Preformed DSA Research Articles

Single fixed low-dose rituximab as induction therapy suppresses de novo donor-specific anti-HLA antibody production in ABO compatible living kidney transplant recipients.

This study was conducted to evaluate de novo donor-specific anti-human leukocyte antigen (HLA) antibody (dnDSA) production leading to antibody-mediated rejection (ABMR) after rituximab induction in non-sensitized ABO-compatible living kidney transplantation (ABO-CLKTx). During 2008–2015, 318 ABO-CLKTx were performed at the Department of Surgery III at Tokyo Women’s Medical University Hospital. To reduce confounding factors, we adopted a propensity score analysis, which was applied with adjustment for age, gender, duration of pretransplant dialysis, HLA mismatch count, preformed DSA, non-insulin-dependent diabetes mellitus, immunosuppressive treatment, and estimated glomerular filtration rate (eGFR) on postoperative day 7. Using a propensity score matching model (1:1, 115 pairs), we analyzed the long-term outcomes of 230 ABO-CLKTx recipients retrospectively. Recipients were classified into a rituximab-treated (RTX-KTx, N = 115) group and a control group not treated with rituximab (C-KTx, N = 115). During five years, adverse events, survival rates for grafts and patients, and incidence of biopsy-proven acute rejection (BPAR) and dnDSA production for the two groups were monitored and compared. All recipients in the RTX-KTx group received rituximab induction on preoperative day 4 at a single fixed low dose of 100 mg; the CD19+ B cells were eliminated completely before surgery. Of those recipients, 13 (11.3%) developed BPAR; 1 (0.8%) experienced graft loss. By contrast, of C-KTx group recipients, 25 (21.7%) developed BPAR; 3 (2.6%) experienced graft loss. The RTX-KTx group exhibited a significantly lower incidence of BPAR (P = .041) and dnDSA production (13.9% in the RTX-KTx group vs. 26.9% in the C-RTx group, P = .005). Furthermore, lower incidence of CMV infection was detected in the RTX-KTx group than in the C-KTx group (13.9% in the RTX-KTx group vs. 27.0% in the C-KTx group, P = .014). No significant difference was found between groups for several other factors: renal function (P = .384), graft and patient survival (P = .458 and P = .119, respectively), and the respective incidences of BK virus infection (P = .722) and leukopenia (P = .207). During five-year follow-up, single fixed low-dose rituximab therapy is sufficient for ensuring safety, reducing rejection, and suppressing dnDSA production for immunological low-risk non-sensitized ABO-CLKTx.

P4.46: PEDIATRIC INTESTINAL RETRANSPLANTATION: OUTCOMES IN A REFERAL CENTER

Purpose: Intestinal retransplantation is required in more than 10% of long term survivors, especially in those who underwent isolated intestinal transplantation. However, no clear guidelines are available regarding indications, type of graft and immunological management. Our aim is to report our experience after retransplantation in children. Material and methods: A retrospective study of pediatric intestinal retransplantation performed in our center in the last 15 years (2003–2018) was conducted. Epidemiological and anthropometric data, number of grafts received by each patient and the type of these with or without liver inclusion, causes of graft loss and recorded DSA data were analyzed. Results: A total of 18 patients were retransplanted (male predominance, mean age of 5.5yrs), 12 of them twice and 6 three times, totaling 42 grafts implanted. The liver was included in the graft in 22 (4 liver-intestinal -CLSB- and 18 multivisceral -MV-). Overall survival after 5yrs was similar in children undergoing transplantation compared to our general series (63 vs 73%). Regarding the type of graft, survival after 1 and 5yrs for isolated bowel and liver-including grafts was 20% and 0% vs. 68 and 61% respectively (p<0.05). Preformed DSA were present in 6 cases (14%), 3 of them lost their second grafts. And 2 patients (5%) developed de novo DSA and lost their non-liver included grafts (SB). Conclusions: Retransplantation in children had long term outcomes similar to the main series. Liver-including grafts showed much better survival. Preformed or the novo DSA had negative impact on graft survival that was attenuated by the presence of the liver. Further studies are warranted to provide better understanding of the role DSA on these patients.

Risk Factors for Cellular and Antibody-Mediated Rejections in the First-Year Post-Transplant: A Population-Based Study

Purpose To analyze the differences between risk factors for acute cellular (ACR) and antibody-mediated rejections (AMR) in a large cohort of highly phenotyped heart-transplant recipients. Methods We performed a multicenter, retrospective, observational study (Pitie-Salpetriere, HEGP, 2004-2016, n=1250 patients). We included all patients with at least one endomyocardial biopsy (EMB): n=1053 patients, 13677 EMB Results All patients received induction therapy (ATG = 92%). Combined transplantations were performed in 53 patients and 324 were transplanted with pre-formed DSA (31.1%). ACR were diagnosed in 458 patients (43.5%, 896 EMB) and AMR in 82 patients (7.8%, 138 EMB, figure). Risk factors for ACR included recipient's age (HR=0.990 per 1-year increment, p=0.005), type of transplantation (combined heart-kidney and heart-liver compared to isolated heart: HR = 0.243 and 0.161 respectively, p Conclusion We found 6 factors independently associated with the risk of ACR or AMR. Two factors were common between ACR and AMR (recipient's age, pre-formed DSA). These findings might help clinician to customize post-transplant follow-up.

Identification of Risk Factors for Biopsy-Proven Rejection during the First Year Post Heart Transplantation

Purpose To analyze the incidence and the risk factors for biopsy-proven rejections during the first year following heart transplantation in a large cohort of highly phenotyped heart-transplant recipients. Methods We performed a multicenter, retrospective, observational study (Pitie-Salpetriere, HEGP, 2004-2016, n=1250 patients). We included all patients with at least one endomyocardial biopsy (EMB): n=1053 patients, 13677 EMB Results All patients received induction therapy, mostly with ATG (92%). Cyclosporine was the most prescribed calcineurin inhibitor (92%). Combined transplantations were performed in 53 patients (heart-kidney: n=30; heart-liver: n=23). Almost one-third of patients were transplanted with pre-formed DSA (n=324; 31.1%). Biopsy-proven rejections were diagnosed in 489 patients (46.4%) representing 1,009 EMB (7.3%, mostly low-grade cellular rejections: 1R1B or 1R2: n=724). Risk factors for rejection included recipient's age (HR=0.988 per 1-year increment, p=0.002), type of transplantation (combined heart-kidney and heart-liver compared to isolated heart: HR = 0.336 and 0.204 respectively, p Conclusion Recipient age, type of transplantation (combined vs non-combined), pre-formed DSA, number of HLA mismatches and the type of induction were significantly associated with the risk of allograft rejection during the first-year post heart transplantation. These findings might help clinician to customize immunosuppression regimen and post-transplant follow-up.

P112 Clearance of pre-formed donor specific antibodies after liver transplantation is specific for DSA and cross-reactive antigens

Aim Liver transplant recipients with pre-formed HLA DSA are reported to “clear” antibodies post-transplant. We sought to determine if the reduction in HLA antibodies was specific for donor antigens or represented a global decline in alloantibodies. Methods We retrospectively analyzed the post-transplant DSA course for 27 liver allograft recipients transplanted with pre-formed HLA DSA, measured by single antigen assay. Results In 16 patients, DSA was persistently seen at last follow-up (although weaker than pre-transplant in all patients); another 2 patients also developed de novo DSA to other donor HLA. In the remaining 9 patients, DSA declined dramatically post-transplant and was not detected at last follow-up. We asked whether third party antibodies also declined (i.e. global immunosuppression) or whether the patterns were specific for DSA and cross-reactive antigens. In 6 of 9 patients whose DSA declined, some non-DSA remained strong while the third party antibodies that reduced were clearly in cross-reactive groups with the donor specific antibodies. In one of the remaining patients, all of their antibodies disappeared post-transplant, in another the DSA did not fit into a CREG, and in the last patient DSA was to HLA-C locus. One example is given in the figure below, where strong antibodies persisted to third party A1 CREG; but DSA to A32 and antibodies to other A10/A19 CREG antigens declined post-transplant. In the same patient DSA to DQ6 disappeared during follow-up, as did non-DSA DQ5 antibodies; but antibodies to DQ2 persisted. Conclusions Overall, liver allograft recipients experienced a global reduction in HLA antibodies, but the most profound effect was on DSA and CREG-related antigens. These cases suggest that liver clearance of HLA antibodies is specific for donor reactive antibodies and cross-reactive clones.

Cardiogenic shock due to antibody-mediated rejection is associated with high short-term mortality after heart transplantation

Data are scarce about the prognosis of heart allograft antibody-mediated rejection (AMR) with hemodynamic involvement. To evaluate the prognosis of cardiogenic shock due to antibody mediated rejection. We performed a retrospective, single center, observational study. We included all consecutive patients with (1) definite diagnosis of antibody-mediated (AMR) rejection treated at our center and (2) cardiogenic shock due to AMR. We analyzed survival after AMR and causes of death. Patients alive after AMR were followed to analyze recurrences of AMR, graft function and cardiac allograft vasculopathy. Finally, 17 patients met inclusion criteria. Four patients were transplanted with MFI > 1000 preformed DSA. Median time between heart transplantation and AMR was 21.5 months (10–64 months). AMR was mostly due to high-level class II de novo DSA (76% of patients). Despite aggressive immunosuppressive therapies, in-hospital and 1-year mortality were respectively as high as 76 and 82%. All patients discharged from hospital normalized LVEF during follow-up. However, 2 patients were diagnosed with recurrent subclinical AMR: one died suddenly at home and the other presented rapidly progressive cardiac allograft vasculopathy ( Fig. 1 ). Despite aggressive immunosuppressive therapies, prognosis after cardiogenic shock due to AMR is poor with 76% in-hospital mortality. However, follow-up of patients discharged from hospital revealed normalization of LVEF.

Donor-Recipient Matching Based on Predicted Indirectly Recognizable HLA Epitopes Independently Predicts the Incidence of De Novo Donor-Specific HLA Antibodies Following Renal Transplantation.

De novo donor-specific HLA antibodies (dnDSA) are recognized as a risk factor for premature allograft failure. Determinants of DSA specificity are generated via the indirect allorecognition pathway. Here, we present supportive data for the relevance of predicted indirectly recognizable HLA epitopes (PIRCHE) to predict dnDSA following kidney transplantation. A total of 2787 consecutive kidney transplants performed between 1995 and 2015 without preformed DSA have been analyzed. De novo DSA were detected by single antigen bead assay. HLA epitope mismatches were determined by the HLAMatchmaker and PIRCHE approach and correlated in uni- and multivariate analyses with 10-year allograft survival and incidence of dnDSA. The PIRCHE-II score moderately predicted allograft survival. However, the predictive value of elevated PIRCHE-II scores >9 for the incidence of dnDSA was statistically significant (p < 0.001). In a multivariate Cox regression analysis adjusted for antigen mismatch and HLAMatchmaker epitopes, the PIRCHE-II score could be identified as an independent risk factor for dnDSA. The PIRCHE-II score independently from the antigen mismatch and HLAMatchmaker epitopes could be revealed as being a strong predictor for dnDSA. PIRCHE may help to identify acceptable mismatches with decreased risk of dnDSA and thus improve long-term renal allograft survival.

Influences of Pre-formed Donor-Specific Anti–Human Leukocyte Antigen Antibodies in Living-Donor Renal Transplantation: Results With Graft Immunocomplex Capture Fluorescence Analysis

BackgroundAdvances in immunosuppressants enable organ transplantation for sensitized patients. However, influences of pre-formed donor-specific anti–human leukocyte antigen (HLA) antibodies (DSA) have not been fully understood in renal transplantation (RT). On the other hand, immunocomplex capture fluorescence analysis (ICFA) is a reliable method to detect donor-specific anti-HLA antibodies and HLA antigen complexes. Graft ICFA can detect DSA in an allograft (g-DSA). MethodsTo elucidate the consequences of pre-formed DSA, 198 patients who underwent living-donor RT were enrolled for this study (observation period: 57.8 ± 34.9 months); 187 patients in the DSA− group (excluding ABO-incompatible cases) and 11 patients in the DSA+ group. Before RT, all DSA+ patients had undergone rituximab administration and plasmapheresis. For a graft ICFA, the biopsy specimen (1 × 105 cells) was dissolved, and HLA antigens were captured by anti-HLA beads. Finally, DSA-HLA complexes were detected by means of PE-conjugated anti-human IgG antibodies and analyzed by use of a Luminex system. A ratio (sample/blank beads, mean of fluorescence intensity) was calculated: ≥1.0 was determined as positive g-DSA. ResultsThere were no significant differences in 5-year graft survival (87.9%/100% in the DSA−/DSA+ groups, respectively). In terms of antibody-mediated rejection (AMR), within 1 month after RT, pathologically determined AMR occurred 3.2% and 63.4% in the DSA− and DSA+ groups, respectively (P < .0001). However, interestingly, more than half of them (57.1%) indicated only subclinical AMR, that is, no fluctuation of S-Cr. As representative of 2 cases of subclinical AMR, g-DSA deposition could be confirmed (1.15 ± 0.04) at 1 hour after reperfusion by graft ICFA. Furthermore, g-DSA shifted to 2.20 ± 0.98 at 3 weeks after transplantation, along with a decline in s-DSA mean of fluorescence intensity (1718-506.5). ConclusionsAlthough pathologically determined AMR occurred more frequently in pre-formed DSA+ recipients, it can be argued that a successful de-sensitization protocol inhibits further production of DSA and graft destruction.

(927) - Heart Transplantation in Patients with Low or High Titer Pre-Formed DSA Is Safe When Performed with a Specific Desensitization Program

(927) - Heart Transplantation in Patients with Low or High Titer Pre-Formed DSA Is Safe When Performed with a Specific Desensitization Program

Risk factors and outcomes of delayed graft function in renal transplant recipients receiving a steroid sparing immunosuppression protocol.

AIMTo analyse the risk factors and outcomes of delayed graft function (DGF) in patients receiving a steroid sparing protocol.METHODSFour hundred and twenty-seven recipients of deceased donor kidney transplants were studied of which 135 (31.6%) experienced DGF. All patients received monoclonal antibody induction with a tacrolimus based, steroid sparing immunosuppression protocol.RESULTSFive year patient survival was 87.2% and 94.9% in the DGF and primary graft function (PGF) group respectively, P = 0.047. Allograft survival was 77.9% and 90.2% in the DGF and PGF group respectively, P < 0.001. Overall rejection free survival was no different between the DGF and PGF groups with a 1 and 5 year rejection free survival in the DGF group of 77.7% and 67.8% compared with 81.3% and 75.3% in the PGF group, P = 0.19. Patients with DGF who received IL2 receptor antibody induction were at significantly higher risk of rejection in the early post-transplant period than the group with DGF who received alemtuzumab induction. On multivariate analysis, risk factors for DGF were male recipients, recipients of black ethnicity, circulatory death donation, preformed DSA, increasing cold ischaemic time, older donor age and dialysis vintage.CONCLUSIONAlemtuzumab induction may be of benefit in preventing early rejection episodes associated with DGF. Prospective trials are required to determine optimal immunotherapy protocols for patients at high risk of DGF.

OR 25 Immunotherapy induced rapid reduction of preformed dsa in highly sensitized recipients of visceral transplantation

Background DSA reduction with immunotherapy could increase the possibility of successfully transplanting highly sensitized patients (HSP). We report our center’s experience with visceral transplantation (VTX) in HSP. Methods 49 patients underwent VTX during 2008-2014 at our institution. Of these, 12 were HSP, 1 patient was excluded due to incomplete data. 11 patients with pre-Tx cPRA >80% had 12 VTX. Pre-formed DSA (pDSA) reduction was calculated by the percentage drop in MFI values between pre and post-transplant period. Patients with pDSA received Alemtuzumab, Steroids, and/or Bortezomib/IVIG pretreatment. All patients were maintained on tacrolimus immunosuppression. Three month, one year, and two year rejection related graft failure (RGF), rejection related mortality (RM) and graft survival (GS) were analyzed. Results 11/12 (92%) of HSP were females. 9/12 (75%) of HSP had pDSA. 7/9 (78%) VTXs with pDSAs had strong DSAs (MFI >5000) and 4/9 (44%) of these were also C1q positive (Table 1a). The 12 VTXs were divided into 3 groups. G1 group (n = 3) had no pDSA and had negative cross-match (XM) . Patients in this group had neither RGF, nor RM with the best GS at 100%, 67%, and 67% respectively. G2 group (n = 6) had pDSA, and >70% post-immunotherapy reduction in pDSA in 2 weeks and >80% in 4 weeks. 1/6 (17%) patient in G2 had positive CDCXM with C1q positive DSA. 5/6 (83%) of G2 had positive FlowXM. None of the G2 patients had RGF or RM. GS in this group was 83%, 67%, and 33% respectively. G3 group (n = 3) had pDSA, and Conclusions pDSA has a negative impact on VTX. Rapid reduction of strong pDSAs with immunotherapy can lower the risk of RGF and RM and improve GS in HSP of VTX. C1q positive pDSA associated with positive CDCXM is characterized by slow DSA reduction and is related to inferior results. Download high-res image (490KB) Download full-size image

LBP20: RELEVANCE OF STRONG POSITIVE DONOR SPECIFIC ANTIBODIES AND FLOW CROSSMATCH WHILE C1q TEST NEGATIVE IN RENAL TRANSPLANTATION

Aim In this study we present a case of a kidney recipient with pre-formed high titer anti donor DSA to one allelic mismatch antigen of a sibling donor. Pre-transplant the DSA appeared to be clinically significant based on strong positive crossmatch (XM) testing in both T- and B cell XM. Methods HLA typing is performed by SSO for low/intermediate resolution and SSP for high resolution. DSA is determined by One Lambda BScreen Single Antigen and C1q Screen assay. T and B cell IgG XM is performed by CANTO flow cytometer, where our cut-off for positive XM is determined based on normal human studies. Results Both donor recipient pairs were typed using low resolution class I and high resolution class II HLA typing. Five out of the twelve alleles (−A ∗ 31, B ∗ 51, DRB1 ∗ 11:01, DQA1 ∗ 05:05 and DQB1 ∗ 03:01) were considered the mismatched antigen based on the available data. Flow crossmatch result was Positive for bothT (+209 MCS) & B (+224 MCS) cell IgG flow crossmatch. DSA titer for the allelic mismatched antigen, B ∗ 51 was 10360 MFI. C1q-SA was despite high DSA titer and crossmatch clear negative. Anti- A ∗ 31, DRB1 ∗ 11:01, DQA1 ∗ 05:05 and DQB1 ∗ 03:01 were negative based on our cut off. The Kidney was transplanted and functioned immediately and continues to function well one month after transplantation. No modifications have been made in the standard immunosuppressive protocol used in our hospital (Induction: Basiliximab and Methylprednisolon; Maintenance: Mycophenolate mofetil, Prednisone and Tacrolimus) and the patient is without any signs of AMR. Conclusion The interpretation of DSA, flow crossmatch in this case was more accurate and predictive when result of solid phase antibody testing was considered along with the high resolution HLA typing. However, further studies and more data in C1q are needed allow renal transplantation and clarify the clinical relevance in patients with high DSA titers with positive crossmatch and negative C1q Testing.

OR29: ALEMTUZUMAB, BORTEZOMIB, AND INTRAVENOUS IMMUNOGLOBULIN PRE-TREATMENT REDUCES THE RISK OF ACUTE ANTIBODY MEDIATED REJECTION AFTER INTESTINAL TRANSPLANT IN HIGHLY SENSITIZED PATIENTS

Aim Strong pre-formed HLA DSA (p-DSA) is detrimental to short and long-term visceral allograft survival particularly of the liver-free allograft. This limits the opportunity to obtain an immunologically compatible graft for highly sensitized patients with clinical urgency for intestinal transplant. Here we report our preliminary and short term outcome following successful intestinal transplant under alemtuzumab (ALT), bortezomib (BTZ) and IVIG pretreatment. Methods Four patients with cPRA = 100% underwent intestinal transplantation. All patients had strong (MFI ⩾ 5000) p-DSA (either C1q + or −) and +T/B cell flow cytometer cross-match (FCXM) only or both FCXM and cytotoxicity cross-match (CDCXM). Luminex single antigen and C1q assays were done for DSA detection. All patients received ALT (30 mg, iv single dose), BTZ (1.3 mg/m[2], iv /week for 4 weeks) and IVIG (2 gm/kg, iv, single dose) pretreatment combined with tacrolimus based immunosuppression. Results Two out of 4 patients were transplanted in the presence of single strong p-DSA (-C1q), both with a T/B cell +FCXM and -CDCXM. The third patient was transplanted in the presence of multiple strong class I and II p-DSAs. In this patient, class I antibodies were +C1q and were both T/B cell +FCXM/+CDCXM. The MFI of DSA turned negative ( Fig. 1 A). To date, graft survivals are 8, 21 and 39 weeks respectively. The fourth patient was transplanted with multiple strong p-DSAs, one of which was +C1q. This patient had +FCXM and -CDCXM at the time of transplant. There was no reduction in C1q or DSA during the early post-operative day (POD). A negative C1q antibody turned positive in 1.5 weeks with graft lost ( Fig. 1 B). Biopsy confirmed +C4d. Conclusions Rapid reduction of p-DSAs under ALT, BTZ, and IVIG pretreatment can reduce the risk of antibody mediated rejection after isolated intestine transplant. Close monitoring of reduction in DSA trend during the early POD may help to adjust treatment strategies and improve clinical outcomes.

Early Excellent Outcomes of Flow Cross-Match Negative Kidney Transplant Recipients With Pre-Formed IgG DSA, C1q Negative With or Without Peri-Operative IVIg Infusion.

It is not clear if patients with a negative cross-match but with DSA need additional therapy at the time of transplant to prevent an amnestic reaction that could lead to early AMR. Based on our experience with a high dose IVIg desensitization program, we have conducted a stepwise deletion of peri-operative IVIg according to the DSA MFI. Intervention: All patients received ATG induction and triple therapy maintenance immunosuppression with tac, MMF and steroids. All patients were T and B flow cross-match compatible. The first 10 pts had at least one current DSA with a MFI > 1000. They received IVIg 2G/Kg at the time of Tx and 3 weeks later. The threshold to received IVIg was increased to 1500 MFI for the next 10 pts and subsequently to 2000 MFI (3 pts thus far). All patients were monitored post Tx for DSA with single antigen bead immunoassay for IgG and C1q. We compared the outcomes at 1 year of pts who received the IVIg and those who didn't according to the titration scheme. Results: From 4/08 to 9/13, thirty-two patients were treated according to this protocol. Twenty three patients with DSA MFI > 1000 received IVIg (group1) and 9 patients with MFI < 2000 did not receive IVIg (group2). Patient and graft survival were 100% at 1 year in both groups. Only one graft was lost in group 1, five years post-Tx after 2 years lost to follow-up. There were no episodes of cellular or humoral rejection in either group. Of group 1, 16/23 became DSA negative post Tx at least for 1 year, 6 remained DSA IgG positive and 1 intermittently became DSA IgG and C1q positive. In group 2: 8/9 became DSA negative, one pt remained DSA IgG. The 1-year creatinine was 1.37 ± 0.49 for group1 (n=20) and 1.06 ± 0.33 (n=9) for group 2. In conclusion, recipients of kidney transplant with negative flow cross-match but with IgG DSA up to MFI of 1500 may not need peri-operative IVIg to prevent early AMR. We are currently pursuing our evaluation with increasing MFI cut off. DISCLOSURES:Tyan, D.: Other, One Lambda, licensed product.

Peri-operative third party red blood cell transfusion in renal transplantation and the risk of antibody-mediated rejection and graft loss

Historic red blood cell transfusion (RBCT) may induce anti-HLA antibody which, if donor specific (DSA), is associated with increased antibody-mediated rejection (AMR). Whether post-operative RBCT influences this risk is unknown.We examined the RBCT history in 258 renal transplant recipients stratified according to prevalent recipient HLA antibody (DSA, Non-DSA or No Antibody).AMR occurred more frequently in patients who received RBCT both pre and post transplant compared with all other groups (Pre+Post-RBCT 21%, Pre-RBCT 4%, Post-RBCT 6%, No-RBCT 6%, HR 4.1 p=0.004). In the 63 patients who received Pre+Post-RBCT, 65% (13/20) with DSA developed AMR compared with 0/6 in the Non-DSA group and 2/37 (5%) in the No-Antibody group (HR 13.9 p<0.001). In patients who received No-RBCT, Pre-RBCT or Post-RBCT there was no difference in AMR between patients with DSA, Non-DSA or No-Antibody. Graft loss was independently associated with Pre+Post-RBCT (HR 6.5, p=0.001) AMR (HR 23.9 p<0.001) and Non-AMR (6.0 p=0.003) after adjusting for DSA and delayed graft function.Re-exposure to RBCT at the time of transplant is associated with increased AMR only in patients with preformed DSA, suggesting that RBCT provides additional allostimulation. Patients receiving Pre+Post-RBCT also had an increased risk of graft loss independently of AMR or DSA. Both pre and post procedural RBCT in renal transplantation is associated with modification of immunological risk and warrants additional study.

A novel ELISA-based crossmatch procedure to detect donor-specific anti-HLA antibodies responsible for corneal allograft rejections

Previous studies had shown that donor-specific anti-HLA antibodies may highly influence the survival rate of corneal allografts, although the anterior chamber generally represents an immune‐privileged compartment of the eye. We postulated that the introduction of a novel crossmatch procedure for the detection of donor-specific anti-HLA antibodies in recipients awaiting a corneal graft would be adequate to investigate their influence on the outcome of the graft survival. The Antibody Monitoring System (AMS) HLA class I & II crossmatch ELISA was adapted for the use of material from the outer scleral rim instead of blood lymphocytes to isolate the donors' HLA molecules. In case of detectable donor-specific anti-HLA class I and/or class II antibodies (DSA) this result was confirmed using an identification ELISA to specify the detectable recipient's anti-HLA antibodies. PCR-based genetic tissue typing of the donors was performed also using their outer scleral rims. 45 recipients of corneal grafts were analyzed for DSA prior to or after grafting, respectively. 75% of the recipients with preformed DSA exhibited immunological complications up to the complete graft loss in four cases during the first two months. In contrast 77% of the recipients without DSA did not show any complications during the follow up period of averagely 18months. Only two cases of graft loss were observed in this group after 17 and 23months, respectively. The results demonstrate the impact of preventing donor-specific anti-HLA antibodies which are for the first time reliably detectable in any laboratory's daily work using the adapted AMS-ELISA.

141 Age Greater Than 70 Is Associated with Diminished Post-Lung Transplant Survival

was performed. Results: There was no correlation between pre-transplant DSA and the development of BOS (BOS stage 2/3 vs 0/1) p 0.22, or stages of BOS p 0.48. Similarly, there was no correlation between any of the pretransplant clinical factors and the development of BOS in patients surviving greater than one year in the presence of preformed DSA or non-DSA. Graft ischaemic time, HLA-mismatches, underlying disease and number of infections requiring hospital admission after transplantation did not have significant impact on the presence of BOS. Conclusions: Whilst pre-transplant donor specific HLA antibodies are known to have a deleterious effect on graft outcome in the early posttransplant period, it would appear from this preliminary data that in patients who survive despite having DSA there is no effect on subsequent development of BOS.

Incidence and Prediction of Early Antibody-Mediated Rejection due to Non-Human Leukocyte Antigen-Antibodies

Antibody-mediated rejection (AMR) is responsible for a large proportion of early allograft losses. While preformed donor-specific human leukocyte antigen (HLA)-antibodies (HLA-DSA) are accountable for the majority of these episodes, non-HLA-DSA are also involved. However, data on the incidence of early AMR due to non-HLA-DSA are currently lacking. This study evaluated (i) the incidence of early AMR due to non-HLA-DSA -- defined by exclusion of circulating HLA-DSA detected by flow beads -- and (ii) the association with donor-specific major histocompatibility complex class I chain-related gene (MICA)-antibodies (MICA-DSA) and angiotensin-receptor antibodies. A retrospective cohort (n=279) risk stratified by complement-dependent cytotoxicity crossmatches (CDC-XM era) and a prospective cohort (n=154) risk stratified by virtual crossmatching using flow beads (virtual-XM era) were investigated. In the CDC-XM era 25/279 patients (9%) developed early AMR, but only 3/154 patients (2%) in the virtual-XM era (P=0.004). The incidence of early AMR due to HLA-DSA was significantly higher in the CDC-XM era than in virtual-XM era (18/279 patients [6.5%] vs. 0/154 patients [0%]; P=0.0005). However, the incidence of early AMR presumably due to non-HLA-DSA remained unchanged in these two cohorts (7/279 patients [2.5%] vs. 3/154 patients [2%]; P=1.0) consistent with a persisting gap in the ability to identify preformed DSA. Overall, 10/433 patients (2.3%) experienced early AMR presumably due to non-HLA-DSA. None of these 10 patients had angiotensin-receptor antibodies, at most 3/10 patients had MICA-DSA, while the antibodies remained unexplained in 7/10 cases. Early AMR due to non-HLA-DSA is a rare event, which is still difficult to predict by currently available assays.

Clinical Relevance of Preformed HLA Donor-Specific Antibodies in Kidney Transplantation

This study analyzes the influence of preformed DSA, identified by HLA-specific ELISA assays, on graft survival and evaluates the incidence of antibody-mediated rejection (AMR) in patients with and without pregraft desensitization. Kidney graft survival at 8 years was significantly worse in patients with DSA (n = 43) than in those without DSA (n = 194)(p = 0.03). The incidence of AMR in patients with DSA is 9-fold higher than in patients without DSA (p < 0.001) and their graft survival is significantly worse than in DSA patients without AMR and in non-DSA patients (p = 0.005). The prevalence for AMR in patients with DSA detected on historic serum is 32.3% in nondesensitized patients and 41.7% in desensitized patients. The risk for AMR is significantly more elevated in patients with strongly positive DSA (score 6-8) compared to those with DSA score 4 (p < 0.001), and in patients with historic DSA+/CXM+ compared to those with DSA+/CXM- (p = 0.01). The presence of preformed DSA is strongly associated with graft loss in kidney transplants, related to an increased risk of AMR. Our findings demonstrate the importance of detection and characterization of DSA before transplantation. Stratification of this risk could be used to determine kidney allocation and to devise specific strategies for these patients.