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Background: A considerable proportion of patients awaiting kidney transplantation is immunized by previous transplantation(s). We investigated how allograft nephrectomy (Nx) and withdrawal of maintenance immunosuppression (WD-MIS) in patients with a failed renal allograft contribute to allosensitization., Methods: HLA antibodies (HLAabs) were analyzed before and after Nx and/or WD-MIS using a single antigen bead assay. Patients were grouped as follows: (A) Nx and concomitant WD-MIS (n = 28), (B) Nx (n = 14) and (C) WD-MIS (n = 12). In a subgroup of patients, the epitope specificity of HLAabs was determined by adsorption and elution of sera with recombinant single HLA allele-expressing cell lines., Results: Following Nx and/or WD-MIS, HLAabs were detectable in 100, 100 and 92% of patients in Groups A, B and C, respectively. In patients of all groups, de novo donor-specific HLAabs (DSAs) were found. After Nx, an increase in the breadth [percent panel reactive antibody (%PRA)] and mean fluorescence intensity of class I HLAabs was predominant. In contrast, an increase of class II HLAabs prevailed following WD-MIS. Experimental analysis of the epitope specificities revealed that 64% of the class I HLAabs classically denoted as non-DSA were donor epitope-specific HLAabs (DESA)., Conclusions: Both Nx and WD-MIS contribute to alloimmunization with differing patterns concerning class I and II HLAabs. Nx preferentially increased class I HLAabs and most of the observed class I HLAabs were DESA. Considering that class I, but not class II, HLA molecules are constitutively expressed, our results support the hypothesis that the increase of HLAabs following Nx might have been caused by removal of the adsorbing donor tissue (sponge hypothesis)., (© The Author 2016. Published by Oxford University Press on behalf of ERA-EDTA. All rights reserved.)

Background: We have found antibodies against denatured HLA class I antigens in the serum of allograft recipients which were not significantly associated with graft failure. It is unknown whether transplant recipients also have denatured HLA class II and MICA antibodies. The effects of denatured HLA class I, class II, and MICA antibodies on long-term graft outcome were further investigated based on their ability to fix complement c1q., Materials and Methods: In this 4-year retrospective cohort study, post-transplant sera from 975 kidney transplant recipients were tested for antibodies against denatured HLA/MICA antigens and these antibodies were further classified based on their ability to fix c1q., Results: Thirty percent of patients had antibodies against denatured HLA class I, II, or MICA antigens. Among them, 8.5% and 21.5% of all patients had c1q-fixing and non c1q-fixing antibodies respectively. There was no significant difference on graft survival between patients with or without antibodies against denatured HLA/MICA. However, when these antibodies were further classified according to their ability to fix c1q, patients with c1q-fixing antibodies had a significantly lower graft survival rate than patients without antibodies or patients with non c1q-fixing antibodies (p=0.008). In 169 patients who lost renal grafts, 44% of them had c1q-fixing antibodies against denatured HLA/MICA antigens, which was significantly higher than that in patients with functioning renal transplants (25%, p<0.0001). C1q-fixing antibodies were more significantly associated with graft failure caused by AMR (72.73%) or mixed AMR/CMR (61.9%) as compared to failure due to CMR (35.3%) or other causes (39.2%) (p=0.026)., Conclusions: Transplant recipients had antibodies against denatured HLA class I, II, and MICA antigens. However, only c1q-fixing antibodies were associated with graft failure which was related to antibody mediated rejection., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Background: Many patients develop de novo donor-specific anti-human leukocyte antigen antibodies (dnDSA) after transplantation. Despite development of dnDSA, not all patients will immediately fail. This study analyzes dnDSA intensity and longitudinal trends as prospective clinical parameters to assess subsequent allograft function., Methods: Twenty-four patients with dnDSA onset in the first 2 years after transplantation received antibody monitoring by LABScreen single antigen beads. Estimated glomerular filtration rate (eGFR) was recorded at time of dnDSA onset and up to 24 months thereafter. The dnDSA mean fluorescence intensity (MFI) of the stable function patient group (n=8; eGFR decline ≤ 25%) was compared with the impaired function patient group (n=16; eGFR decline>25%) using first year peak MFI (pMFI), eight month MFI change (ΔMFI), and eighteen month MFI trend (MFI slope)., Results: Both groups showed similar dnDSA characteristics (time to onset after transplantation, class I/II distribution, and initial MFI). Between groups, MFI trends were analyzed. Impaired patients showed a higher pMFI during the first year (median pMFI, 13,055 vs. 2,397; P=0.007). Longitudinal analysis revealed that ΔMFI was strongly associated with dysfunction. Both a ΔMFI increase greater than 20% as well as a stronger increase (ΔMFI>50%) were followed by graft dysfunction in almost all patients and could significantly differentiate between stable and impaired function patients (P=0.001 and P=0.04, respectively)., Conclusion: Our study suggests that tracking dnDSA intensity, particularly in the early period after onset, is important to estimate the impact of dnDSA on the allograft and could, therefore, determine help on how best to monitor patients with dnDSA.

Background: With standard IgG donor-specific anti-HLA antibody (DSA) testing, it is unclear which immunoglobulin-G (IgG) DSA positive patients will fail. We looked further into the immune response by studying immunoglobulin-M (IgM) and IgG subclass 3 (IgG3) DSA to determine if these identify the IgG DSA patients at highest risk for allograft loss., Methods: In 189 consecutively transplanted primary renal allograft recipients, sera were collected sequentially pre- and posttransplant. Of the 189, 179 patients had sera available to retrospectively test for anti-HLA IgG, IgM, and IgG3 antibodies via LABScreen single-antigen bead assay and were included in the study. All patients had a negative crossmatch. Per patient, all DSA (IgM, IgG3, and IgG) refers to the same serologic specificity., Results: Overall, 100 (56%) patients developed an alloimmune response (IgM or IgG DSA positive, or both). Ninety-five patients developed IgM DSA and 47 patients developed IgG DSA. IgM DSA was detected in 42 of 47 patients with IgG DSA. IgM DSA alone did not increase the allograft loss risk, whereas IgG DSA did (P=0.002). Once IgG DSA appeared, IgM DSA persisted in 33 patients and an isotype switch to IgG3 positive DSA occurred in 25 patients. Patients with IgM persistent IgG3 positive DSA (n=19) were more likely to have allograft failure than those without (P=0.02)., Conclusion: This study shows the evolution of the humoral immune response from IgM to IgG DSA posttransplant. We found that development of IgM persistent IgG3 positive DSA identifies the most dangerous IgG DSA subpopulation.

The development of donor specific antibodies (DSA) post transplant has been associated with chronic rejection and graft failure. In a longitudinal study, we have shown that increases in DSA precede rejection by months, thus allowing time for intervention. We hypothesized that mycophenolic acid (MPA) dose increases may reduce and/or stabilize DSA strength and also preserve renal function. Thirty stable DSA positive kidney transplant recipients participated in this Institutional Review Board approved, exploratory, open-label, single center study to assess the efficacy of MPA dose escalation in patients with DSA. MPA escalation was well tolerated and most patients were able to take higher doses for at least two years (duration of the study). In addition, MPA escalation is safe and participants had no significant side effects such as cytomegalovirus and BK infections. Long-term allograft survival of the MPA escalation group was superior when compared with the control group (p = 0.018). This pilot study indicates that escalation of MPA is safe and may stabilize DSA. In addition, five-year follow up demonstrates improved long-term survival with MPA escalation compared with DSA positive recipients receiving the standard of care. Additional studies using larger cohorts are warranted.

Background: Approximately 7% to 9% of patients with donor-specific anti-human leukocyte antigen (HLA) antibodies (DSA) fail within 1 year post-DSA onset. However, little is known as to how this DSA-associated failure temporally progresses. This longitudinal study investigates DSA's temporal relationship to allograft dysfunction and identifies predictors of allograft function's progressive deterioration post-DSA., Methods: A cohort of 175 non-HLA identical patients receiving their first transplant between March 1999 and March 2006 were analyzed. Protocol testing for DSA via single antigen beads was done before transplantation and at 1, 3, 6, 9, and 12 months after transplantation then annually. Estimated glomerular filtration rate (eGFR) was analyzed before and after DSA onset., Results: Forty-two patients developed DSA and had adequate eGFR information for analysis. Before DSA onset, the 42 patients had stable eGFR. By 1 year post-DSA, the cohort's eGFR was significantly lower (P<0.001); however, 30 of 42 had stable function. Twelve patients had failure or early allograft dysfunction (eGFR decline >25% from DSA onset). Those who failed early (by 1 year post-DSA) had more antibody-mediated rejection than stable patients (P=0.03). Late failures (after 1 year post-DSA) were predictable with evidence of early allograft dysfunction (eGFR decline >25% by 1 year post-DSA; P<0.001). Early allograft dysfunction preceded late failure by nearly 1 year., Conclusions: DSA is temporally related to allograft function deterioration. However, in many cases, late allograft failures are preceded by early allograft dysfunction. Therefore, monitoring for early allograft dysfunction provides treating physicians with a window of opportunity for treatment or continued monitoring.

Objectives: The clinical relevance of pre-transplant "low-level" donor specific anti-HLA antibodies (DSAs) in crossmatch negative kidney transplant recipients remains unclear. To determine what level of DSA associates with antibody mediated rejection (AMR) could be the way to measure the clinical relevance of pre-transplant "low-level" donor specific anti-HLA antibodies (DSAs) in crossmatch negative kidney transplant recipients., Design and Methods: A retrospective analysis of 221 patients from October 2008 to December 2009 was included in this study. Sera were obtained pre-transplant and two weeks post-transplant and tested for DSA using LABScreen single antigen beads., Results: Among the 221 patients, 11 experienced AMR within 200days after transplant (5%). Pre-transplant DSA was associated with AMR at multiple mean fluorescence intensity (MFI) cutoffs (500, 1000, 2000, 3000, 5000; p=0.003, 0.001, 0.007, 0.003, and 0.003, respectively). No correlation was seen between acute T-cell mediated rejection (CMR) and pre-transplant DSA at any of the same MFI cutoffs. There was an increased risk of AMR with higher levels of pre-transplant DSA. Finally, an increase in DSA MFI from pre- to two weeks post-transplant was indicative of a higher probability of AMR., Conclusion: Overall, this data supports using the single antigen bead to detect "low-level" DSA both pre- and post- as having a positive and persistent DSA may be predictive of higher AMR rates and poorer graft survival., (Copyright © 2013 The Canadian Society of Clinical Chemists. All rights reserved.)

Donor-specific antibodies (DSA) in sera of sensitized transplant patients are often produced against the specific epitopes on mismatched HLA antigens. In this study, we selected sera from 30 kidney transplant patients with DSA and AMR to define DQ epitopes. Using adsorption and elution assays, we identified 18 antibody reaction patterns to define 6 new epitopes and to confirm 12 previously defined epitopes. In one patient case, one mismatched antigen produced 3 different antibodies and, in another, antibodies were produced against the alpha and beta chains of the same antigen. For some sera, a single epitope can explain reactions for 27 of the 29 DQ beads in the single antigen panel. Several studies highlighted the prevalence of anti-DQ antibodies. In 2011, Almeshari et al. observed DQ DSA in 34/46 (74%) of rejection episodes - 44 patients had DSA and 20 lost their graft due to AMR. Other studies have shown a high prevalence of anti-DQ antibodies and an association with adverse effects on the graft. We conclude that analysis of the epitopes of the DQ antibodies using Adsorption/Elution and testing on single antigen DQ beads helps to better understand the specificities and cross-reactions of DQ antibodies in transplant patients., (Copyright © 2013 American Society for Histocompatibility and Immunogenetics. Published by Elsevier Inc. All rights reserved.)

Background: Anti-HLA-DQ antibodies are the predominant HLA class II donor-specific antibodies (DSAs) after transplantation. Recently, de novo DQ DSA has been associated with worse allograft outcomes. The aim of this study was to determine the further complement-binding characteristics of the most harmful DQ DSA., Methods: Single-antigen bead technology was used to screen 284 primary kidney transplant recipients for the presence of posttransplantation DQ DSA. Peak DSA sera of 34 recipients with only de novo DQ DSA and of 20 recipients with de novo DQ plus other DSAs were further analyzed by a modified single-antigen bead assay using immunoglobulin (Ig)-G subclass-specific reporter antibodies and a C1q-binding assay., Results: Compared with recipients who did not have DSA, those with de novo persistent DQ-only DSA and with de novo DQ plus other DSAs had more acute rejection (AR) episodes (22%, P=0.005; and 36%, P=0.0009), increased risk of allograft loss (hazards ratio, 3.7, P=0.03; and hazards ratio, 11.4, P=0.001), and a lower 5-year allograft survival. De novo DQ-only recipients with AR had more IgG1/IgG3 combination and C1q-binding antibodies (51%, P=0.01; and 63%, P=0.001) than patients with no AR. Furthermore, the presence of C1q-binding de novo DQ DSA was associated with a 30% lower 5-year allograft survival (P=0.003)., Conclusions: The presence of de novo persistent, complement-binding DQ DSA negatively impacts kidney allograft outcomes. Therefore, early posttransplantation detection, monitoring, and removal of complement-binding DQ might be crucial for improving long-term kidney transplantation outcomes.

The development of anti-donor humoral responses after transplantation associates with higher risks for acute rejection and 1-year graft survival in adults, but the influence of humoral immunity on transplant outcomes in children is not well understood. Here, we studied the evolution of humoral immunity in low-risk pediatric patients during the first 2 years after renal transplantation. Using data from 130 pediatric renal transplant patients randomized to steroid-free (SF) or steroid-based (SB) immunosuppression in the NIH-SNSO1 trial, we correlated the presence of serum anti-HLA antibodies to donor HLA antigens (donor-specific antibodies) and serum MHC class 1-related chain A (MICA) antibody with both clinical outcomes and histology identified on protocol biopsies at 0, 6, 12, and 24 months. We detected de novo antibodies after transplant in 24% (23% of SF group and 25% of SB group), most often after the first year. Overall, 22% developed anti-HLA antibodies, of which 6% were donor-specific antibodies, and 6% developed anti-MICA antibody. Presence of these antibodies de novo associated with significantly higher risks for acute rejection (P=0.02), chronic graft injury (P=0.02), and decline in graft function (P=0.02). In summary, antibodies to HLA and MICA antigens appear in approximately 25% of unsensitized pediatric patients, placing them at greater risk for acute and chronic rejection with accelerated loss of graft function. Avoiding steroids does not seem to modify this incidence. Whether serial assessments of these antibodies after transplant could guide individual tailoring of immunosuppression requires additional study.

Background: To date, limited information is available describing the incidence and impact of de novo donor-specific anti-human leukocyte antigen (HLA) antibodies (dnDSA) in the primary renal transplant patient. This report details the dnDSA incidence and actual 3-year post-dnDSA graft outcomes., Methods: The study includes 189 consecutive nonsensitized, non-HLA-identical patients who received a primary kidney transplant between March 1999 and March 2006. Protocol testing for DSA via LABScreen single antigen beads (One Lambda) was done before transplantation and at 1, 3, 6, 9, and 12 months after transplantation then annually and when clinically indicated., Results: Of 189 patients, 47 (25%) developed dnDSA within 10 years. The 5-year posttransplantation cumulative incidence was 20%, with the largest proportion of patients developing dnDSA in the first posttransplantation year (11%). Young patients (18-35 years old at transplantation), deceased-donor transplant recipients, pretransplantation HLA (non-DSA)-positive patients, and patients with a DQ mismatch were the most likely to develop dnDSA. From DSA appearance, 9% of patients lost their graft at 1 year. Actual 3-year death-censored post-dnDSA graft loss was 24%., Conclusion: We conclude that 11% of the patients without detectable DSA at transplantation will have detectable DSA at 1 year, and over the next 4 years, the incidence of dnDSA will increase to 20%. After dnDSA development, 24% of the patients will fail within 3 years. Given these findings, future trials are warranted to determine if treatment of dnDSA-positive patients can prevent allograft failure.

Donor specific human leukocyte antigen (HLA) antibodies (DSA) are a significant cause of allograft failure. However, it has been reported that some DSA negative patients still experience allograft failure. In addition, some DSA positive patients maintain good graft function for >20 years. These findings suggest that while DSA is a cause of failure, it is not the sole risk factor for graft dysfunction and that the presence of DSA alone may not predict the time course of graft failure. Here, we report the predictive value of a proprietary panel of four biomarkers in long-term renal allograft outcome. A total of 310 consecutive patients, who received kidney transplants between 1999 and 2012, were included in this study. Recipient sera was tested for HLA antibodies and biomarkers at 3, 6, 12, 24, and 36 months post-transplant. HLA antibodies were identified using Labscreen single antigen beads. The biomarker combination (BMC) test consisted of a proprietary panel of 4 biomarkers and was performed using Luminex. Sera were defined as positive when any one of the 4 biomarkers became detectable. Sera of normal healthy people were used as negative controls. Graft survival analyses were performed and compared between different patient groups based on the positivity of DSA and BMC. Our results indicate that 57% of DSA negative patients and 54% of DSA positive patients had detectable biomarkers. There was no significant difference in BMC positive patients between the DSA positive and negative groups, which suggests that presence of BMC is not associated with HLA DSA. DSA positive patients had a 10% lower 10-year graft survival rate than patients without DSA, while BMC positive patients had a 25% lower 10-year graft survival rate than patients without detectable BMC. When DSA negative patients were divided into two groups based on the positivity of BMC, BMC positive patients had a 20% lower 10-year graft survival rate compared to BMC negative patients (p<0.05). Similarly, when DSA positive patients were divided into two groups based on the positivity of BMC, BMC positive patients had a 30% lower 10-year graft survival rate compared to BMC negative patients (p<0.01). When both DSA and BMC testing results were considered, DSA and BMC double positive patients had the lowest and double negative patients had the highest graft survival rates. The survival rates for the BMC alone and DSA alone positive groups were in between (p<0.001). Multivariate Cox models confirmed that BMC was an independent risk factor for graft failure, with a higher hazard ratio than DSA (BMC=2.60 versus DSA=1.64). In conclusion, serum BMC is an independent predictor of graft failure. BMC was more significantly associated with graft failure than DSA. In combination with DSA, BMC better predicted graft outcome than DSA or BMC alone.

Background: Experience with tolerance protocols has shown that none is perfect and that each escape from tolerance must be identified early to prevent graft failure. In addition, some test is needed for patients who are weaned off immunosuppression (IS) to forewarn of weaning failure. The usual measures of function--such as serum creatinine levels--are not sensitive enough to detect rejection in a timely manner., Methods: A study was carried out on 72 patients who received living-donor kidney transplants with clonal deletion protocol (total lymphoid irradiation or bortezomib), and followed with reduced doses of maintenance IS. Every month or every 2 months, a test was performed for donor-specific antibodies (DSA) using Luminex mixed and/or single antigen beads., Results: After transplantation, DSA developed in 17% of the patients at 6 months, 41% at 1 year, and 57% at 2 years, with 95% confidence limits of 10%, 28%; 30%, 55%; and 44%, 71%, respectively. Fifty-three percent of patients weaned IS to less than 10 mg prednisone daily experienced DSA within 3 months. Furthermore, prednisone dose (per 2.5 mg) and years after transplantation were inversely associated with DSA production (risk ratio 0.92 [95% confidence limits: 0.85, 0.99], and 0.70 [0.49, 1.00])., Conclusions: DSA monitoring is highly effective for detecting escape from tolerance and reemergence of the immune response in weaned patients. DSA appearance was inversely proportional to the level of maintenance drugs in the weaning process. Measurement of DSA on a monthly basis is adequate for detection of the change in immune reactivity.

Objectives: To determine the prevalence, allele specificity, and intensity of anti-MICA antibodies in long-term renal graft recipients and to investigate their association with impaired renal function., Materials and Methods: Sixty-eight long-term (> 10 y) renal graft recipients were divided into 2 groups: (1) patients with impaired renal function (serum creatinine ≥ 2 mg/dL, n=6); (2) patients with normal renal function (serum creatinine < 176.8 μmol/L, n=62). Anti-MICA antibodies were tested using Luminex single antigen beads assays and the frequency, specificity, and intensity of these antibodies were compared between 2 patient groups., Results: MICA antibodies were detected in 33% of impaired renal function patients and 15% of normal renal function patients (P > .05). Anti-MICA*027 antibodies were found in 11.76% of patients, whereas antibody to MICA*012 was found in 2.94% of patients. Interestingly, among all antibody specificities, MICA*001,*004, *007, *009, *012, and *018 were found more frequently in impaired renal function patients than in normal renal function patients. The peak mean fluorescence intensity levels of MICA antibodies in impaired renal function patients were significantly higher than those in normal renal function patients (P < .05)., Conclusions: Our data suggest that increased prevalence and intensity of anti-MICA antibodies are associated with impaired renal graft function in long-term renal graft recipients and some MICA antibodies might be more important than others in mediating graft rejection.

Background: Evidence of the short-term effect of bortezomib on donor-specific human leukocyte antigen (HLA) antibody (DSA) removal capacity has emerged. However, no published data characterize the durability of DSA response. Here, we report the long-term DSA response results on renal transplant patients treated with bortezomib., Methods: In this single-center study, 26 living-donor renal transplant patients with a positive level of de novo DSA were preemptively treated with bortezomib (1.3 mg/m × 4 doses). A total of 15 patients received bortezomib as part of a combination regimen; 11 received bortezomib alone. Weekly serial measurements of HLA antibody were noted before, during, and after treatment using single-antigen beads., Results: At a median follow-up of 25.8 months posttreatment, allograft function remained good in each of the patients. Following treatment, 96% of the patients achieved at least a partial response. Eighteen patients (69%) experienced a complete response followed by a period of DSA remission. Ten patients had DSA relapse after remission, at a median of 3.8 months. The remaining eight patients are still in remission at 14 months posttreatment (median). Patients with remission enjoyed better allograft functional stability than those who relapsed (P=0.023). After bortezomib therapy, the addition of a calcineurin inhibitor or mycophenolate mofetil was predictive for maintaining a DSA remission (hazard ratio 0.09, 95% confidence interval 0.01-0.76)., Conclusions: Bortezomib therapy consistently provides reduction in DSA and in many a DSA remission may occur. However, sustaining remission is likely necessary to improve allograft stability.

The non-donor-specific anti-HLA-Ia antibodies correlate significantly with lower graft survival in organ transplant patients. Based on our earlier findings that anti-HLA-E murine monoclonal antibodies (MEM-E/02 and 3D12) reacted with different HLA-Ia alleles and the peptides shared by HLA-E and HLA class, Ia alleles inhibited the HLA-Ia reactivity of the anti-HLA-E antibodies in normal non-alloimmunized males, the possibility of that anti-HLA-E IgG may account for the non-donor-specific anti-HLA-Ia antibodies in the allograft recipients was examined by multiplex-Luminex®-immunoassay. About 73% of renal and 53% of liver transplant patients' sera with high level of anti-HLA-E IgG showed reactivity to different non-donor HLA-Ia alleles. About 50% renal and 52% liver allograft recipients' sera with low level of anti-HLA-E IgG had no reactivity to any HLA-Ia alleles; however, the IgG isolated from the same sera with protein-G columns showed the presence of anti-HLA-E IgG with HLA-Ia reactivity. Furthermore, both recombinant HLA-E and the IgG-free serum containing soluble HLA-E (sHLA-E) inhibited HLA-Ia reactivity of anti-HLA-E murine monoclonal IgG significantly. The data suggest that the HLA-Ia reactivity of the anti-HLA-E antibody accounts for the non-donor-specific anti-HLA-Ia antibodies. It is proposed that the sHLA-E heavy chain, shed in circulation after organ transplantation, may expose cryptic epitopes of HLA-E to elicit anti-HLA-E IgG antibodies, which may cross react with HLA-Ia alleles due to the peptide sequences shared between them. This study provides a new explanation for the presence of non-donor-specific antibodies for non-existing HLA-Ia alleles, frequently observed and correlated with survival in organ transplant recipients.

Background: The status of sensitization in kidney transplant recipients in the last 10 years and the trend of induction and maintenance therapy in patients of different panel-reactive antibody (PRA) levels have not been analyzed. The aim of this study was to investigate the current status of pre-transplant sensitization and its association with graft outcome., Methods: A total of 155 570 kidney transplants reported to United Network for Organ Sharing (UNOS) during 2000 - 2009 were included in this study. We investigated the current status of pre-transplant sensitization and its association with graft outcome, and also compared the usage trend of 16 induction agents and 7 maintenance immunosuppressants in patients at different PRA levels. The difference of distributions of categorical variables between groups was investigated using the chi-square test. Unpaired t test or one-way analysis of variance (ANOVA) were used for numerical variables. The survival rates of transplant recipients were estimated using Kaplan-Meier methods and significance was determined by Log-rank test. Two-side P value < 0.05 was considered statistically significant. All statistical analyses were performed using STATA 10 with all available updates as of March 2010 (StataCorp LP, College Station, Texas 77845, USA)., Results: Despite the fact of the decreased percentages of kidney transplant recipients with presensitization history, the mean PRA levels of all kidney recipients has been increasing in the last 7 years, which was possibly due to the introduction of more sensitive antibody testing techniques. The percentage of patients with treated rejection episodes within one year post-transplant were significantly higher in sensitized patients (PRA = 50% - 100%:14.3% and PRA = 1% - 49%:13.9%) than in non-sensitized patients (12.4%). Both 1- and 5-year graft survival rates improved in the last 10 years; this was more significant in high PRA patients. Thymoglobulin was the most commonly used induction agent in last 10 years. Its users increased from 10% to 46% in non-sensitized patients, from 12% to 57% in PRA 1% - 49% patients, and from 19% to 63% in PRA 50% - 100% patients. The users of Campath, intravenous immunoglobulin (IVIG), and Rituximab have been increasing and reached 16%, 20%, and 11% in highly sensitized patients. In the last 5 years, steroid-free patients were 33% - 36%, 30% - 37%, and 10% - 25% for PRA 0, 1% - 49%, and 50% - 100% respectively. Almost 90% of patients were on Prograf at discharge. It seems that Myfortic users have been increasing since 2005 and it may soon replace mycophenolate mofetil (MMF) if long-term follow-up study conforms its safety and efficacy., Conclusions: Application of sensitive antibody testing techniques increased the mean PRA levels of transplant recipients in spite of a decreased percentage of sensitized recipients. Induction and maintenance therapy differed in patients at different PRA levels.

Background: Proteasome inhibition abrogates donor-specific anti-human leukocyte antigen (HLA) antibody (DSA) in patients posttransplant. However, its effects on protective humoral immunity to vaccine antigens remain unknown. Herein, we report on bortezomib's safety regarding protective immunity in patients who have experienced HLA antibody reduction/removal., Methods: Thirteen living donor renal transplant patients were treated with bortezomib one to two cycles (1.3 mg/m² × 4 doses) and plasmapheresis in 2008 to remove HLA antibodies posttransplant. Serial measurements of HLA antibody were conducted weekly before, during, and after treatment by means of single antigen bead on Luminex (One Lambda Inc., Canoga Park, CA). Measles and tetanus toxoid IgGs were measured quantitatively by using ELISA (American Research Products Inc., Belmont, MA)., Results: All patients treated with bortezomib/plasmapheresis resulted in a primary DSA reduction of more than 50%. In 10 of 13 patients, complete DSA removal (to below 1000 mean fluorescent intensity) occurred. At 1 year posttreatment, antibody intensity remains significantly depressed in the group as a whole. Despite the significant effect on antibody production, tetanus toxoid and measles IgG levels remained unchanged and above the level of protection at 1 year posttreatment., Conclusion: These data indicate that proteasome inhibitors plus plasmapheresis results in prolonged reduction of HLA antibodies while leaving protective immunity intact.

Background: Induction agents have been shown to reduce the rate of acute rejection. They have not been clearly shown to improve graft and patient survival., Methods: United Network for Organ Sharing registry data were analyzed to show the status of induction therapy in the United States and to determine the effect of induction therapy on long-term graft and patient survival., Results: Since establishment of the United Network for Organ Sharing renal transplant registry, there have been three distinct eras of induction regimen: (1) the low-induction, old antibody era, 1987 to 1993, when antilymphocyte globulin and muromonab-CD3 were the major agents; (2) a high-induction, transitional era, 1994 to 2002, when basiliximab (1998), daclizumab (1998), and rabbit antithymocyte globulin (rATG; 1999) replaced antilymphocyte globulin and muromonab-CD3, with maintenance agents also used; (3) the high-induction, modern antibody era, 2003 to present, with most patients receiving rATG, basiliximab, daclizumab, or alemtuzumab (2003). Induction recipients had higher graft and patient survival rates than nonrecipients in all categories of organ transplant. The improvement was statistically significant in kidney, liver, and lung transplants, although liver and lung recipients had a lower percentage of patients receiving induction than did kidney patients. Kidney transplant recipients on alemtuzumab with steroids had the lowest risk of graft failure, followed by those on alemtuzumab alone, rATG with steroids, rATG alone, and then basiliximab with steroids. Improvement was not statistically significant with daclizumab (alone or with steroids), basiliximab alone, or steroids alone., Conclusion: Induction immunosuppression improved graft and patient outcome for most organ transplants. Depleting agents (alemtuzumab and rATG)--especially in combination with steroids--seem to be more efficient in preventing renal graft failure than nondepleting agents (basiliximab and daclizumab).

Objective: To determine whether short-term improvement in pancreas graft survival with simultaneous pancreas-kidney (SPK) transplants translated into improved long-term survival, then to examine the implications of that determination., Research Design and Methods: We analyzed data for 14,311 diabetic patients who received a first SPK transplant between October 1987 and November 2007, using Kaplan-Meier analysis for graft survival rates and Cox regression analysis for year-of-transplant effect., Results: Overall, from 1995 to 2004, 5-year pancreas graft survival stayed about the same (70-71%). Limiting analysis to grafts that survived more than 1 year, 5-year survival from 1987 to 2004 ranged from 80 to 84%. With 1987-1989 as reference, the adjusted hazard ratio for graft failure by year of transplant increased to 1.49 (95% CI 0.97-2.30) in 2000-2004., Conclusions: Long-term pancreas graft survival has remained unchanged despite the dramatic decreases in technical failures and early acute rejection rates that have contributed to prolonged SPK graft survival.

Background: The common endpoint in the treatment of antibody-mediated rejection (AMR) is functional reversal (creatinine levels). Reduction of human leukocyte antigen (HLA) antibody strength is not commonly considered as an essential endpoint for AMR resolution. The purpose of this study was to determine whether reduction in HLA antibody intensity in patients with histologic AMR reversal influences long-term renal allograft survival., Methods: Renal allograft recipients were included if he or she had a biopsy diagnosis of AMR (between August 2000 and October 2008) and serial evaluation for HLA antibodies prebiopsy and postbiopsy. Antibody reduction was defined as mean fluorescence intensity decrease more than 50% in highest intensity antibody after AMR therapy and the absence of new antibody formation. Patients were treated with plasmapheresis, thymoglobulin/OKT3, and corticosteroids. Survival analysis was performed using STATA/MP v10 (College Station, TX)., Results: Twenty-eight patients were analyzed. Antibody reduction failed to occur in 22 of 28 cases. Baseline characteristics were similar between groups. Antibody nonresponders had significantly shorter allograft survival time (61.4 months) compared with antibody responders (no failures) (P=0.04, log-rank test)., Conclusions: In conclusion, failure to significantly reduce antibody levels and prevent new formation was strongly predictive of allograft loss. This observation suggests that the therapeutic intervention that reduces antibody production may prolong graft survival in transplantation.

We investigate the status of sensitization in kidney transplant recipients and analyze the trend of induction and maintenance therapy in patients of different PRA levels. Despite the fact of the decreased percentages of kidney transplant recipients with presensitization history, the mean PRA levels of all kidney recipients has been increasing in the last 7 years, which is possibly due to the introduction of more sensitive antibody testing techniques or the tendency for kidney allocation organization and clinicians to give priority to patients with elevated PRA once a compatible donor kidney becomes available. The percentage of patients with treated rejection episodes within one year post-transplant were significantly higher in sensitized patients (PRA = 50-100:14.3%, and PRA = 1-49%: 13.9%) than in non-sensitized patients (12.4%). Both 1- and 5-yr graft survival rates have improved in the last 10 years; this was more significant in high PRA patients. Thymoglobulin was the most commonly used induction agent in last 10 years. Its users increased from 10% to 46% in non-sensitized patients, from 12% to 57% in PRA = 1-49% patients, and from 19% to 63% in PRA = 50-100% patients. The users of Campath, IVIg, and Rituximab have been increasing and reached 16%, 20%, and 11% in highly sensitized patients. In the last 5 years, steroid-free patients were 33-36%, 30-37%, and 10-25% for patients with PRA levels of 0, 1-49, and 50-100 respectively. Almost 90% of patients were on Prograf at discharge. Myfortic users have been increasing since 2005 and it may soon replace MMF if long-term follow-up study confirms its safety and efficacy.

Background: Evidence shows posttransplant antibodies lead to renal allograft failure, but does the time elapsed between transplantation and antibody development impact allograft survival? This is the first study showing importance of when antibodies appear., Methods: Serial sera were collected during 17 years (1991-2008) from two groups of patients, one whose allograft failed due to chronic rejection containing 25 patients (230 sera) and a control group consisting of 25 graft functioning patients (305 sera) who were matched by transplant date to a patient whose graft failed., Results: The median follow-up for failure patients was 7.1+/-4.8 years and 11.8+/-4.4 years for controls. Human leukocyte antigens alloantibodies appeared in 24 of 25 (96%) of the failed patients and 48% of the controls (P<0.0001). Time to antibodies also differed between groups. Fifteen (60%) patients from the failure group developed antibodies by 1 year compared with none in the control group. Hazard ratio of antibodies present in 1-year posttransplant from multivariate analysis for allograft loss was 7.77 (P<0.001).Ten-year renal allograft survival in early antibody developers (<1 year) was 27% vs. 80% in the late antibody developers., Conclusions: Overall, human leukocyte antigens antibody development within 1-year posttransplantation markedly lowers allograft survival compared with later antibody development. Therefore, monitoring early antibodies is useful.

Background: We investigated the effects of intact and beta2m-free heavy chain (HC)-specific human leukocyte antigen (HLA) class I antibodies on long-term graft survival., Methods: HLA class I mixed antigen beads were used to detect intact and beta2m-free HC-specific antibodies, whereas elution buffer-treated beads were used to detect antibodies against beta2m-free HC. Donor-specific antibodies (DSAs) were identified using single-antigen beads. Complement-dependent cytotoxicity assays were performed to determine the cytotoxicity of DSA., Results: Three hundred seventy-nine of 994 of patients (38%) had antibodies against intact HLA and beta2m-free HC. There was no survival rate difference between antibody-positive and -negative groups. When the 379 antibody-positive patients were further tested with beta2m-free HC-coated beads, 179 of them with antibodies only against intact form of antigens had a 4-year graft survival rate of 76%, which is significantly lower than that of 200 patients with antibodies against beta2m-free HC of HLA antigens (88%, P=0.0056). Patients with intact antigen specific DSAs had a significantly lower graft survival rate as compared with those with no DSAs (70% vs. 89%, P=0.0073). More patients with strong donor-specific cytotoxic antibodies lost allografts than those with weak-cytotoxic or noncytotoxic antibodies. However, cytotoxic activity of DSA was not correlated to antibody level., Conclusions: We concluded that intact antigen-specific antibodies, especially DSAs, are predictive of graft failure. DSAs were not always cytotoxic. Strong cytotoxic activity of DSA was associated with a higher rate of graft loss but not correlated to the antibody level. Antibodies against beta2m-free HC negatively interfere with the predictive value of intact antigen-specific antibodies.

Background: Current treatments for autoantibody-mediated diseases (i.e., systemic lupus erythematosus) and alloantibodies (in transplant) are minimally effective. Although they deplete naïve B cells, plasmablasts, and transiently reduce antibody concentrations, they are minimally effective against long-lived, antibody-producing plasma cells. In transplantation, plasma cells produce antibodies directed against human leukocyte antigen (HLA) antigens causing poor allograft survival. We report the first clinical experience with a plasma cell depleting therapy, bortezomib, to abrogate anti-HLA antibodies in transplantation (outside of rejection) in an attempt to improve long-term allograft survival., Methods: Eleven patients with anti-HLA alloantibodies were treated with bortezomib. All patients underwent plasmapheresis to aid in removal of antibodies and to determine the effect of bortezomib. Serial measurements of anti-HLA antibody levels were conducted weekly by single antigen bead on Luminex platform., Results: Bortezomib treatment elicited substantial reduction in both donor-specific antibody (DSA) and non-DSA levels. Antibodies were directed against DSA in 8 of 11 cases. Mean time to antibody appearance was 2 months posttransplant. Within 22 days (median) from treatment initiation, 9 of 11 patients' antibody levels dropped to less than 1000 mean fluorescence intensity. Of two patients without successful depletion, all had peak mean fluorescence intensity more than 10,000. At a mean follow-up of approximately 4 months posttreatment, all patients have stable graft function. Minimal transient side effects were noticed with bortezomib in the form of gastrointestinal toxicity, thrombocytopenia, and paresthesias., Conclusions: Bortezomib therapy effectively abrogates anti-HLA antibodies. Hence, removal of antibodies, by proteasome inhibition, represents a new treatment strategy for transplantation and may have benefit in autoimmune-related disease.

Background: Although the incidence of early acute rejection could have been diminished in the past, the long-term renal allograft survival could not benefit from the introduction of more effective immunosuppressive regimens mainly aiming at cellular rejection mechanisms. The cause of chronic rejection is still discussed controversially. Here, we demonstrate to what extent human leukocyte antigen (HLA) antibodies (HLAab) posttransplant contribute to late graft outcome., Methods: A total of 1014 deceased kidney transplant recipients transplanted at the Charité hospital were monitored in a cross-sectional manner for the development of HLAab using Luminex Single Antigen beads. Patients with stable kidney function at a median of 5-years posttransplant were tested once for HLAab and monitored for 5.5 years after testing., Results: Thirty percent of recipients showed HLAab. Donor-specific antibodies (DSA) were found in 31% of antibody positive patients. The presence of DSA was associated with a significantly lower graft survival of 49% vs. 83% in the HLAab negative group (P< or =0.0001). Non-DSAs also had an adverse effect on graft survival (70% vs. 83%; P=0.0001). In a prospective analysis of 195 patients with repeatedly no detectable HLAab, the survival probability was 94% as opposed to 79% survival among patients who developed HLAab de novo after the first testing (P=0.05)., Conclusions: We confirmed that HLAab produced even late after transplantation are detrimental to graft outcome. DSA were proven to have a strong adverse impact on graft survival. The results indicate that a posttransplant HLAab monitoring routine could be appropriate to improve long-term results.

The purpose of this study was to determine the effect of early corticosteroid cessation on the occurrence of de novo human leukocyte antigen (HLA) antibody posttransplant. Renal transplant recipients (n=37) were randomized to early corticosteroid withdrawal at day 7 posttransplant (n=21 patients), or to chronic steroids (n=16), all in combination with thymoglobulin as induction agent, tacrolimus and mycophenolic acid as maintenance therapy. To establish the time course of HLA antibody appearance, sera collected pretransplant and for up to 5 years posttransplant were screened for the appearance of HLA antibodies. In this 5-year longitudinal study, only one patient in the control group developed a de novo donor-specific HLA antibody. We conclude that renal transplant recipients on steroid withdrawal by the end of week 1 are not at higher risk for developing HLA antibodies compared with a standard steroid regimen up to 5 years posttransplant.

Many transplant protocols now use high-intensity induction immunosuppression immediately post-transplant, followed by weaning from immunosuppressives. This chapter reports on the risks of post-transplant immunosuppression reduction/removal revealed by a close monitoring of HLA antibodies in 14 patients who recently received living-donor kidney grafts under clonal deletion at the IKDRC-ITS in Ahmedabad, India. While all 14 patients still have good function at a median follow-up of 13.5 months (7.3-20.1), only two had no dysfunction or antibody appearance. Antibodies appeared in 10 of the other 12, and serum creatinine rose in six of the 12. In light of the high rate of immune activation following immunosuppression reduction/withdrawal, it is clear that close monitoring is required to ensure safety in these patients. The tests that are most effective as early warning of immune activation in these kidney recipients were elevation of serum creatinine and antibodies monitoring, the latter being, perhaps, the more important for reasons discussed in this chapter.

Background: Transplant patients often produce human leukocyte antigen (HLA) antibodies against their donors and produce more specificities than can be accounted for by HLA antigen mismatches. We theorize that the presence of extra, otherwise unexplainable specificities could be accounted for if antibodies reacted to more than one epitope (primary and mimetic) on distinct HLA molecules. The theory states that mimetic epitopes consist of the same three amino acids that comprise the primary, sterically placed approximately the same distance apart as are the corresponding amino acids of the primary., Methods: A mimetic epitope table containing all primary epitopes and corresponding mimetic epitopes was built. Then, the HLA specificities of monoclonal and single patient antibodies were determined. These specificities that could not be defined by unique position or amino acid epitopes alone were then used to query the mimetic epitope table., Results: A single antibody from a transplant patient and three monoclonal antibodies produced reactions that can best be explained as the result of one antibody reacting to the same amino acids at two distinct sites on the molecule. Those position and amino acid combinations (pos/aa) are the primary and mimetic epitopes. Using computerized methods of searching, mimetic epitopes were found in five additional kidney transplant patients who produced nondonor-specific antibodies in addition to donor-specific antibodies., Conclusions: Epitopes on the HLA molecule that mimic the primary epitope have been found. We suggest that these mimetic epitopes explain the additional antibodies often found on HLA immunization resulting from allograft rejections, pregnancies, and transfusions.

Background: Epitopes are the sites to which antibodies bind. Both alpha and beta peptide chains of the human leukocyte antigen-DQ heterodimers (DQA1 and DQB1, respectively) contain polymorphic regions. We can identify DQA1 and DQB1 epitopes by DQ single antigen beads assay of the antibodies, correlating the beads' reaction patterns with either DQA1 or DQB1 alleles., Methods: Sera from 74 transplant patients and 35 mouse DQB1 monoclonal antibodies were tested with DQ single antigen beads for their DQ allelic and serological specificities. Epitopes were defined by amino acids shared by the positive antigens of the antibodies. Unique amino acids were identified as potential epitope sites by comparing the peptide sequences of all human leukocyte antigen class II alleles. For the absorption or elution, patient's serum sample was absorbed by a homozygous B-lymphoblast cell line of specific DQ typing, the eluted antibody then tested with single antigen beads to demonstrate that the antibody reacted to a single epitope shared by multiple DQ antigens., Results: Three DQA1 and 15 DQB1 epitopes were identified. We found that 21 patients produced antibodies against one of the DQA1 epitopes; 27 patients produced antibodies against one of the DQB1 epitopes., Conclusion: The DQA1 and DQB1 epitopes identified here seem to be immunogenic and to elicit DQ antibodies. For the DQB1 epitopes, multiple DQ serological specificities that were detected in the serum of a transplant patient could be explained as a single donor-specific DQ antibody reacting to a mismatched DQ epitope of the donor. Ten examples are shown here.

1. Immunological responses cause antibodies to be produced, and transplant patients often produce HLA antibodies against their donors' HLA. But in many cases, more antibodies are produced than just those against mismatched HLA. Many patients with zero mismatched transplants also produce antibodies, and some of these grafts fail. This study used graft survival analysis that contemplated the primary epitopes against the donors' alleles and the corresponding mimetic epitopes, less those shared with the patients' alleles. 2. A table of primary epitopes and corresponding mimetic epitopes was built in a previous study. For each case studied, here, a subset of the table was selected for all of the donor's HLA alleles. The subset was reduced by eliminating all primary and mimetic epitopes shared by the patient's own HLA alleles. The remaining mimetic and primary epitopes-the maximum pool of epitopes (MaxPEpi)-produced epitope frequency (Efr) for each allele. Using a zero-HLA-AB-mismatch subset (n = 16) of the Greenville and Holland kidney transplant data set (n = 451), the Efr for each allele in the MaxPEpi of each case was tallied and compared with the corresponding observed antibody specificities. The average frequencies were 2,869 and 1,975, respectively, and the t-test was significant at the p-value of less than 0.00001, indicating that the higher the frequency, the more likely that the corresponding allele appears as a part of the specificity. 3. The trends were used to perform survival analysis on zero-mismatch kidney transplant cases in the 2007 UNOS data file. Different variables of MaxPEpi parameters and cutoff points were tried with half of the deceased-donor cases in the file, survival rates for those below and above each cutoff point were compared by the log-rank test. The average Efr with a cutoff of 3,700 yielded the best 10-year graft survival in this subgroup of the 2007 UNOS cases. The results were supported and verified by analyzing the second half of the deceased-donor cases and the living-donor cases in the 2007 UNOS data set. 4. With the 2007 file living-donor cases, 10-year graft survival rate was 86.2% for cases below the cutoff, and 85.4% for those above, and the difference was significant at a p-value less than 0.01. A larger difference was observed for 2007 living-donor cases, excluding identical sibling transplants, but the difference was not statistically significant due to the low number of cases. 5. The same trends were found for 10-year survivals with cases in the 2008 data, but were not statistically significant. With living donors, the rates below and above the cutoff were 85.7% and 85.2%, respectively, and with deceased donors, 72.7% and 72.1%, respectively, p < 0.125. As with the 2007 data, cases in the 2008 file with living donors--excluding identical sibling transplants--had survival rates of 91.7% and 79.0%, respectively, but the 12.7% difference was not statistically significant because of the low number of cases.

This is a long-term retrospective case-control study. Serial sera were collected over 17 years (1991-2008) from two groups comprised of 29 patients with allograft failure (250 sera) and 25 controls with functioning grafts (305 sera), each control matched by transplant date to one failure-group patient, and all patients tested with single antigen beads. The median follow-up for failure-group patients was 7.3 +/- 4.7 years and 11.8 +/- 4.4 years for controls. HLA alloantibodies appeared in 28 of the 29 failure-group patients (97%) and in 12 of the 25 controls (48%) (p < 0.0001). DSA and non-DSA that appeared alone--without any DSA detected-were both associated with graft failure (p = 0.001, p = 0.01). DSA against HLA-DQ antigen was found in 13 of 17 graft-failed patients who had received DQ-incompatible transplants (76%) compared with only one of 11 similarly DQ-mismatched control patients (9%) (p < 0.001). The strength of strong DSA (defined as MFI > 5000) was higher in graft-failed patients than in graft-functioning patients. The time it took for antibodies to develop also differed between groups. HLA antibodies were formed sooner in the failure group compared with the controls (1.7 versus 3.7 years, P < 0.01). Fifteen of the failure group patients developed antibodies within one year while none in the control group did. In conclusion, our study reinforces the observation that circulating de novo HLA alloantibodies predict adverse long-term kidney allograft outcomes. The significant negative impact of all alloantibodies calls for clinicians to monitor patients and implement removal therapy when alloantibody is first detected. This may prove a key step in the ongoing attempt to prevent chronic rejection and prolonging renal allograft survival.

It is apparent from calculations that for paired kidney donation programs, a national program will provide optimum benefit. To obviate major problems associated with donors traveling long distances, we propose shipping donor kidneys. Evidence is provided from the United Network for Organ Sharing (UNOS) Kidney Transplant Registry, that 14,873 immediate functioning kidneys from deceased head-trauma donors with an average cold ischemia time of 18.3 h had 85.7% three-yr graft survival compared with 87.8% survival of 23 369 transplants from living donors with 2.4 h of ischemia. Grafts from 10,368 deceased donors with 13-24 h cold ischemia time had three-yr graft survival of 82.6% compared with 84.2% for 1153 transplants with up to six h cold ischemia time. After adjusting for major confounding factors, cold ischemia does not significantly influence graft survival. We conclude that shipment of donor kidneys can be performed safely and will significantly increase paired donor transplants.

Background: The goals of this study are first to determine the epitope specificity of donor specific antibody (DSA) in the serum of alloimmunized transplant patients with a failed renal graft; and second to understand the correlation between the development of DSA and nondonor specific antibody (NDSA)., Methods: The sera of 35 pretransplant panel reactive antibody (PRA)-negative patients with failed allografts were examined with single-antigen (SA) luminex beads to identify human leukocyte antigen (HLA)-A and -B antibodies. Potential HLA antibody epitopes were identified by using computer software and verified by absorption and elution from single-antigen cell lines., Results: Twenty-seven patients developed donor-specific HLA-A and/or -B antibodies, while the remaining eight patients had only nondonor-specific HLA-A and/or -B antibodies. The DSA-positive patients also had a long list of NDSA. Sixty-eight percent of the reactions found in 27 recipients with DSA were attributable to 66 epitopes on the mismatched donor HLA molecule. All 39 NDSA in eight patients with only NDSA shared 17 epitopes within positive allele specificities. By absorption and elution using recombinant cell lines having a single HLA specificity, we confirmed the epitopes involved in three patients., Conclusion: Development of most NDSA in patients with failed allografts is likely due to sharing epitopes with DSA and/or other NDSA.

Background: Although patients positive for HIV were once thought to be unsuitable candidates for kidney transplantation, their increasing numbers with end-stage renal disease (ESRD) and the introduction of highly active antiretroviral therapy has indicated that they should no longer be excluded for transplantation. To counteract suggestions that human immunodeficiency virus (HIV) patients received suboptimal kidneys, we provide studies of kidneys transplanted from the same donor into patients with and without HIV., Methods: United Network for Organ Sharing kidney transplant data between 1997 and 2004 were analyzed. Graft and patient survival of 38 HIV patients who had received a renal transplant were compared with the survival of 38 recipients who had received a graft from the same donor., Results: The 38 HIV-positive recipients were younger (49.0 vs. 52.3 years, P=0.14) and had lower peak panel-reactive antibodies (PRA; 5.1% vs. 15.6%, P=0.07) when compared with their bilateral donor to HIV-negative recipients. Sirolimus was used more frequently in HIV patients than in non-HIV patients (36.8% vs. 23.7%, P=0.09). The serum creatinine at 1, 3, and 5 years posttransplantation were higher in HIV patients when compared to non-HIV patients. Although not statistically significant, graft survival was higher among HIV-positive patients compared with their negative controls (76.1% vs. 65.1% at 5 years, P=0.21), as was patient survival (91.3% vs. 87.3% at 5 years, P=0.72). More grafts failed due to death with a functioning graft than rejection in HIV-positive patients., Conclusion: This study supports the position that there is no longer an ethical question surrounding the use of kidneys for HIV-positive patients.

Antibodies to MICA and MICB antigens were sought in the sera of 139 kidney transplant recipients. MICA*001, *002, *007, *008, and MICB*002 antigens were produced in Escherichia coli and then tested using enzyme-linked immunosorbent assay plates. Among 35 normal sera, 6% had MIC antibodies, and among 14 sera from pregnant women, 21% had MIC antibodies. Among 34 patients with functioning transplants with human leukocyte antigen (HLA) antibodies, 24% had MIC antibodies, and 19% of 32 patients without HLA antibodies had MIC antibodies. Among 46 patients who lost grafts with HLA antibodies, 26% had MIC antibodies, and among 27 failed patients without HLA antibodies, 37% had MIC antibodies. We conclude that antibodies to MIC are produced in the course of immunization by pregnancies and kidney transplants. They also occurred more frequently in rejected patients (30%) than in those with functioning grafts (21%).

1. From the analysis of 266 Greenville kidney recipients, we found that almost every patient who had graft failure had HLA antibodies (93%), while less than half of patients with currently functioning graft had antibodies (46%). 2. The difference between failed grafts and successful grafts was even greater for de novo antibodies (60% vs. 14%) and greatest for donor-specific antibodies (75% vs. 9%). 3. The incidence of HLA-DQ antibodies was surprisingly high, and the majority was donor-specific. Now that the improved detection beads are available, the effect of DQ antibodies on transplants should be further studied. 4. MICA antibodies were found in 12% of total 266 patients, and found to be more frequent (21%) in patients with graft failure than in patients with successful graft (7%). Almost all patients with MICA antibodies also had HLA antibodies. 5. From the sequential sera testing, we were able to see that, in most cases, antibodies are produced long before the failure and before the elevation of serum creatinine. 6. Periodic testing of sera by single antigen beads enable us to distinguish de novo antibodies from preformed antibodies and to determine whether they are donor-specific. This is important since de novo DSA are most detrimental to graft.

390 serum samples taken from a series of 28 patients who lost their grafts were retrospectively investigated for antibodies by single antigen HLA Class I&II and MICA Luminex beads. In 20 patients with immunological failure, 10 patients had DSA, 16 had epitope-related NDSA, and 16 had NDSA or MICA antibodies. In 16 which increased antibodies leading up to the graft failure, DSA were found in 7 patients. However, in 9 patients who similarly rejected their grafts, 6 were epitope-related DSA and 3 were NDSA. We were unable to establish why NDSA might be associated with the failure, but this was clearly the case in these patients. Of the 25 patients whose grafts were rejected, 23 (92%) produced antibody prior to failure. Our results strongly suggest that antibody monitoring is important as a means of detecting chronic rejection before a rise in SCr. Early detection of antibody would allow for appropriate and timely interventions to save a graft.

Out of 30 patients who rejected their kidney grafts, HLA or MICA antibodies were found in 26 patients. Among the four patients in whom antibodies could not be found, two patients did not have any HLA mismatch with their mother donor, and 1 had died with a functioning graft. There was only one other patient who chronically rejected but did not have antibodies. The antibodies were shown to appear before rejection of the grafts, and before the increase in serum creatinine. In most instances, the de novo antibodies increased in strength with time. Donor specific antibodies were found in 12 patients, two had non-donor-specific antibodies. In the absence of other antibodies, six had DQ antibodies and six had MICA antibodies, and one had DP antibodies.

Background: The ABO incompatible kidney transplants have been performed successfully from large numbers of living donors and some blood type A2 deceased donors. However, there are few reports to support the feasibility of ABO incompatible transplants from non-A2 deceased donors. This problem was examined in the United Network for Organ Sharing (UNOS) data file., Patients and Methods: The UNOS Registry data of kidney transplants performed between 1995 and 2003 from 256 centers was utilized for Kaplan-Meier curves and log-rank tests to compare graft and functional graft survival rates., Results: Deceased donor transplants from all ABO incompatible donors had the same graft survival rates, as that from ABO compatible donors regardless of whether the blood group incompatibility was A (A1), A2 or B. Graft survival from 201 ABO incompatible donors was 66.9% at 5 yr, compared with 66.7% for ABO compatible donors (p = 0.83). Non-A2 incompatible donors also yielded comparable survival rates to ABO compatible donors. From living donors, ABO incompatible donors yielded significantly lower graft survival rates than the ABO compatible group, although long-term graft survival rates of those who survived >1 yr did not differ significantly., Conclusion: Except higher initial graft loss possibility because of insufficient removal of antibodies, non-A2 kidneys yielded equivalent graft survival rates to ABO compatible transplants. In addition to A2 incompatibilities, Blood group A1 and B incompatibilities can also be considered in ABO incompatible transplants.

After the analysis of the UNOS Renal Transplant Registry database, with more than 138,000 cases, we showed that: 1. The important improvement seen in the last 20 years is basically only in the short-term graft survivals, with the best advances in the first 6 months after transplant. Unfortunately, the graft loss rate after the first year post-transplant remains the same as observed 10 years ago. The development of new immunosuppression has not been sufficient in avoiding chronic graft rejection. 2. Ten year graft survival has remained essentially the same in the period after 1996 as the reference period of 1987 to 1995. The year at which the transplant had been performed had absolutely no effect in the past 5. As to trends, there was an important shift to older recipients and older donors which has been notable in the 30 years. Living unrelated and spouse donors have increased 10 years on the long-term survival of HLA identical sibling donor, parental and cadaver donor grafts. 3. The factors which had a small effect before, and continued to have a similar small effect were: regrafts, high PRA, and sex of recipient. 4. The factors which had a large effect before and continued to influence long-term survival were: living versus cadaver donor, age of the recipient, age of the donor, original disease of the recipient, race of the recipient, immediate kidney function and delayed graft function. HLA matching had approximately a 15% difference in best and worst mismatches in the earlier and 10% in the later periods. 6. Cold ischemia time has relatively little effect on graft survival or percent of nonfunctional kidneys in all groups.

1. The prevalence of HLAabs was 25% in the entire population, with a similar distribution for class I and class II. 2. For kidney transplant recipients the prevalence of HLAabs was 25.7 % and the prevalence of de novo HLAabs was 19.8 % in patients without preformed antibodies. 3. Patients with less than 5 years from transplant to blood sampling had lesser HLAabs. The WS were on 2002 and 2004, and then those patients were transplanted after 1997 when TAC was routinely used in some centers. 4. In patients transplanted after 1992, the prevalence of HLAabs was lower and it is consistent with the incorporation of the new IS drugs (TAC was introduced around 1994). 5. For kidney transplantation, the use of cyclosporine plus rapamune had the strongest protector effect for the presence of HLAabs, followed by tacrolimus and cyclosporine group. 6. This protective effect is diminished in patients with female gender, history of previous transplant, and pre-transplant positive PRA. 7. For kidney transplantation, the presence of HLAabs was associated with a higher rate of graft loss (P < 0.0001, HR = 2.26, 95% CI 2.11 - 3.36) but not with patient survival. 8. Patients with liver transplantation showed the lowest prevalence of HLAabs with only 9.6 % and similar distribution for both classes. The presence of HLAabs was not associated with patient or graft survival in Liver Transplantation. 9. For heart and lung transplantation, a posttransplant positive test for HLAabs was associated with worse graft survival, similar to kidney transplantation, with a HR = 3.05 and 3.87 respectively.

A total of 1,043 serum samples of kidney allograft recipients with functioning grafts at a mean of 6 years after transplantation were tested once for HLA abs and monitored for graft and patient survival for 4 years after testing. All grafts were transplanted between 1984 and 2004 at the Charité hospital (Berlin, Germany). 1. To our knowledge this is the first study with more than 1,000 patients designed to elucidate the impact of DSA and NDSA on late renal graft outcome. True donor specificity of HLA abs in 30% of antibody positive patients was determined by using a single HLA antigen coated bead assay. 2. The long-term graft survival was adversely impacted by the presence of HLA abs directed against mismatched HLA antigens and to a lesser extent also by NDSA. This difference was shown to be leveled at late stages of graft damage, suggesting NDSA as a predictor of adsorbed DSA killing the graft. 3. The deleterious effect of preexisting and de novo formed HLA abs on long-term graft function was shown to be equal. Despite a negative crossmatch at the time of transplantation sensitized recipients and those who produced HLA abs de novo posttransplant showed the same increased risk of late graft failure. 4. Among patients with DSA and additional NDSA the majority of 74% of assigned potential HLA class I epitopes were shared among DSA and NDSA suggesting mismatched donor antigens as the cause of detected NDSA.

This study applied the single antigen microsphere technology to the retrospective analysis of sequential post-transplant serum samples in the context of the patient's clinical course. Detailed information on nine of the study patients was presented as representative of the larger cohort and illustrative of different patterns of anti-HLA antibody development and different clinical scenarios that culminated in graft failure. Our major observations are summarized as follows: 1. These data confirm the high sensitivity of the single antigen bead method: In some patients, DSA and NDSA that were undetected by standard methods were found pre-transplant and in sequential post-transplant samples. 2. The precise role that anti-HLA antibody plays in a particular rejection are complicated in cases in which humoral rejection is not diagnosed in the biopsy: The possible involvement of ADCC and mechanisms involving an indirect role for antibody in the rejection process should be carefully investigated. 3. Although anti-HLA antibodies are associated with graft rejection, the time interval between detection and rejection can vary dramatically between patients. Both DSA and NDSA can be adsorbed by the graft and erratically detected in the circulation, in some cases remaining undetected until nephrectomy. 4. Anti-HLA antibody strengths often fluctuate widely over a patient's clinical course, with de novo DSA generally of greater strength than de novo NDSA. 5. In addition to DSA, we have observed the consistent induction of diverse, cross-reactive NDSA. This occurs not only during the post-transplant course but also after graft failure, when immunosuppression is tapered prior to nephrectomy. Our data support further studies to evaluate the value of prospective monitoring of anti-HLA antibodies to better understand the place of anti-HLA antibodies in acute rejection. This may improve our ability to reverse some acute rejection episodes. Since acute rejection has been considered a predictor of late graft loss via chronic allograft nephropathy, understanding and modifying the antibody response is critical to extending the longevity of transplanted organs. Finally, since the strong sensitization to NDSA will seriously hamper the ability to identify a compatible donor for a future transplant, these data reinforce the importance of minimizing HLA mismatches between the donor and the recipient.

With the development of single DP-antigen beads, antibodies to DP could clearly be segregated from other HLA antibodies. We studied 323 sera of patients from four different centers with functioning or rejected kidney grafts. DP antibodies were found in 5.1% of 138 patients with functioning grafts, and 19.5 % of 185 patients with rejected grafts (P<0.001)). 42.9% and 63.9% of the DP antibody positive patients had DR/DQ antibodies among the functioning and rejected group. Among patients who did not have Class I and DR/DQ antibodies, 13% of those who rejected a graft had DP antibodies, compared to 3.5% of patients with functioning grafts (P<0.05). DP 0301 had the highest specificity frequency in the rejected group. In conclusion, HLA-DP antibodies were detected at a higher frequency in patients who have rejected their grafts than those with functioning grafts. For regrafts, DP tissue typing is recommended to avoid presensitized DP alleles.

In this large collaborative study, 2,231 transplanted patients with functioning kidneys were tested for HLA antibodies, then examined 2 years later for graft survival. Among 478 patients with antibodies, 15.1% failed in 2 years, compared to 6.8% failure in 1,753 patients without antibodies (P=0.00000002). Cytotoxicity testing correlated better with outcome than flow cytometry or ELISA testing on HLA coated beads, possibly because it detected non-HLA antigens. When the patients were further broken down into those with serum creatinine at the time of testing of 0.5-1.9, 4.4% of antibody patients failed at 2 years, compared to 4.3% of patients without antibodies. This 0.1% difference increased among patients with serum creatinine values of 2.0-2.9 to 17.9%, and among those with 3.0-3.9 to 16.3%. We conclude that HLA antibodies posttransplantation is predictive of subsequent graft failure, and its predictive value can be enhanced among patients with higher serum creatinine values.

The involvement of immunologic and nonimmunologic events in long-term kidney allograft failure is difficult to assess. The development of HLA antibodies after transplantation is the witness of ongoing reactivity against the transplant, and several studies have suggested that the presence of HLA antibodies correlates with poor graft survival. However, they have not discriminated between donor-specific (DS) and non-specific (NDS) antibodies. A total of 1229 recipients of a kidney graft, transplanted between 1972 and 2002, who had over a 5-yr period a prospective annual screening for HLA antibodies with a combination of ELISA, complement-dependent cytotoxicity, and flow cytometry tests were investigated; in 543 of them, the screening was complete from transplantation to the fifth year postgrafting. Correlations were established between the presence and the specificity of the antibodies and clinical parameters. A total of 5.5% of the patients had DS, 11.3% had NDS, and 83% had no HLA antibodies after transplantation. NDS antibodies appeared earlier (1 to 5 yr posttransplantation) than DS antibodies (5 to 10 yr). In multivariate analysis, HLA-DR matching, pretransplantation immunization, and acute rejection were significantly associated with the development of both DS and NDS antibodies and also of DS versus NDS antibodies. The presence of either DS or NDS antibodies significantly correlated with lower graft survival, poor transplant function, and proteinuria. Screening of HLA antibodies posttransplantation could be a good tool for the follow-up of patients who receive a kidney transplant and allow immunosuppression to be tailored.