Keywords: Anti-human leukocyte antigen-donor specific antibody; Kidney transplantation; Luminex Single Antigen assay; Allograft biopsy

Transcription

1 ORIGINAL ARTICLE Koren J Intern Med 18;33: Clinicl significnce of the presence of ntihumn leukocyte ntigen-donor specific ntibody in kidney trnsplnt recipients with llogrft dysfunction Byung H Chung 1,2, Jeong Ho Kim 1,2, Bum Soon Choi 1,2, Cheol Whee Prk 1,2, Ji-Il Kim 3, In Sung Moon 3, Yong-Soo Kim 1,2, Yeong Jin Choi 4, Eun-Jee Oh 1,5, nd Chul Woo Yng 1,2 1 Trnsplnt Reserch Center, 2 Division of Nephrology, Deprtment of Internl Medicine, Deprtments of 3 Surgery, 4 Hospitl Pthology, nd 5 Lbortory Medicine, College of Medicine, Seoul St. Mry s Hospitl, The Ctholic University of Kore, Seoul, Kore Received : Mrch 3, 16 Revised : My 19, 16 Accepted : My 28, 16 Correspondence to Chul Woo Yng, M.D. Deprtment of Internl Medicine, College of Medicine, Seoul St. Mry s Hospitl, The Ctholic University of Kore, 222 Bnpo-dero, Seocho-gu, Seoul 6591, Kore Tel: Fx: E-mil: yngch@ctholic.c.kr Bckground/Aims: This study investigted the clinicl significnce of detecting nti-humn leukocyte ntigen-donor specific ntibody (HLA-DSA) in kidney trnsplnt recipients (KTRs) requiring indiction biopsy owing to llogrft dysfunction. Methods: We nlyzed the presence of HLA-DSA in 21 KTRs who took indiction biopsy. We divided these cses into two groups, (n = 52) nd HLA-DSA ( ) (n = 158) group, nd compred the clinicl chrcteristics, pthologicl findings, nd clinicl outcomes of the two groups. Results: The rtes of retrnsplnt, pretrnsplnt sensitiztion, nd HLA-mismtch were significntly higher in group thn in HLA-DSA ( ) group (p <.5 for ech comprison). In histologic finding, ll types of rejections were more frequent in the former group. Besides, scores of both the T-cell injury mrkers such s tubulitis, interstitil inf lmmtion, nd vsculitis nd ntibody-medited injury mrkers such s peritubulr C4d deposition nd microvsculr inflmmtion (glomerulitis plus peritubulr cpillritis) were higher in group (p <.5 for ech). Notbly, llogrft outcomes were worse in group. Further, multivrite nlysis showed tht presence of HLA- DSA, dvnced interstitil fibrosis/tubulr trophy (interstitil fibrosis plus tubulr trophy 2), nd llogrft rejection in biopsy were independent risk fctors for llogrft filure. Conclusions: The results of this study showed tht presence of HLA-DSA in cse of llogrft dysfunction dversely influences llogrft outcome, nd its detection, irrespective of the result of the llogrft biopsy, necessittes intensive monitoring nd tretment. Keywords: Anti-humn leukocyte ntigen-donor specific ntibody; Kidney trnsplnttion; Luminex Single Antigen ssy; Allogrft biopsy Copyright 18 The Koren Assocition of Internl Medicine This is n Open Access rticle distributed under the terms of the Cretive Commons Attribution Non-Commercil License ( by-nc/3./) which permits unrestricted noncommercil use, distribution, nd reproduction in ny medium, provided the originl work is properly cited. pissn eissn

2 The Koren Journl of Internl Medicine Vol. 33, No. 1, Jnury 18 INTRODUCTION The mrked increse over time in detection rte of nti-humn leukocyte ntigen-donor specific ntibody (HLA-DSA) is significntly ssocited with the decline in llogrft function posttrnsplnt nd the subsequent llogrft filure [1,2]. It is well known tht HLA- DSA plys criticl role in the progression of chronic ntibody-medited tissue injury, which in turn, is the most importnt cuse of lte llogrft filure in kidney trnsplnt recipients (KTRs) [3,4]. Therefore, detection nd monitoring of HLA-DSA my help in predicting llogrft outcomes nd plnning proper mngement to prevent decline of llogrft function in KTRs. Most previous studies investigting the role of HLA- DSA performed seril mesurement of HLA-DSA in cliniclly stble ptients without llogrft dysfunction nd showed tht its presence is ssocited with deteriortion of llogrft function nd subsequent rejection [5-7]. Notbly, the clinicl significnce of HLA-DSA in ptients with llogrft dysfunction hs not been fully investigted. For exmple, in cses of llogrft dysfunction not dignosed s ntibody medited rejection (AMR) but dignosed s T cell medited rejection (TCMR) or clcineurin inhibitor (CNI) toxicity, the clinicl significnce of detecting HLA-DSA hs not yet been fully evluted. In this bckground, we investigted the clinicl significnce of detecting HLA-DSA in cses with llogrft dysfunction. First, we exmined the presence of HLA-DSA using Luminex Single Antigen (LSA) ssy in ll these cses t the time of indiction llogrft biopsy, which ws performed owing to kidney llogrft dysfunction. Second, we nlyzed the ssocition between the presence of HLA-DSA nd specific histologicl nd pthologicl findings. Finlly, we evluted whether detection of HLA-DSA significntly influences llogrft outcome during follow-up. METHODS Bseline chrcteristics of ptients nd procedure of indiction biopsy Between Februry 1 nd Februry 13, totl of 21 KTRs took llogrft biopsy due to llogrft dysfunction t Seoul St. Mry s Hospitl. The definition of llogrft dysfunction in this study ws serum cretinine level of % higher thn the bseline vlue or more. Bseline chrcteristics of ptients re given in Tble 1. The procedure of biopsy nd the technique of histologicl dignosis were bsed on previous study [8]. In brief, 16-guge biopsy gun ws used under ultrsonic locliztion. Indirect immunofluorescence stining ws performed using monoclonl ntibodies ginst complement protein C4d (Biogenesis, Poole, Englnd; dilution, 1:5) for detecting C4d deposition. C4d positivity ws defined s diffuse (> 5%) nd liner stining of peritubulr cpillries. Microvsculr inflmmtion (MVI) score ws clculted by dding glomerulitis (g) nd peritubulr cpillritis (ptc) scores. Histopthologicl dignosis ws bsed on the revised Bnff working clssifiction [9-11]. Pretrnsplnt desensitiztion protocol for pretrnsplnt highly sensitized ptients According to our center s pretrnsplnt desensitiztion protocol, the trget HLA-DSA vlue t the time of kidney trnsplnt (KT) ws wek or negtive level (medin fluorescence intensity [MFI] < 5,) by LSA ssy [12]. In ptients with moderte to strong HLA-DSA vlues (MFI > 5,), rituximb t dose of 375 mg/m2 (MbTher, Genentech Inc., Sn Frncisco, CA, USA) ws dministered 2 to 3 weeks before trnsplnttion, nd plsmpheresis/intrvenous immunoglobulin (PP/ IVIG) therpy ws initited 13 dys prior to trnsplnttion nd dministered every other dy. In ddition, we initited immune suppressnt (IS) tretment 7 dys prior to trnsplnttion in these ptients. HLA-DSA nd crossmtch (XM) testing ws performed 2 dys prior to the trnsplnt. When HLA-DSA decresed to negtive or wek levels nd XM testing showed negtive conversion, KT ws performed. If HLA-DSA results were moderte to strong, or if the XM ws positive, we performed dditionl PP/IVIG three times nd subsequently retested the ptient for HLA-DSA nd XM. Detection of HLA-DSA nd HLA typing We exmined the presence of HLA-DSA using LSA ssy in ll cses t the time of llogrft biopsy. As described in previous studies, LSA ssy for HLA-DSA ws performed s per the mnufcturer s instructions using Lifecodes LifeScreen Deluxe kits (Tepnel Lifecodes Corp., Stmford, CT, USA) [12,13]. In brief, microbeds

3 Chung BH, et l. HLA-DSA t llogrft dysfunction Tble 1. Comprison of clinicl chrcteristics between nd HLA-DSA ( ) group Chrcteristic (n = 52) HLA-DSA ( ) (n = 158) p vlue Age t biopsy, yr 43.7 ± ± Mle sex 27 (51.9) 99 (62.7).11 Primry renl disese Chronic glomerulonephritis 25 (48.1) 52 (32.9) Dibetes mellitus 6 (11.5) 13 (8.2) Hypertension 3 (5.8) 17 (1.8).7 ADPKD 3 (5.8) 3 (1.9) Others 15 (28.8) 73 (46.2) Posttrnsplnt month 22.4 ± ± 65.7 <.5 ABO IKT 7 (13.5) 17 (1.8).62 Donor type LRD 22 (42.3) 81 (51.3) LURD 7 (13.5) 32 (.3).38 DD 23 (44.2) 45 (28.5) Induction therpy.4 Anti-thymocyte globulin 17 (32.7) 29 (18.4) Bsiliximb 35 (67.3) 129 (81.6) Min IS Tcrolimus 45 (86.5) 135 (85.4) Cyclosporine 6 (11.5) 19 (12.).85 Azthioprine 2 (1.3).85 Rpmune 1 (1.9) 2 (1.3) Desensitiztion therpy RTX/PP/IVIG 17 (32.7) 7 (4.4) <.1 RTX 15 (9.5) <.1 No desentiztion 35 (67.3) 136 (86.1) Re-trnsplnttion 13 (25.) 7 (4.4) <.1 HLA mismtch number 3.6 ± ± 1.8 <.5 High PRA (> 5%) 17 (32.7) 22 (13.9) <.1 17 (32.1) 15 (9.5) <.5 Positive crossmtch test 9 (17.3) 8 (5.1) <.5 Vlues re presented s men ± SD or number (%). HLA-DSA, nti-humn leukocyte ntigen-donor specific ntibody; ADPKD, utosoml dominnt polycystic kidney disese; ABO IKT, ABO incomptible kidney trnsplnttion; LRD, living relted donor; LURD, living unrelted donor; DD, decesed donor; IS, immune suppressnt; RTX, rituximb; PP, plsmpheresis; IVIG, intrvenous immunoglobulin; PRA, pnel rective ntibody. coted with purified HLA clss I/clss II glycoproteins were incubted with 12.5 μl of ptient s serum in 96 well pltes for 3 minutes. After three wshes with vcuum mnifold, the beds were incubted with 5 μl of 1:1 dilution of R-phycoerythrin-conjugted got nti-humn immunoglobulin G for 3 minutes. After wshing, the test smples were nlyzed using the Quick-Type User s Mnul Reserch Use Only progrm, version 2.4 of the LABScn1 flow cytometer (Luminex Corp., Austin, TX, USA); both positive nd negtive controls

4 The Koren Journl of Internl Medicine Vol. 33, No. 1, Jnury 18 were included. The positive criterion ws MFI level of > 1,. HLA typing ws performed in ll ptients nd donors using the DNA moleculr typing method. Reverse sequence-specific oligonucleotide probes nd RELI TM SSO HLA-A, B, C, DR, DQ Typing Kit (Dynl Biotech Ltd., Bromborough, Englnd) were used. In cse, in ptient, the nti-hla ntibody detected by LSA ssy corresponded to the HLA-type of the donor, it ws clssified s HLA-DSA. The results were presented s MFI nd were clssified into four levels bsed on the pek vlue of the detected HLA-DSA in smple: strong, > 1,; moderte, 5, to 1,; wek, 1, to 5,; nd negtive, < 1,. Assessment of clinicl outcomes All subjects were divided into two groups, nd HLA-DSA ( ), bsed on the presence of HLA-DSA t the time of indiction biopsy. We compred clinicl outcomes between the two groups. The primry outcome of this study ws the impct of the presence of HLA-DSA t the time of indiction biopsy on the llogrft outcome. Secondry outcome ws the rte of detection of HLA- DSA, the fctors ssocited with the development of HLA-DSA, nd the ssocition between HLA-DSA nd the histologicl findings of the llogrft biopsy. Clinicl informtion ws collected using retrospective chrt review. Allogrft function ws ssessed using the Modified Diet in the Renl Disese formul [14]. This study ws pproved by the Institutionl Review Bord of Seoul s Seoul St. Mry s Hospitl (KC11RCMI687). Sttisticl nlysis Sttisticl nlysis ws performed using SPSS version 16. (SPSS Inc., Chicgo, IL, USA). Dt were presented s men ± SD or counts nd percentges, depending on the type of dt. For continuous vribles, mens were compred using Student t test. For ctegorized vribles, Person chi-squre test nd Fisher exct test were used. Allogrft survivl ws nlyzed using Kpln-Meier method with log-rnk test. It ws censored in cse of ptient s deth with functioning llogrft. Cox regression nlysis ws used for multivrite nlysis to evlute risk fctors for llogrft filure. Results were considered sttisticlly significnt if the p vlue ws below.5. RESULTS Detection of HLA-DSA nd distribution of HLA-DSA subtypes HLA-DSA ws detected in totl of 52 cses (24.8%) out of 21 cses. HLA-DSA clss I ws detected in 17 cses (8.1%); HLA-DSA clss II ws found in 41 cses (19.5%); nd six cses (2.9%) showed both clss I nd II HLA- DSA. The most common HLA-DSA subtype ws HLA- DSA-DR (26 cses, 12.4%) followed by HLA-DSA-DQ (19 cses, 9.%) nd HLA-DSA-B (12 cses, 5.7%). Wek HLA-DSA ws detected in 35 cses (16.7%), five cses (2.4%) showed moderte HLA-DSA, nd 12 cses (5.7%) showed strong HLA-DSA. In comprison between the two groups, (n = 52) nd HLA-DSA ( ) (n = 158), no significnt difference in clinicl chrcteristics such s ge t biopsy, sex, primry renl disese, donor type, type of min IS, nd ABO incomptibility to donor ws detected. The posttrnsplnt durtion ws significntly longer in HLA-DSA ( ) group compred to HLA- DSA (+) group (45.7 ± 65.7 vs ± 41., p <.5) (Tble 1). Regrding pretrnsplnt immunologicl chrcteristics, significnt differences were found between the two groups. The HLA-mismtch number, incidence of retrnsplnttion, number of ptients with high pnel rective ntibody, nd HLA-DSA t bseline were significntly higher in group compred to HLA-DSA ( ) group (Tble 1). Compring llogrft function nd pthologicl findings of nd HLA-DSA ( ) groups At the time of llogrft biopsy, there ws no significnt difference in llogrft function between the two groups (HLA-DSA [+] group, 29.3 ± 14.5 ml/min/1.73 m 2 vs. HLA- DSA [ ] group, 31.6 ± 15.4 ml/min/1.73 m 2, p >.5). In contrst, pthologicl dignoses/findings bsed on Bnff clssifiction differed significntly between the two groups [9-11]. The incidence rtes of not only totl rejection (HLA-DSA [+], 8.8%, 42/52 vs. HLA-DSA [ ], 3.4%, 48/158) (Fig. 1A), but lso AMR (HLA-DSA [+], 28.8%, 15/52 vs. HLA-DSA [ ], %, /158) nd TCMR (HLA-DSA [+], 48.1%, 25/52 vs. HLA-DSA [ ], 3.4%, 48/158) were significntly higher in group compred to HLA- DSA ( ) group (p <.5 for ech comprison). In contrst, the incidence of CNI toxicity ws significntly higher in the HLA-DSA ( ) group (24.7%, 39/158) thn in the HLA

5 Chung BH, et l. HLA-DSA t llogrft dysfunction Rejection A HLA-DSA ( ) B C D E F Figure 1. Compring llogrft biopsy finding nd the distribution of Bnff score in the nti-humn leukocyte ntigen-donor specific ntibody (HLA-DSA) (+) nd HLA-DSA ( ) groups. (A) Incidence of totl llogrft rejection ws significntly higher in group compred to HLA-DSA ( ) group. Scores of (B) tubulitis nd (C) interstitil inflmmtion were higher in the group thn in the HLA-DSA ( ) group. No significnt difference ws detected in the distribution of (D) interstitil fibrosis, (E) tubulr trophy, nd (F) glomerulosclerosis scores between the nd HLA-DSA ( ) groups. p <.5 vs. HLA-DSA ( ) group. DSA (+) group (9.6%, 5/52, p <.5) (Tble 2). When we compred the biopsy findings of KTRs who progressed to llogrft filure between nd HLA-DSA ( ) group, AMR showed incresing tendency s cuse for llogrft filure in group, but it did not rech sttisticl significnce (Tble 3). Compring distribution of Bnff nd microvsculr inflmmtion scores Regrding distribution of Bnff score, t (tubulitis), i (interstitil inflmmtion) scores were higher in the HLA- DSA (+) group thn in the HLA-DSA ( ) group (p <.5) (Fig. 1B nd 1C). In contrst, ci (interstitil fibrosis), ct (tubulr trophy) nd cg (glomerulr sclerosis) scores did not differ significntly between the two groups (p >.5) (Fig. 1D-1F). Higher scores of histologicl mrkers ssocited with ctivtion of humorl immunity such s peritubulr C4d deposition, g, v (vsculitis), nd ptc scores were distributed in group compred to HLA-DSA ( ) group (p <.5) (Fig. 2A-2D). Lstly, higher MVI score, clculted by dding g nd ptc scores, ws distributed in group (p <.5) (Fig. 2E). Compring llogrft outcomes between HLA-DSA (+) nd HLA-DSA ( ) groups Fig. 3 compres clinicl outcomes between the HLA- DSA (+) nd HLA-DSA ( ) groups. The incidence of recurrent or newly developed cute rejection within 6 months (19.2% [1/52] vs. 6.3% [1/158]) nd tht of steroid-resistnt rejection (32.7% [17/52] vs. 7.6% [12/158]) were significntly higher in group compred to HLA-DSA ( ) group (p <.5 for ech) (Fig. 3A nd 3B). When only rejection cses were included, incidence of recurrent or newly developed rejection with

6 The Koren Journl of Internl Medicine Vol. 33, No. 1, Jnury 18 Tble 2. Comprison of histologic dignosis between group nd HLA-DSA ( ) group Vrible (n = 52) HLA-DSA ( ) (n = 158) p vlue TCMR 25 (48) 48 (3) <.1 Acute or chronic AMR 17 (33) <.1 CNI toxicity 5 (1) 39 (25) <.1 Recurrent GN 18 (11) <.1 Borderline chnge 2 (4) 18 (11) <.1 Acute tubulr necrosis 8 (5) <.1 BKVAN 1 (2) 6 (4) <.1 Norml nd others 4 (8) 29 (18) <.1 Vlues re presented s number (%). HLA-DSA, nti-humn leukocyte ntigen-donor specific ntibody; TCMR, T cell medited rejection; AMR, ntibody medited rejection; CNI, clcineurin inhibitor; GN, glomerulonephritis; BKVAN, BK virus ssocited nephropthy. Tble 3. Comprison of histologic dignosis in ptients with llogrft filure between group nd HLA-DSA ( ) group Vrible (n = 17) HLA-DSA ( ) (n = 26) p vlue TCMR 11 (65) 15 (58) <.1 Acute or chronic AMR 4 (24) <.1 CNI toxicity 1 (6) 5 (19) <.1 BKVAN 1 (2) 6 (4) <.1 Recurrent GN 4 (15) <.1 Vlues re presented s number (%). HLA-DSA, nti-humn leukocyte ntigen-donor specific ntibody; TCMR, T cell medited rejection; AMR, ntibody medited rejection; CNI, clcineurin inhibitor; BKVAN, BK virus ssocited nephropthy; GN, glomerulonephritis. in 6 months (21.4% [9/42] vs. 12.5% [6/48]) showed n incresing tendency (p =.8) (Fig. 3C), wheres incidence of steroid-resistnt rejection (.5% [17/42] vs. 25% [12/48]) showed significnt increse (p <.5) (Fig. 3D). Therefore, llogrft survivl rte ws lower in HLA-DSA (+) group thn in HLA-DSA ( ) group not only in the entire cohort, but lso in the rejection nd non-rejection groups (p <.5 for ech) (Fig. 3E-3G). Multivrite nlysis showed tht presence of HLA-DSA, especilly clss I, dvnced Interstitil fibrosis/tubulr trophy (IF/TA; ci + ct 2), nd llogrft rejection in llogrft tissue were independent risk fctors for llogrft filure (Tble 4). DISCUSSION Appliction of Luminex technology in mesuring HLA- DSA enbles ccurte determintion of humorl immunity in KTRs [6,15,16]. In our previous study, we reported the clinicl usefulness of pretrnsplnt HLA-DSA mesured by Luminex technique in predicting posttrnsplnt outcome [12,17]. In this study, we investigted the clinicl usefulness of mesuring posttrnsplnt HLA-DSA, especilly in cses of llogrft dysfunction, nd found tht it is ssocited with higher incidence of rejection nd more ctive pthologicl findings despite similr llogrft function. We lso found it to be n independent risk fctor for llogrft filure. To strt with, we studied the distribution of HLA-DSA ccording to clss nd strength nd found tht the pttern of posttrnsplnt HLA-DSA ws different from tht of pretrnsplnt HLA-DSA [12,17]. Before trnsplnt, the frequencies of clss I nd clss II HLA-DSA were the sme; however, during the posttrnsplnt period,

7 Chung BH, et l. HLA-DSA t llogrft dysfunction A HLA-DSA ( ) B C D E Figure 2. Compring histologicl mrkers ssocited with humorl immunity between the donor specific nti-humn leukocyte ntigen ntibody (HLA-DSA) (+) nd HLA-DSA ( ) groups. (A) C4d, (B) glomerulitis, (C) vsculitis, nd (D) peritubulr cpillritis scores were higher in the group thn in the HLA-DSA ( ) group. (E) In ddition, the proportion of cses with significnt microvsculr inflmmtion score ( 2) ws higher in the group. p <.5 vs. HLA-DSA ( ) group. HLA-DSA clss II, especilly HLA-DSA-DR nd HLA- DSA-DQ, were more frequently detected. This finding is consistent with results of previous studies tht reported up-regultion of clss II HLA-DSA de novo fter trnsplnttion [7,18]. Regrding strength of HLA-DSA, most posttrnsplnt cses, like the pretrnsplnt ones, showed wek HLA-DSA [12,17]. Next, we divided cses into two groups, nd HLA-DSA ( ), bsed on the presence of HLA-DSA detected t the time of llogrft biopsy. We compred the clinicl nd immunologicl chrcteristics between the two groups nd found tht the incidence of pretrnsplnt sensitiztion ws significntly higher in HLA-DSA (+) group. This is consistent with the findings of the previous studies tht showed the role of pretrnsplnt sensitiztion in de novo ppernce of posttrnsplnt HLA- DSA [19,]. Interestingly, in our study, posttrnsplnt durtion ws longer in HLA-DSA ( ) group in contrst with the results of the previous studies which showed grdul increse in de novo detection of HLA-DSA with time fter KT [5,7,21]. This my be due to the fct tht we studied cses with llogrft dysfunction, wheres previous studies minly included cliniclly stble ptients. In ptients without HLA-DSA, the min cuse of llogrft dysfunction is not llogrft rejection but other resons such s CNI toxicity, s lso shown in our study, nd compred to llogrft rejection, llogrft dysfunction requires longer time to develop [22]. Therefter, we compred pthologicl findings of the llogrft tissue. As expected, the incidence of llogrft rejection ws significntly higher in group. Interestingly, not only the incidence of AMR, but lso tht of TCMR ws significntly higher in group compred to HLA-DSA ( ) group. It is well known tht complex interctions between T cells nd B cells re involved in the ctivtion of immune system, which

8 The Koren Journl of Internl Medicine Vol. 33, No. 1, Jnury A B C HLA-DSA ( ) D Allogrft survivl rte (%) 8 Allogrft survivl rte (%) 8 HLA-DSA ( ) HLA-DSA ( ) HLA-DSA ( ) Postbiopsy month Postbiopsy month Postbiopsy month E F G Allogrft survivl rte (%) 8 Figure 3. Compring clinicl outcomes fter llogrft biopsy between the donor specific nti-humn leukocyte ntigen ntibody (HLA-DSA) (+) nd HLA-DSA ( ) groups. Compring incidence of (A) repeted rejection within 6 months of llogrft biopsy nd (B) steroid-resistnt rejection between the nd HLA-DSA ( ) groups in the entire cohort. Compring incidence of (C) repeted rejection within 6 months of llogrft biopsy nd (D) steroid-resistnt rejection between the HLA- DSA (+) nd HLA-DSA ( ) groups mong the rejection cses. Compring llogrft survivl rte fter biopsy between the HLA- DSA (+) nd HLA-DSA ( ) groups (E) in the entire cohort, (F) mong the rejection cses, nd (G) mong the non-rejection cses. p <.5 vs. HLA-DSA ( ) group. results in llogrft rejection; therefore, T cell ctivtion could be involved in de novo development of HLA-DSA or vice vers [23-27]. Besides, in our study, CNI toxicity ws more frequent in the HLA-DSA ( ) group. This my be becuse llogrft dysfunction requires longer time to develop posttrnsplnttion resulting in greter exposure to CNI in this group [22]. Regrding pthologicl findings bsed on Bnff clssifiction, the histologicl mrkers ssocited with ctivtion of locl humorl immune system such s C4d score nd MVI score, the combintion of g nd ptc scores, incresed in the group, which indictes frequent development of HLA-DSA-induced llogrft tissue injury in this group [28,29]. In ddition, scores of t nd i were higher in the group thn in the HLA- DSA ( ) group, which suggests incresed rte of TCMR in this group. In previous studies bout the role of HLA- DSA in stble KTRs without llogrft dysfunction, the progression of chronic tissue injury mostly presenting s chronic AMR, ws the most dominnt finding [5-7]. In contrst, ct, ci, nd cg scores were similrly distributed in the two groups indicting tht dvncement

9 Chung BH, et l. HLA-DSA t llogrft dysfunction Tble 4. Significnt risk fctors ssocited with llogrft filure Vrible Univrite Multivrite HR 95% CI p vlue HR 95% CI p vlue Age Donor type (DD) Posttrnsplnt month Pretrnsplnt sensitiztion Min IS Retrnsplnt HLA-DSA t biopsy Strong HLA-DSA HLA-DSA clss I < HLA-DSA clss II Advnced IF/TA Rejection HR, hzrd rtio; CI, confidence intervl; DD, decesed donor; IS, immune suppressnt; HLA-DSA, nti-humn leukocyte ntigen-donor specific ntibody; IF/TA, interstitil fibrosis tubulr trophy. of chronic chnge is not chrcteristic findings when HLA-DSA ws detected in KTRs with llogrft dysfunction in contrst to cses with stble llogrft function. Finlly, we investigted the impct of HLA-DSA in cses with llogrft dysfunction on future clinicl outcomes nd found tht these outcomes were significntly worse in group in terms of recurrent rejection, steroid resistnce, nd lower survivl rte of llogrft following llogrft dysfunction. Interestingly, similr results were found in the sub-nlysis involving only the rejection group or only the non-rejection group. Additionlly, multivrite nlysis showed tht presence of HLA-DSA, especilly clss I ntibody, dvnced IF/TA, nd finding of llogrft rejection in biopsy were independent risk fctors for llogrft filure. These findings suggest tht presence of HLA-DSA my be ssocited with dverse llogrft outcome irrespective of histologicl findings in cses with llogrft dysfunction. Previous studies show tht HLA-DSA could induce ctivtion of locl immune system in llogrft tissue resulting in progression of llogrft tissue inflmmtion, corroborted by ctive pthologicl findings of our study [3]. Notbly, cses with such injuries my be ssocited with dverse clinicl outcomes such s recurrence of rejection, resistnce to steroid pulse therpy, nd finlly llogrft filure. Indeed, TCMR or AMR ccounts for 88% of the cuse for llogrft filure in group, which is higher thn tht of HLA-DSA ( ) group (57%) (Tble 3). The limittion of this study is tht we did not study the longitudinl chnge in HLA-DSA before nd fter the llogrft biopsy; hence, we could not ssess the ctul point of time of development of HLA-DSA nd llogrft dysfunction. Second, group lredy showed higher pre-trnsplnt immunologic risk, which my induce bis during nlysis. Lstly, we could not ssess the cliniclly significnt vlue of MFI becuse the strength of HLA-DSA in our study did not hve significnt impct on llogrft outcome. This is in contrst with the sitution before KT, where only the strong HLA-DSA is not cliniclly relevnt [11,12,31]. The reson is uncler, but it my be possible tht even wek HLA- DSA my develop strong immunogenicity under mintennce immunosuppression. In conclusion, presence of HLA-DSA in cses with llogrft dysfunction is significntly ssocited with ccompnied rejection, ctive pthologicl findings, nd histologicl mrkers for ntibody-medited injury. In ddition, presence of HLA-DSA is n independent risk fctor for llogrft filure. Hence, we suggest tht mesurement of HLA-DSA should be mndtorily performed in KTRs with llogrft dysfunction. Moreover, in cse HLA-DSA is detected, strict monitoring nd intensive

10 The Koren Journl of Internl Medicine Vol. 33, No. 1, Jnury 18 immunosuppressive therpy my be required even when llogrft biopsy does not show llogrft rejection. KEY MESSAGE 1. Posttrnsplnt detection rte of nti-humn leukocyte ntigen-donor specific ntibody (HLA-DSA) ws higher in cses with pretrnsplnt sensitiztion. 2. Detection of HLA-DSA in cses with llogrft dysfunction is ssocited with ctive pthologicl findings, nd histologicl mrkers for ntibody-medited injury. 3. Presence of HLA-DSA in cses with llogrft dysfunction dversely inf luences llogrft outcome irrespective of the result of the llogrft biopsy. Conflict of interest No potentil conflict of interest relevnt to this rticle ws reported. Acknowledgments This study ws supported by grnt (HI13C1232) of the Koren Helth Technology R&D Project, Ministry of Helth nd Welfre, Republic of Kore. REFERENCES 1. Wiebe C, Gibson IW, Blydt-Hnsen TD, et l. Evolution nd clinicl pthologic correltions of de novo donor-specific HLA ntibody post kidney trnsplnt. Am J Trnsplnt 12;12: Lee PC, Terski PI, Tkemoto SK, et l. All chronic rejection filures of kidney trnsplnts were preceded by the development of HLA ntibodies. Trnsplnttion 2;74: Sellres J, de Freits DG, Mengel M, et l. Understnding the cuses of kidney trnsplnt filure: the dominnt role of ntibody-medited rejection nd nondherence. Am J Trnsplnt 12;12: Cosio FG, Gloor JM, Sethi S, Stegll MD. Trnsplnt glomerulopthy. Am J Trnsplnt 8;8: Everly MJ, Rebellto LM, Hisch CE, et l. Incidence nd impct of de novo donor-specific llontibody in primry renl llogrfts. Trnsplnttion 13;95: Dieplinger G, Ditt V, Arns W, et l. Impct of de novo donor-specific HLA ntibodies detected by Luminex solid-phse ssy fter trnsplnttion in group of 88 consecutive living-donor renl trnsplnttions. Trnspl Int 14;27: Dieplinger G, Everly MJ, Rebellto LM, et l. Chnges in successive mesures of de novo donor-specific nti-humn leukocyte ntigen ntibodies intensity nd the development of llogrft dysfunction. Trnsplnttion 14;98: Chung BH, Lee JY, Kng SH, et l. Comprison of clinicl outcome between high nd low bseline nti-abo ntibody titers in ABO-incomptible kidney trnsplnttion. Ren Fil 11;33: Solez K, Colvin RB, Rcusen LC, et l. Bnff 7 clssifiction of renl llogrft pthology: updtes nd future directions. Am J Trnsplnt 8;8: Hs M, Sis B, Rcusen LC, et l. Bnff 13 meeting report: inclusion of C4d-negtive ntibody-medited rejection nd ntibody-ssocited rteril lesions. Am J Trnsplnt 14;14: Cooper JE, Grll J, Chn L, Wisemn AC. Clinicl significnce of post kidney trnsplnt de novo DSA in otherwise stble grfts. Clin Trnspl 11: Chung BH, Choi BS, Oh EJ, et l. Clinicl impct of the bseline donor-specific nti-humn leukocyte ntigen ntibody mesured by Luminex single ntigen ssy in living donor kidney trnsplnt recipients fter desensitiztion therpy. Trnspl Int 14;27: Chung BH, Joo YY, Lee J, et l. Impct of ABO incomptibility on the development of cute ntibody-medited rejection in kidney trnsplnt recipients presensitized to HLA. PLoS One 15;1:e Armstrong KA, Cmpbell SB, Hwley CM, Nicol DL, Johnson DW, Isbel NM. Obesity is ssocited with worsening crdiovsculr risk fctor profiles nd proteinuri progression in renl trnsplnt recipients. Am J Trnsplnt 5;5: Vld G, Ho EK, Vsilescu ER, et l. Relevnce of different ntibody detection methods for the prediction of ntibody-medited rejection nd decesed-donor kidney llogrft survivl. Hum Immunol 9;7: Cro-Oles JL, Gonzlez-Escribno MF, Gonzlez-Roncero FM, et l. Clinicl relevnce of HLA donor-specific

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