5 days (no DCreg infusion; n = 6 historical controls) and 29 days with control unpulsed DCreg (n = 2), to 56 days with donor Ag-pulsed DCreg (n = 5) was associated with evidence of modulated host CD4+ and CD8+ T cell responses to donor Ag and attenuation of systemic IL-17 production. Circulating anti-donor antibody (Ab) was not detected until CTLA4 Ig withdrawal. One monkey treated with donor Ag-pulsed DCreg rejected its graft in association with progressively elevated anti-donor Ab, 525 days posttransplant (160 days after withdrawal of immunosuppression). These findings indicate a modest but not statistically significant beneficial effect of donor Ag-pulsed autologous DCreg infusion on NHP graft survival when administered with a minimal immunosuppressive drug regimen.

Preemptive CD20+ B cell Depletion Attenuates Cardiac Allograft Vasculopathy in CD154-Treated Monkeys.

Anti-CD154 monotherapy is associated with antidonor allo-antibody (Ab) elaboration, cardiac allograft vasculopathy (CAV), and allograft failure in preclinical primate cell and organ transplant models. In the context of calcineurin inhibitors (CNI), these pathogenic phenomena are delayed by preemptive "induction" B cell depletion. αCD154 (IDEC-131)-treated cynomolgus monkey heart allograft recipients were given peritransplant rituximab (αCD20) alone or with rabbit antihuman thymocyte globulin. Relative to previously reported reference groups, αCD20 significantly prolonged survival, delayed Ab detection, and attenuated CAV within 3 months in αCD154-treated recipients (αCD154 + αCD20 graft median survival time > 90 days, n = 7, vs 28 days for αCD154 alone (IDEC-131), n = 21; P = 0.05). Addition of rabbit antihuman thymocyte globulin to αCD154 (n = 6) or αCD154 + αCD20 (n = 10) improved graft protection from graft rejection and failure during treatment but was associated with significant morbidity in 8 of 16 recipients (6 infections, 2 drug-related complications). In αCD20-treated animals, detection of antidonor Ab and relatively severe CAV were anticipated by appearance of CD20 cells (>1% of lymphocytes) in peripheral blood and were associated with low αCD154 trough levels (below 100 μg/mL). These observations support the hypothesis that efficient preemptive "induction" CD20 B cell depletion consistently modulates pathogenic alloimmunity and attenuates CAV in this translational model, extending our prior findings with calcineurin inhibitors to the context of CD154 blockade.

Restrictive allograft syndrome after lung transplantation: new radiological insights

To describe the CT changes in patients with restrictive allograft syndrome (RAS) after lung transplantation, before and after clinical diagnosis. This retrospective study included 22 patients with clinical diagnosis of RAS. Diagnosis was based on a combination of forced expiratory volume (FEV1) decline (≥20%) and total lung capacity (TLC) decline (≥10%). All available CT scans after transplantation were analyzed for the appearance and evolution of lung abnormalities. In 14 patients, non-regressing nodules and reticulations predominantly affecting the upper lobes developed an average of 13.9months prior to the diagnosis of RAS. Median graft survival after onset of non-regressing abnormalities was 33.5months, with most patients in follow-up (9/14). In eight patients, a sudden appearance of diffuse consolidations mainly affecting both upper and lower lobes was seen an average of 2.8months prior to the diagnosis of RAS. Median graft survival was 6.4months after first onset of non-regressing abnormalities, with graft loss in most patients (6/8). RAS has been previously described as a homogenous group. However, our study shows two different groups of RAS-patients: one with slow progression and one with fast progression. The two groups show different onset and progression patterns of CT abnormalities. • RAS is the newest discovered form of chronic lung allograft dysfunction (CLAD). • RAS is not a homogenous group, as survival varies greatly between patients. • In this study, we see two different CT onset and progression patterns. • These two different CT patterns also correlate with a different survival rate.

Long-term results of allogenic penetrating limbo-keratoplasty: 20 years of experience.

AimThe objective of the study was to evaluate the long-term results of allogenic penetrating limbo-keratoplasy. This method allows simultaneous transplantation of a corneal graft and limbal stem cells of the donor by means of eccentric trephination of the donor button.MethodThe data of 192 consecutive cases of allogenic penetrating limbo-keratoplasty from 1995 to 2015 were reviewed. These had been performed exclusively in eyes with complete failure of the limbal stem cells, in combination with deep corneal scarring. Indications were predominantly eye burns, inflammatory conditions, and congenital aniridia. Graft survival and rejection rates were assessed using Kaplan-Meier analysis.ResultsFollow-up averaged 2.1±2.2 years. Median graft survival was 3.4 years in eye burns, 3.9 years in inflammatory disease, and 3.2 years in congenital aniridia. Median survival was 3.9 years in the heterogenous group of other indications.ConclusionAllogenic limbo-keratoplasty is a suitable option used to treat patients with bilateral complete failure of the limbal stem cells and deep opacification of the central cornea. The main reasons for graft failure are the loss of graft-limbal stem cell functioning and endothelial graft rejection.

Long-Term Outcome of Kidney Transplantation in Recipients with Focal Segmental Glomerulosclerosis.

FSGS can recur after kidney transplantation and is associated with poor graft outcomes. We aimed to assess the incidence of FSGS recurrence post-transplant and determine the effect of graft source on recurrence and graft survival in patients with biopsy-proven FSGS. Using the Australian and New Zealand Dialysis and Transplant Registry, we assessed incidence of FSGS, the influence of donor type on the risk of FSGS recurrence, and graft loss in recipients with ESRD caused by primary FSGS using Kaplan-Meier and logistic regression analyses. Between 1992 and 2011, 736 first kidney transplants were performed in 666 adults and 70 children (≤20 years old) with biopsy-proven primary FSGS. FSGS recurred in 76 (10.3%) patients. Younger age (P<0.001), nonwhite ethnicity (P=0.02), and having a live donor (P=0.02) were independent risk factors associated with recurrence. Median graft survival was significantly better for live donor compared with deceased donor grafts (14.8 versus 12.1 years; P<0.01). Disease recurrence predicted poor graft outcomes, with 52% (95% confidence interval, 40% to 63%) 5-year graft survival in the recurrence group compared with 83% (95% confidence interval, 79% to 86%) in the group without recurrent disease (P<0.001). FSGS recurrence after kidney transplantation was more common in live donor kidneys. Despite this, graft survival in live donor recipients was significantly better for both children and adults with FSGS. We propose that live donor transplantation should not be avoided in patients with FSGS.

Ribonuclease (RNase) Prolongs Survival of Grafts in Experimental Heart Transplantation.

BackgroundCell damage, tissue and vascular injury are associated with the exposure and release of intracellular components such as RNA, which promote inflammatory reactions and thrombosis. Based on the counteracting anti‐inflammatory and cardioprotective functions of ribonuclease A (RNase A) in this context, its role in an experimental model of heart transplantation in rats was studied.Methods and ResultsInbred BN/OrlRj rat cardiac allografts were heterotopically transplanted into inbred LEW/OrlRj rats. Recipients were intravenously treated every other day with saline or bovine pancreatic RNase A (50 μg/kg). Toxic side effects were not found (macroscopically and histologically). Heart tissue flow cytometry and quantitative morphological analyses of explanted hearts at postoperative day 1 or postoperative day 4 showed reduced leukocyte infiltration, edema, and thrombus formation in RNase A‐treated rats. In allogeneic mixed lymphocyte reactions, RNase A decreased the proliferation of effector T cells. RNase A treatment of rats resulted in prolonged median graft survival up to 10.5 days (interquartile range 1.8) compared to 6.5 days (interquartile range 1.0) in saline treatment (P=0.001). Treatment of rats with a new generated (recombinant) human pancreatic RNase 1 prolonged median graft survival similarly, unlike treatment with (recombinant) inactive human RNase 1 (each 50 μg/kg IV every other day, 11.0 days, interquartile range 0.3, versus 8.0 days, interquartile range 0.5, P=0.007).ConclusionsUpon heart transplantation, RNase administration appears to present a promising and safe drug to counteract ischemia/reperfusion injury and graft rejection. Furthermore, RNase treatment may be considered in situations of critical reperfusion after percutaneous coronary interventions or in cardiac surgery using the heart–lung machine.

Predictors of survival in restrictive chronic lung allograft dysfunction after lung transplantation

Chronic lung allograft dysfunction (CLAD) is the main factor limiting long-term survival after lung transplantation. Besides bronchiolitis obliterans syndrome, a restrictive phenotype of CLAD (rCLAD) exists, which is associated with poor prognosis after diagnosis. However, survival determinants for rCLAD remain to be elucidated. Our aim in this study was to establish parameters predicting survival in patients with rCLAD. All patients diagnosed with rCLAD in 2 lung transplant centers were assessed in a retrospective manner. Various clinical parameters [demography, pulmonary function, bronchoalveolar lavage (BAL), histopathology, radiology and blood differentials] at rCLAD diagnosis were correlated with graft survival using unadjusted and adjusted analysis. A total of 53 patients with rCLAD were included with a median graft survival after diagnosis of 1.1 years. Univariate analysis demonstrated that lower-lobe-dominant or diffuse infiltrates on chest computed tomography, presence of an identifiable trigger before rCLAD onset, lymphocytic bronchiolitis, increased BAL neutrophilia, increased BAL eosinophilia and increased blood eosinophils were associated with inferior graft survival after rCLAD diagnosis. Multivariate analysis confirmed the association of location of infiltrates and blood eosinophilia on graft survival. In this study we have identified parameters associated with graft survival after rCLAD diagnosis that may be useful to predict prognosis.

Chimeric 2C10R4 anti-CD40 antibody therapy is critical for long-term survival of GTKO.hCD46.hTBM pig-to-primate cardiac xenograft.

Preventing xenograft rejection is one of the greatest challenges of transplantation medicine. Here, we describe a reproducible, long-term survival of cardiac xenografts from alpha 1-3 galactosyltransferase gene knockout pigs, which express human complement regulatory protein CD46 and human thrombomodulin (GTKO.hCD46.hTBM), that were transplanted into baboons. Our immunomodulatory drug regimen includes induction with anti-thymocyte globulin and αCD20 antibody, followed by maintenance with mycophenolate mofetil and an intensively dosed αCD40 (2C10R4) antibody. Median (298 days) and longest (945 days) graft survival in five consecutive recipients using this regimen is significantly prolonged over our recently established survival benchmarks (180 and 500 days, respectively). Remarkably, the reduction of αCD40 antibody dose on day 100 or after 1 year resulted in recrudescence of anti-pig antibody and graft failure. In conclusion, genetic modifications (GTKO.hCD46.hTBM) combined with the treatment regimen tested here consistently prevent humoral rejection and systemic coagulation pathway dysregulation, sustaining long-term cardiac xenograft survival beyond 900 days.

(79) - Early CD20+ B Cell Depletion Attenuates Cardiac Allograft Vasculopathy in Anti-CD154 Treated Non-Human Primate Model

Anti-CD154 (αCD154) monotherapy is associated with the development of anti-donor alloantibodies, cardiac allograft vasculopathy (CAV), and allograft failure in preclinical primate cell and organ transplant models. Evidence suggests that peri-transplant B cell depletion may remove an efficient source of donor antigen-specific costimulation, thereby attenuating development of anti-donor alloantibodies and subsequent pathogenic phenomena. Cynomolgus monkeys underwent heterotopic intraabdominal cardiac transplantation, and recipients were treated with either αCD154 (IDEC-131, n=21), αCD20 (Rituximab, n=2), αCD154 + αCD20 (n=7), or αCD154 + αCD20 + rabbit anti-human thymocyte immune globulin (rATG, n=10). Primary graft survival was defined as time to the first rejection episode. All biopsies and explant tissue specimens were assessed for CAV using the 2005 ISHLT revised criteria. Serum BAFF (B-cell activating factor of the tumor necrosis factor family) levels were analyzed by ELISA. Combined αCD154 + αCD20 treatment significantly prolonged graft median survival time (MST >90 days), delayed anti-donor alloantibody production, and attenuated CAV compared to αCD154 monotherapy (MST 28 days, p=0.05). Addition of rATG to αCD154 + αCD20 also prolonged graft MST to >90 days, but it was associated with a 50% incidence of treatment-attributed morbidity. Incidence of severe acute rejection (>1R) and CAV severity within 90 days was reduced in αCD20 treated animals relative to reference groups (p<0.05 for both endpoints). In αCD20 treated recipients, detection of anti-donor IgM and IgG alloantibody and relatively severe CAV were anticipated by appearance of CD20 cells in peripheral blood and was associated with trough αCD154 levels <100 μg/mL. Serum BAFF levels were higher in αCD20 treated animals (d7: 3762±181pg/mL) compared to untreated controls (628±507 pg/mL) and, similarly, in combined therapy (d7: 2687±829 pg/mL) vs. αCD154 controls (371±80 pg/mL). Efficient preemptive CD20+ B cell depletion in combination with selective CD154 inhibition consistently modulates pathogenic alloimmunity and attenuates CAV in this non-human primate heterotopic heart transplantation model; however, the addition of T cell depletion to the combined therapy was associated with significant morbidity in this model.

MP32-01 PERCUTANEOUS NEPHROSTOMY PLACEMENT AFTER RENAL TRANSPLANTATION: INDICATIONS AND IMPLICATIONS FOR LONG-TERM GRAFT SURVIVAL

You have accessJournal of UrologyTransplantation & Vascular Surgery: Renal Transplantation & Vascular Surgery II1 Apr 2016MP32-01 PERCUTANEOUS NEPHROSTOMY PLACEMENT AFTER RENAL TRANSPLANTATION: INDICATIONS AND IMPLICATIONS FOR LONG-TERM GRAFT SURVIVAL Jeffrey Pearl, Joshua Finkle, Mikin Patel, Thuong Van Ha, Jeffrey Leef, Kenneth Ogan, and Jonathan Lorenz Jeffrey PearlJeffrey Pearl More articles by this author , Joshua FinkleJoshua Finkle More articles by this author , Mikin PatelMikin Patel More articles by this author , Thuong Van HaThuong Van Ha More articles by this author , Jeffrey LeefJeffrey Leef More articles by this author , Kenneth OganKenneth Ogan More articles by this author , and Jonathan LorenzJonathan Lorenz More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2016.02.1296AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Urologic intervention after renal transplantation is a rare requirement often utilizing a multidisciplinary approach between interventional radiologists and urologists. Frequently, prior to definitive treatment of transplant ureteral obstruction or urinary leak, urinary diversion by percutaneous nephrostomy is required. The purpose of our study is to identify indications and factors predictive of long-term graft survival in patients requiring percutaneous nephrostomy placement after renal transplantation. METHODS A retrospective chart review was performed to identify renal transplant patients who underwent subsequent nephrostomy placement between July 1, 2005 and June 30, 2015. Patient and procedure related variables, imaging of the transplant graft, and graft survival data were collected. Kaplan-Meier analysis was performed to assess our primary outcome, which was the relationship between factors surrounding nephrostomy placement and graft survival. RESULTS 51 patients received percutaneous nephrostomy tubes with a technical success rate of 98% and complication rate of 27%. Indications included ureteral stricture in 63%, urolithiasis in 25%, and urine leak in 12%. Median graft survival was 11.9 years (IQR 4.7 to 17.9 years) versus 10.5 years as reported in the general transplant population. Factors associated with worse graft survival included age greater than 60 years (HR 2.9 [CI 1.3-6.1, p<.05]), moderate or greater hydronephrosis (HR 3.8 [CI 1.5-9.8, p<.01]), serum creatinine (SCr) greater than 2 ng/dL (HR 5.3 [CI 1.3-22.4, p<.05]), no post-procedural drop in SCr (HR 2.8 [CI 1.3-6.0, p<.01]), and a nephrostomy related complication (HR 2.7 [CI 1.2-6.3, p<.05]). Patients with ureteral stricture had worse graft survival than those with stones (HR 6.4 [CI 1.9-21.7, p<.01]). Patients with stricture underwent either dilation by interventional radiology (28%), endourologic treatment (12.5%), open repair (9%), had graft failure or death prior to repair (12.5%), or did not undergo repair (38%). There was not a significant difference in graft survival between types of stricture repair. CONCLUSIONS Renal transplant patients requiring nephrostomy have similar graft survival compared to the general transplant population. In our population, ureteral stricture was the most common indication for nephrostomy tube, and a variety of approaches were implemented for repair without significant differences in graft survival. Multiple patient-related factors may be predictive of long-term graft survival in this setting. © 2016FiguresReferencesRelatedDetails Volume 195Issue 4SApril 2016Page: e427-e428 Advertisement Copyright & Permissions© 2016MetricsAuthor Information Jeffrey Pearl More articles by this author Joshua Finkle More articles by this author Mikin Patel More articles by this author Thuong Van Ha More articles by this author Jeffrey Leef More articles by this author Kenneth Ogan More articles by this author Jonathan Lorenz More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Post-transplant survival in idiopathic pulmonary fibrosis patients concurrently listed for single and double lung transplantation

Lung transplantation is a widely accepted treatment for patients with end-stage lung disease related to idiopathic pulmonary fibrosis (IPF). However, there are conflicting data on whether double lung transplant (DLT) or single lung transplant (SLT) is the superior therapy in these patients. The purpose of this study was to determine whether actuarial post-transplant graft survival among IPF patients concurrently listed for DLT and SLT is greater for recipients undergoing the former or the latter. The United Network for Organ Sharing provided de-identified patient-level data. Analysis included lung transplant candidates with IPF listed between January 1, 2001 and December 31, 2009 (n = 3,411). The study population included 1,001 (29.3%) lung transplant recipients concurrently listed for DLT and SLT, all ≥18 years of age. The primary outcome measure was actuarial post-transplant graft survival, expressed in years. Among the study population, 433 (43.26%) recipients underwent SLT and 568 (56.74%) recipients underwent DLT. The analysis included 2,722.5 years at risk, with median graft survival of 5.31 years. On univariate (p = 0.317) and multivariate (p = 0.415) regression analyses, there was no difference in graft survival between DLT and SLT. Among IPF recipients concurrently listed for DLT and SLT, there is no statistical difference in actuarial graft survival between recipients undergoing DLT vs SLT. This analysis suggests that increased use of SLT for IPF patients may increase the availability of organs to other candidates, and thus increase the net benefit of these organs, without measurably compromising outcomes.

Graft Outcomes in Kidney Transplantation Alone with Concomitant Cirrhosis

aim: The influence of cirrhosis on kidney graft survival post-transplantation has not been well defined. As previously demonstrated, lower rates of patient overall survival (31%) were in conjunction with lower rates of 5-year graft survival (31%) in cirrhotic patients who received a kidney transplant alone. Unfortunately, there is scarce data on the graft survival in patients who receive a kidney transplant alone with concomitant cirrhosis. Methods: Among 131 patients with kidney transplantations at Methodist Specialty Hospital in San Antonio, TX between January 1999 and December 2011 with serological evidence of hepatitis B or C, 12 patients were found to have concomitant cirrhosis at time of transplantation. Rates of survival at 3-years were compared to the national average and overall for Methodist Specialty Hospital. Results: In the 12 patients, cirrhosis etiologies included hepatitis B (n=2), hepatitis C (n=7) and cryptogenic (n=3). All were confirmed by histology at or within 6 months prior to kidney transplantation. The median graft survival was 8.1 years and graft survival at 3- and 5-years were 75% and 58%, respectively. Conclusion: Preliminary data suggests that cirrhotic patients might be considered for combined liver and kidney transplantation. Graft survival at 3 years is far below the national average (85%) and Methodist Hospital overall (89%). Study also demonstrates the benefit of a liver biopsy with or without portal pressure measurement prior to transplantation in patients with viral hepatitis and/or other hepatic disease to accurately stage fibrosis.

Targeting JAK2 By Gene Knockout or Pacritinib Treatment Reduces Gvhd and Xenograft Rejection By Promoting Induced Treg Differentiation

Janus kinase 2 (JAK2) signal transduction is a critical mediator of immune response. JAK2 activation promotes T-cell allosensitization, as well as Th1 and Th17 differentiation. JAK2-mediated STAT3 phosphorylation limits the generation of induced regulatory T cells (iTregs) by disrupting interactions between STAT5 and Foxp3. JAK2 is implicated in the onset of graft-versus-host disease (GVHD), which is a significant cause of transplant-related mortality after allogeneic hematopoietic cell transplantation (allo-HCT). Using murine models of allo-HCT we show here that transfer of donor JAK2-/- T cells is associated with significantly less GVHD, compared with wild-type or JAK2 replete donors (Figure 1, P =.003 and 0.01, respectively). Th1 differentiation among JAK2-/- T-cells is dramatically decreased, compared with controls. Conversely, iTreg polarization and stability are significantly increased among the JAK2 deficient T cells. To investigate the effect of pharmacologic JAK2 inhibition on T-cell alloresponses, pacritinib (supplied by CTI BioPharma), was chosen as it does not induce myelosuppression or increase risk for opportunistic infections in myelofibrosis patients - unlike JAK1/JAK2 inhibitors. Pacritinib potently inhibits JAK2, but also has suppressive activity toward JAK3, CSF1R, and IRAK1. Pacritinib was administered at 100mg/kg twice a day by oral gavage for 4 weeks beginning on the day of MHC-disparate allo-HCT, significantly reducing GVHD in recipient mice compared with methylcellulose vehicle control. In allogeneic mixed leukocyte reactions using human cells, pacritinib (2.5μM) significantly reduces T-cell proliferation after 5 days of culture (P <.0001). In vitro studies verified pacritinib eliminates STAT3 activity by IL-6 in human CD4+ T-cells, while permitting IL-2 induced STAT5 phosphorylation despite its effects on JAK3. This offers a platform to reduce Th17 development, while promoting iTreg populations. As such, pacritinib dramatically suppresses Th17 differentiation reflected by RORgammaT expression among naïve CD4+ T cells stimulated by allogeneic dendritic cells (DC), compared with DMSO (P <.0001). iTregs were generated with allogeneic DCs in the presence of pacritinib or DMSO for 5 days, and then cultured with self T-cell responders and fresh DCs without additional drug exposure. The suppressive potency of either pacritinib- or DMSO-treated iTregs was similar, suggesting JAK2 is not required for iTreg function. We then evaluated whether JAK2 inhibition could prevent human skin graft rejection in an NSG mouse xenograft model. A 1x1 cm split-thickness human skin graft was transplanted onto the animal dorsally. The mice rested for 30 days after surgery to permit engraftment, then received 5x106 allogeneic peripheral blood mononuclear cells (PBMC) by intraperitoneal injection. Pacritinib was administered at the same dose and schedule for 2 weeks beginning at time of PBMC injection as tolerated by the NSG mice. The treatment significantly delayed allograft rejection by the human donor PBMCs, compared with vehicle control (Figure 2: Median graft survival 32.5 v 51 days, Day +60 survival 0% v 33.3%, pooled data from 2 experiments, n=5-6 mice per arm, P =.0011). Collectively, these data clearly identify JAK2 as a therapeutic target to control donor alloreactivity and promote iTreg responses after HCT or solid organ transplantation.*BCB and DB contributed equally to this work. [Display omitted] [Display omitted] DisclosuresSinger:CTI BioPharma, Corp: Employment, Equity Ownership.

Pre-clinical heterotopic intrathoracic heart xenotransplantation: a possibly useful clinical technique.

As a step towards clinical cardiac xenotransplantation, our experimental heterotopic intrathoracic xenotransplantation model offers a beating and ejecting donor heart while retaining the recipient's native organ as a backup in case of graft failure. Clinically applicable immunosuppressive regimens (IS) were investigated first, then treatments known to be effective in hypersensitized patients or those with recalcitrant rejection reactions. Consecutive experiments were carried out between 2009 and 2013. Twenty-one genetically modified pigs (GGTA1-knockout/hCD46/± thrombomodulin, in one case HLA-E instead) were used as donors. In all experiments, two cycles of immunoabsorption reduced preformed antibodies. Recipient baboons were divided into two groups according to IS regimen: In group one (n = 10), pre-treatment started either one (anti-CD20) or four weeks (anti-CD20 plus the proteasome inhibitor bortezomib) prior to transplantation. The extended conventional (as for allotransplantation) immunosuppressive maintenance regimen included anti-thymocyte globuline, tacrolimus, mycophenolate mofetil, methylprednisolone and weekly anti-CD20. In group two (n = 11), myeloablative pre-treatment as in multiple myeloma patients (long and short regimens) was added to extended conventional IS; postoperative total thoracic and abdominal lymphoid irradiation (TLI; single dose of 600 cGY) was used to further reduce antibody-producing cells. In the perioperative course, the surgical technique was safely applied: 19 baboons were weaned off extracorporeal circulation and 17 extubated. Nine animals were lost in the early postoperative course due to causes unrelated to surgical technique or IS regimen. Excluding these early failures, median graft survival times of group 1 and 2 were 18.5 (12-50) days and 16 (7-35) days. Necropsy examination of group 1 donor organs revealed hypertrophy of the left ventricular wall in the six longer-lasting grafts; myocardial histology confirmed pre-clinical suspicion of humoral rejection, which was not inhibited by the extended conventional IS including intensified treatments, and signs of thrombotic microangiopathy. Grafts of group 2 presented with only mild-to-moderate features of humoral rejection and thrombotic microangiopathy, except in one case of delayed rejection on day 17. The other experiments in this group were terminated because of untreatable pulmonary oedema, recurring ventricular fibrillation, Aspergillus sepsis, as well as a combination of a large donor organ and late toxic side effects due to TLI. Longer-term results were difficult to achieve in this model due to the IS regimens used. However, we conclude that heterotopic intrathoracic heart transplantation may be an option for clinical xenotransplantation.

Spiegelmer Inhibition of MCP-1/CCR2--Potential as an Adjunct Immunosuppressive Therapy in Transplantation.

The rejection process remains the key unsolved issue after transplantation of disparate tissue. The CC chemokine monocyte chemoattractant protein-1 (MCP-1/CCL2) has been reported to be involved in the process of alloimmune interaction. Spiegelmers are l-oligonucleotides that can be designed to bind to pharmacologically relevant target molecules. Here, we tested a high-affinity Spiegelmer-based MCP-1 inhibitor (mNOX-E36) in an allogeneic heart transplant model. Fully vascularized allogeneic heterotopic heart transplantations from BALB/c to C57BL/6 mice were performed. Mice were either treated with the anti-MCP-1-Spiegelmer (mNOX-E36) in monotherapy or in combination with subtherapeutic doses of cyclosporine A (CsA) (10 mg/kgBW/day) for 10 days. Controls received equivalent doses of a non-functional Spiegelmer (revmNOX-E36). Graft survival of allogeneic heart transplants was slightly but significantly prolonged under mNOX-E36 monotherapy (median graft survival 10 day ± 0.7) compared to revmNOX-E36 (median graft survival 7 day ± 0.3; P = 0.001). A synergistic beneficial effect could be seen when mNOX-E36 was administered in combination with subtherapeutic doses of CsA (18 day ± 2.8 versus 7 day ± 0.3; P < 0.0001). Levels of inflammatory cytokines and 'alarmins' were significantly reduced, and the number of F4/80(+) cells was lower under combination therapy (1.8% ± 1.3%; versus 14.6% ± 4.4%; P = 0.0002). This novel inhibitor of the MCP-1/CCR2 axis (mNOX-E36), which has already proven efficacy and tolerability in early clinical trials, alleviates acute rejection processes in allogeneic transplantation especially when combined with subtherapeutic doses of CsA. Thus, mNOX-E36 may have potential as an adjunct immunomodulatory agent.

Recurrent membranoproliferative glomerulonephritis type I after kidney transplantation: a 17-year single-center experience.

Most previously published studies of patients with membranoproliferative glomerulonephritis type I are small or have short follow-up period. We report the outcome of a fairly large cohort of patients followed up for nearly 10 years. Retrospective cohort study. Graft survival, recurrence rate and risk factors for recurrence were analyzed for 43 patients transplanted between the years 1995 and 2012. At a mean overall follow-up of 118 ± 61 months (median, 127.8; range, 4.9-217), 12 patients lost their graft (28%). Death-censored actuarial 15-year graft survival rate was 56%. Membranoproliferative glomerulonephritis recurred in eight patients (19%) at a median time of 15.4 months (range, 4.4-70 months). Recurrence led to graft loss in seven patients (88%) within a median of 11.6 months (range, 1.3-54 months) from diagnosis. Median graft survival was 30.5 months for recurrence (range, 7-86). Actuarial 15-year graft survival was 71% for nonrecurrent. The risk for recurrence was higher for patients with human leukocyte antigen (HLA) B49 (odds ratio, 16.9; 95% confidence interval, 1.1-246; P=0.038) and HLA DR4 (odds ratio, 15.9; 95% confidence interval, 1.07-237; P=0.044) alleles. A trend toward increased risk was found with shorter duration of dialysis before transplantation. Four of 16 (25%) living-related versus none of the living-unrelated donors' recipients recurred. The HLA B49, acute tubular necrosis after transplantation, previous transplantations, and Arab origin were all associated with decreased graft and patient survival. Patients without recurrence in the first years should expect an excellent graft survival. Nonrelated living donors should be preferred. The HLA B49 and DR4 alleles may increase the risk for recurrence.

Effect of selective CD28 blockade with FR104 alone or combined with CD154 blockade in monkey cardiac allograft transplantation. (TRAN2P.967)

Abstract Selective CD28 blockade, which preserves CTLA-4-dependent immune regulation, is a promising approach to induce tolerance. FR104, a novel CD28 antagonist, was evaluated in monkey heart transplant recipients that received αCD28 alone (n=3, FR104) or in combination with αCD154 (n=5, hu5C8), dosed to achieve receptor coverage for 84 days; results were compared to αCD154 alone (n=4). Acute (AR) and chronic (CAV) rejection scores were quantified histologically. Median graft survival time (MST) with FR104 (167d), hu5C8 (133d), and FR104+hu5C8 (173d) was not statistically different between groups. Rejection was seen in 1 of 3 animals with FR104 monotherapy despite CD28 receptor saturation, but not with additional CD154 blockade. CAV was decreased with FR104 (0.7±0.3) and FR104+hu5C8 (0.5±0.3) compared to hu5C8 (1.6±0.7 p&amp;lt;0.05). AR tended to be lower with FR104 (0.5±0.4, p=0.047) or combined therapy (0.8±0.4, p=0.07) on biopsy/explanted functional grafts relative to hu5C8 (1.4±0.6). Blood Tregs frequency on d7-21 was increased (fold change vs. baseline) with FR104 (1.5±0.2), hu5C8 (1.4±0.3), and FR104+hu5C8 (2.3±0.1). CD28 saturation became undetectable by d150 in blood and LN; subsequent graft failure was associated with alloantibodies. We conclude that selective non-activating CD28 blockade protects primate cardiac allografts from pathogenic alloimmunity, an effect enhanced by simultaneous CD154 blockade, supporting the hypothesis that CD28 blockade is potentially tolerogenic.

The Clinical Course of Kidney Transplant Recipients After 20 Years of Graft Function

There is a growing population of kidney transplant recipients who have survived 20 years with a functioning graft. This study identified the factors associated with prolonged survival and described the clinical course of recipients after two decades of transplant function. All recipients transplanted in Northern Ireland between 1968 and 1993 were included (n = 706) and data were collected prospectively. At 20 years, 25% had a functioning transplant; in multivariate analysis younger recipient age and living donation were associated with 20-year survival. The median recipient survival beyond two decades was 13.3 years; cancer was the commonest cause of death. De novo malignancy developed in 37% of recipients and cardiovascular disease in 27% after 20 years of graft function. The median graft survival after 20 years was 9.3 years; 69% of graft loss was due to death with a functioning transplant. Advances in kidney transplantation have improved the long-term survival of both graft and recipient. After two decades the majority of patients die with a functioning graft. The focus of management in long-term survivors may need to be on the prevention of cancer and cardiovascular disease to allow further improvements in graft and recipient survival.

Combination of C-X-C motif chemokine 9 and C-X-C motif chemokine 10 antibodies with FTY720 prolongs the survival of cardiac retransplantation allografts in a mouse model.

The upregulation of chemokine genes and the subsequent T-lymphocyte recruitment to the graft are early events in the development of acute cardiac transplant rejection or cardiac allograft vasculopathy. In the present study, a combined immunosuppressive regimen of C-X-C motif chemokine 9 (CXCL9) antibody (Ab), CXCL10 Ab and FTY720 was used in order to reduce the infiltration of memory T lymphocytes and prolong graft survival in a retransplantation murine model. BALB/c donor hearts were transplanted heterotopically into C57BL/6 mice at day 28 after skin transplantation. The mice were divided into four groups: i) Control (normal saline), ii) CXCL9 Ab and CXCL10 Ab [150 μg; once daily (qd); intraperitoneal (ip)], iii) FTY720 (0.2 mg/day; qd; ip) and iv) combined (2 mg/kg/day; qd; ip). Measurements of the median survival time of the cardiac grafts, histological examination, reverse transcription-quantitative polymerase chain reaction analysis, enzyme-linked immunosorbent assay and a mixed lymphocyte reaction were performed. The median graft survival time of the combined group was prolonged (9.3 days) compared with that of the control group (3.5 days) (P<0.001). Histological examination revealed that the combined treatment group graft rejection pathological score was 0.50, while the control group score was 3.62 (P<0.001). In addition, the gene expression level of interleukin (IL)-2 was significantly lower and the levels of IL-10 and transforming growth factor-β (TGF-β) were significantly higher in the combined group compared with those in the control group (P<0.001). Furthermore, the serum concentration levels of IL-2 and interferon-γ (IFN-γ) were significantly lower (P<0.001) and the concentration of IL-10 was significantly higher (P<0.05) in the combined group compared with those in the control group. In the mixed lymphocyte reaction, T-cell proliferation was found to be significantly lower in the combined treatment group than that in the control group (P<0.001). In conclusion, treatment with CXCL9 Ab and CXCL10 Ab or FTY720 reduced the graft infiltration of inflammatory cells, inhibited T-cell proliferation and prolonged graft survival. The combined treatment regimen of CXCL9 Ab, CXCL10 Ab and FTY720 was found to significantly reduce the infiltration of inflammatory cells in the graft and prolong graft survival.