[Show abstract][Hide abstract] ABSTRACT: BACKGROUND:
Development of noninvasive molecular assays to improve disease diagnosis and patient monitoring is a critical need. In renal transplantation, acute rejection (AR) increases the risk for chronic graft injury and failure. Noninvasive diagnostic assays to improve current late and nonspecific diagnosis of rejection are needed. We sought to develop a test using a simple blood gene expression assay to detect patients at high risk for AR.
METHODS AND FINDINGS:
We developed a novel correlation-based algorithm by step-wise analysis of gene expression data in 558 blood samples from 436 renal transplant patients collected across eight transplant centers in the US, Mexico, and Spain between 5 February 2005 and 15 December 2012 in the Assessment of Acute Rejection in Renal Transplantation (AART) study. Gene expression was assessed by quantitative real-time PCR (QPCR) in one center. A 17-gene set-the Kidney Solid Organ Response Test (kSORT)-was selected in 143 samples for AR classification using discriminant analysis (area under the receiver operating characteristic curve [AUC] = 0.94; 95% CI 0.91-0.98), validated in 124 independent samples (AUC = 0.95; 95% CI 0.88-1.0) and evaluated for AR prediction in 191 serial samples, where it predicted AR up to 3 mo prior to detection by the current gold standard (biopsy). A novel reference-based algorithm (using 13 12-gene models) was developed in 100 independent samples to provide a numerical AR risk score, to classify patients as high risk versus low risk for AR. kSORT was able to detect AR in blood independent of age, time post-transplantation, and sample source without additional data normalization; AUC = 0.93 (95% CI 0.86-0.99). Further validation of kSORT is planned in prospective clinical observational and interventional trials.
The kSORT blood QPCR assay is a noninvasive tool to detect high risk of AR of renal transplants. Please see later in the article for the Editors' Summary.
[Show abstract][Hide abstract] ABSTRACT: Monitoring of renal graft status through peripheral blood (PB) rather than invasive biopsy is important as it will lessen the risk of infection and other stresses, while reducing the costs of rejection diagnosis. Blood gene biomarker panels were discovered by microarrays at a single center and subsequently validated and cross-validated by QPCR in the NIH SNSO1 randomized study from 12 US pediatric transplant programs. A total of 367 unique human PB samples, each paired with a graft biopsy for centralized, blinded phenotype classification, were analyzed (115 acute rejection (AR), 180 stable and 72 other causes of graft injury). Of the differentially expressed genes by microarray, Q-PCR analysis of a five gene-set (DUSP1, PBEF1, PSEN1, MAPK9 and NKTR) classified AR with high accuracy. A logistic regression model was built on independent training-set (n = 47) and validated on independent test-set (n = 198)samples, discriminating AR from STA with 91% sensitivity and 94% specificity and AR from all other non-AR phenotypes with 91% sensitivity and 90% specificity. The 5-gene set can diagnose AR potentially avoiding the need for invasive renal biopsy. These data support the conduct of a prospective study to validate the clinical predictive utility of this diagnostic tool.
American Journal of Transplantation 10/2012; 12(10):2710-2718. · 6.19 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: To determine whether steroid avoidance in pediatric kidney transplantation is safe and efficacious, a randomized, multicenter trial was performed in 12 pediatric kidney transplant centers. One hundred thirty children receiving primary kidney transplants were randomized to steroid-free (SF) or steroid-based (SB) immunosuppression, with concomitant tacrolimus, mycophenolate and standard dose daclizumab (SB group) or extended dose daclizumab (SF group). Follow-up was 3 years posttransplant. Standardized height Z-score change after 3 years follow-up was -0.99 ± 2.20 in SF versus -0.93 ± 1.11 in SB; p = 0.825. In subgroup analysis, recipients under 5 years of age showed improved linear growth with SF compared to SB treatment (change in standardized height Z-score at 3 years -0.43 ± 1.15 vs. -1.07 ± 1.14; p = 0.019). There were no differences in the rates of biopsy-proven acute rejection at 3 years after transplantation (16.7% in SF vs. 17.1% in SB; p = 0.94). Patient survival was 100% in both arms; graft survival was 95% in the SF and 90% in the SB arms (p = 0.30) at 3 years follow-up. Over the 3 year follow-up period, the SF group showed lower systolic BP (p = 0.017) and lower cholesterol levels (p = 0.034). In conclusion, complete steroid avoidance is safe and effective in unsensitized children receiving primary kidney transplants.
American Journal of Transplantation 06/2012; 12(10):2719-2729. · 6.19 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Steroid avoidance is safe and effective in children receiving kidney transplants in terms of graft function and survival, but the effects on allograft histology are unknown. In this multicenter trial, 130 pediatric renal transplant recipients were randomized to steroid-free (SF; n = 60) or steroid-based (SB; n = 70) immunosuppression, and underwent renal allograft biopsies at the time of graft dysfunction and per protocol at implantation and 6, 12 and 24 months after transplantation. Clinical follow-up was 3 years posttransplant. Subclinical acute rejection was present in 10.6% SF versus 11.3% SB biopsies at 6 months (p = 0.91), 0% SF versus 4.3% SB biopsies at 1 year (p = 0.21) and 0% versus 4.8% at 2 years (p = 0.20). Clinical acute rejection was present in 13.3% SF and 11.4% SB patients by 1 year (p = 0.74) and in 16.7% SF and 17.1% SB patients by 3 years (p = 0.94) after transplantation. The cumulative incidence of antibody-mediated rejection was 6.7% in SF and 2.9% in SB by 3 years after transplantation (p = 0.30). There was a significant increase in chronic histological damage over time (p < 0.001), without difference between SF and SB patients. Smaller recipient size and higher donor age were the main risk factors for chronic histological injury in posttransplant biopsies.
American Journal of Transplantation 06/2012; 12(10):2730-2743. · 6.19 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Despite early promising patient and graft outcomes with steroid-free (SF) immunosuppression in pediatric kidney transplant recipients, data on long-term safety and efficacy results are lacking. We present our single-center experience with 129 consecutive pediatric kidney transplant recipients on SF immunosuppression, with a mean follow-up of 5 years. Outcomes are compared against a matched cohort of 57 concurrent recipients treated with steroid-based (SB) immunosuppression. In the SF group, 87% of kidney recipients with functioning grafts remain corticosteroid-free. Actual intent-to-treat SF (ITT-SF) and still-on-protocol SF patient survivals are 96% and 96%, respectively, actual graft survivals for both groups are 93% and 96%, respectively and actual death-censored graft survivals for both groups are 97% and 99%, respectively. Unprecedented catch-up growth is observed in SF recipients below 12 years of age. Continued low rates of acute rejection, posttransplant diabetes mellitus (PTDM), hypertension and hyperlipidemia are seen in SF patients, with sustained benefits for graft function. In conclusion, extended enrollment and longer experience with SF immunosuppression for renal transplantation in low-risk children confirms protocol safety, continued benefits for growth and graft function, low acute rejection rates and reduced cardiovascular morbidity.
American Journal of Transplantation 06/2009; 9(6):1362-72. · 6.19 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We report 1-year outcomes of a randomized study of Rituximab versus standard-of-care immunosuppression (Thymoglobulin and/or pulse steroids) for treatment of biopsy confirmed, acute transplant rejection with B-cell infiltrates, in 20 consecutive recipients (2-23 years). Graft biopsies, with Banff and CADI scores, CD20 and C4d stains, were performed at rejection and 1 and 6 months later. Peripheral blood CMV, EBV and BK viral loads, graft function, DSA, immunoglobulins, serum humanized antichimeric antibody (HACA) and Rituximab, and lymphocyte counts were monitored until 1 year posttreatment. Rituximab infusions were given with a high index of safety without HACA development and increased infections complications. Rituximab therapy resulted in complete tissue B-cell depletion and rapid peripheral B-cell depletion. Peripheral CD19 cells recovered at a mean time of approximately 12 months. There were some benefits for the recovery of graft function (p = 0.026) and improvement of biopsy rejection scores at both the 1- (p = 0.0003) and 6-month (p < 0.0001) follow-up biopsies. Reappearance of C4d deposition was not seen on follow-up biopsies after Rituximab therapy, but was seen in 30% of control patients. There was no change in DSA in either group, independent of rejection resolution. This study reports safety and suggests further investigation of Rituximab as an adjunctive treatment for B-cell-mediated graft rejection.
American Journal of Transplantation 09/2008; 8(12):2607-17. · 6.19 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: With the increasing adoption of steroid-sparing immunosuppression protocols in renal transplantation, it is important to evaluate any adverse effects of steroid avoidance on graft function. Early graft function, measured by CrCl was retrospectively studied in 158 consecutive pediatric renal transplant recipients from 1996 to 2005, receiving either steroid-free or steroid-based immunosuppression. Patients receiving steroid-free immunosuppression vs. steroid-based immunosuppression had no difference change in CrCl (DeltaCrCl) in the first week post-transplantation (p = 0.12). When stratified by corticosteroid usage, patients with higher tacrolimus trough levels (> or =14 ng/mL) had slower graft function recovery in the first week post-transplantation than those with lower tacrolimus trough levels (p = 0.008) in the steroid-free group only. Despite initial slower graft function recovery in this subgroup, there was no negative impact on graft function in the steroid-free group; in fact steroid-free patients trended towards better CrCl at six months (p = 0.047) and 12 months (p < 0.001) post-transplant than the steroid-based group. With the improved immunological outcomes with steroid avoidance, close surveillance should be performed of tacrolimus levels to avoid levels >14 ng/mL. In patients with slow recovery of early graft function, short-term perioperative steroids may be considered.
[Show abstract][Hide abstract] ABSTRACT: To describe the evolution, risk factors and impact of nonimmune histological injury after pediatric kidney transplantation, we analyzed 245 renal allograft protocol biopsies taken regularly from the time of transplantation to 2 years thereafter in 81 consecutive rejection-free pediatric recipients of an adult-sized kidney. Isometric tubular vacuolization was present early after transplantation was not progressive, and was associated with higher tacrolimus pre-dose trough levels. Chronic tubulo-interstitial damage and tubular microcalcifications were already noted at 3 months, were progressive and had a greater association with small recipient size, male donor gender, higher donor age and female recipient gender, but not with tacrolimus exposure. Renal function assessment showed that older recipients had a significant increase in absolute glomerular filtration rate with time after transplantation, which differed from small recipients who showed no increase. It is concluded that progressive, functionally relevant, nonimmune injury is detected early after adult-sized kidney transplantation in pediatric recipients. Renal graft ischemia associated with the donor-recipient size discrepancy appears to be a greater risk factor for this chronic histological injury, suggesting that the exploration of additional therapeutic approaches to increase allograft perfusion could further extend the graft survival benefit of adult-sized kidneys transplanted into small children.
American Journal of Transplantation 12/2007; 7(11):2504-14. · 6.19 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Transient or intermittent plasmapheresis with concurrent immunosuppressive therapy is thought to be beneficial in the treatment of recurrent focal segmental glomerulosclerosis (FSGS) in the early post-transplant period. The results of long-term (6-year) plasmapheresis therapy, in a 9-year-old female with an immediate recurrence of FSGS [urinary protein/urinary creatinine (UP/UC)=17.7] after cadaveric renal transplant, are presented. A 4-week plasmapheresis course induced a decline in the proteinuria, but a relapse occurred after cessation of plasmapheresis. Addition of protein A column therapy led to a further decrease in the proteinuria, to a non-nephrotic range. Long-term control of the nephrotic syndrome was established using a chronic treatment regimen consisting of a single-volume plasmapheresis, followed by a protein A column treatment, performed on sequential days every 3-4 weeks. Mean UP/UC values decreased to 1.15+/-0.9. A course of cyclophosphamide was successfully used to control a worsening of proteinuria 4 years post transplant. Although sequential renal biopsies demonstrated progressive glomerular sclerosis, the patient's mean calculated creatinine clearance only modestly declined from 78.3 ml/min per 1.73 m2, at the time of transplantation, to 62.7 ml/min per 1.73 m2, 6 years later. This patient demonstrated dependence on plasmapheresis/protein A column therapy to maintain a clinical remission of her FSGS recurrence. While long-term plasmapheresis and protein A column therapy in combination with immunosuppressive therapy reversed the effects of uncontrolled nephrosis and possibly facilitated long-term renal allograft survival, the glomerular sclerosis continued to progress.
[Show abstract][Hide abstract] ABSTRACT: Corticosteroids have been a cornerstone of immunosuppression for four decades despite their adverse side effects. Past attempts at steroid withdrawal in pediatric renal transplantation have had little success. This study tests the hypothesis that a complete steroid-free immunosuppressive protocol avoids steroid dependency for suppression of the immune response with its accompanying risk of acute rejection on steroid withdrawal.
An open labeled prospective study of complete steroid avoidance immunosuppressive protocol was undertaken in 10 unsensitized pediatric recipients (ages 5-21 years; mean 14.4 years) of first renal allografts. Steroids were substituted with extended daclizumab use, in combination with tacrolimus and mycophenolate mofetil. Protocol biopsies were performed in the steroid-free group at 0, 1, 3, 6, and 12 months posttransplantation. Clinical outcomes were compared to a steroid-based group of 37 matched historical controls.
Graft and patient survival was 100% in both groups. Clinical acute rejection was absent in the steroid-free group at a mean follow-up time of 9 months (range 3-13.7 months). Protocol biopsies in the steroid-free group (includes 10 patients at 3 months, 7 at 6 months, and 4 at 12 months) revealed only two instances of mild (Banff 1A) subclinical rejection (reversed by only a nominal increase in immunosuppression) and no chronic rejection. At 6 months the steroid-free group had no hypertension requiring treatment (P=0.003), no hypercholesterolemia (P=0.007), and essentially no body disfigurement (P=0.0001). Serum creatinines, Schwartz GFR, and mean delta height Z scores trended better in the steroid-free group. In the steroid-free group, one patient had cytomegalovirus disease at 1 month and three had easily treated herpes simplex stomatitis, but with no significant increase in bacterial infections or rehospitalizations over the steroid-based group. The steroid-free group was more anemic early posttransplantation (P=0.004), suggesting an early role of steroids in erythrogenesis; erythropoietin use normalized hematocrits by 6 months.
Complete steroid-free immunosuppression is efficacious and safe in this selected group of children with no early clinical acute rejection episodes. This protocol avoids the morbid side effects of steroids without increasing infection, and may play a future critical role in avoiding noncompliance, although optimizing renal function and growth.
[Show abstract][Hide abstract] ABSTRACT: Differentiating etiologies of transplant dysfunction without biopsy and optimizing therapy for acute rejection by predicting steroid resistance will reduce patient morbidity. Granulysin is a cytolytic molecule released by CTL and NK cells and coexpressed with effectors of acute allograft rejection, like perforin and granzymes. Granulysin mRNA and protein expression were studied in peripheral blood lymphocytes (PBL; n = 61 total, n = 10 with intercurrent infections) and biopsy tissue from adult and children renal transplant recipients (n = 97) by competitive quantitative-reverse transcriptase-PCR (QC-RT-PCR) and immunohistochemistry. Differences in cell phenotypes were studied in steroid sensitive and resistant acute rejection biopsies. Granulysin was studied in phytohemagglutinin (PHA) stimulated cell lines (donor PBL and CD45RO(+) T cells) by FACS, Western blotting, and RT-PCR after pretreating with cyclosporine A (CSA), azathioprine, mycophenolic acid, and steroids. Granulysin mRNA was significantly increased in patient PBL and transplant biopsies during acute rejection (p < 0.0001) and infection (p < 0.001). Rejecting biopsies alone (n = 53) had mononuclear cell granulysin staining. Steroid resistant biopsies (n = 25) had denser granulysin staining (>2 cells/high power field) and CD45RO(+) lymphocytes, when compared with steroid sensitive (n = 28) rejecting tissue. Granulysin levels were unchanged after azathioprine and mycophenolic acid treatment, decreased after treating activated PBL with steroids and cyclosporine A (CSA), and paradoxically, increased (p < 0.05) after treating CD45RO(+) CTL with CSA. Elevated PBL granulysin is a peripheral marker for acute rejection and infection and dense granulysin staining a tissue marker for steroid resistance. Memory CTL abound in steroid resistant grafts and may have a markedly different response to CSA immunotherapy, suggesting a possible mechanism for steroid resistance.
Human Immunology 02/2001; 62(1):21-31. · 2.30 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: BACKGROUND: Infants with end-stage renal disease are at highest risk for early graft loss and mortality of any subgroup undergoing renal transplantation. This study evaluates the influence of donor tissue mass and acute tubular necrosis (ATN) on graft survival and incidence of acute rejection episodes in infant and small child recipients of living donor (LD) and cadaver (CAD) adult-size kidneys (ASKs), pediatric CAD kidneys and combined kidney-liver transplants. Methods. Kidney transplants in infants and small children at a single center and those reported to the UNOS Scientific Renal Transplant Registry were analyzed. At Stanford, multi-variate analysis was conducted on 45 consecutive renal allograft recipients weighing < or = 15 kg, mean weight 11.2 +/- 2.6 kg. The UNOS Registry results in age groups 0-2.5 (n=548) and 2.5-5 years (n=743) were compared with age groups 6-12, 13-18, and the lowest risk adult group of 19-45 years. STANFORD RESULTS. Graft survival was 97.8 +/- 0.0 at 2 years and 84.6 +/- 0.1% at 8 years. The incidence of biopsy proven rejection was 8.8% in the first 3 months and 15.5% over the 8-year follow-up. None of the pediatric CAD kidneys had ATN. Rejection episodes were restricted to the pediatric CAD kidneys alone (3/3), with no kidney rejections in the combined pediatric CAD kidney-liver transplants (0/6; P=0.003). Four ASK transplants had ATN (1 postoperative and 3 late), and all predisposed to subsequent acute rejection episodes (4/4), whereas there were no rejection episodes in ASK transplants without ATN (0/32; P<0.001). At 3 years posttransplantation, mean serum creatinines were worse in ASKs with ATN (1.5 vs. 0.9 mg/dL; P<0.001) and in all grafts with rejection episodes (1.2 vs. 0.9 mg/dL; P<0.05). UNOS RESULTS: Among the 5 age groups studied, significantly better (P<0.001) long-term graft survival rates were observed in allograft recipients in the 2 youngest age groups with ASKs without ATN: 82 +/- 3% and 81 +/- 3% for LD and 70 +/- 7% and 78 +/- 4% for CAD recipients in the 0-2.5 and 2.5- to 5-year age groups, respectively, at 6 years after transplantation. Moreover, the projected graft half-lives after the 1st year in the LD groups without ATN were at least equivalent to those of HLA-identical sibling recipients ages 19-45 years: 26.3 +/- 5 and 29.3 +/- 6 years for the 0- to 2.5- and 2.5- to 5-year age groups, respectively, and 23.3 +/- 1 years for HLA-identical transplants. The graft half-lives for CAD recipients without ATN ages 0-2.5 and 2.5-5 yearswere equivalent or better than those for LD transplants without ATN in recipients aged 19-45 years: 15.4+/- 7 and 23.7 +/- 8 years versus 15.0 +/- 0.3 years. Mean serum creatinines were superior in the 2 younger recipient age groups compared with older age groups. CONCLUSIONS: Increased donor tissue mass of the ASK or kidney-liver transplants, in the absence of ATN, seems to confer a protective effect to infant and small child recipients of these allografts. This is manifested by a prolonged rejection-free state in the single center experience and enhanced graft survival and function in the UNOS analysis, comparable to HLA identical sibling transplants for LD infant and small child recipients and to LD adult results for CAD infant and small child recipients. To optimize this protective effect by whatever mechanism, absolute avoidance of ATN is essential in infant recipients of ASK or combined kidney-liver transplants.
[Show abstract][Hide abstract] ABSTRACT: Infants make up the most high-risk, difficult to care for subgroup undergoing kidney transplantation, with the lowest 1- and 2-year graft survival rates of any other age group. The principal causes of graft loss have been graft thrombosis, primary nonfunction, technical error, and irreversible acute rejection.
Infants undergoing kidney transplantation can achieve near 100% graft survival at 2 years following surgery, despite their very high-risk status.
Analysis of 45 consecutive kidney transplants performed in patients weighing less than or equal to 15 kg during an 8-year period beginning August 1991. Patients included complex referrals from throughout the United States and all received transplants and were cared for by the same pediatric kidney transplantation team.
Mean weight at transplantation was 11. 2 kg. Renal failure was due to congenital or urologic causes in the majority (53%) of cases. Size-discrepant adult-sized kidney grafts were transplanted in 80% of patients; 64% received live-donor grafts; 78% were receiving dialysis prior to transplantation; and 27% had extremely small bladders (<20 cm(3)) requiring modification of the ureteral implantation. Excluding 1 transplant-unrelated death, graft and patient survival at 2 years was 100%. Eight-year patient and graft survival rates (for our combined live and cadaver donor series) were 89.6% and 84.6%, respectively. This compares favorably with much lower graft survival in low-risk adult recipients. Delayed graft function occurred in only 1 patient (2%). Rate of incidence of rejection was 9.3% within 2 years of transplantation and the overall rejection rate was 15.5%. No graft was lost to vascular thrombosis, primary nonfunction, technical error, or acute rejection. The mean creatinine level was 53.04 micromol/L (0.6 mg/dL) and 61.9 micromol/L (0.7 mg/dL) at 1 and 2 years, respectively, and 88.4 micromol/L (1.0 mg/dL) at 3, 4, and 5 years after transplantation.
One hundred percent 2-year and excellent 8-year graft survival rates can be achieved in what has historically been the highest-risk and most difficult to care for patient subgroup undergoing kidney transplantation.
Archives of Surgery 10/2000; 135(9):1063-8; discussion 1068-9. · 4.10 Impact Factor