The study explores relation of preformed DSA and incidence of ABMR with its effect on renal transplant outcome. In 5 years duration ,462 transplanted patients were included. DSA assay was through LUMINEX positive while CDC was negative .It was found that:
. 8.7% of DSA positive (20% of all included patients) developed ABMR at 1 year..
. ABMR occurred only with MFI more than 3000
.ABMR rate was correlated with decreased 5 year graft survival.. Without ABMR graft survival incidence was comparable in both positive and negative DSA.
.induction with ATG in DSA positive patients made ABMR incidence similar to negative DSA patients.
Post Tx DSA monitoring is important with ATG induction to considered when indicated
Alyaa Ali
3 years ago
Preformed HLA donor specific antibodies have been shown to increase the risk of antibody mediated rejection and have a deleterious effect on kidney graft survival even in the presence of a negative complement dependent cytotoxicity cross match or flow cytometry cross match
the study showed increased prevalence of AMR at 1 year post-transplant and significantly reduced 5 year death censured graft survival in patient with positive DSA
DSA number and strength correlated significantly with AMR as only DSA MFI over 3000 were associated with higher incidence of AMR
Nadia Ibrahim
3 years ago
In your own words, summarise this article.
Anti HLA Ab includes (DSA, non DSA) and is screened via solid phase immune essay ( luminex) technique
DSA could be positive in patient serum despite negative CDC cross matching and/or flow cytometry crossmatches , and was found to be associated with kidney graft loss and poor impact on the overall graft survival, in relation with AMR .However other DSA positive recipients did not.
What is the clinical relevance of DSA? Is positive DSA superior on negative CDC to predict graft rejection and survival? And why not all DSA positive patients develop AMR?
In order to answer these questions a study was conducted on KTX rciepients with positive anti-HLA IgG antibodies DSA via Luminex technique and negative CDC , HLA antibody detection was done by single antigen bead (SAB) assay, the strength was based on the MFI of one SAB.
Patients with high number of HLA mismatches underwent Induction protocol immunosuppression with anti-IL-2 receptor monoclonal antibody (Basiliximab) and ( ATG) .
All enrolled recipients had similar triple maintenance immunosuppression, consisting of oral tacrolimus (FK-506), mycophenolate mofetil (MMF) and methylprednisolone (MP)/prednisolone.
follow up of the graft outcome was done
results showed that DSA main effect will depend on its specificity and strength [3]. Associated with AMR and poor graft survival within five years. DSA ability to mediate endothelial injury through complement-dependent and independent mechanisms, mainly through acute and/or chronic AMR [2].
Hence it is of crucial value to identify DSA characteristics as HLA class, number , and strength by mean fluorescence intensity (MFI) to help improve immunologicalrisk stratification of sensitized patients and their immunesuppression protocol. [1]
Please reflect on your practice if possible
Pretransplant assessment of recipients with Anti-HLA antibodies screening and DSA detection by solid-phase assays hand to hand with the cell based CMX,helps careful selection of highly sensitized patients and stratification of the proper desentisation protocol to eliminate the expected AMR , and whether the patient would benefit from protocol biopsies.
References:
1] Mohan S, Palanisamy A, Tsapepas D, Tanriover B, Crew RJ, Dube G, et al. Donorspecific antibodies adversely affect kidney allograft outcomes. J Am Soc Nephrol 2012;23(12):2061–71.
(2) Zhang X, Reed EF. Effect of antibodies on endothelium. Am J Transplant 2009;9(11): 2459–65.
(3] Jindra PT, Zhang X, Mulder A, et al. Anti-HLA antibodies can induce endothelial cell survival or proliferation depending on their concentration. Transplantation 2006; 82(1 Suppl.):S33–5.
AMAL Anan
3 years ago
Cell mediated rejection vs Antibody mediated rejection.
AMAL Anan
3 years ago
In waiting list patients we test anti-HLA IgG antibodies every three month by luminex. Positive positive underwent SAB to determine specifity. Cut-off for positive reactions if MFI > 1000.
Typings for HLA-A* , B* , DRB1* done for all deceased donors by PCR before donor selection .
We performed virtual cross match , we consider DSA with high MFI.
CDC carried out and considered positive when cell death exceeded 20%.
Induction therapy done either by basiliximab anti-IL2 receptor monoclonal (anti CD25) 20 mg twice at day 0 and day 4. Or ATG (polyclonal antithymocyte globulin) 3 mg/kg 5-7 day which in sensitised patients or high numbers of HLA mismatch , this induction followed by triple maintenance therapy ( oral tacrolimus , MMF and methyl prednisone) FK started with dose 0.1-0.15 mg/kg/d adjusted to maintain trough level between 8-12 ng/ml during first month , between 7-10 ng/ml during 2-3 month and between 5-8 ng/ml thereafter.
MMF started at dose 2000 mg/d then decreased to 1000-1500mg/d during first non postoperative depend on white cell count . Methylprednisone given at dose of 500 , 250 , 125 mg/d on day 1-2 and days 3-4 after operation respectively.we started oral prednisolone on day 5 by 20 mg then tapered to 5-10 mg/d within 2-3 month after transplant.
Delayed graft function: define to those require dialysis in first week post transplant. GFR estimated using MDRD.
Allograft rejection diagnosed by biopsy proven rejection which evaluated by LM and IF staining for C4d which classified according to Banff classification.
Mild acute cellular rejection (ACR grade 1) treated by pulse steroid therapy 500 mg MP for 3days with increasing maintenance immunosuppressive medication.
Other ACR treated with ATG .Patient with AMR treated by 4 session of plasmapheresis, Every other day, after each one IVIG 100 mg/kg then after last plasmapheresis session , patient will receive high dose of IVIG 2g/kg divided in four daily dose followed by one dose of rituximab 375 mg/m2 and similar dose of IVIG 2g/kg repeated after 1 month.
Close association between AMR in DSA and poor graft survival.
Studies shown only DSA positive with AMR has reduced 5 y graft survival. DSA positive with AMR patients had similar graft survival in comparison to DSA negative patients. Although other studies shown reduced graft survival in patients with DSA positive even without AMR.
DSA positive patients who received ATG induction in complications to basiliximab shown less AMR development.
The use of ATG plus IVIG induction therapy in patients with DSA ( below 5000 MFI) give benefits of successful kidney transplantation with similar rejection rate and graft survival with or without DSA.
References:
Mohan S, Palanisamy A, Tsapepas D, Tanriover B, Crew RJ, Dube G, et al. Donorspecific antibodies adversely affect kidney allograft outcomes. J Am Soc Nephrol 2012;23(12):2061–71.
[2] Tait BD, Susal C, Gebel HM, et al. Consensus guidelines on the testing and clinical
management issues associated with HLA and non-HLA antibodies in transplantation.
Transplantation 2013;95(1):19–47.
[3] Amico P, Honger G, Mayr M, et al. Clinical relevance of pretransplant donor-specific
HLA antibodies detected by single-antigen flow-beads. Transplantation 2009;
87(11):1681–8.
[4] Caro-Oleas JL, Gonzalez-Escribano MF, Gonzalez-Roncero FM, Acevedo-Calado MJ,
Cabello-Chaves V, Gentil-Govantes MÁ, et al. Clinical relevance of HLA donorspecific antibodies detected by single antigen assay in kidney transplantation.
Nephrol Dial Transplant 2012;27(3):1231–8.
[5] Lefaucheur C, Loupy A, Hill GS, et al. Preexisting donor-specific HLA antibodies
predict outcome in kidney transplantation. J Am Soc Nephrol 2010;21(8):1398–406.
[6] Lefaucheur C, Suberbielle-Boissel C, Hill GS, et al. Clinical relevance of preformed
HLA donor-specific antibodies in kidney transplantation. Am J Transplant 2008;
8(2):324–31.
[7] Otten HG, Verhaar MC, Borst HP, Hené RJ, van Zuilen AD. Pretransplant donorspecific HLA class-I and -II antibodies are associated with an increased risk for kidney graft failure. Am J Transplant 2012;12(6):1618–23.
Nasrin Esfandiar
3 years ago
To evaluate correlation of preformed DSA with graft outcome, 462 patients were investigated. Among them 95 patients (20.6%) had positive anti-H LA antibodies and 40 (8.7%) had DSA. AMR was significantly higher in patients with DSA.
DSA with MFI >3000 were associated with AMR and more graft loss even with a negative CDC-XM or FCXM. Majority of induction therapy in sensitized patients without AMR was performed with ATG vs Basiliximab.
Ahmed mehlis
3 years ago
This study CDC-XM negative 462 patient in one TX center in 5 years .
Induction therapy was used in 413 patients (89.4%),
1. Basiliximab ..
2 ATG ..
Then triple maintenance immunosuppressive therapy
●
367 (79.4%) patients had no anti-HLA
antibodies (HLA−) detectable in pretransplant serum.
A positive anti-HLA antibody
screening was present in 95 (20.6%) patients, with 40 of them having antibodies directed
against donor HLA molecules (HLA+ DSA+), while the remaining 55 patients had not(HLA+ DSA−).
●Acute rejection was particularly more common in HLA+ DSA+ patients (37.5%) in comparison with HLA+
DSA− (14.5%) and HLA− patients (10.6%)
Moreover, acute rejection developed earlier in HLA+ DSA+ patients (median
11 days) than in HLA+ DSA− (median 71 days) and HLA− patients (median 35 days).
●The prevalence of AMR increased significantly with the presence of DSA MFI value
over 3000.
Wessam Moustafa
3 years ago
This article showed the relevance of preformed DSAs regarding incidence of AMR and graft survival .
They concluded that there is strong correlation between preformed DSAs and subsequent AMR ,
another interesting conclusion was that patients who had Pre formed DSAs without occurrence of AMR has similar graft survival to Those who don’t have AMR ,
Which means that occurance of the AMR is essential for the adverse outcomes related to DSAs .
They also found that induction using ATG decreases incidence of AMR in patients with Pre formed DSAs .
They studies characteristics of DSAs that included strength and no. And they found strong correlation between them and AMR .
MFI less than 3000 was associated with Lower risk of AMR Than those with more than 5000 MFI .
Limitations of the study include lack of flow cytometry data , no protocol biopsies, limitations regarding SAB assays and MFI values.
Mohammed Sobair
3 years ago
DSA be increase the risk of antibody-mediated rejection and have a deleterious effect
on kidney graft survival, even in the presence of a negative CDC) and/or flow cytometry
cross matches.
HLA antibody detection by single antigen bead assay is much more sensitive than cell
based cross matches.
Identification of DSA characteristics associated with AMR occurrence would have
important clinical implications for highly sensitized patient management
,immunosuppressive strategy optimization and kidney graft recipients.
This study, aimed to analyze the relationship between preformed DSA and kidney graft
outcomes, in the presence of a negative CDC crossmatch.
Anti-HLA antibodies screening and specificity:
Pre-transplant anti-HLA IgG antibodies were tested in patient sera collected every 3
months by multiplex microsphere based on Luminex Xmap® Technology
To determinate the specificity of the HLA antibodies, single-antigen bead (SAB) assays
(LabScreen Single Antigen Bead, were performed in patients with a positive screening.
Donor HLA typing and virtual crossmatch:
Samples of all deceased donors were routinely typed before recipient selection in loci
HLA-A*, B* and DRB1* using PCR) amplification with specific sequence primers (SSP;
Olerup SSP®).
With the information of the donor HLA typing, we performed a virtual crossmatch.
In each individual DSA, the strength was based on the MFI of one SAB. In the case of
several DSA against different HLA-antigens, we usually considered the DSA with the
highest MFI; occasionally, we also used the cumulative strength of all DSA by adding the
individual MFI values, as stated in the results.
CDC crossmatch:
carried out using the standard NIH technique, not enhanced with anti-human globulin.
Induction protocol and maintenance immunosuppression:
Induction therapy was used in the study .with an anti-IL-2 receptor monoclonal antibody
(Basiliximab Novartis®, 20 mg twice at day 0 and 4) or a polyclonal anti-thymocyte
globulin (ATG Fresenius®, 3 mg/kg for 5–7 days).
All enrolled recipients had similar triple maintenance immunosuppression, consisting of
oral tacrolimus, mycophenolate mofetil (MMF) and methylprednisolone
(MP)/prednisolone.
Result of the study cohort of kidney graft recipients, transplanted with a negative CDC
crossmatch in peak and current sera, the presence of preformed DSA was clearly
associated AMR occurrence and prevalence, also show a significantly reduced 5-years
death censured graft survival in patients with DSA.
Mujtaba Zuhair
3 years ago
This article studied graft survival in patients with pretransplant DSA and negative CDC.
and found that :
patients with DSA had higher incidence of AMR.
Patients with DSA and AMR had higher risk of chronic antibody mediated rejection.
Patients with DSA and AMR had lower 5 years graft survival . Patients with DSA without AMR had graft survival similar to patients with no DSA.
patients with DSA who treated with ATG had lower incidence of AMR than patients whom received basiliximab.
DSA with MFI more than 3000 and the sum of all DSA more than 11000 is associated with higher risk of AMR.
Heba Wagdy
3 years ago
Preformed DSA are associated with poor graft outcome and increased risk of antibody mediated rejection
Studies showed various results about the impact of DSA on graft survival depending on level of DSA, different characteristics of DSA, occurrence of AMR or not and whether the DSA is complement dependent or not, while other studies showed that DSA may have protective effect through promoting graft accommodation.
The study analyzes the impact of DSA with negative CDC on graft outcome and showed that:
preformed DSA with negative CDC was associated with high incidence of AMR and significantly decreased 5 year death censored graft survival
patients with DSA who developed AMR had detectable DSA and had more decline in eGFR than those who didn’t develop AMR
patients with DSA who received ATG as induction had lower rate of AMR than those who received Basiliximab
Only DSA with MFI >3000 were associated with increased incidence of AMR
Limitations:
FCXM results were not available and they are important especially in sensitized patients
Protocol biopsies were not performed
limitations of SAB assay and the reported MFI values
anti-HLA DQ was not detected at allele level
Conclusion:
Preformed DSA associated with increased incidence of AMR and poor graft outcome.
Anti-HLA antibodies screening and DSA detection by solid phase assays are valuable tools to determine DSA strength and characterization which may improve immunological risk stratification of sensitized patients
saja Mohammed
3 years ago
SUMMARY: Indroduction :
Performed anti-HLA–AB and its characteristic with impact on organ transplant still need further clarification ,the Anti-HLA AB effect on renal graft depend on its class, intensity and complement binding capacity. Study aim:
To assess the impact of performed anti-HLA AB on renal transplant outcome and to identify the predictive value of DSA characteristics for AMR occurrence by using single antigen bead Luminex assay with positive MFI cutoff value of 1000 and above.
Results:
They included 462 recipients of KTX with negative CDCXM ,no flow cytometry XM duringthe period between20072012 from single center,thepatients demographics and clinical characteristics were homogenous between the 3 groups ,majority received ATG induction IS (55%) ,and all have similler maintenance IS protocol with triple therapy ( tacrolimus MMF , steroids ).
This study clearly shows the presence of preformed anti HLA AB associated with AMR whic occure in 35% of non-sensitized patients ,its similar to the results others studies , with moreAMR in the first year after transplantation with a significantly reduced 5-years death censured graft survival in patients with DSA with p value (0.006), also they found that only DSA MFI over 3000 were associated with higher incidence of AMR, and 55% of the patients use ATG induction protocol with lower rate of AMR
Also,only DSA+ AMR+ patients had a significantly reduced 5-year graft survival (68.8%); DSA+ AMR− patients (96.0%).
persistence of DSA after 1 years associated with increased risk of chronic antibody-mediated lesions which is more frequently in DSA+ AMR+ group with lower estimated GFR at the last visit than DSA+ AMR− group, the DSA -HLA class was not associated with AMR as in others [3,8] although anti-HLA-DQ was not determined at the allelic level in this study. In fact,they found that the DSA number and strength correlated significantly with the risk of AMR.
Conclusion:
The presence of anti HLA, DSA BY numbers and intensity significantly associated higher risk of AMR with negative impact onthe renal graft survival, DSA characterization has crucial importance in the management of sensitized patients.
Limitations:
No data about the FCXM
No protocol biopsies.
The SAB MFI cutoff value interpretaion. No donor typing at anti-HLA-C and/or -DP antibodies not available
The anti-HLA-DQ was not determined at the allelic level in every case.
During my practise we use lumenix SAB assay for screening and monitoring of DSAs with MFI cutoff value of 1000 and above , with CDCXM, currnetly no access to FCXM assay , no protocal biopsy.
Tahani Hadi
3 years ago
The aim of this study is to present the effect of DSA on transplanted kidney in patients with different figures of presensetization or not and effect of CDC results and flow cytometry XM whether negative or positive. female with history of pregnancy, history of blood transfusion and previous transplantation all those groups considered as sensitized patients.
The study followed 462 patients underwent kidney transplantation with their immunological tests mainly by luminex test,and single Ag beads assay to evaluate anti HLA-Ab with MFI more than 1000.
Generally most studies showed that there is direct relationship between presence of DSA and incidence of AMR and graft survival but this depends on the multiple factors like HLA class,it’s range and amount of MFI .
The patients that are screened in this study are 462 patients with negative pre transplant B and T cells CDC crossmatch and the patients had been divided into groups according to presence or absence of HLA and DSA with different levels of MFI of more than 1000 and more than 3000.
The results of the study showed the rate of acute rejection is higher in those group with positive of both DSA and HLA and even earlier rejection posttransplant comparing with other group more specifically AMR which is correlated to percentage of DSA and MFI and for most of those patients ATG are used in the induction therapy ,regarding graft survival with prolonged follow up showed that the patients with DSA positive and history of pos6AMR occurrence had poor graft survival for less than 5 years.
Asmaa Khudhur
3 years ago
the identification of DSA characteristics asso- ciated with AMR occurrence would have important clinical implications for highly sensitized patient management, immunosuppressive strategy optimization and kidney graft recipients’ posttransplant follow-up individualization.
A study done to analyze the relationship between preformed DSA and kidney graft outcomes.462 patients (89%) for the final analysis was taken . All patients were transplanted with a negative pretransplant T- and B-lymphocyte CDC crossmatch in current and peak sera. No data on flow-cytometric crossmatches was available
Anti-HLA antibodies screening and specificity
Pre-transplant anti-HLA IgG antibodies were tested in patient sera collected every 3 months while in the waiting list, by multiplex microsphere based on Luminex Xmap® Technology .The cut-off for positive samples was the Normalized Background (NBG) ratio .To determinate the specificity of the HLA antibodies, single-antigen bead (SAB) were performed in patients with a positive screening. a cut-off for a positive reaction were set in MFI value of ≥1000.
Donor HLA typing and virtual crossmatch
Samples of all deceased donors were routinely typed before recipient selection in loci HLA-A*, B* and DRB1* using polymerase chain reaction (PCR) amplification with specific sequence primers (SSP).HLA-DQ antigens with strong linkage disequilibrium were primarily inferred from the HLA-DR antigens .If a potential donor-specific HLA-DQ antibody was present, HLA-DQ antigens of the donor were verified by SSP DNA-typing .
After donor HLA typing, we performed a virtual crossmatch (virtual XM).
Then CDC crossmatch was carried out using the standard NIH technique, not enhanced with anti-human globulin.
Induction therapy was used in 413 patients (89.4%), with an anti-IL-2 receptor monoclonal antibody (Basiliximab Novartis®, 20 mg twice at day 0 and 4) or a polyclonal anti-thymocyte globulin (ATG Fresenius®, 3 mg/kg for 5–7 days).
Rejection diagnosis and treatment
Allograft rejection was defined as biopsy proven rejection (specimens were evaluated by light microscopy and immunofluorescence staining for C4d) and classified according to Banff classification as updated in 2013. Mild acute cellular rejection (ACR Banff grade I) was treated with pulse steroids (500 mg MP for 3 days) and increased maintenance immunosuppression. All other ACRs were treated with ATG. Patients with AMR were treated with plasmapheresis every other day and IvIg 100 mg/kg after each session.
followed by one dose of rituximab (375 mg/m2); a similar dose of IvIg (2 g/kg) was repeated 1 month later.
They found that the the presence of preformed DSA was clearly associated AMR occurrence.
With poor graft and patients survival at 1 and 5 year .
They found no adverse effect on graft outcomes associated with the presence of non-DSA anti-HLA antibodies in comparison with non- sensitized patients, as others have reported
they found a close association between AMR in DSA+ patients and poor graft survival.
DSA + AMR + patients had a significantly reduced 5-year graft survival .DSA + AMR − patients had similar graft survival in comparison with DSA− patients .
chronic antibody-mediated lesions evolved more frequently in DSA + AMR + patients.
Regarding immunosuppressive therapy , they found that DSA + patients receiving ATG induction in comparison with basiliximab showed a trend for developing less AMR .
In contrary ,Interestingly, in a cohort of 67 kidney graft recipients with preformed DSA, no difference in AMR incidence was found between patients transplanted without any induction and those receiving basiliximab or daclizumab .
In this study DSA HLA class was not associated with AMR as in others.
In contrast, DSA number and strength correlated significantly with AMR.
They demonstrated that only DSA MFI over 3000 were asso- ciated with higher incidence of AMR , and sDSA MFI threshold level (11,000) that had good specificity.
Limitations of this study :
First, the lack of available information about flow cytometry crossmatch results may affect its clinical application,
Second, no protocol biopsies were undertaken in this cohort
Third, the known limitations of SAB assay and their reported MFI values should be considered while interpreting the results.
Finally, as donor specificity of anti-HLA-DQ was not determined at the allelic level in every case (i.e., when the likelihood was ≥90%), rare false assignments as DSA cannot be excluded.
A detailed DSA characterization has a pivotal role in the management of HLA- sensitized patients, improving our ability to reduce the burden of AMR.
Wael Hassan
3 years ago
-anti HLA ab in 95 patient of 462 only 40 patient had DSA develop AMR 35% more than those without
-DSA with MFI>3000 significantly associated with AMR
-DSA increase risk of AMR and decrease graft survival even in negative CDC
-presence of DSA not mean no hope for kidney transplantation but it mean that recepient need aggressive induction and maintenence immunosuppresant
-in this study we search for relationship between DSA in negative CDC and AMR
-462 patient with negative CDC (2007-2012)
-anti HLA ab detected by SAB (DSA consider positive if MFI >1000
-donor HLA typing & virsual crossmatch for deceased donor done to detect DSA
-induction done with IL2 antagonist or ATG (in highly senstized patient or high HLA mismatch
-maintenence with tacrolimus, MMF & methylpredinsilone
-follow up patient till end of 2013
-rejection
*if mild ACR use pulse steroid & augment maintenace
*if more serious use ATG
*in AMR 4 session of plasmaphariese every other day with small dose IVIG after each one then after 4th session give high dose IVIG devided into 4 doses then retuximab then IVIG again after 1 month
-data gathering & comparison of MFI value in patient with DSA pre & 12 month after transplantation
-graft survival ,DGF ,episodes oof AMR , time of rejection , acute rejection & death with functioning graft all this data in comparison between DSA + & others
*DGF 25% in +ve DSA &19% in -ve
*acute rejection 37% vs 10%
*AMR 35% vs 0.8%
*days for rejection 11 vs 35
*graft failed 12% vs 3%
*death with functioning graft 2.5% vs 3.3%
finally this cohort study show that
1-presence of DSA associated with high risk of AMR (MFI >3000)
2-significance reduce in graft survival
*I saw patient with MFI >4000 and AMR occur after 10 months treated with IVIG & plasmaphariese then after 6 month occur again but patient refuse ttt
his maintenance immunosuppressant (tac,mmf,methylpredinisilone) & induction had done with ATG
Theepa Mariamutu
3 years ago
Analysis of preformed donor specific anti-HLA antibodies characteristics for prediction if antibody- mediated rejection in Kidney transplantation
Preformed HLA DSAs have shown to increase the risk of AMR and have deleterious effect on kidney graft survival even in the presence of negative CDCXM and FCXM
Usage of SAB to define DSA has been subject of debate. Differences in opinions regarding the effect of low level of DSA in kidney graft outcomes. Some studies has shown DSA characteristics, HLA class, number and strength (MFI) associated with kidney outcome
This study aimed to analyse the relationship between performed DSA and kidney graft outcomes in DCDXM and FCXM negative patients.
AMR
• Prevalance at 1 year -35%
• Compare with Lefaucher- 32.3%
• Other studies 55% incidence at 200 days but with protocol biopsies
• 55% of the current study patients received ATG – AMR free survival -65%
Death censored graft survival
• Reduced ( 84.8%) with DSA vs non -DSA( 94.9%)
• Amico et al- overall reduced 5 year censored graft survival ( 76.5% vs 89%)
o Possibly higher incidence of clinical AR
o No ATG induction and 59% no induction
• Willicombe et al – kidney graft survival DSA + 86.7% vs DSA – 94.3% in 3 years but induced with alemtuzumab and Tac mono therapy
Association AMR in DSA + and poor graft function
• Only DSA+AMR+ showed reduced 5 year graft survival 68.8%
• DSA+AMR- – 96%, DSA- -94.9%
• Other studies showed lower graft function in DSA+ patients with or without AMR
• DSA + AMR+ : remained detectable DSA at 1 year – had lower eGFR than DSA+AMR-
DSA characteristics and IS
• DSA+ :received ATG had lesser AMR than Basiliximab ( p=
• DSA-: no differences in AMR Basiliximab/Daclizumab vs no induction
• MFI below 5000 and DSA+: usage of ATG and IVIG had similar graft survival among DSA+ and DSA-
• DSA class is not associated with AMR in surenthiran study
• DSA number and strength (MFI) correlate with AMR
o MFI >3000 – higher incidence of AMR
Reem Younis
3 years ago
-This study, aimed to analyze the relationship between performed DSA and renal allograft outcome, in presence of negative CDC crossmatch which is not enhanced with anti-human globulin.
-Performed human leukocyte antigen (HLA) donor-specific antibodies (DSA) increase the risk of antibody-mediated rejection (AMR) and decrease renal allograft survival.
-DSA detection by single antigen bead (SAB) is the most sensitive than cell-based cross matches.
-517 patients (2007-2012) were involved in this study,55 patients were excluded, leaving 462 for final analysis.
-All patients were pretransplant negative T and B lymphocyte CDC crossmatch.
-Pretransplant anti-HLA IgG antibodies were tested in patient sera.SAB was performed in patients with positive screening.
-Deceased donors were typed in loci HLA-A, B, DRB1. High-resolution typing was performed when it was necessary to establish whether the anti-HLA antibodies were DSA.
-Virtual crossmatch was done.
-Induction therapy was used in 413 with Basiliximab or ATG.
-All patients had similar triple maintenance immunosuppressive (Tacrolimus, MMF, prednisolone)
-Living donor-recipient (89) started tacrolimus and MMF 7 days before transplantation, and no desensitization was carried out in these patients.
-Allograft rejection was defined as biopsy-proven rejection and classified according to Banff classification as an update in 2013.
-The study follow-up is about 39.6months.
-AMR was common in the HLA+DSA+ group especially in deceased donors and common with DSA MFI value ˃3000 while acute cellular rejection occurrence was similar between groups(DSA+ and DSA-).
-After one -year eGFR was similar between groups.
– DSA +ve AMR +ve had a significant reduce 5-year graft survival and lower eGFR.
-Death-censored kidney graft survival at 5-year in DSA +ve without AMR was similar to DSA -ve patients.
-According to this study, DSA +ve patients receiving ATG induction have a trend for developing less AMR. Limitations of the study :
1.No information about flow cytometry crossmatch results.
2.No protocol biopsies were undertaken.
3. Limitation of the SAB assay and their reported MFI values should be considered while interpreting our results.
4. Donor specificity of anti-HLA-DQ was not determined at the allelic level in every case.
-In my practice, yes, there is an association between DSA and AMR but we have no studies for that.
Abdullah Raoof
3 years ago
preformed ab ( sensitization ) can be formed by exposure to
1- previous transplant
2- pregnancy
3- blood transfusion
preformed ab is closely associated with graft loss through ab mediated rejection
ab types are
HLA ab
non HLA ab
DSA ab
non DSA ab
transplant outcome with negative cdc xm and negative anti HLA ab is beter than negative CDC XM and +ve ANTI HLA ab .. furthur more transplant outcome is beter in HLA ab +VE / DSA negative than HLA ab +ve / DSA +VE patient .
DSA associated with graft outcome through
1- intensity oF hDSA ab ( highest ab level – MFI MORE THAN 3000 ) or sDSA ( SUM OF DSA ab of more than – 11000 )
2- multiple DSA target .
there is no difference in graft outcome according HLA class DSA .
there is no clear cut of negative DSA as it depend on the lab –
Weam Elnazer
3 years ago
The purpose of this research, which included 462 transplant patients, was to determine if premade DSA had any influence on graft survival.
When DSA was performed, it was shown to be related to an increase in AMR in individuals who had negative CDC crossmatch.
patients with positive DSA had a lower graft survival rate. MFI greater than 3000 was found to be associated with a statistical significance.
Patient’s with a positive DSA test had an AMR of more than 35 %. Following 5 years of follow-up.
DSAs that have been performed have been demonstrated to increase the probability of antibody-mediated rejection (AMR) and have a detrimental influence on long-term graft survival.
Single antigen bead (SAB) assays for HLA antibody detection are substantially more sensitive than cell-based cross matches, allowing for the determination of antibody specificity.
Limitation:
-The group did not undergo any protocol biopsies.
– SAB assay limitations and reported MFI values
-Anti-HLA-DQ donor specificity was unknown
-The lack of data on flow cytometry cross-match findings.
Impact on clinical practice:
Patients with preformed DSA need close monitoring after transplantation. and the higher the titre of preformed DSA, the worse the graft outcome and the risk of rejection.
Rehab Fahmy
3 years ago
Impact of preformed DSA characteristics on kidney graft outcomes.
*Preformed human leukocyte antigen (HLA) donor specific antibodies (DSA) have been shown to increase the risk of antibody-mediated rejec- tion (AMR) and have a deleterious effect on kidney graft survival, even in the presence of a negative complement-dependent cytotoxicity (CDC) and/or flow cytometry crossmatches .
*Some have reported that low level DSA exert no effect on kidney grafts outcomes , while others have associated DSA characteristics as HLA class , number , and strength as read by mean fluorescence intensity (MFI) with kidney graft outcomes.
* Several studies showed that it is the occurrence of AMR associated with DSA that jeopardizes kidney graft survival, with DSA positive patients without AMR having similar survival as compared with those without DSA .This contradictory effect may result from the DSA ability to mediate endothelial injury through complement-dependent and independent mechanisms, mainly through acute and/or chronic AMR, but also, under certain conditions, to promote graft accommoda- tion through up-regulation of complement regulatory or cytoprotective proteins .
*Aimof the study: aimed to analyze the relationship between preformed DSA and kidney graft outcomes, in the presence of a negative CDC crossmatch. Furthermore, we sought to explore the predictive value of DSA characteristics for AMR occurrence.
*Methods: 462 patients that received a kidney graft in our unit, between 2007 and 2012, pre-transplant sera were analyzed by Luminex screening assay to determine the presence of anti-human leukocyte antigen (HLA) antibodies and single-antigen bead assay [positive if mean fluorescence intensity (MFI) ≥ 1000] to assign anti-HLA specificities.
*Results:
1-Anti-HLA antibodies were present in 95 patients (20.6%), but only 40 (8.7%) had DSA.
2-Antibody- mediated rejection (AMR) at 1-year was significantly higher in patients with DSA (35.0%) than in those without them (0.9%) .
3- Only DSA with a MFI of >3000 were significantly associated with AMR occurrence ,MFI of >4900 in the highest DSA bead had a high sensitivity (85.7%) and that the sum of all DSA beads MFI >11,000 had a high specificity (92.3%) for AMR prediction.
4-Anti-thymocyte globulin versus basiliximab induction was more frequent in DSA + AMR − (65.4%) versus DSA + AMR + (34.6%) patients . Five-year censored graft survival was lower in DSA+ than in DSA− patients (respectively, 84.8% versus 94.9%,
5-survival was only reduced in DSA+ AMR+ (68.8%) versus DSA+ AMR− (96.0%) patients .
*Conclusion:
Preformed DSA is associated with kidney graft loss, in relation with AMR occurrence. DSA strength may be used to improve immunological risk stratification of sensitized patients and their clinical management.
Good study as they do correlation between DSA and AMR ,Survival,Induction therapy.
Strong point also :using χ2 test in statistical analysis of variable data
Our practice : According to MFI we choose Pt with DSA positive but low MFI ,no updated registry to notice statistically significant AMR or not ,,usually we go with ATG induction in those cases
Mohamed Fouad
3 years ago
Preformed donor specific antibodies (DSA) have been shown to increase the risk of antibody-mediated rejection (AMR) and have a negative impact on long term graft survival. HLA antibody detection by single antigen bead (SAB) assay is much more sensitive than cell-based crossmatches, allowing for the definition of antibody specificity.
DSA characteristics that determine their effect on long term graft survival depends on HLA class (Antibodies against class I or class II), number and strength of DSAs.
This retrospective study analysed anti HLA results from462 patients that received a kidney graft between 2007 and 2012.The aim of this study to determine the impact of preformed DSA (detected by Luminex assays with negative CDC) characteristics on kidney graft outcomes. The positive value if mean fluorescence intensity (MFI) ≥ 1000.
Results of the study revealed: The presence of preformed DSA was clearly associated AMR occurrence. Similar results obtained in negative CDC crossmatch transplants. Prevalence of AMR at 1-year posttransplant in patients with DSA was high (35.0%).
The study limitations:
1-lack of available information about flow cytometry crossmatch results.
2-No protocol biopsies were done.
3-The limitations of SAB assay and their reported MFI values has to be considered when interpreting the results.
4-The donor specificity of anti-HLA-DQ was not determined at the allelic level in every case.
Thank you for mentioning the limitations in your own words What about your own practice
Last edited 3 years ago by Ala Ali
Amit Sharma
3 years ago
1. In your own words, summarise this article.
Analysis of preformed donor-specific anti-HLA antibodies characteristics for prediction of antibody-mediated rejection in kidney transplantation
The study involved analysis of kidney transplant recipients in a single center performed over 5 years vis a vis the presence of prior presence of donor specific antibodies (DSA) and their effects on the graft prognosis.
DSA, even in presence of negative CDCXM, have shown to be associated with antibody mediated rejection (AMR) and poor graft survival and this study tried to document effect of pre-formed antibodies, in such patients, on the graft with respect to type of antibodies as well as strength of antibodies.
A total of 462 patients were evaluated and out of them 95 (20%) had pre-formed antibodies with 42% of those having DSAs. The patients were divided into 3 groups: patients with DSAs, patients with non-DSA antibodies, and patients without any pre-formed antibodies.
Acute rejection (overall as well as episodes of AMR alone) was more common and developed earlier in patients with DSAs than those with non-DSA antibodies. AMR prevalence correlated with MFI of DSA with 0% in patients with MFI less than 3000 and 60% in patients with MFI more than 6000. A single antibody MFI of >4900 and total MFI strength of >11000 was associated with higher risk of AMR. DSA positive patients who received ATG as induction had lower AMR as compared to those receiving basiliximab as induction. In the study, no correlation could be established between DSA class and AMR.
Although the eGFR at one year was similar in the 3 groups, presence of DSA was associated with reduced 5 year graft survival (85%) as compared to those with non-DSA antibodies (89%) and patients without antibodies (94%). Death censored graft survival at 5 years in patients with DSA experiencing AMR was lower (68.8%) than those who did not develop AMR (96%) as well as patients without DSA (96%).
The study had its limitations including non-availability of flowcytometry crossmatch and protocol biopsies as well as donor specificity of anti HLA DQ (as donor HLA DQ typing was not done).
The study concluded that pre-formed DSAs are associated with increased incidence of AMR and poor graft survival.
2. Please reflect on your practice if possible.
In patients with prior history of sensitization, SAB for detecting preformed antibodies is performed in our unit and in patients with pre-formed antibodies, we give induction using ATG with triple drug immunosuppression (Tacrolimus, MMF and steroids) and follow post transplant antibody levels to take pre-emptive action prior to clinical deterioration for better management of renal transplant recipients.
Last edited 3 years ago by Amit Sharma
Hamdy Hegazy
3 years ago
This a retrospective study involving 462 renal transplant recipients from 2007-2012 where pre-transplant sera were analyzed by Luminex for anti-HLA antibodies and SAB assay. MFI> 1000 was considered as positive.
The aim of the study is to analyze the relationship between preformed DSA and kidney graft outcomes in the presence of negative CDC crossmatch.
Presence of preformed DSA even with negative cross-match was clearly associated with AMR occurrence.
AMR at 1-year post-Tx was higher in those with positive DSA (35%), however it was very low in those with negative DSA (0.9%).
AMR-free survival at 1-year was 65% for patients with DSA+ve who had induction immunosuppression with ATG.
5-year graft survival was lower in those with +ve DSA/+ve AMR (68.8%) compared to those with +ve DSA/-ve AMR (96%).
5-year graft survival was 84.8% in those with DSA +ve, in comparison to 94% in those with negative DSA.
DSAs increase AMR and lower graft survival even with negative cross-match.
Results:
Acute rejection was common in HLA+DSA+ patients (37.5%), HLA +DSA- (14.5%), HLA-ve(10.6%).
AMR was higher in DSA+ve/HLA +ve (35%).
Acute cellular rejection was similar between different groups.
Acute rejection developed earlier in DSA+ve/HLA+ve (median 11 days), HLA+ve/DSA-ve (median 71 days), HLA -ve (median 35 days).
The higher DSA MFI(>10,000), the earlier of AMR ( first 11 days).
Study limitations:
1- Lack of information about flow cytometry cross-match results may affect its clinical application.
2- No protocol biopsy was undertaken in this cohort.
3- Limitations of SAB assay and their reported MFI values.
4- Donor specificity of anti-HLA-DQ was not determined at the allelic level in every case.
Clinical application:
1- Serial monitoring of DSA especially in highly sensitized patients or previously sensitized ones.
2- Detailed DSA characterization has an important role in the management of HLA-sensitized patients, improving our ability to reduce the burden of AMR.
Huda Al-Taee
3 years ago
This study aimed to analyze the relationship between preformed DSA and kidney graft outcomes in the presence of negative CDC crossmatch.
The study involve 462 patients who received kidney transplantation with negative CDC XM( both T & B cells).Anti HLA antibodies tested by luminex were found in 20% of patients and only 8% had DSA. No data available about FCXM. Ethical approval gained from the institutional review board at centro hospitalar do porto.
The study found that ABMR at 1 year was higher in patients with DSA than in those without. DSAs with MFI> 3000 were associated with ABMR occurrence. ATG induction was more frequent in DSA+ ABMR – patients. five year graft survival was lower in DSA+ patients, although survival was only reduced in DSA+ ABMR+ patients as compared to DSA+ AMR- patients. DSA HLA class was not associated with ABMR.
Limitation of this study:
lack of information about FCXM
no protocol biopsies was taken
the limitations of SAB assay and their MFI values should be considered while interpreting the results.
rare false assignments as DSA can not be excluded as donor specificity of anti HLA-DQ was not determined at the allelic level in every case.
Mahmud Islam
3 years ago
Preformed HLA antigens specific antibodies (DSA) was shown to have a negative effect on graft survival. some studies showed that low levels may be of low importance. this is dependent on both DSA level and specificity. In this study performed DSA in case of negative CDC-CXM was evaluated in terms of relation to graft survival in addition to its predictive value for ABMR.
In this 5 years study (2007-2012), 462 (out of 517) were included. pateints with LUMINEX MFI>1000 were accepted . All patients had CDC negative CMX. serial assays each 3 months were performed. SBA assays added when when indicated.. Flow cytometry data was not available. Low resolution mach for HLA-A*, -B* and DRB1 using PCR amplification and SSP low resolution used. High resolution was evaluated in special cased. . 4 patients were excluded because of HLA-c positive or negative ppi.
Virtual CMX was performed using the results of HLA typing.
ATG was used in highly sensetized patients and or basiliximab (total 413/462; %89) as induction therapy. maintenance triple immune suppression using Tacrolimus/MMF and MP was used.
all regections were proved by biopsy. mild cases treated with pulse MP (500mg) plus elevetaion of target levels of immunosuppression. server cases were treated with ATG and plasmaphresesi/IVIG followed by Rituximab.
In this cohort study presence of preformed DSA’s was associated with ABMR.
acute rejection was more prominent in both HLA+ and DSA + cases. graft survival was a little bit lower in DSA – group compared with DSA + (84.8% vs 94.9%). ABMR was higher in case with DSA>3000.
In clinical practice one should follow DSA serilaly especially in highly sensetized or in case with previous history of sensetization.
Ban Mezher
3 years ago
The detection of preformed DSA is very important in stratification of rejection risk, it shown that worse effect on graft outcome even CDCXM & FCXM are both negative. Therefor determination of DSA characteristics associated with detection of AMR had important clinical implication in sensitized patient, immunosuppression strategy & in post transplantation follow-up.
This study follow 462 recipients with negative CDCXM, preformed DSA ( by Luminex, SAP) & VXM. The patient followed to the end of 2013, death or graft loss. Induction with ATG (for high HLA mismatch & allo immunized patients) & anti-IL-2 Abs & maintenance immunosuppression with MMF, CNI & low dose steroid. Graft rejection diagnosed based on histopathological changes of graft biopsy. The study found the following:
there is strong association between AMR & presence of preformed DSA inspire negative CDCXM
patient with ATG induction had more AMR free survival at 1 year of transplantation
death censured graft survival decrease in patient with DSA
DSA+/AMR+ recipients had decreased 5 years survival when compared to DSA+/AMR- recipient ( similar to DSA-ve recipients.)
DSA+/AMR+ recipients with persistent DSA beyond first year post transplantation was associated with lower GFR comparing to DSA+/AMR- recipients.
level of DSA associated with increased risk of AMR, if MFI < 3000 the res decreased.
The study had several limitation including:
no FCXM result available
protocol biopsy not done.
known limitation of SAP
HLA-DQ Abs not measured.
Shereen Yousef
3 years ago
Summary of this article
This article presents a study of 462 kidney transplant recipients, between 2007 and 2012, all patients had negative CDC-XM for both B and T CELLS .
pre-transplant sera were analyzed by Luminex screening assay to determine the presence of HLA antibodies and single-antigen bead assay [positive if mean fluorescence intensity (MFI) ≥ 1000] to assign anti-HLA specificities.
The aim was to analyse the effect of preformed DSA graft outcomes.
It was shown that
-Preformed HLA donor specific antibodies (DSA) increases risk of AMR and associated with poor kidney graft survival, even with negative CDC .
detection of HLA by single antigen bead assay is more sensitive than cell-based crossmatches.
Not only the presence of DSAs affect the graft but t was shown that that low level DSA exert no effect on kidney grafts outcomes also DSA characteristics as HLA class , number , and strength as read by mean fluorescence intensity (MFI) all has its effecton graft outcome.
Although some studies showed that the occurrence of AMR not the mere presence of DSAs that affects the graft and whether this DSAs can initiate endothelial injury or not .
Prevalence of AMR at 1-year posttransplant in our patients with DSA was high.
In a cohort of ATG-induction improved AMR-free survival at 1-year as it was 65.1% for patients with DSA .
There was a significant reduction in 5-years graft survival in recipients with positive DSAS possibly related with a higher incidence of clinical acute rejection
The study also shows that only DSA+ AMR+ patients had a significantly reduced 5-year graft survival while ; DSA+ AMR− patients (96.0%) had similar graft survival in comparison with DSA− patients (94.9%).
There for the occurrence of chronic antibody-mediated rejection is more frequently in DSA+ AMR+ patients.
In comparison of induction therapy ATG was better than basiliximab eitj less AMR .
ATG and IVIG induction in patients with DSA (below 5000 MFI) has been shown to allow for successful kidney transplantation with similar rejection rates and graft survival between DSA+ and DSA− patients .
DSA MFI values below 3000 compared to higher MFI values were associated with improved 5-year death-censored graft survival .
Limitation of the study
No available FCXM .
No protocol biopsies were done .
limitations of SAB assay and MFI values.
anti-HLA-DQ was not determined at the allelic level in every case .
Esmat MD
3 years ago
Performed HLA DSAs have been associated with increasing the risk of AMR and lower graft survival even in the presence of negative CDC and FCXM.
HLA DSA detection by SAB is more sensitive and specific compared to cell-based crossmatches.
Although some studies have reported low level of DSAs has no significant effect on graft survival, others have reported the association between DSA characteristics such as HLA class, class, number, and strength (MFI) with kidney allograft outcome.
DSAs that lead to AMR have had a significant effect on reducing graft survival by complement mediated and non-mediated endothelial injury through acute and chronic AMR.
DSA characteristics may have important implications for highly sensitized patients, immunosuppressive strategies, and post-transplant follow up individualization.
This study has analyzed the graft outcomes between anti HLA+DSA+ and anti HLA+DSA- recipients. The incidence of AMR has been more in DSA+ compared to DSA- patients (37.5% versus 14.5%), and acute cellular rejection was the same.
This study has demonstrated significant increase in AMR with DSA MFI of>3000, the highest sensitivity for AMR prediction with a MFI of>4900 in the highest MFI bead, and highest specificity for AMR prediction with the sum of all DSA beads MFI>11000.
Induction therapy with rATG compared to Basiliximab has been related to lower incidence of AMR in DSA+ recipients (65% versus 34%).
There was no adverse effect of non-DSA HLA antibodies in comparison with non-sensitized patients.
DSA+AMR+ with persistence detectable DSA had lower eGFR than DSA+AMR- patients. In addition, chronic AMR has evolved more frequently in DSA+AMR+ patients.
Five-year censored graft survival was lower in DSA+ than in DSA-patients.
Dalia Eltahir
3 years ago
DSAs has been associated with increased risk of AMR and decrease graft survival, even if the CDC and/ or flow cytometry cross matches were negative. The main effect of DSA depends on its specificity and strength as defined by MFI. So, defining DSA characteristics associated with AMR occurrence has an important clinical role in the management of a highly sensitized patient, immunosuppressive protocol and patient’s follow up post-transplantation. In the present cohort, the presence of DSA was associated with AMR occurrence, and affected the graft survival, although the CDC crossmatch was negative. The prevalence of AMR was high at 1-year post-transplantation and DSA remained detectable in patient receiving ATG leading to a chronic antibody-mediated . Other studies use protocol biopsy; this incidence was much higher. However, ATG induction increases the AMR-free survival at 1 year to 65% for DSA positive recipients. As well, ATG induction had been shown to improve the 5-years graft survival in DSA positive recipients.
The present study found no adverse effect on graft outcomes associated with the presence of non-DSA anti-HLA antibodies in comparison with nonsensitized patients, as others have reported.
DSA+ patients whose developed AMR have significantly reduced 5-year graft survival; while DSA+ patients who do not develop AMR had graft survival similar to those with negative DSA.
the difference between the DSA positive recipients in developing AMR, point to exploring the importance of DSA characteristics and immunosuppression more likely associated with its occurrence. ATG has been associated with less AMR in DSA + patients as compared with basiliximab. furthermore, ATG and intravenous immunoglobulin induction treatment give a comparable graft survival between DSA positive (below 5000MFI) and DSA negative . interestingly, others have shown that DSA MFI values below 3000 compared to higher MFI values were associated with improved 5-year death-censored graft survival
Limitation duto First, the lack of available FCXM . Second, no protocol biopsies were taken . Third, the known limitations of SAB assay. Finally, as donor specificity of anti-HLA-DQ was not determined at the allelic level in every case .
In our centre we don’t transplant patient with positive DSA .Many patient came with AMR later ,they develop De novo DSA.
Mahmoud Hamada
3 years ago
In your own words, summarise this article.
· This study enrolled 462 transplant patients, the aim of it was to assess the effect of preformed DSA on graft survival.
· Preformed DSA was associated with increased AMR among patients with negative CDC crossmatch
· AMR for patients with positive DSA was > 35%.
· There was a reduced graft survival after 5 years among patients with positive DSA.
· MFI > 3000 was associated with a significant increased AMR. Also, high DSA MFI had predicted graft survival with an AUC of 0.86.
· DSA positive, AMR positive has significant poor graft survival (5-years) 87%
· DSA negative = DSA positive with AMR negative survival was similar average 95%.
Hinda Hassan
3 years ago
This study is conducted on 462 kidney recipients with a negative complement-dependent cytotoxicity (CDC) cross-match but positive luminex. It confirmed the association between preformed DSA and rejection. Not all DSA affect graft as DSA can affect the graft negatively through endothelial injury and positively they can upregulate protective mechanisms . the effect is attributed to the HLA class , the number and intensity .
In this study not all antibodies present are DSA ,DSA are 8.7% only. DSA are associated significantly with higher rate of 1 year AMR(35.0%). This is only for DSA of MFI(N3000) or higher. The study suggest that ATG in DSA positive is better than bailiximab as it is associated with less AMR (weak significance)
There was no statically significant difference between those with who had DSA versus those who have non DSA antibodies regarding : DGF, eGFR at one year and death with functioning graft. Time to acute rejection is shorter in DSA group and the number of failed graft is more.
Though DSA associated with AMR have higher number, intensity and MFI sum, there was no statically significant difference between DSA causing AMR and DSA not causing AMR in following : DSA class, previous sensitization, or induction
Death censored graft survival is shorter with presence of DSA, DSA lead to statically significant lesser 5 year censored graft loss and less survival .
The drawbacks of this study are the unavailability of FCXM , SAB limitations , no protocol biopsy were conducted and donor DQ were not identified
In our country we do not transplant across DSA barriers. the cost of treatment are nationally secured through a national program. This is why we are trying to concentrate on patients who can make the most benefit from the transplant .
Jamila Elamouri
3 years ago
Analysis of preformed donor-specific anti-HLA antibodies characteristics
for prediction of antibody-mediated rejection in kidney transplantation
The presence of DSAs has been associated with increased risk of AMR and deleterious graft survival, even if the CDC and/ or flow cytometry cross matches were negative. SAB assay is more sensitive than cell-based crossmatches in defining DSA specificity. The main effect of DSA depends on its specificity and strength as defined by MFI. So, defining DSA characteristics associated with AMR occurrence has an important clinical role in the management of a highly sensitized patient, immunosuppressive protocol and patient’s follow up post-transplantation. In the present cohort, the presence of DSA was clearly associated with AMR occurrence, and affected the graft survival, although the CDC crossmatch was negative. In spite of the majority of the patient in the study receiving ATG induction; the prevalence of AMR was high at 1-year post-transplantation and DSA remained detectable a clue to chronic antibody-mediated lesions in these recipients. Other studies use protocol biopsy; this incidence was much higher. However, ATG induction increases the AMR-free survival at 1 year to 65% for DSA positive recipients. As well, ATG induction had been shown to improve the 5-years graft survival in DSA positive recipients.
The present study found no adverse effect on graft outcomes associated with the presence of non-DSA anti-HLA antibodies in comparison with nonsensitized patients, as others have reported.
DSA+ patients whose developed AMR have significantly reduced 5-year graft survival; while DSA+ patients who do not develop AMR had graft survival similar to those with negative DSA.
the difference between the DSA positive recipients in developing AMR, point to exploring the importance of DSA characteristics and immunosuppression more likely associated with its occurrence. ATG has been associated with less AMR in DSA + patients as compared with basiliximab. furthermore, ATG and intravenous immunoglobulin induction treatment give a comparable graft survival between DSA positive (below 5000MFI) and DSA negative patients.
The DSA HLA class has no role in AMR development, while, the DSA number and strength are significantly associated with AMR. Especially if DSA MFI is over 3000. The study reveals that detect hDSA MFI threshold level (4900) represent good sensitivity and sDSA MFI level (11,000) had good specificity, So, both together can be used to improve AMR prediction. The association of DSA strength with the occurrence of AMR has been studied by many investigators, some reporting no association, while others showed a significant association which could be related to the variability in MFI cutoff levels between the different centres.
anti-HLA antibodies screen and DSA detection by solid-phase assays are valuable tools, support the transplantation decision in sensitized patients with or without desensitization protocols as well, the applicability of protocol biopsy for selected patients.
The type of DSA whether class 1 or 2 has an effect on the AMR. AMR due to class 1 is usually early and aggressive, while class 2 is usually late and less acute.
Mohamad Habli
3 years ago
Preformed DSA have been shown to be associated with increase the risk of AMR and eventually risk of graft failure.
The implementation of solid phase assays in the clinical practice has been associated with improvement in DSA detection and identification of specificities and characteristics of antibodies. HLA antibody detection by SAB assay is well known to be more sensitive than cell based crossmatches.
Some studies reported that low level DSA based on MFI levels have no adverse renal effect, while others studies reported renal rejection and graft failure based on DSA characteristics as HLA class, number, and strength reflected by MFI.
This article reports the results of study done to evaluate the incidence of preformed anti-HLA antibodies by Luminex and SAB assay screening techniques and their association with renal allograft outcomes.
Screening for HLA antibodies were tested in patient every 3 months by Luminex Technology. If initial screening is positive then, SAB assay was performed to determinate the specificity of the HLA antibodies.
Deceased donors were routinely typed for loci HLA-A*, B* and DRB1. If a DSA-DQ antibody was present, HLA-DQ antigens of the donor were verified by SSP. Patients with anti-HLA-C and/or -DP antibodies were excluded.
89.4% of recipients in this review received anti-IL-2 receptor monoclonal antibody as Induction therapy and the remaining received ATG being considered high risk for rejection.
Results of the study
-Acute rejection was particularly more common in HLA+ DSA+ patients 37.5% in comparison with HLA+ DSA− 14.5% and HLA− patients (10.6%). Acute rejection developed earlier in HLA+ DSA+ patients than in HLA+ DSA−.
-DSA+ patients had highest DSA bead with a median MFI of 10,273, while those with later AMR had a median MFI of 5331.
-Death-censored kidney graft survival was detected with a 5-years survival of 94.9% in DSA− versus 84.8% in DSA+ patients. Patient survival was similar.
-Frequent use of ATG versus Basiliximab induction in patients with DSA that did not experience AMR (65.4% and 34.6%) was noticeable.
-At 12-months posttransplant, DSA remained detectable in 40% of DSA+ AMR− and 81.8% DSA+ AMR+ patients.
-Only DSA MFI over 3000 were associated with higher incidence of AMR with good sensitivity 85.7% and MFI threshold level 11,000 that had good specificity 92.3%.
Conclusion
SAB assay is more sensitive and specific in identifying donor specific antibodies. This is clinically translated in better matching between donor and recipients and better planning for the induction and maintenance therapy for at risk recipients. The presence of anti-HLA antibodies is independent risk factor for antibody medicated rejection even in the absence of donor specific antibodies.
Luminex is a standard technique for screening for pre-transplant preformed antibodies. If positive proceeding with SAB id of great benefit on allograft survival.
Preformed DSA is associated with increased risk of AMR and graft loss, so its detection is very important, it was found that negative CDC or FCXM might be associated with detection of preformed DSA by solid phase assay SAB (single antigen bead).
Graft survival of the patients with AMR without DSA and patients with DSA and no AMR are the same, its effect on graft survival is related to its strength and specific phenotype.
Management of the patient with DSA with desensitization protocol is very important issue to reduce or avoid AMR posttransplant.
in my practice:
if CDC or FCXM negative will proceed to transplantation, even if there is history of sensitization
Sherif Yusuf
3 years ago
Incidence of occurrence of early ABMR is markedly increased in patient with preformed DSA and those who received desensitization thus lowering graft survival.
The significance of positive luminex negative FCM (PLNF) is debatable some studies found no significant difference when compared to negative luminex, negative FCM in terms of incidence of AR at 1 year, graft failure at 1 and 5 years, patient survival at 1 and 5 years,
Other studies including the current study found an increased incidence of ABMR and graft loss in patients with PLNC when compared to those with negative luminex
Several studies found that the presence of DSA alone may not decrease graft survival and do so only if associated with biopsy proved ABMR and this may be explained by the fact that although DSA can produce damage through complement and non-complement dependent injury but at the same time it can up regulate complement regulatory proteins leading to graft accommodation
Factors affecting significance of preformed DSA
1. Strength of DSA (MFI):
DSA best detected by Luminex which can detect very week DSA with MFI of 300, different thresholds for detection of DSA was set with MFI cutoff ranging from > 300 in some centers to > 1500 in others, MFI dose not correlate well with cross matching, some patients have low MFI and positive cross match, others have high MFI and negative cross match
Moreover, patients with biopsy proven ABMR, DSA MFI of >3000 have lower graft survival than those with MFI of ≤3000, on the other hand patients without biopsy- proven rejection DSA MFI dose not correlate well with graft survival
In the current study it was found that patients with ABMR has 7.4 higher probability of having DSA with MFI > 11000 (good specificity), and patients without ABMR has 5 higher probability of having DSA with MFI < 4900 (good sensitivity)
2.Complement fixing ability of DSA :
Complement fixing AB (cause positive CDC ) are more clinically significant than non-complement fixing AB since it is associated with ABMR and C4d staining which affect badly graft survival
Moreover it is correlated well with cross matching but not with intensity of DSA this means there may be a DSA with high intensity but is not complement fixing.
3. Specificity of DSA :
In another ward, The specific type of HLA Ag to which DSA is formed, graft survival in patients with DSA directed to both HLA class I and II is lower than when DSA is directed to either class I or II.(HLA class)
Moreover DSAs directed against HLA-A, HLA-B and HLA-DR are associated with earlier AMR when compared to DSA directed to HLADQ, also DSA directed against HLA C and HLA DP may have clinical significance (HLA type and number)
In my practice
All cases i have participated in had negative cross match so we did not do DSA to detect PLNC cases
See scenario 4 Sherif They may develop de novo DSA
Assafi Mohammed
3 years ago
Summary of the Article: Study type: This is a retrospective analysis Study Background:
The relevance of preformed donor specific antibodies (DSA) detected by Luminex assays, with a negative complement-dependent cytotoxicity (CDC) crossmatch, remains unsettled in kidney transplantation (KT).
Preformed human leukocyte antigen (HLA) donor specific antibodies (DSA) have been shown to increase the risk of antibody-mediated rejection (AMR) and have a deleterious effect on kidney graft survival, even in the presence of a negative complement-dependent cytotoxicity (CDC) and/or flow cytometry crossmatches.
HLA antibody detection by single antigen bead (SAB) assay is much more sensitive than cell- based crossmatches, allowing for the definition of antibody specificity. However, the clinical relevance of DSA defined by SAB has been the matter of an ongoing debate. Some have reported that low level DSA exert no effect on kidney grafts outcomes ,while others have associated DSA characteristics as HLA class , number , and strength as read by mean fluorescence intensity (MFI) with kidney graft outcomes.
Ultimately, DSA main effect will depend on its specificity and strength.
Aim of the study: to analyze the impact of preformed DSA characteristics on kidney graft outcomes. Study Methods:
In 462 patients that received a kidney graft in the unit of study group, between 2007 and 2012, pre-transplant sera were analyzed by Luminex screening assay to determine the presence of anti-human leukocyte antigen (HLA) antibodies and single-antigen bead assay [positive if mean fluorescence intensity (MFI) ≥ 1000] to assign anti-HLA specificities.
All patients were transplanted with a negative pretransplant T- and B-lymphocyte CDC crossmatch in current and peak sera.
No data on flow-cytometric crossmatches was available for this study.
The analysis was performed using HLA fusion® software (One Lambda Inc.) and a cut-off for a positive reaction were set in MFI value of ≥1000.
High resolution typing was performed when it was necessary in order to establish whether the anti-HLA antibodies were DSA. Patients with only anti-HLA-C and/or -DP antibodies were not considered (n = 4) in the analysis, as no donor typing at these loci was available.
In each individual DSA, the strength was based on the MFI of one SAB. In the case of several DSA against different HLA-antigens, the study group usually considered the DSA with the highest MFI; occasionally, they used the cumulative strength of all DSA by adding the individual MFI values, as stated in the results.
Induction therapy was used in 413 patients (89.4%), with (Basiliximab Novartis®, 20 mg twice at day 0 and 4) or a polyclonal anti-thymocyte globulin (ATG Fresenius®, 3 mg/kg for 5–7 days).
All enrolled recipients had similar triple maintenance immunosuppression, consisting of oral tacrolimus (FK-506), mycophenolate mofetil (MMF) and methylprednisolone (MP)/prednisolone.
FK-506 was started at a dose of 0.1–0.15 mg/kg/day, and the dose was adjusted to maintain a trough level in whole blood between 8 and 12 ng/ml during the first month postoperatively, between 7 and 10 ng/ml during 2–3 months after transplant and between 5 and 8 ng/ml thereafter. MMF was started at a dose of 2000 mg/day, with the dose decreasing to 1000–1500 mg/day during the first month postoperatively, depending on white blood cells count. Methylprednisolone was administered intravenously at doses of 500, 250 and 125 mg/day on the day of transplantation, days 1–2 and days 3–4 after the operation, respectively. Oral prednisolone was started on day 5 after the operation at the dose of 20 mg, being then tapered to 5–10 mg/day within 2–3 months after transplant. Living donor recipients (n = 89) were prescribed FK-506 and MMF 7 days before transplant. No desensitization protocol was carried out in these patients.
Study Results: 1.Patient characteristics according with HLA antibodies detection
In study sample of 462 kidney graft recipients:
367 (79.4%) patients had no anti-HLA antibodies (HLA−) detectable in pretransplant serum.
A positive anti-HLA antibody screening was present in 95 (20.6%) patients, with 40 of them having antibodies directed against donor HLA molecules (HLA+ DSA+), while the remaining 55 patients had not (HLA+ DSA−). Only 2 out of 40 HLA+ DSA+ patients had no presensitizing events recorded.
As expected, presensitizing events and female gender were more common in HLA+ DSA− and HLA+ DSA+ patients, in comparison with HLA− patients (P < 0.001).
HLA-sensitized patients presented more frequently a PRA level N 20% (P <0.001), had a longer time on dialysis (P< 0.001) and were less likely to receive a graft from a woman (P = 0.050) or a living donor (P = 0.004).
No significant differences between groups were found regarding recipient and donor age, HLA mismatches and cold ischemia time.
Lastly, induction therapy was used in all HLA+ DSA+ patients, with ATG use predominating over basiliximab (P< 0.001).
2.Transplant outcomes according with HLA antibodies detection
Acute rejection was particularly more common in HLA+ DSA+ patients (37.5%) in comparison with HLA+ DSA− (14.5%) and HLA− patients (10.6%) (P< 0.001). This difference resulted from the high incidence of antibody-mediated rejection in HLA+ DSA+ patients (n = 14/40, 35.0%), while acute cellular rejection occurrence was similar between groups (P = 0.322).
Acute rejection developed earlier in HLA+ DSA+ patients (median 11 days) than in HLA+ DSA− (median 71 days) and HLA− patients (median 35 days) (P = 0.018).
DSA+ patients experiencing AMR in the first 11 days after transplant showed, in the highest DSA bead (hDSA), a median MFI of 10,273 (6944–13,919) while those with later occurring (N11 days) AMR had a median hDSA MFI of 5331 (4265– 6317) (P = 0.020).
Incidence of AMR in DSA+ graft recipients from a deceased donor was 33.3% (n = 12) and 50% (n = 2) in living donor graft recipients (P = 0.602), with no significant difference in the number of HLA mismatches between them (3.97 ± 1.21 and 3.25 ± 1.71 respectively; P = 0.465).
At 12-months post-transplant, kidney graft eGFR was similar between groups (P = 0.168). Nevertheless, a significant detrimental effect of DSA presence on death-censored kidney graft survival was detected, with a 5-years survival of 94.9% in DSA− versus 84.8% in DSA+ patients (P = 0.006). Patient survival was similar (P = 0.585).
3.Antibody-mediated rejection association with DSA characteristics and immunosuppression
DSA+ patients with AMR presented a higher number of DSA beads (P = 0.033) and stronger DSA, considering the highest DSA bead MFI (P < 0.001) or the sum of all DSA beads MFI (P = 0.001). A trend towards a more frequent use of ATG versus Basiliximab induction in patients with DSA that did not experience AMR (65.4% and 34.6% respectively, P = 0.072) was noticeable. No difference was detected concerning DSA HLA class or presensitizing events.
The prevalence of AMR increased significantly with the presence of DSA MFI value over 3000: 0.9% in DSA− patients, 0% in DSA+ patients with a MFI of < 3000, 38.5% in those with a MFI of 3000–5999 and 60.0% in those with a MFI of ≥6000 (P< 0.001, χ2 for trend).
4.Kidney graft outcomes in DSA+ patients with or without antibody-mediated rejection
At 12-months post-transplant, DSA remained detectable (only 36 patients had available data) in 10 (40%) DSA+ AMR− and 9 (81.8%) DSA+ AMR+ patients (P = 0.031). Considering these 19 patients, median hDSA MFI in pre-transplant and at 12-months post-transplant sera were, respectively, 2725 (1709–5603) and 2212 (1383–2630) in DSA+ AMR− patients (P = 0.093); 6354 (4913–10,572) and 4567 (3545–6123) in DSA+ AMR+ patients (P = 0.260).
Death-censored graft survival at 5 years in DSA+ patients without AMR occurrence (96.0%) was similar to DSA− (94.9%) patients (P = 0.888).
DSA + patients who experienced AMR had significantly lower 5-years graft survival (68.8%).
In 34 DSA+ patients that remain with a functioning graft, estimated GFR at the last visit was 37.0 ± 19.0 ml/min and 53.8 ± 17.1 ml/min in patients with and without AMR occurrence, respectively (P = 0.028).
Study Discussion
In this study ,transplanted with a negative CDC crossmatch in peak and current sera, the presence of preformed DSA was clearly associated AMR occurrence.
Prevalence of AMR at 1-year post-transplant in our patients with DSA was high (35.0%). Lefaucher et al. demonstrated 32.3% prevalence for AMR in nondesensitized patients with DSA.{http://refhub.elsevier.com/S0966-3274(15)00003-9/rf0030}. Others have shown a 55% incidence of AMR at 200-days posttransplant, but these investigators performed protocol biopsies, thus reporting both clinical and subclinical AMR episodes [http://refhub.elsevier.com/S0966-3274(15)00003-9/rf0015]. A majority (55.0%) of patients in this study with DSA received induction with ATG. In a cohort of ATG-induced patients, AMR-free survival at 1-year was 65.1% for patients with DSA [http://refhub.elsevier.com/S0966-3274(15)00003-9/rf0085].
Study results also show a significantly reduced 5-years death- censured graft survival in patients with DSA (84.8% versus 94.9%, P = 0.006). Amico et al. reported an overall lower 5-year censored graft survival (76.5% and 89% in patients with and without DSA, respectively), possibly related with a higher incidence of clinical acute rejection (33.4%) in comparison to our cohort (13.4%)[http://refhub.elsevier.com/S0966-3274(15)00003-9/rf0015]
This group used no ATG induction, with 59% of patients receiving no induction therapy. A similar kidney graft survival of 86.7% and 94.3% in DSA+ and DSA− patients respectively, but with a shorter follow-up (3-years), was described in a cohort of transplanted patients with a negative CDC and T-cell flow crossmatches, induced with alemtuzumab and under tacrolimus as maintenance monotherapy [http://refhub.elsevier.com/S0966-3274(15)00003-9/rf0045]. This somewhat inferior graft survival, given the shorter follow-up period, in comparison with this study results may be explained by the differences in immunosuppression strategies.
The study found no adverse effect on graft outcomes associated with the presence of non-DSA anti-HLA antibodies in comparison with non- sensitized patients, as others have reported {http://refhub.elsevier.com/S0966-3274(15)00003-9/rf0020}{http://refhub.elsevier.com/S0966-3274(15)00003-9/rf0090}
DSA + AMR + patients remained with DSA detectable 1- year posttransplant more frequently and had lower estimated GFR at the last visit than DSA + AMR − ones. These results lead to speculate that chronic antibody-mediated lesions evolved more frequently in DSA + AMR + patients. Loupy et al.(http://refhub.elsevier.com/S0966-3274(15)00003-9/rf0095) showed, in a comparative study of two desensitization protocols in patients with preformed DSA, that persistence of DSA at 1-year was accompanied by lower estimated GFR and higher histological scores for transplant glomerulopathy and chronic AMR, both ominous signs for graft survival [http://refhub.elsevier.com/S0966-3274(15)00003-9/rf0100,http://refhub.elsevier.com/S0966-3274(15)00003-9/rf0105%5D.
The study found that DSA + patients receiving ATG induction in comparison with basiliximab showed a trend for developing less AMR (P = 0.072).
The use of ATG and intravenous immunoglobulin induction treatment in patients with DSA (below 5000 MFI) has been shown to allow for successful kidney transplantation with similar rejection rates and graft survival between DSA + and DSA − patients [http://refhub.elsevier.com/S0966-3274(15)00003-9/rf0110].
DSA number and strength correlated significantly with AMR. In particular, study demonstrated that only DSA MFI over 3000 were associated with higher incidence of AMR and using ROC curves we were able to detect hDSA MFI threshold level (4900) that presented good sensitivity (85.7%) and sDSA MFI threshold level (11,000) that had good specificity (92.3%), thus complementing each other for an improved AMR prediction. Considering the strongest likelihood ratios from study results, patients with AMR had a probability 7.4 times higher of having a sDSA MFI of N11,000; instead, patients without AMR had a probability 5 times higher of having a hDSA MFI of b4900.
Interestingly, others have shown that DSA MFI values below 3000 compared to higher MFI values were associated with improved 5-year death-censored graft survival (respectively, 86.7% and 54.6%)
First limitation; the lack of available information about flow cytometry crossmatch results may affect its clinical application, particularly in highly sensitized patients, in whom the close analysis of both flow cytometry and virtual crossmatch is important for their management.
Second limitation; no protocol biopsies were undertaken in this cohort.
Finally, as donor specificity of anti-HLA-DQ was not determined at the allelic level in every case (i.e., when the likelihood was ≥90%), rare false assignments as DSA cannot be excluded.
In our practice, it was noticed the strong association between DSA+ and occurrence of AMR, just an observations need to be studied in an organized and systematic way.
(DSA) have been shown to increase the risk of antibody-mediated rejection (AMR) and have a deleterious effect on kidney graft survival, even in the presence of a negative cross matches . Some have reported that low level DSA exert no effect on kidney grafts outcomes . DSAs effects depend also on the their characteristics as HLA class , number , and strength as read by mean fluorescence intensity (MFI). Several studies showed that the occurrence of AMR associated with DSA jeopardizes kidney graft survival . DSA positive patients without AMR having similar survival as compared with those without DSA . This contradictory effect may result from the DSA ability to mediate endothelial injury through complement-dependent and independent mechanisms, mainly through acute and/or chronic AMR, but also, under certain conditions, to promote graft accommodation through up-regulation of complement regulatory or cytoprotective
proteins . Ultimately, DSA main effect will depend on its specificity
and strength. .
This research is designed to study the impact of preformed DSA on kidney graft . 462 patients that received a kidney graft are enrolled in the study . Luminex screening assay is used to determine the presence of anti-human leukocyte antigen (HLA) antibodies and single-antigen bead assay [positive if mean fluorescence intensity (MFI) ≥ 1000] to assign anti-HLA specificities. Anti-HLA antibodies were present in 95 patients (20.6%), but only 40 (8.7%) had DSA. Results of the cross matched negative DSA patients were analyzed . anti body mediated rejection (AMR) at 1-year was found to be higher in patients with DSA (35.0%) than in those without them . DSA with a MFI of N3000 were significantly associated with AMR occurrence. Five-year censored graft survival was lower in DSA positive patient ,although survival was only reduced in those who have AMR .
In my practice ,I notice that, there is increased in the frequency of AMR and chronic allograft nephropathy in DSA positive compared to negative group of patient . also the graft survival is decreased . we a study to confirm this .
HLA donor specific antibodies (DSA) presence increase the risk of antibody-mediated rejection (AMR) and affects kidney graft survival.
The clinical importance of DSA detected by SAB has been the matter of a debate.
The noticing of DSA characteristics associated with AMR occurrence have important clinical aspects for highly sensitized patient management, immunosuppressive strategy and kidney graft recipients’ posttransplant follow-up
This study addresses the relation between DSA and kidney transplantation outcomes in patients with negative CDC
The current study included 517 patients whom underwent renal transplantation with 6 consecutive years and followed them
Pre-transplant anti-HLA IgG antibodies were tested by multiplex microsphere based on Luminex Xmap Technology and SAB was done for positive cases
Donor HLA typing of HLA A ,B DR through PCR by Specific sequence primer and DQ was typed excluding DP and C due to lack of the needed materials for that ; all these data was virtualy cross matched.
The CDC crossmatch results were considered positive when cell death exceeded 20%.
DSA strength depended on MFI of one SAB.
Induction therapy was used in 413 patients, with an anti-IL-2 receptor monoclonal antibody (Basiliximab ) or a polyclonal anti-thymocyte globulin.
All enrolled recipients had similar triple maintenance immunosuppression, consisting of oral tacrolimus , mycophenolate mofetil (MMF) and methylprednisolone (MP)/prednisolone.
Acute rejection was more common in HLA+ DSA+ patients (37.5%) in comparison with HLA+ DSA− (14.5%) and HLA− patients (10.6%). This difference resulted from the high incidence of antibody-mediated rejection in HLA+ DSA+ patients, while acute cellular rejection occurrence was similar between groups.
Incidence of AMR in DSA+ graft recipients from a deceased donor was 33.3% and 50% in living donor graft recipients , with no significant difference in the number of HLA mismatches between them.
DSA+ patients with AMR presented a higher number of DSA beads and stronger DSA.
The presence of preformed DSA was associated AMR even in negative CDC crossmatch transplants . Prevalence of AMR at 1-year posttransplant in the studied patients with DSA was high.
There was close association between AMR in DSA+ patients and poor graft survival.
They found that DSA+ patients receiving ATG induction in comparison with basiliximab showed a trend for developing less AMR.
No difference in AMR incidence was found between patients transplanted without any induction and those receiving basiliximab or daclizumab,
DSA number and strength correlated significantly with AMR and DSA MFI over 3000 were associated with higher incidence of AMR
The pitfalls of the study was that a positive flow crossmatch, in the absence of DSA,had an effect on kidney graft outcomes ,no protocol biopsies were undertaken in the study . Histological screening is crucial in the management HLA-incompatible kidney transplantation, and, since 2013, they started to perform it.
The limitations of SAB assay and their reported MFI values should be considered while interpreting the results. As donor specificity of anti-HLA-DQ was not determined at the allelic level in every case rare false assignments as DSA cannot be excluded.
this article was a review of a study of the relationship of preformed DSA and the outcome of renal transplants in terms of incidence of anti-body mediated rejection and the overall graft survival. this study was carried out over a period of 5 years, from 2007 to 2012, for a 462 ESRD patients treated with renal transplantation.
preformed DSA was detected by Luminex assay but it was -ve by CDC. the clinical relevance of this type of DSA is always a matter of debate as it is less potent than DSA with +ve CDC and also not necessarily associated with worse graft outcome.
the study declared the following :
a- DSA was +ve in 20 % of all included patients but only 8.7 % of them developed
anti-body mediated rejection (AMR) at 1 year.
b- only DSA with MFI of more than 3000, were associated with AMR.
c- AMR rate was associated with reduced 5 year graft survival.
d- 5 year graft survival in DSA +ve patients without AMR, was almost similar to
those patients with -ve DSA.
e- ATG induction in DSA+ve patients decreased the incidence of AMR to be
similar to that in DSA-ve patients.
2- Please reflect on your practice if possible.
of course, I will adhere to the conclusion of this study regarding ATG induction in those with DSA +ve patients then to monitor the development and titre of these DSA post-transplantation.
DSA have been shown to increase the risk of antibody-mediated rejection and have a deleterious effect on kidney graft survival, even in the presence of a negative complement-dependent cytotoxicity and/or flow cytometry crossmatches. SAB assay is much more sensitive.
The study aim was to analyse the relationship between preformed DSA and kidney graft outcomes, in the presence of a negative CDC crossmatch.
517 consecutive adult patients were investigated.
All patients were transplanted with a negative pretransplant T- and B-lymphocyte CDC crossmatch in current and peak sera.
Screening for HLA antibody was done by Luminex Xmap.
To determinate the specificity of the
HLA antibodies, single-antigen bead assays were performed in patients with a positive screening and a cut-off for a positive reaction were set in MFI value of ≥1000.
Samples of all deceased donors were routinely typed before recipient selection in loci HLA-A*, B* and DRB1. HLA-DQ antigens with strong linkage disequilibrium were primarily inferred from the
HLA-DR antigens. Patients with only anti-HLA-C and/or -DP antibodies were not considered in the analysis, as no donor typing at these loci was available. Induction protocol and maintenance immunosuppression
Induction therapy was used in 413 patients (89.4%)-
Basiliximab 20 mg twice at day 0 and 4 or
polyclonal anti-thymocyte globulin 3 mg/kg for 5–7 days).
ATG was primarily used in allo-sensitized patients or in the presence of high number of HLA mismatches. Triple maintenance immunosuppression-
Tacrolimus at a dose of 0.1–0.15 mg/kg/day, and the dose was adjusted to maintain a trough level in whole blood between 8 and 12 ng/ml during the first month postoperatively, between 7 and 10 ng/ml during 2–3 months after transplant and between 5 and 8 ng/ml thereafter
Mycophenolate mofetil started at a dose of 2000 mg/day, with the dose decreasing to 1000–1500 mg/day during the first month postoperatively, depending on white blood cells count.
Methylprednisolone was administered intravenously at doses of 500, 250 and 125 mg/day on the day of transplantation, days 1–2 and days 3–4 after the operation, respectively.
Oral prednisolone was started on day 5 after the operation at the dose of 20 mg, being then tapered to 5–10 mg/day within 2–3 months after transplant.
Living donor recipients (n = 89) were prescribed Tacrolimus and MMF 7 days before transplant. No desensitization protocol was carried out in these patients.
The presence of preformed DSA was clearly associated with AMR occurrence. Prevalence of AMR at 1-year posttransplant in our patients with DSA was high (35.0%). Significantly reduced 5-years death censured graft survival in patients with DSA. Found no adverse effect on graft outcomes associated with the presence of non-DSA anti-HLA antibodies in comparison with non-sensitized patients. y DSA+ AMR+ patients had a significantly reduced 5-year graft survival (68.8%); DSA+ AMR− patients (96.0%) had similar graft survival in comparison with DSA− patients (94.9%). Persistence of DSA at 1-year is accompanied by lower estimated GFR and higher histological scores for transplant glomerulopathy and chronic AMR. DSA+ patients receiving ATG induction in comparison with basiliximab showed a trend for developing less AMR. DSA HLA class was not associated with AMR , In contrast, DSA number and strength correlated significantly with AMR. Patients with AMR had a probability 7.4 times higher of having a sDSA MFI of >11,000; instead, patients without AMR had a probability 5 times higher of having a Hdsa MFI of <4900. Limitations –
lack of available information about flow cytometry crossmatch results may affect its clinical application, particularly in highly sensitized patients, in whom the close analysis of both flow cytometry and virtual crossmatch is important for their management.
No protocol biopsies done.
Known limitations of SAB assay and their reported MFI values.
Rare false assignments as DSA cannot be excluded. Conclusion –
Close relationship between the presence of preformed DSA and an increase incidence of AMR in kidney transplantation, with subsequent adverse impact on graft survival.
Anti-HLA antibodies screening and DSA detection by solid-phase assays are valuable tools, together with cell-based crossmatches.
Please reflect on your practice if possible.
Only CDC crossmatch used to be done and HLA typing.
The study explores relation of preformed DSA and incidence of ABMR with its effect on renal transplant outcome. In 5 years duration ,462 transplanted patients were included. DSA assay was through LUMINEX positive while CDC was negative .It was found that:
. 8.7% of DSA positive (20% of all included patients) developed ABMR at 1 year..
. ABMR occurred only with MFI more than 3000
.ABMR rate was correlated with decreased 5 year graft survival.. Without ABMR graft survival incidence was comparable in both positive and negative DSA.
.induction with ATG in DSA positive patients made ABMR incidence similar to negative DSA patients.
Preformed HLA donor specific antibodies have been shown to increase the risk of antibody mediated rejection and have a deleterious effect on kidney graft survival even in the presence of a negative complement dependent cytotoxicity cross match or flow cytometry cross match
the study showed increased prevalence of AMR at 1 year post-transplant and significantly reduced 5 year death censured graft survival in patient with positive DSA
DSA number and strength correlated significantly with AMR as only DSA MFI over 3000 were associated with higher incidence of AMR
In your own words, summarise this article.
Anti HLA Ab includes (DSA, non DSA) and is screened via solid phase immune essay ( luminex) technique
DSA could be positive in patient serum despite negative CDC cross matching and/or flow cytometry crossmatches , and was found to be associated with kidney graft loss and poor impact on the overall graft survival, in relation with AMR .However other DSA positive recipients did not.
What is the clinical relevance of DSA? Is positive DSA superior on negative CDC to predict graft rejection and survival? And why not all DSA positive patients develop AMR?
In order to answer these questions a study was conducted on KTX rciepients with positive anti-HLA IgG antibodies DSA via Luminex technique and negative CDC , HLA antibody detection was done by single antigen bead (SAB) assay, the strength was based on the MFI of one SAB.
Patients with high number of HLA mismatches underwent Induction protocol immunosuppression with anti-IL-2 receptor monoclonal antibody (Basiliximab) and ( ATG) .
All enrolled recipients had similar triple maintenance immunosuppression, consisting of oral tacrolimus (FK-506), mycophenolate mofetil (MMF) and methylprednisolone (MP)/prednisolone.
follow up of the graft outcome was done
results showed that DSA main effect will depend on its specificity and strength [3]. Associated with AMR and poor graft survival within five years. DSA ability to mediate endothelial injury through complement-dependent and independent mechanisms, mainly through acute and/or chronic AMR [2].
Hence it is of crucial value to identify DSA characteristics as HLA class, number , and strength by mean fluorescence intensity (MFI) to help improve immunologicalrisk stratification of sensitized patients and their immunesuppression protocol. [1]
Please reflect on your practice if possible
Pretransplant assessment of recipients with Anti-HLA antibodies screening and DSA detection by solid-phase assays hand to hand with the cell based CMX,helps careful selection of highly sensitized patients and stratification of the proper desentisation protocol to eliminate the expected AMR , and whether the patient would benefit from protocol biopsies.
References:
1] Mohan S, Palanisamy A, Tsapepas D, Tanriover B, Crew RJ, Dube G, et al. Donorspecific antibodies adversely affect kidney allograft outcomes. J Am Soc Nephrol 2012;23(12):2061–71.
(2) Zhang X, Reed EF. Effect of antibodies on endothelium. Am J Transplant 2009;9(11): 2459–65.
(3] Jindra PT, Zhang X, Mulder A, et al. Anti-HLA antibodies can induce endothelial cell survival or proliferation depending on their concentration. Transplantation 2006; 82(1 Suppl.):S33–5.
Cell mediated rejection vs Antibody mediated rejection.
In waiting list patients we test anti-HLA IgG antibodies every three month by luminex. Positive positive underwent SAB to determine specifity. Cut-off for positive reactions if MFI > 1000.
Typings for HLA-A* , B* , DRB1* done for all deceased donors by PCR before donor selection .
We performed virtual cross match , we consider DSA with high MFI.
CDC carried out and considered positive when cell death exceeded 20%.
Induction therapy done either by basiliximab anti-IL2 receptor monoclonal (anti CD25) 20 mg twice at day 0 and day 4. Or ATG (polyclonal antithymocyte globulin) 3 mg/kg 5-7 day which in sensitised patients or high numbers of HLA mismatch , this induction followed by triple maintenance therapy ( oral tacrolimus , MMF and methyl prednisone) FK started with dose 0.1-0.15 mg/kg/d adjusted to maintain trough level between 8-12 ng/ml during first month , between 7-10 ng/ml during 2-3 month and between 5-8 ng/ml thereafter.
MMF started at dose 2000 mg/d then decreased to 1000-1500mg/d during first non postoperative depend on white cell count . Methylprednisone given at dose of 500 , 250 , 125 mg/d on day 1-2 and days 3-4 after operation respectively.we started oral prednisolone on day 5 by 20 mg then tapered to 5-10 mg/d within 2-3 month after transplant.
Delayed graft function: define to those require dialysis in first week post transplant. GFR estimated using MDRD.
Allograft rejection diagnosed by biopsy proven rejection which evaluated by LM and IF staining for C4d which classified according to Banff classification.
Mild acute cellular rejection (ACR grade 1) treated by pulse steroid therapy 500 mg MP for 3days with increasing maintenance immunosuppressive medication.
Other ACR treated with ATG .Patient with AMR treated by 4 session of plasmapheresis, Every other day, after each one IVIG 100 mg/kg then after last plasmapheresis session , patient will receive high dose of IVIG 2g/kg divided in four daily dose followed by one dose of rituximab 375 mg/m2 and similar dose of IVIG 2g/kg repeated after 1 month.
Close association between AMR in DSA and poor graft survival.
Studies shown only DSA positive with AMR has reduced 5 y graft survival. DSA positive with AMR patients had similar graft survival in comparison to DSA negative patients. Although other studies shown reduced graft survival in patients with DSA positive even without AMR.
DSA positive patients who received ATG induction in complications to basiliximab shown less AMR development.
The use of ATG plus IVIG induction therapy in patients with DSA ( below 5000 MFI) give benefits of successful kidney transplantation with similar rejection rate and graft survival with or without DSA.
References:
Mohan S, Palanisamy A, Tsapepas D, Tanriover B, Crew RJ, Dube G, et al. Donorspecific antibodies adversely affect kidney allograft outcomes. J Am Soc Nephrol 2012;23(12):2061–71.
[2] Tait BD, Susal C, Gebel HM, et al. Consensus guidelines on the testing and clinical
management issues associated with HLA and non-HLA antibodies in transplantation.
Transplantation 2013;95(1):19–47.
[3] Amico P, Honger G, Mayr M, et al. Clinical relevance of pretransplant donor-specific
HLA antibodies detected by single-antigen flow-beads. Transplantation 2009;
87(11):1681–8.
[4] Caro-Oleas JL, Gonzalez-Escribano MF, Gonzalez-Roncero FM, Acevedo-Calado MJ,
Cabello-Chaves V, Gentil-Govantes MÁ, et al. Clinical relevance of HLA donorspecific antibodies detected by single antigen assay in kidney transplantation.
Nephrol Dial Transplant 2012;27(3):1231–8.
[5] Lefaucheur C, Loupy A, Hill GS, et al. Preexisting donor-specific HLA antibodies
predict outcome in kidney transplantation. J Am Soc Nephrol 2010;21(8):1398–406.
[6] Lefaucheur C, Suberbielle-Boissel C, Hill GS, et al. Clinical relevance of preformed
HLA donor-specific antibodies in kidney transplantation. Am J Transplant 2008;
8(2):324–31.
[7] Otten HG, Verhaar MC, Borst HP, Hené RJ, van Zuilen AD. Pretransplant donorspecific HLA class-I and -II antibodies are associated with an increased risk for kidney graft failure. Am J Transplant 2012;12(6):1618–23.
To evaluate correlation of preformed DSA with graft outcome, 462 patients were investigated. Among them 95 patients (20.6%) had positive anti-H LA antibodies and 40 (8.7%) had DSA. AMR was significantly higher in patients with DSA.
DSA with MFI >3000 were associated with AMR and more graft loss even with a negative CDC-XM or FCXM. Majority of induction therapy in sensitized patients without AMR was performed with ATG vs Basiliximab.
This study CDC-XM negative 462 patient in one TX center in 5 years .
Induction therapy was used in 413 patients (89.4%),
1. Basiliximab ..
2 ATG ..
Then triple maintenance immunosuppressive therapy
●
367 (79.4%) patients had no anti-HLA
antibodies (HLA−) detectable in pretransplant serum.
A positive anti-HLA antibody
screening was present in 95 (20.6%) patients, with 40 of them having antibodies directed
against donor HLA molecules (HLA+ DSA+), while the remaining 55 patients had not(HLA+ DSA−).
●Acute rejection was particularly more common in HLA+ DSA+ patients (37.5%) in comparison with HLA+
DSA− (14.5%) and HLA− patients (10.6%)
Moreover, acute rejection developed earlier in HLA+ DSA+ patients (median
11 days) than in HLA+ DSA− (median 71 days) and HLA− patients (median 35 days).
●The prevalence of AMR increased significantly with the presence of DSA MFI value
over 3000.
This article showed the relevance of preformed DSAs regarding incidence of AMR and graft survival .
They concluded that there is strong correlation between preformed DSAs and subsequent AMR ,
another interesting conclusion was that patients who had Pre formed DSAs without occurrence of AMR has similar graft survival to Those who don’t have AMR ,
Which means that occurance of the AMR is essential for the adverse outcomes related to DSAs .
They also found that induction using ATG decreases incidence of AMR in patients with Pre formed DSAs .
They studies characteristics of DSAs that included strength and no. And they found strong correlation between them and AMR .
MFI less than 3000 was associated with Lower risk of AMR Than those with more than 5000 MFI .
Limitations of the study include lack of flow cytometry data , no protocol biopsies, limitations regarding SAB assays and MFI values.
DSA be increase the risk of antibody-mediated rejection and have a deleterious effect
on kidney graft survival, even in the presence of a negative CDC) and/or flow cytometry
cross matches.
HLA antibody detection by single antigen bead assay is much more sensitive than cell
based cross matches.
Identification of DSA characteristics associated with AMR occurrence would have
important clinical implications for highly sensitized patient management
,immunosuppressive strategy optimization and kidney graft recipients.
This study, aimed to analyze the relationship between preformed DSA and kidney graft
outcomes, in the presence of a negative CDC crossmatch.
Anti-HLA antibodies screening and specificity:
Pre-transplant anti-HLA IgG antibodies were tested in patient sera collected every 3
months by multiplex microsphere based on Luminex Xmap® Technology
To determinate the specificity of the HLA antibodies, single-antigen bead (SAB) assays
(LabScreen Single Antigen Bead, were performed in patients with a positive screening.
Donor HLA typing and virtual crossmatch:
Samples of all deceased donors were routinely typed before recipient selection in loci
HLA-A*, B* and DRB1* using PCR) amplification with specific sequence primers (SSP;
Olerup SSP®).
With the information of the donor HLA typing, we performed a virtual crossmatch.
In each individual DSA, the strength was based on the MFI of one SAB. In the case of
several DSA against different HLA-antigens, we usually considered the DSA with the
highest MFI; occasionally, we also used the cumulative strength of all DSA by adding the
individual MFI values, as stated in the results.
CDC crossmatch:
carried out using the standard NIH technique, not enhanced with anti-human globulin.
Induction protocol and maintenance immunosuppression:
Induction therapy was used in the study .with an anti-IL-2 receptor monoclonal antibody
(Basiliximab Novartis®, 20 mg twice at day 0 and 4) or a polyclonal anti-thymocyte
globulin (ATG Fresenius®, 3 mg/kg for 5–7 days).
All enrolled recipients had similar triple maintenance immunosuppression, consisting of
oral tacrolimus, mycophenolate mofetil (MMF) and methylprednisolone
(MP)/prednisolone.
Result of the study cohort of kidney graft recipients, transplanted with a negative CDC
crossmatch in peak and current sera, the presence of preformed DSA was clearly
associated AMR occurrence and prevalence, also show a significantly reduced 5-years
death censured graft survival in patients with DSA.
This article studied graft survival in patients with pretransplant DSA and negative CDC.
and found that :
Preformed DSA are associated with poor graft outcome and increased risk of antibody mediated rejection
Studies showed various results about the impact of DSA on graft survival depending on level of DSA, different characteristics of DSA, occurrence of AMR or not and whether the DSA is complement dependent or not, while other studies showed that DSA may have protective effect through promoting graft accommodation.
The study analyzes the impact of DSA with negative CDC on graft outcome and showed that:
preformed DSA with negative CDC was associated with high incidence of AMR and significantly decreased 5 year death censored graft survival
patients with DSA who developed AMR had detectable DSA and had more decline in eGFR than those who didn’t develop AMR
patients with DSA who received ATG as induction had lower rate of AMR than those who received Basiliximab
Only DSA with MFI >3000 were associated with increased incidence of AMR
Limitations:
FCXM results were not available and they are important especially in sensitized patients
Protocol biopsies were not performed
limitations of SAB assay and the reported MFI values
anti-HLA DQ was not detected at allele level
Conclusion:
Preformed DSA associated with increased incidence of AMR and poor graft outcome.
Anti-HLA antibodies screening and DSA detection by solid phase assays are valuable tools to determine DSA strength and characterization which may improve immunological risk stratification of sensitized patients
SUMMARY:
Indroduction :
Performed anti-HLA–AB and its characteristic with impact on organ transplant still need further clarification ,the Anti-HLA AB effect on renal graft depend on its class, intensity and complement binding capacity.
Study aim:
To assess the impact of performed anti-HLA AB on renal transplant outcome and to identify the predictive value of DSA characteristics for AMR occurrence by using single antigen bead Luminex assay with positive MFI cutoff value of 1000 and above.
Results:
They included 462 recipients of KTX with negative CDCXM ,no flow cytometry XM duringthe period between20072012 from single center,thepatients demographics and clinical characteristics were homogenous between the 3 groups ,majority received ATG induction IS (55%) ,and all have similler maintenance IS protocol with triple therapy ( tacrolimus MMF , steroids ).
This study clearly shows the presence of preformed anti HLA AB associated with AMR whic occure in 35% of non-sensitized patients ,its similar to the results others studies , with moreAMR in the first year after transplantation with a significantly reduced 5-years death censured graft survival in patients with DSA with p value (0.006), also they found that only DSA MFI over 3000 were associated with higher incidence of AMR, and 55% of the patients use ATG induction protocol with lower rate of AMR
Also,only DSA+ AMR+ patients had a significantly reduced 5-year graft survival (68.8%); DSA+ AMR− patients (96.0%).
persistence of DSA after 1 years associated with increased risk of chronic antibody-mediated lesions which is more frequently in DSA+ AMR+ group with lower estimated GFR at the last visit than DSA+ AMR− group, the DSA -HLA class was not associated with AMR as in others [3,8] although anti-HLA-DQ was not determined at the allelic level in this study. In fact,they found that the DSA number and strength correlated significantly with the risk of AMR.
Conclusion:
The presence of anti HLA, DSA BY numbers and intensity significantly associated higher risk of AMR with negative impact onthe renal graft survival, DSA characterization has crucial importance in the management of sensitized patients.
Limitations:
No data about the FCXM
No protocol biopsies.
The SAB MFI cutoff value interpretaion.
No donor typing at anti-HLA-C and/or -DP antibodies not available
The anti-HLA-DQ was not determined at the allelic level in every case.
During my practise we use lumenix SAB assay for screening and monitoring of DSAs with MFI cutoff value of 1000 and above , with CDCXM, currnetly no access to FCXM assay , no protocal biopsy.
The aim of this study is to present the effect of DSA on transplanted kidney in patients with different figures of presensetization or not and effect of CDC results and flow cytometry XM whether negative or positive. female with history of pregnancy, history of blood transfusion and previous transplantation all those groups considered as sensitized patients.
The study followed 462 patients underwent kidney transplantation with their immunological tests mainly by luminex test,and single Ag beads assay to evaluate anti HLA-Ab with MFI more than 1000.
Generally most studies showed that there is direct relationship between presence of DSA and incidence of AMR and graft survival but this depends on the multiple factors like HLA class,it’s range and amount of MFI .
The patients that are screened in this study are 462 patients with negative pre transplant B and T cells CDC crossmatch and the patients had been divided into groups according to presence or absence of HLA and DSA with different levels of MFI of more than 1000 and more than 3000.
The results of the study showed the rate of acute rejection is higher in those group with positive of both DSA and HLA and even earlier rejection posttransplant comparing with other group more specifically AMR which is correlated to percentage of DSA and MFI and for most of those patients ATG are used in the induction therapy ,regarding graft survival with prolonged follow up showed that the patients with DSA positive and history of pos6AMR occurrence had poor graft survival for less than 5 years.
the identification of DSA characteristics asso- ciated with AMR occurrence would have important clinical implications for highly sensitized patient management, immunosuppressive strategy optimization and kidney graft recipients’ posttransplant follow-up individualization.
A study done to analyze the relationship between preformed DSA and kidney graft outcomes.462 patients (89%) for the final analysis was taken . All patients were transplanted with a negative pretransplant T- and B-lymphocyte CDC crossmatch in current and peak sera. No data on flow-cytometric crossmatches was available
Anti-HLA antibodies screening and specificity
Pre-transplant anti-HLA IgG antibodies were tested in patient sera collected every 3 months while in the waiting list, by multiplex microsphere based on Luminex Xmap® Technology .The cut-off for positive samples was the Normalized Background (NBG) ratio .To determinate the specificity of the HLA antibodies, single-antigen bead (SAB) were performed in patients with a positive screening. a cut-off for a positive reaction were set in MFI value of ≥1000.
Donor HLA typing and virtual crossmatch
Samples of all deceased donors were routinely typed before recipient selection in loci HLA-A*, B* and DRB1* using polymerase chain reaction (PCR) amplification with specific sequence primers (SSP).HLA-DQ antigens with strong linkage disequilibrium were primarily inferred from the HLA-DR antigens .If a potential donor-specific HLA-DQ antibody was present, HLA-DQ antigens of the donor were verified by SSP DNA-typing .
After donor HLA typing, we performed a virtual crossmatch (virtual XM).
Then CDC crossmatch was carried out using the standard NIH technique, not enhanced with anti-human globulin.
Induction therapy was used in 413 patients (89.4%), with an anti-IL-2 receptor monoclonal antibody (Basiliximab Novartis®, 20 mg twice at day 0 and 4) or a polyclonal anti-thymocyte globulin (ATG Fresenius®, 3 mg/kg for 5–7 days).
Rejection diagnosis and treatment
Allograft rejection was defined as biopsy proven rejection (specimens were evaluated by light microscopy and immunofluorescence staining for C4d) and classified according to Banff classification as updated in 2013. Mild acute cellular rejection (ACR Banff grade I) was treated with pulse steroids (500 mg MP for 3 days) and increased maintenance immunosuppression. All other ACRs were treated with ATG. Patients with AMR were treated with plasmapheresis every other day and IvIg 100 mg/kg after each session.
followed by one dose of rituximab (375 mg/m2); a similar dose of IvIg (2 g/kg) was repeated 1 month later.
They found that the the presence of preformed DSA was clearly associated AMR occurrence.
With poor graft and patients survival at 1 and 5 year .
They found no adverse effect on graft outcomes associated with the presence of non-DSA anti-HLA antibodies in comparison with non- sensitized patients, as others have reported
they found a close association between AMR in DSA+ patients and poor graft survival.
DSA + AMR + patients had a significantly reduced 5-year graft survival .DSA + AMR − patients had similar graft survival in comparison with DSA− patients .
chronic antibody-mediated lesions evolved more frequently in DSA + AMR + patients.
Regarding immunosuppressive therapy , they found that DSA + patients receiving ATG induction in comparison with basiliximab showed a trend for developing less AMR .
In contrary ,Interestingly, in a cohort of 67 kidney graft recipients with preformed DSA, no difference in AMR incidence was found between patients transplanted without any induction and those receiving basiliximab or daclizumab .
In this study DSA HLA class was not associated with AMR as in others.
In contrast, DSA number and strength correlated significantly with AMR.
They demonstrated that only DSA MFI over 3000 were asso- ciated with higher incidence of AMR , and sDSA MFI threshold level (11,000) that had good specificity.
Limitations of this study :
First, the lack of available information about flow cytometry crossmatch results may affect its clinical application,
Second, no protocol biopsies were undertaken in this cohort
Third, the known limitations of SAB assay and their reported MFI values should be considered while interpreting the results.
Finally, as donor specificity of anti-HLA-DQ was not determined at the allelic level in every case (i.e., when the likelihood was ≥90%), rare false assignments as DSA cannot be excluded.
A detailed DSA characterization has a pivotal role in the management of HLA- sensitized patients, improving our ability to reduce the burden of AMR.
-anti HLA ab in 95 patient of 462 only 40 patient had DSA develop AMR 35% more than those without
-DSA with MFI>3000 significantly associated with AMR
-DSA increase risk of AMR and decrease graft survival even in negative CDC
-presence of DSA not mean no hope for kidney transplantation but it mean that recepient need aggressive induction and maintenence immunosuppresant
-in this study we search for relationship between DSA in negative CDC and AMR
-462 patient with negative CDC (2007-2012)
-anti HLA ab detected by SAB (DSA consider positive if MFI >1000
-donor HLA typing & virsual crossmatch for deceased donor done to detect DSA
-induction done with IL2 antagonist or ATG (in highly senstized patient or high HLA mismatch
-maintenence with tacrolimus, MMF & methylpredinsilone
-follow up patient till end of 2013
-rejection
*if mild ACR use pulse steroid & augment maintenace
*if more serious use ATG
*in AMR 4 session of plasmaphariese every other day with small dose IVIG after each one then after 4th session give high dose IVIG devided into 4 doses then retuximab then IVIG again after 1 month
-data gathering & comparison of MFI value in patient with DSA pre & 12 month after transplantation
-graft survival ,DGF ,episodes oof AMR , time of rejection , acute rejection & death with functioning graft all this data in comparison between DSA + & others
*DGF 25% in +ve DSA &19% in -ve
*acute rejection 37% vs 10%
*AMR 35% vs 0.8%
*days for rejection 11 vs 35
*graft failed 12% vs 3%
*death with functioning graft 2.5% vs 3.3%
finally this cohort study show that
1-presence of DSA associated with high risk of AMR (MFI >3000)
2-significance reduce in graft survival
*I saw patient with MFI >4000 and AMR occur after 10 months treated with IVIG & plasmaphariese then after 6 month occur again but patient refuse ttt
his maintenance immunosuppressant (tac,mmf,methylpredinisilone) & induction had done with ATG
Analysis of preformed donor specific anti-HLA antibodies characteristics for prediction if antibody- mediated rejection in Kidney transplantation
Preformed HLA DSAs have shown to increase the risk of AMR and have deleterious effect on kidney graft survival even in the presence of negative CDCXM and FCXM
Usage of SAB to define DSA has been subject of debate. Differences in opinions regarding the effect of low level of DSA in kidney graft outcomes. Some studies has shown DSA characteristics, HLA class, number and strength (MFI) associated with kidney outcome
This study aimed to analyse the relationship between performed DSA and kidney graft outcomes in DCDXM and FCXM negative patients.
AMR
• Prevalance at 1 year -35%
• Compare with Lefaucher- 32.3%
• Other studies 55% incidence at 200 days but with protocol biopsies
• 55% of the current study patients received ATG – AMR free survival -65%
Death censored graft survival
• Reduced ( 84.8%) with DSA vs non -DSA( 94.9%)
• Amico et al- overall reduced 5 year censored graft survival ( 76.5% vs 89%)
o Possibly higher incidence of clinical AR
o No ATG induction and 59% no induction
• Willicombe et al – kidney graft survival DSA + 86.7% vs DSA – 94.3% in 3 years but induced with alemtuzumab and Tac mono therapy
Association AMR in DSA + and poor graft function
• Only DSA+AMR+ showed reduced 5 year graft survival 68.8%
• DSA+AMR- – 96%, DSA- -94.9%
• Other studies showed lower graft function in DSA+ patients with or without AMR
• DSA + AMR+ : remained detectable DSA at 1 year – had lower eGFR than DSA+AMR-
DSA characteristics and IS
• DSA+ :received ATG had lesser AMR than Basiliximab ( p=
• DSA-: no differences in AMR Basiliximab/Daclizumab vs no induction
• MFI below 5000 and DSA+: usage of ATG and IVIG had similar graft survival among DSA+ and DSA-
• DSA class is not associated with AMR in surenthiran study
• DSA number and strength (MFI) correlate with AMR
o MFI >3000 – higher incidence of AMR
-This study, aimed to analyze the relationship between performed DSA and renal allograft outcome, in presence of negative CDC crossmatch which is not enhanced with anti-human globulin.
-Performed human leukocyte antigen (HLA) donor-specific antibodies (DSA) increase the risk of antibody-mediated rejection (AMR) and decrease renal allograft survival.
-DSA detection by single antigen bead (SAB) is the most sensitive than cell-based cross matches.
-517 patients (2007-2012) were involved in this study,55 patients were excluded, leaving 462 for final analysis.
-All patients were pretransplant negative T and B lymphocyte CDC crossmatch.
-Pretransplant anti-HLA IgG antibodies were tested in patient sera.SAB was performed in patients with positive screening.
-Deceased donors were typed in loci HLA-A, B, DRB1. High-resolution typing was performed when it was necessary to establish whether the anti-HLA antibodies were DSA.
-Virtual crossmatch was done.
-Induction therapy was used in 413 with Basiliximab or ATG.
-All patients had similar triple maintenance immunosuppressive (Tacrolimus, MMF, prednisolone)
-Living donor-recipient (89) started tacrolimus and MMF 7 days before transplantation, and no desensitization was carried out in these patients.
-Allograft rejection was defined as biopsy-proven rejection and classified according to Banff classification as an update in 2013.
-The study follow-up is about 39.6months.
-AMR was common in the HLA+DSA+ group especially in deceased donors and common with DSA MFI value ˃3000 while acute cellular rejection occurrence was similar between groups(DSA+ and DSA-).
-After one -year eGFR was similar between groups.
– DSA +ve AMR +ve had a significant reduce 5-year graft survival and lower eGFR.
-Death-censored kidney graft survival at 5-year in DSA +ve without AMR was similar to DSA -ve patients.
-According to this study, DSA +ve patients receiving ATG induction have a trend for developing less AMR.
Limitations of the study :
1.No information about flow cytometry crossmatch results.
2.No protocol biopsies were undertaken.
3. Limitation of the SAB assay and their reported MFI values should be considered while interpreting our results.
4. Donor specificity of anti-HLA-DQ was not determined at the allelic level in every case.
-In my practice, yes, there is an association between DSA and AMR but we have no studies for that.
preformed ab ( sensitization ) can be formed by exposure to
1- previous transplant
2- pregnancy
3- blood transfusion
preformed ab is closely associated with graft loss through ab mediated rejection
ab types are
transplant outcome with negative cdc xm and negative anti HLA ab is beter than negative CDC XM and +ve ANTI HLA ab .. furthur more transplant outcome is beter in HLA ab +VE / DSA negative than HLA ab +ve / DSA +VE patient .
DSA associated with graft outcome through
1- intensity oF hDSA ab ( highest ab level – MFI MORE THAN 3000 ) or sDSA ( SUM OF DSA ab of more than – 11000 )
2- multiple DSA target .
there is no difference in graft outcome according HLA class DSA .
there is no clear cut of negative DSA as it depend on the lab –
The purpose of this research, which included 462 transplant patients, was to determine if premade DSA had any influence on graft survival.
When DSA was performed, it was shown to be related to an increase in AMR in individuals who had negative CDC crossmatch.
patients with positive DSA had a lower graft survival rate. MFI greater than 3000 was found to be associated with a statistical significance.
Patient’s with a positive DSA test had an AMR of more than 35 %. Following 5 years of follow-up.
DSAs that have been performed have been demonstrated to increase the probability of antibody-mediated rejection (AMR) and have a detrimental influence on long-term graft survival.
Single antigen bead (SAB) assays for HLA antibody detection are substantially more sensitive than cell-based cross matches, allowing for the determination of antibody specificity.
Limitation:
-The group did not undergo any protocol biopsies.
– SAB assay limitations and reported MFI values
-Anti-HLA-DQ donor specificity was unknown
-The lack of data on flow cytometry cross-match findings.
Impact on clinical practice:
Patients with preformed DSA need close monitoring after transplantation. and the higher the titre of preformed DSA, the worse the graft outcome and the risk of rejection.
Impact of preformed DSA characteristics on kidney graft outcomes.
*Preformed human leukocyte antigen (HLA) donor specific antibodies (DSA) have been shown to increase the risk of antibody-mediated rejec- tion (AMR) and have a deleterious effect on kidney graft survival, even in the presence of a negative complement-dependent cytotoxicity (CDC) and/or flow cytometry crossmatches .
*Some have reported that low level DSA exert no effect on kidney grafts outcomes , while others have associated DSA characteristics as HLA class , number , and strength as read by mean fluorescence intensity (MFI) with kidney graft outcomes.
* Several studies showed that it is the occurrence of AMR associated with DSA that jeopardizes kidney graft survival, with DSA positive patients without AMR having similar survival as compared with those without DSA .This contradictory effect may result from the DSA ability to mediate endothelial injury through complement-dependent and independent mechanisms, mainly through acute and/or chronic AMR, but also, under certain conditions, to promote graft accommoda- tion through up-regulation of complement regulatory or cytoprotective proteins .
*Aimof the study: aimed to analyze the relationship between preformed DSA and kidney graft outcomes, in the presence of a negative CDC crossmatch. Furthermore, we sought to explore the predictive value of DSA characteristics for AMR occurrence.
*Methods: 462 patients that received a kidney graft in our unit, between 2007 and 2012, pre-transplant sera were analyzed by Luminex screening assay to determine the presence of anti-human leukocyte antigen (HLA) antibodies and single-antigen bead assay [positive if mean fluorescence intensity (MFI) ≥ 1000] to assign anti-HLA specificities.
*Results:
1-Anti-HLA antibodies were present in 95 patients (20.6%), but only 40 (8.7%) had DSA.
2-Antibody- mediated rejection (AMR) at 1-year was significantly higher in patients with DSA (35.0%) than in those without them (0.9%) .
3- Only DSA with a MFI of >3000 were significantly associated with AMR occurrence ,MFI of >4900 in the highest DSA bead had a high sensitivity (85.7%) and that the sum of all DSA beads MFI >11,000 had a high specificity (92.3%) for AMR prediction.
4-Anti-thymocyte globulin versus basiliximab induction was more frequent in DSA + AMR − (65.4%) versus DSA + AMR + (34.6%) patients . Five-year censored graft survival was lower in DSA+ than in DSA− patients (respectively, 84.8% versus 94.9%,
5-survival was only reduced in DSA+ AMR+ (68.8%) versus DSA+ AMR− (96.0%) patients .
*Conclusion:
Preformed DSA is associated with kidney graft loss, in relation with AMR occurrence. DSA strength may be used to improve immunological risk stratification of sensitized patients and their clinical management.
Good study as they do correlation between DSA and AMR ,Survival,Induction therapy.
Strong point also :using χ2 test in statistical analysis of variable data
Our practice : According to MFI we choose Pt with DSA positive but low MFI ,no updated registry to notice statistically significant AMR or not ,,usually we go with ATG induction in those cases
Preformed donor specific antibodies (DSA) have been shown to increase the risk of antibody-mediated rejection (AMR) and have a negative impact on long term graft survival. HLA antibody detection by single antigen bead (SAB) assay is much more sensitive than cell-based crossmatches, allowing for the definition of antibody specificity.
DSA characteristics that determine their effect on long term graft survival depends on HLA class (Antibodies against class I or class II), number and strength of DSAs.
This retrospective study analysed anti HLA results from 462 patients that received a kidney graft between 2007 and 2012.The aim of this study to determine the impact of preformed DSA (detected by Luminex assays with negative CDC) characteristics on kidney graft outcomes. The positive value if mean fluorescence intensity (MFI) ≥ 1000.
Results of the study revealed: The presence of preformed DSA was clearly associated AMR occurrence. Similar results obtained in negative CDC crossmatch transplants. Prevalence of AMR at 1-year posttransplant in patients with DSA was high (35.0%).
The study limitations:
1-lack of available information about flow cytometry crossmatch results.
2-No protocol biopsies were done.
3-The limitations of SAB assay and their reported MFI values has to be considered when interpreting the results.
4-The donor specificity of anti-HLA-DQ was not determined at the allelic level in every case.
Thank you for mentioning the limitations in your own words
What about your own practice
1. In your own words, summarise this article.
Analysis of preformed donor-specific anti-HLA antibodies characteristics for prediction of antibody-mediated rejection in kidney transplantation
The study involved analysis of kidney transplant recipients in a single center performed over 5 years vis a vis the presence of prior presence of donor specific antibodies (DSA) and their effects on the graft prognosis.
DSA, even in presence of negative CDCXM, have shown to be associated with antibody mediated rejection (AMR) and poor graft survival and this study tried to document effect of pre-formed antibodies, in such patients, on the graft with respect to type of antibodies as well as strength of antibodies.
A total of 462 patients were evaluated and out of them 95 (20%) had pre-formed antibodies with 42% of those having DSAs. The patients were divided into 3 groups: patients with DSAs, patients with non-DSA antibodies, and patients without any pre-formed antibodies.
Acute rejection (overall as well as episodes of AMR alone) was more common and developed earlier in patients with DSAs than those with non-DSA antibodies. AMR prevalence correlated with MFI of DSA with 0% in patients with MFI less than 3000 and 60% in patients with MFI more than 6000. A single antibody MFI of >4900 and total MFI strength of >11000 was associated with higher risk of AMR. DSA positive patients who received ATG as induction had lower AMR as compared to those receiving basiliximab as induction. In the study, no correlation could be established between DSA class and AMR.
Although the eGFR at one year was similar in the 3 groups, presence of DSA was associated with reduced 5 year graft survival (85%) as compared to those with non-DSA antibodies (89%) and patients without antibodies (94%). Death censored graft survival at 5 years in patients with DSA experiencing AMR was lower (68.8%) than those who did not develop AMR (96%) as well as patients without DSA (96%).
The study had its limitations including non-availability of flowcytometry crossmatch and protocol biopsies as well as donor specificity of anti HLA DQ (as donor HLA DQ typing was not done).
The study concluded that pre-formed DSAs are associated with increased incidence of AMR and poor graft survival.
2. Please reflect on your practice if possible.
In patients with prior history of sensitization, SAB for detecting preformed antibodies is performed in our unit and in patients with pre-formed antibodies, we give induction using ATG with triple drug immunosuppression (Tacrolimus, MMF and steroids) and follow post transplant antibody levels to take pre-emptive action prior to clinical deterioration for better management of renal transplant recipients.
This a retrospective study involving 462 renal transplant recipients from 2007-2012 where pre-transplant sera were analyzed by Luminex for anti-HLA antibodies and SAB assay. MFI> 1000 was considered as positive.
The aim of the study is to analyze the relationship between preformed DSA and kidney graft outcomes in the presence of negative CDC crossmatch.
Presence of preformed DSA even with negative cross-match was clearly associated with AMR occurrence.
AMR at 1-year post-Tx was higher in those with positive DSA (35%), however it was very low in those with negative DSA (0.9%).
AMR-free survival at 1-year was 65% for patients with DSA+ve who had induction immunosuppression with ATG.
5-year graft survival was lower in those with +ve DSA/+ve AMR (68.8%) compared to those with +ve DSA/-ve AMR (96%).
5-year graft survival was 84.8% in those with DSA +ve, in comparison to 94% in those with negative DSA.
DSAs increase AMR and lower graft survival even with negative cross-match.
Results:
Acute rejection was common in HLA+DSA+ patients (37.5%), HLA +DSA- (14.5%), HLA-ve(10.6%).
AMR was higher in DSA+ve/HLA +ve (35%).
Acute cellular rejection was similar between different groups.
Acute rejection developed earlier in DSA+ve/HLA+ve (median 11 days), HLA+ve/DSA-ve (median 71 days), HLA -ve (median 35 days).
The higher DSA MFI(>10,000), the earlier of AMR ( first 11 days).
Study limitations:
1- Lack of information about flow cytometry cross-match results may affect its clinical application.
2- No protocol biopsy was undertaken in this cohort.
3- Limitations of SAB assay and their reported MFI values.
4- Donor specificity of anti-HLA-DQ was not determined at the allelic level in every case.
Clinical application:
1- Serial monitoring of DSA especially in highly sensitized patients or previously sensitized ones.
2- Detailed DSA characterization has an important role in the management of HLA-sensitized patients, improving our ability to reduce the burden of AMR.
This study aimed to analyze the relationship between preformed DSA and kidney graft outcomes in the presence of negative CDC crossmatch.
The study involve 462 patients who received kidney transplantation with negative CDC XM( both T & B cells).Anti HLA antibodies tested by luminex were found in 20% of patients and only 8% had DSA. No data available about FCXM. Ethical approval gained from the institutional review board at centro hospitalar do porto.
The study found that ABMR at 1 year was higher in patients with DSA than in those without. DSAs with MFI> 3000 were associated with ABMR occurrence. ATG induction was more frequent in DSA+ ABMR – patients. five year graft survival was lower in DSA+ patients, although survival was only reduced in DSA+ ABMR+ patients as compared to DSA+ AMR- patients. DSA HLA class was not associated with ABMR.
Limitation of this study:
Preformed HLA antigens specific antibodies (DSA) was shown to have a negative effect on graft survival. some studies showed that low levels may be of low importance. this is dependent on both DSA level and specificity. In this study performed DSA in case of negative CDC-CXM was evaluated in terms of relation to graft survival in addition to its predictive value for ABMR.
In this 5 years study (2007-2012), 462 (out of 517) were included. pateints with LUMINEX MFI>1000 were accepted . All patients had CDC negative CMX. serial assays each 3 months were performed. SBA assays added when when indicated.. Flow cytometry data was not available. Low resolution mach for HLA-A*, -B* and DRB1 using PCR amplification and SSP low resolution used. High resolution was evaluated in special cased. . 4 patients were excluded because of HLA-c positive or negative ppi.
Virtual CMX was performed using the results of HLA typing.
ATG was used in highly sensetized patients and or basiliximab (total 413/462; %89) as induction therapy. maintenance triple immune suppression using Tacrolimus/MMF and MP was used.
all regections were proved by biopsy. mild cases treated with pulse MP (500mg) plus elevetaion of target levels of immunosuppression. server cases were treated with ATG and plasmaphresesi/IVIG followed by Rituximab.
In this cohort study presence of preformed DSA’s was associated with ABMR.
acute rejection was more prominent in both HLA+ and DSA + cases. graft survival was a little bit lower in DSA – group compared with DSA + (84.8% vs 94.9%). ABMR was higher in case with DSA>3000.
In clinical practice one should follow DSA serilaly especially in highly sensetized or in case with previous history of sensetization.
The detection of preformed DSA is very important in stratification of rejection risk, it shown that worse effect on graft outcome even CDCXM & FCXM are both negative. Therefor determination of DSA characteristics associated with detection of AMR had important clinical implication in sensitized patient, immunosuppression strategy & in post transplantation follow-up.
This study follow 462 recipients with negative CDCXM, preformed DSA ( by Luminex, SAP) & VXM. The patient followed to the end of 2013, death or graft loss. Induction with ATG (for high HLA mismatch & allo immunized patients) & anti-IL-2 Abs & maintenance immunosuppression with MMF, CNI & low dose steroid. Graft rejection diagnosed based on histopathological changes of graft biopsy. The study found the following:
The study had several limitation including:
Summary of this article
This article presents a study of 462 kidney transplant recipients, between 2007 and 2012, all patients had negative CDC-XM for both B and T CELLS .
pre-transplant sera were analyzed by Luminex screening assay to determine the presence of HLA antibodies and single-antigen bead assay [positive if mean fluorescence intensity (MFI) ≥ 1000] to assign anti-HLA specificities.
The aim was to analyse the effect of preformed DSA graft outcomes.
It was shown that
-Preformed HLA donor specific antibodies (DSA) increases risk of AMR and associated with poor kidney graft survival, even with negative CDC .
detection of HLA by single antigen bead assay is more sensitive than cell-based crossmatches.
Not only the presence of DSAs affect the graft but t was shown that that low level DSA exert no effect on kidney grafts outcomes also DSA characteristics as HLA class , number , and strength as read by mean fluorescence intensity (MFI) all has its effecton graft outcome.
Although some studies showed that the occurrence of AMR not the mere presence of DSAs that affects the graft and whether this DSAs can initiate endothelial injury or not .
Prevalence of AMR at 1-year posttransplant in our patients with DSA was high.
In a cohort of ATG-induction improved AMR-free survival at 1-year as it was 65.1% for patients with DSA .
There was a significant reduction in 5-years graft survival in recipients with positive DSAS possibly related with a higher incidence of clinical acute rejection
The study also shows that only DSA+ AMR+ patients had a significantly reduced 5-year graft survival while ; DSA+ AMR− patients (96.0%) had similar graft survival in comparison with DSA− patients (94.9%).
There for the occurrence of chronic antibody-mediated rejection is more frequently in DSA+ AMR+ patients.
In comparison of induction therapy ATG was better than basiliximab eitj less AMR .
ATG and IVIG induction in patients with DSA (below 5000 MFI) has been shown to allow for successful kidney transplantation with similar rejection rates and graft survival between DSA+ and DSA− patients .
DSA MFI values below 3000 compared to higher MFI values were associated with improved 5-year death-censored graft survival .
Limitation of the study
No available FCXM .
No protocol biopsies were done .
limitations of SAB assay and MFI values.
anti-HLA-DQ was not determined at the allelic level in every case .
Performed HLA DSAs have been associated with increasing the risk of AMR and lower graft survival even in the presence of negative CDC and FCXM.
HLA DSA detection by SAB is more sensitive and specific compared to cell-based crossmatches.
Although some studies have reported low level of DSAs has no significant effect on graft survival, others have reported the association between DSA characteristics such as HLA class, class, number, and strength (MFI) with kidney allograft outcome.
DSAs that lead to AMR have had a significant effect on reducing graft survival by complement mediated and non-mediated endothelial injury through acute and chronic AMR.
DSA characteristics may have important implications for highly sensitized patients, immunosuppressive strategies, and post-transplant follow up individualization.
This study has analyzed the graft outcomes between anti HLA+DSA+ and anti HLA+DSA- recipients. The incidence of AMR has been more in DSA+ compared to DSA- patients (37.5% versus 14.5%), and acute cellular rejection was the same.
This study has demonstrated significant increase in AMR with DSA MFI of>3000, the highest sensitivity for AMR prediction with a MFI of>4900 in the highest MFI bead, and highest specificity for AMR prediction with the sum of all DSA beads MFI>11000.
Induction therapy with rATG compared to Basiliximab has been related to lower incidence of AMR in DSA+ recipients (65% versus 34%).
There was no adverse effect of non-DSA HLA antibodies in comparison with non-sensitized patients.
DSA+AMR+ with persistence detectable DSA had lower eGFR than DSA+AMR- patients. In addition, chronic AMR has evolved more frequently in DSA+AMR+ patients.
Five-year censored graft survival was lower in DSA+ than in DSA-patients.
DSAs has been associated with increased risk of AMR and decrease graft survival, even if the CDC and/ or flow cytometry cross matches were negative. The main effect of DSA depends on its specificity and strength as defined by MFI. So, defining DSA characteristics associated with AMR occurrence has an important clinical role in the management of a highly sensitized patient, immunosuppressive protocol and patient’s follow up post-transplantation. In the present cohort, the presence of DSA was associated with AMR occurrence, and affected the graft survival, although the CDC crossmatch was negative. The prevalence of AMR was high at 1-year post-transplantation and DSA remained detectable in patient receiving ATG leading to a chronic antibody-mediated . Other studies use protocol biopsy; this incidence was much higher. However, ATG induction increases the AMR-free survival at 1 year to 65% for DSA positive recipients. As well, ATG induction had been shown to improve the 5-years graft survival in DSA positive recipients.
The present study found no adverse effect on graft outcomes associated with the presence of non-DSA anti-HLA antibodies in comparison with nonsensitized patients, as others have reported.
DSA+ patients whose developed AMR have significantly reduced 5-year graft survival; while DSA+ patients who do not develop AMR had graft survival similar to those with negative DSA.
the difference between the DSA positive recipients in developing AMR, point to exploring the importance of DSA characteristics and immunosuppression more likely associated with its occurrence. ATG has been associated with less AMR in DSA + patients as compared with basiliximab. furthermore, ATG and intravenous immunoglobulin induction treatment give a comparable graft survival between DSA positive (below 5000MFI) and DSA negative . interestingly, others have shown that DSA MFI values below 3000 compared to higher MFI values were associated with improved 5-year death-censored graft survival
Limitation duto First, the lack of available FCXM . Second, no protocol biopsies were taken . Third, the known limitations of SAB assay. Finally, as donor specificity of anti-HLA-DQ was not determined at the allelic level in every case .
In our centre we don’t transplant patient with positive DSA .Many patient came with AMR later ,they develop De novo DSA.
· This study enrolled 462 transplant patients, the aim of it was to assess the effect of preformed DSA on graft survival.
· Preformed DSA was associated with increased AMR among patients with negative CDC crossmatch
· AMR for patients with positive DSA was > 35%.
· There was a reduced graft survival after 5 years among patients with positive DSA.
· MFI > 3000 was associated with a significant increased AMR. Also, high DSA MFI had predicted graft survival with an AUC of 0.86.
· DSA positive, AMR positive has significant poor graft survival (5-years) 87%
· DSA negative = DSA positive with AMR negative survival was similar average 95%.
This study is conducted on 462 kidney recipients with a negative complement-dependent cytotoxicity (CDC) cross-match but positive luminex. It confirmed the association between preformed DSA and rejection. Not all DSA affect graft as DSA can affect the graft negatively through endothelial injury and positively they can upregulate protective mechanisms . the effect is attributed to the HLA class , the number and intensity .
In this study not all antibodies present are DSA ,DSA are 8.7% only. DSA are associated significantly with higher rate of 1 year AMR(35.0%). This is only for DSA of MFI(N3000) or higher. The study suggest that ATG in DSA positive is better than bailiximab as it is associated with less AMR (weak significance)
There was no statically significant difference between those with who had DSA versus those who have non DSA antibodies regarding : DGF, eGFR at one year and death with functioning graft. Time to acute rejection is shorter in DSA group and the number of failed graft is more.
Though DSA associated with AMR have higher number, intensity and MFI sum, there was no statically significant difference between DSA causing AMR and DSA not causing AMR in following : DSA class, previous sensitization, or induction
Death censored graft survival is shorter with presence of DSA, DSA lead to statically significant lesser 5 year censored graft loss and less survival .
The drawbacks of this study are the unavailability of FCXM , SAB limitations , no protocol biopsy were conducted and donor DQ were not identified
In our country we do not transplant across DSA barriers. the cost of treatment are nationally secured through a national program. This is why we are trying to concentrate on patients who can make the most benefit from the transplant .
Analysis of preformed donor-specific anti-HLA antibodies characteristics
for prediction of antibody-mediated rejection in kidney transplantation
The presence of DSAs has been associated with increased risk of AMR and deleterious graft survival, even if the CDC and/ or flow cytometry cross matches were negative. SAB assay is more sensitive than cell-based crossmatches in defining DSA specificity. The main effect of DSA depends on its specificity and strength as defined by MFI. So, defining DSA characteristics associated with AMR occurrence has an important clinical role in the management of a highly sensitized patient, immunosuppressive protocol and patient’s follow up post-transplantation. In the present cohort, the presence of DSA was clearly associated with AMR occurrence, and affected the graft survival, although the CDC crossmatch was negative. In spite of the majority of the patient in the study receiving ATG induction; the prevalence of AMR was high at 1-year post-transplantation and DSA remained detectable a clue to chronic antibody-mediated lesions in these recipients. Other studies use protocol biopsy; this incidence was much higher. However, ATG induction increases the AMR-free survival at 1 year to 65% for DSA positive recipients. As well, ATG induction had been shown to improve the 5-years graft survival in DSA positive recipients.
The present study found no adverse effect on graft outcomes associated with the presence of non-DSA anti-HLA antibodies in comparison with nonsensitized patients, as others have reported.
DSA+ patients whose developed AMR have significantly reduced 5-year graft survival; while DSA+ patients who do not develop AMR had graft survival similar to those with negative DSA.
the difference between the DSA positive recipients in developing AMR, point to exploring the importance of DSA characteristics and immunosuppression more likely associated with its occurrence. ATG has been associated with less AMR in DSA + patients as compared with basiliximab. furthermore, ATG and intravenous immunoglobulin induction treatment give a comparable graft survival between DSA positive (below 5000MFI) and DSA negative patients.
The DSA HLA class has no role in AMR development, while, the DSA number and strength are significantly associated with AMR. Especially if DSA MFI is over 3000. The study reveals that detect hDSA MFI threshold level (4900) represent good sensitivity and sDSA MFI level (11,000) had good specificity, So, both together can be used to improve AMR prediction. The association of DSA strength with the occurrence of AMR has been studied by many investigators, some reporting no association, while others showed a significant association which could be related to the variability in MFI cutoff levels between the different centres.
anti-HLA antibodies screen and DSA detection by solid-phase assays are valuable tools, support the transplantation decision in sensitized patients with or without desensitization protocols as well, the applicability of protocol biopsy for selected patients.
Well done, Jamila
Keep going
The type of DSA whether class 1 or 2 has an effect on the AMR. AMR due to class 1 is usually early and aggressive, while class 2 is usually late and less acute.
Preformed DSA have been shown to be associated with increase the risk of AMR and eventually risk of graft failure.
The implementation of solid phase assays in the clinical practice has been associated with improvement in DSA detection and identification of specificities and characteristics of antibodies. HLA antibody detection by SAB assay is well known to be more sensitive than cell based crossmatches.
Some studies reported that low level DSA based on MFI levels have no adverse renal effect, while others studies reported renal rejection and graft failure based on DSA characteristics as HLA class, number, and strength reflected by MFI.
This article reports the results of study done to evaluate the incidence of preformed anti-HLA antibodies by Luminex and SAB assay screening techniques and their association with renal allograft outcomes.
Screening for HLA antibodies were tested in patient every 3 months by Luminex Technology. If initial screening is positive then, SAB assay was performed to determinate the specificity of the HLA antibodies.
Deceased donors were routinely typed for loci HLA-A*, B* and DRB1. If a DSA-DQ antibody was present, HLA-DQ antigens of the donor were verified by SSP. Patients with anti-HLA-C and/or -DP antibodies were excluded.
89.4% of recipients in this review received anti-IL-2 receptor monoclonal antibody as Induction therapy and the remaining received ATG being considered high risk for rejection.
Results of the study
-Acute rejection was particularly more common in HLA+ DSA+ patients 37.5% in comparison with HLA+ DSA− 14.5% and HLA− patients (10.6%). Acute rejection developed earlier in HLA+ DSA+ patients than in HLA+ DSA−.
-DSA+ patients had highest DSA bead with a median MFI of 10,273, while those with later AMR had a median MFI of 5331.
-Death-censored kidney graft survival was detected with a 5-years survival of 94.9% in DSA− versus 84.8% in DSA+ patients. Patient survival was similar.
-Frequent use of ATG versus Basiliximab induction in patients with DSA that did not experience AMR (65.4% and 34.6%) was noticeable.
-At 12-months posttransplant, DSA remained detectable in 40% of DSA+ AMR− and 81.8% DSA+ AMR+ patients.
-Only DSA MFI over 3000 were associated with higher incidence of AMR with good sensitivity 85.7% and MFI threshold level 11,000 that had good specificity 92.3%.
Conclusion
SAB assay is more sensitive and specific in identifying donor specific antibodies. This is clinically translated in better matching between donor and recipients and better planning for the induction and maintenance therapy for at risk recipients. The presence of anti-HLA antibodies is independent risk factor for antibody medicated rejection even in the absence of donor specific antibodies.
Luminex is a standard technique for screening for pre-transplant preformed antibodies. If positive proceeding with SAB id of great benefit on allograft survival.
Thanks
Preformed DSA is associated with increased risk of AMR and graft loss, so its detection is very important, it was found that negative CDC or FCXM might be associated with detection of preformed DSA by solid phase assay SAB (single antigen bead).
Graft survival of the patients with AMR without DSA and patients with DSA and no AMR are the same, its effect on graft survival is related to its strength and specific phenotype.
Management of the patient with DSA with desensitization protocol is very important issue to reduce or avoid AMR posttransplant.
in my practice:
if CDC or FCXM negative will proceed to transplantation, even if there is history of sensitization
Incidence of occurrence of early ABMR is markedly increased in patient with preformed DSA and those who received desensitization thus lowering graft survival.
The significance of positive luminex negative FCM (PLNF) is debatable some studies found no significant difference when compared to negative luminex, negative FCM in terms of incidence of AR at 1 year, graft failure at 1 and 5 years, patient survival at 1 and 5 years,
Other studies including the current study found an increased incidence of ABMR and graft loss in patients with PLNC when compared to those with negative luminex
Several studies found that the presence of DSA alone may not decrease graft survival and do so only if associated with biopsy proved ABMR and this may be explained by the fact that although DSA can produce damage through complement and non-complement dependent injury but at the same time it can up regulate complement regulatory proteins leading to graft accommodation
Factors affecting significance of preformed DSA
1. Strength of DSA (MFI):
2.Complement fixing ability of DSA :
3. Specificity of DSA :
In my practice
All cases i have participated in had negative cross match so we did not do DSA to detect PLNC cases
See scenario 4 Sherif
They may develop de novo DSA
Summary of the Article:
Study type: This is a retrospective analysis
Study Background:
Aim of the study: to analyze the impact of preformed DSA characteristics on kidney graft outcomes.
Study Methods:
Study Results:
1.Patient characteristics according with HLA antibodies detection
In study sample of 462 kidney graft recipients:
As expected, presensitizing events and female gender were more common in HLA+ DSA− and HLA+ DSA+ patients, in comparison with HLA− patients (P < 0.001).
HLA-sensitized patients presented more frequently a PRA level N 20% (P <0.001), had a longer time on dialysis (P< 0.001) and were less likely to receive a graft from a woman (P = 0.050) or a living donor (P = 0.004).
No significant differences between groups were found regarding recipient and donor age, HLA mismatches and cold ischemia time.
Lastly, induction therapy was used in all HLA+ DSA+ patients, with ATG use predominating over basiliximab (P< 0.001).
2.Transplant outcomes according with HLA antibodies detection
3.Antibody-mediated rejection association with DSA characteristics and immunosuppression
4.Kidney graft outcomes in DSA+ patients with or without antibody-mediated rejection
Study Discussion
[http://refhub.elsevier.com/S0966-3274(15)00003-9/rf0115].
Study Limitations:
In our practice, it was noticed the strong association between DSA+ and occurrence of AMR, just an observations need to be studied in an organized and systematic way.
Summary please Safi
(DSA) have been shown to increase the risk of antibody-mediated rejection (AMR) and have a deleterious effect on kidney graft survival, even in the presence of a negative cross matches . Some have reported that low level DSA exert no effect on kidney grafts outcomes . DSAs effects depend also on the their characteristics as HLA class , number , and strength as read by mean fluorescence intensity (MFI). Several studies showed that the occurrence of AMR associated with DSA jeopardizes kidney graft survival . DSA positive patients without AMR having similar survival as compared with those without DSA . This contradictory effect may result from the DSA ability to mediate endothelial injury through complement-dependent and independent mechanisms, mainly through acute and/or chronic AMR, but also, under certain conditions, to promote graft accommodation through up-regulation of complement regulatory or cytoprotective
proteins . Ultimately, DSA main effect will depend on its specificity
and strength. .
This research is designed to study the impact of preformed DSA on kidney graft . 462 patients that received a kidney graft are enrolled in the study . Luminex screening assay is used to determine the presence of anti-human leukocyte antigen (HLA) antibodies and single-antigen bead assay [positive if mean fluorescence intensity (MFI) ≥ 1000] to assign anti-HLA specificities. Anti-HLA antibodies were present in 95 patients (20.6%), but only 40 (8.7%) had DSA. Results of the cross matched negative DSA patients were analyzed . anti body mediated rejection (AMR) at 1-year was found to be higher in patients with DSA (35.0%) than in those without them . DSA with a MFI of N3000 were significantly associated with AMR occurrence. Five-year censored graft survival was lower in DSA positive patient ,although survival was only reduced in those who have AMR .
In my practice ,I notice that, there is increased in the frequency of AMR and chronic allograft nephropathy in DSA positive compared to negative group of patient . also the graft survival is decreased . we a study to confirm this .
Thanks
HLA donor specific antibodies (DSA) presence increase the risk of antibody-mediated rejection (AMR) and affects kidney graft survival.
The clinical importance of DSA detected by SAB has been the matter of a debate.
The noticing of DSA characteristics associated with AMR occurrence have important clinical aspects for highly sensitized patient management, immunosuppressive strategy and kidney graft recipients’ posttransplant follow-up
This study addresses the relation between DSA and kidney transplantation outcomes in patients with negative CDC
The current study included 517 patients whom underwent renal transplantation with 6 consecutive years and followed them
Pre-transplant anti-HLA IgG antibodies were tested by multiplex microsphere based on Luminex Xmap Technology and SAB was done for positive cases
Donor HLA typing of HLA A ,B DR through PCR by Specific sequence primer and DQ was typed excluding DP and C due to lack of the needed materials for that ; all these data was virtualy cross matched.
The CDC crossmatch results were considered positive when cell death exceeded 20%.
DSA strength depended on MFI of one SAB.
Induction therapy was used in 413 patients, with an anti-IL-2 receptor monoclonal antibody (Basiliximab ) or a polyclonal anti-thymocyte globulin.
All enrolled recipients had similar triple maintenance immunosuppression, consisting of oral tacrolimus , mycophenolate mofetil (MMF) and methylprednisolone (MP)/prednisolone.
Acute rejection was more common in HLA+ DSA+ patients (37.5%) in comparison with HLA+ DSA− (14.5%) and HLA− patients (10.6%). This difference resulted from the high incidence of antibody-mediated rejection in HLA+ DSA+ patients, while acute cellular rejection occurrence was similar between groups.
Incidence of AMR in DSA+ graft recipients from a deceased donor was 33.3% and 50% in living donor graft recipients , with no significant difference in the number of HLA mismatches between them.
DSA+ patients with AMR presented a higher number of DSA beads and stronger DSA.
The presence of preformed DSA was associated AMR even in negative CDC crossmatch transplants . Prevalence of AMR at 1-year posttransplant in the studied patients with DSA was high.
There was close association between AMR in DSA+ patients and poor graft survival.
They found that DSA+ patients receiving ATG induction in comparison with basiliximab showed a trend for developing less AMR.
No difference in AMR incidence was found between patients transplanted without any induction and those receiving basiliximab or daclizumab,
DSA number and strength correlated significantly with AMR and DSA MFI over 3000 were associated with higher incidence of AMR
The pitfalls of the study was that a positive flow crossmatch, in the absence of DSA,had an effect on kidney graft outcomes ,no protocol biopsies were undertaken in the study . Histological screening is crucial in the management HLA-incompatible kidney transplantation, and, since 2013, they started to perform it.
The limitations of SAB assay and their reported MFI values should be considered while interpreting the results. As donor specificity of anti-HLA-DQ was not determined at the allelic level in every case rare false assignments as DSA cannot be excluded.
Thank you for writing in your own words
a- DSA was +ve in 20 % of all included patients but only 8.7 % of them developed
anti-body mediated rejection (AMR) at 1 year.
b- only DSA with MFI of more than 3000, were associated with AMR.
c- AMR rate was associated with reduced 5 year graft survival.
d- 5 year graft survival in DSA +ve patients without AMR, was almost similar to
those patients with -ve DSA.
e- ATG induction in DSA+ve patients decreased the incidence of AMR to be
similar to that in DSA-ve patients.
2- Please reflect on your practice if possible.
of course, I will adhere to the conclusion of this study regarding ATG induction in those with DSA +ve patients then to monitor the development and titre of these DSA post-transplantation.
Excellent
DSA have been shown to increase the risk of antibody-mediated rejection and have a deleterious effect on kidney graft survival, even in the presence of a negative complement-dependent cytotoxicity and/or flow cytometry crossmatches. SAB assay is much more sensitive.
The study aim was to analyse the relationship between preformed DSA and kidney graft outcomes, in the presence of a negative CDC crossmatch.
517 consecutive adult patients were investigated.
All patients were transplanted with a negative pretransplant T- and B-lymphocyte CDC crossmatch in current and peak sera.
Screening for HLA antibody was done by Luminex Xmap.
To determinate the specificity of the
HLA antibodies, single-antigen bead assays were performed in patients with a positive screening and a cut-off for a positive reaction were set in MFI value of ≥1000.
Samples of all deceased donors were routinely typed before recipient selection in loci HLA-A*, B* and DRB1. HLA-DQ antigens with strong linkage disequilibrium were primarily inferred from the
HLA-DR antigens. Patients with only anti-HLA-C and/or -DP antibodies were not considered in the analysis, as no donor typing at these loci was available.
Induction protocol and maintenance immunosuppression
Induction therapy was used in 413 patients (89.4%)-
Basiliximab 20 mg twice at day 0 and 4 or
polyclonal anti-thymocyte globulin 3 mg/kg for 5–7 days).
ATG was primarily used in allo-sensitized patients or in the presence of high number of HLA mismatches.
Triple maintenance immunosuppression-
Tacrolimus at a dose of 0.1–0.15 mg/kg/day, and the dose was adjusted to maintain a trough level in whole blood between 8 and 12 ng/ml during the first month postoperatively, between 7 and 10 ng/ml during 2–3 months after transplant and between 5 and 8 ng/ml thereafter
Mycophenolate mofetil started at a dose of 2000 mg/day, with the dose decreasing to 1000–1500 mg/day during the first month postoperatively, depending on white blood cells count.
Methylprednisolone was administered intravenously at doses of 500, 250 and 125 mg/day on the day of transplantation, days 1–2 and days 3–4 after the operation, respectively.
Oral prednisolone was started on day 5 after the operation at the dose of 20 mg, being then tapered to 5–10 mg/day within 2–3 months after transplant.
Living donor recipients (n = 89) were prescribed Tacrolimus and MMF 7 days before transplant. No desensitization protocol was carried out in these patients.
The presence of preformed DSA was clearly associated with AMR occurrence. Prevalence of AMR at 1-year posttransplant in our patients with DSA was high (35.0%). Significantly reduced 5-years death censured graft survival in patients with DSA. Found no adverse effect on graft outcomes associated with the presence of non-DSA anti-HLA antibodies in comparison with non-sensitized patients. y DSA+ AMR+ patients had a significantly reduced 5-year graft survival (68.8%); DSA+ AMR− patients (96.0%) had similar graft survival in comparison with DSA− patients (94.9%). Persistence of DSA at 1-year is accompanied by lower estimated GFR and higher histological scores for transplant glomerulopathy and chronic AMR. DSA+ patients receiving ATG induction in comparison with basiliximab showed a trend for developing less AMR. DSA HLA class was not associated with AMR , In contrast, DSA number and strength correlated significantly with AMR. Patients with AMR had a probability 7.4 times higher of having a sDSA MFI of >11,000; instead, patients without AMR had a probability 5 times higher of having a Hdsa MFI of <4900.
Limitations –
lack of available information about flow cytometry crossmatch results may affect its clinical application, particularly in highly sensitized patients, in whom the close analysis of both flow cytometry and virtual crossmatch is important for their management.
No protocol biopsies done.
Known limitations of SAB assay and their reported MFI values.
Rare false assignments as DSA cannot be excluded.
Conclusion –
Close relationship between the presence of preformed DSA and an increase incidence of AMR in kidney transplantation, with subsequent adverse impact on graft survival.
Anti-HLA antibodies screening and DSA detection by solid-phase assays are valuable tools, together with cell-based crossmatches.
Please reflect on your practice if possible.
Only CDC crossmatch used to be done and HLA typing.