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Keywords:

  • Allograft rejection;
  • polyoma-BK virus;
  • polyomavirus-associated nephropathy;
  • reduction of immunosuppression;
  • surveillance biopsies

Abstract

  1. Top of page
  2. Abstract
  3. Introduction
  4. Material and Methods
  5. Results
  6. Discussion
  7. Acknowledgment
  8. Disclosure
  9. References

Early detection of polyomavirus BK (BKV) viremia and reduction of immunosuppression is recommended for preventing polyomavirus-associated nephropathy (PyVAN), but systematic histological evaluations were not performed in previous studies. We routinely screen for decoy cells and, if positive, measure plasma BKV-loads. In a cohort of 203 consecutive renal transplantations performed from 2005–2008, 38 patients (19%) developed BKV-viremia and were treated with reduction of immunosuppression. Based on subsequent allograft biopsy results and peak BKV-viremia, patients were assigned to three groups: (i) definitive PyVAN (n = 13), (ii) presumptive PyVAN defined by plasma BKV-loads of ≥4 log10 copies/ml (n = 17) and (iii) low BKV-viremia (n = 8). Clearance of BKV-viremia was achieved in 35/38 patients (92%) and subsequent clinical rejection occurred in 3/35 patients (8.6%), both without any difference among the groups. Patients with definitive PyVAN had higher peak plasma BKV-loads and required longer time for clearance (8.8 vs. 4.6 vs. 2.9 months; p = 0.001). However, allograft function remained stable from baseline to last follow-up at 34 months (range 18–60) in all three groups with median serum creatinine of 1.6 mg/dl, 1.6 mg/dl and 1.3 mg/dl, respectively. We conclude that screening for BKV-replication and reduction of immunosuppression is an effective strategy to preserve medium-term allograft function even in patients developing definitive PyVAN.


Introduction

  1. Top of page
  2. Abstract
  3. Introduction
  4. Material and Methods
  5. Results
  6. Discussion
  7. Acknowledgment
  8. Disclosure
  9. References

Polyomavirus-associated nephropathy (PyVAN) is a major complication after renal transplantation affecting 1–10% of patients (1). Before noninvasive diagnostic tests were routinely used, PyVAN was mostly diagnosed late in an advanced stage with irreversible functional damage, leading to allograft loss in as many as 90% of cases (1,2). Surveillance biopsies have enabled an earlier diagnosis with improved outcome for a significant number of patients (3).

Since PyVAN is preceded by BKV-viruria and viremia (4,5), several groups have used BKV-viremia as a surrogate marker of PyVAN to reduce immunosuppression (6–10). This preemptive approach has emerged as effective and safe to clear BKV-viremia without functional impairment in most cases (6–9). However, it is currently unknown whether this preemptive strategy also leads to favorable outcomes in patients who develop histologically confirmed definitive PyVAN. In the series of 23 adult patients by Hardinger et al. and the 13 pediatric patients by Ginevri et al., a systematic histological evaluation was not performed in the absence of impaired renal function and no case of definitive PyVAN has been diagnosed (6,7). In a prospective study, Almeras et al. reported the outcome of 13 patients with BKV-viremia (8). Three of 13 developed definitive PyVAN with declining allograft function in two cases. Saad et al. reported on 16 patients with definitive PyVAN and 8 patients with BKV-viremia, but the outcomes of these two groups have not been compared (9). Finally, in a cross-sectional study of 204 kidney transplant recipients at a median time of 3 months posttransplant, Viscount et al. identified four cases of PyVAN among 104 biopsies, but the treatment modalities and outcome was not reported (11).

The aim of this study was to analyze the efficacy and safety of a prospectively defined protocol using urinary decoy cells and BKV-viremia to identify and treat patients with active BKV-infection by reducing maintenance immunosuppression.

Material and Methods

  1. Top of page
  2. Abstract
  3. Introduction
  4. Material and Methods
  5. Results
  6. Discussion
  7. Acknowledgment
  8. Disclosure
  9. References

Patient population

We prospectively collected data from 206 patients transplanted between January 2005 and June 2008 at the University Hospital in Basel with a follow-up until December 31, 2009. Three patients were lost during follow-up, leaving 203 patients for the study with follow-up data of at least 18 months.

Immunosuppressive regimens

Immunosuppressive regimens used during the study period were approved by the local institutional review board. Initial immunosuppression was selected based on the presence/absence of donor-specific HLA-antibodies (HLA-DSA), AB0-blood group compatibility and HLA-matching. Recipients of an HLA-identical allograft did not receive any induction therapy. They started on a triple immunosuppression with tacrolimus (Tac; Prograf, Astellas), mycophenolate-mofetil (MMF; CellCept, Roche) and prednisone (P), which was reduced within the first 3 months to a dual therapy with low dose Tac-MMF (Tac trough levels 4–6 ng/ml).

Recipients of an allograft with one or more HLA-mismatches but no HLA-DSA (i.e. normal risk patient) received an induction therapy with basiliximab (Simulect, Novartis) 20 mg on day 0 and 4, and a triple therapy either with Tac-MMF-P or a steroid-free regimen consisting of Tac, mycophenolate-sodium (Myfortic, Novartis) and sirolimus (Sir; Rapamune, Wyeth). Because mycophenolate-mofetil and mycophenolate-sodium can be regarded as equivalent, no distinction is made and both will be abbreviated with MMF. According to the protocols, immunosuppression was modified and reduced within the first 6 months to establish a dual therapy in the long-term as follows: Tac-MMF (i.e. discontinue prednisone or sirolimus) or Sir-MMF (i.e. discontinue tacrolimus). Target trough levels of tacrolimus were 10–12 ng/ml for the first month, 8–10 ng/ml for months two to three, 6–8 ng/ml for months four to six and 4–6 ng/ml thereafter. Target trough levels of sirolimus were 4–8 ng/ml when used in combination with Tac-MMF and 8–12 ng/ml when used in a dual therapy with MMF. MMF was started at 1000 mg bid (720 mg bid for Myfortic).

Recipients of an allograft with HLA-DSA (i.e. high risk patient) received an induction therapy consisting of a polyclonal anti-T-lymphocyte globulin (ATG; ATG-Fresenius, Fresenius Medical Care) 9 mg/kg bw prior to reperfusion of the allograft and 3 mg/kg bw on day 1–4, as well as intravenous immunoglobulins (IvIg) 0.4 g/kg bw on day 0–4 as previously reported (12,13). Maintenance immunosuppression consisted of Tac-MMF-P. Target tacrolimus trough levels were 10–15 ng/ml for the first month, 8–12 ng/ml for months two to three, 6–10 ng/ml for months four to six and 4–8 ng/ml thereafter. Steroids were tapered to 0.1 mg/kg bw by month three posttransplant and maintained at this level. MMF was started at 1000 mg bid.

AB0-incompatible transplants received one dose of rituximab (Mabthera, Roche) four weeks prior to the transplant, pretransplant immunoabsorption, induction therapy with basiliximab and maintenance immunosuppression with Tac-MMF-P as previously reported (14). Target trough levels of tacrolimus were 10–12 ng/ml for the first month, 8–10 ng/ml for months two to three, 6–8 ng/ml for months four to twelve and 4–6 ng/ml thereafter. Steroids were tapered to 0.1 mg/kg bw by month three posttransplant and maintained at this level for the first year posttransplant. MMF was started at 1000 mg bid.

Monitoring strategy for BKV-viruria and BKV-viremia

Screening for BKV-replication was done according to a standard protocol consisting of urine cytology for decoy cells every two weeks until month 3, then at 6 and 12 months and yearly thereafter (5). Patients with positive decoy cells were tested for plasma BKV-loads by quantitative real-time PCR at the next following visit as described previously (5,15). Sustained BKV-viremia was defined as ≥1000 copies/ml in at least two consecutive measurements. In these patients, BKV-viremia was monitored until cleared.

Evaluation of allograft biopsies

Clinically indicated allograft biopsies were performed when serum creatinine increased by >20% from baseline. Surveillance biopsies were scheduled at 3 and 6 months posttransplant. All biopsy specimens (two cores obtained with a 16 gauge needle) were evaluated by light microscopy, immunofluorescence (C4d, HLA-DR) and immunohistochemistry (SV40 large T-antigen). Findings were graded according to the updated Banff 2007 classification (16). Definitive PyVAN was defined by positive SV40-staining ± viral cytopathic changes. In patients with positive BKV-viremia at the time of the biopsy, tubulointerstitial inflammation was attributed to BKV, even in the absence of cytopathic changes and negative SV40-staining and was treated with reduction of immunosuppression as described below. In patients with negative BKV-viremia, tubulointerstitial inflammation was regarded as rejection.

Definition of BKV replication and nephropathy

Based on histological evaluation and peak plasma BKV-load, three groups were defined (17,18):

  • 1
    definitive PyVAN: Patients with BKV-viremia and an allograft biopsy demonstrating positive SV40-staining ± cytopathic changes.
  • 2
    presumptive PyVAN: Patients with peak BKV-viremia ≥4 log10 copies/ml but no histological features of PyVAN (i.e. negative SV40-staining and no cytopathic changes).
  • 3
    low BKV-viremia: Patients with peak BKV-viremia <4 log10 copies/ml and no histological features of PyVAN.

Treatment protocol

In patients with sustained BKV-viremia (i.e. ≥1000 copies/ml) immunosuppression was reduced as follows:

  • 1
    Step 1: Tac trough levels were set one step lower as intended by the protocol and targeted as predefined for the next time period (e.g. in a normal risk patient at month two posttransplant trough levels were reduced from 8–10 ng/ml to 6–8 ng/ml).
  • 2
    Step 2: If BKV-viremia did not constantly decrease, Tac trough levels were further reduced by one step as predefined for the next time period (e.g. in the example discussed above from 6–8 ng/ml to 4–6 ng/ml).
  • 3
    Step 3: If BKV-viremia did not constantly decrease, the dosing of MMF was reduced in steps of 50% (e.g. from 1000 mg bid to 500 mg bid to 250 mg bid for Cellcept or from 720 mg bid to 360 mg bid to 180 mg bid for Myfortic).

Evaluated outcomes

The following outcomes were evaluated: (i) clearance rate of BKV-viremia, (ii) frequency of clinical and subclinical rejection after clearance of BKV-viremia, (iii) evolution of allograft function during and after BKV-viremia and (iv) allograft and patient survival.

Statistical analysis

We used JMP software version 8.0 (SAS Institute Inc., Cary, NC) for statistical analysis. For categorical data, Fisher's exact test or Pearson's chi-square test were used. Parametric continuous data were analyzed by Student t-tests. For nonparametric continuous data, the Wilcoxon rank sum test was used. If not indicated otherwise, continuous data are given as median (range). Survival analysis was performed by the Kaplan-Meier method and groups compared using the log-rank test. A p-value < 0.05 was considered to indicate statistical significance.

Results

  1. Top of page
  2. Abstract
  3. Introduction
  4. Material and Methods
  5. Results
  6. Discussion
  7. Acknowledgment
  8. Disclosure
  9. References

Characteristics of the patient cohort

Thirty eight of 203 patients (19%) experienced BKV-viremia, 165 patients (81%) did not. Baseline characteristics of these two groups are detailed in Table 1. Male gender was more frequent among patients with BKV-viremia than in patients without BKV-viremia (84% vs. 64%; p = 0.02). There were no further differences between the two groups regarding immunological and nonimmunological parameters as well as initial immunosuppressive therapy.

Table 1.  Baseline characteristics of the whole cohort consisting of 203 patients, grouped by the presence/absence of sustained BKV-viremia
 BKV-viremia (n = 38)No BKV-viremia (n = 165)p-level
Recipient
 Sex, males (%)32 (84)106 (64)0.02
 Age47 (22–72)55 (18–73)0.10
Donor
 Deceased donor, n (%)20 (53)83 (50)0.86
 Age49 (1–77)53 (1–85)0.06
 Sex, females (%)19 (51)86 (53)0.86
Transplant, n with 1st/2nd/3rd32/5/1142/21/20.80
AB0-incompatible transplant, n (%)4 (11)9 (5)0.27
HLA-mismatches
 A, n with 0/1/2 mismatches7/21/1028/87/500.89
 B, n with 0/1/2 mismatches4/14/2017/75/730.61
 DRβ1, n with 0/1/2 mismatches4/20/1427/82/560.66
 Total mismatches, mean ± std.3.8 ± 1.33.6 ± 1.50.64
Induction therapy
 None, n (%)1 (2)7 (4)0.81
 Basiliximab, n (%)31 (82)131 (79) 
 ATG/IvIg, n (%)6 (16)27 (17) 
Initial immunosuppression
 CyA-MMF-P, n (%)03 (2)0.68
 Tac-MMF-P, n (%)26 (68)98 (59) 
 Tac-MMF-Sir, n (%)12 (32)64 (39) 

Categories of active BKV-infection

The patient flow algorithm is detailed in Figure 1. Despite reduction of immunosuppression upon detection of sustained BKV-viremia, 13 patients developed definitive PyVAN giving an overall incidence of 6.4% in the study population. Diagnosis was made at 3.5 months posttransplant (range 2.2–6.9) with median log10 BKV-loads of 5.84 copies/ml (range 4.62–6.68). Seventeen patients developed presumptive PyVAN. In 15 of these 17 patients (88%), 24 biopsies were performed during documented BKV-viremia with median log10 BKV-loads of 3.74 copies/ml (range 2.74–6.66); in 2/17 patients (12%) no allograft biopsies could be obtained during BKV-viremia. Eight patients had only low BKV-viremia. In 4 of these 8 patients (50%), 6 biopsies were performed during documented BKV-viremia with median log10 BKV-loads of 3.37 copies/ml (range 2.32–3.99).

image

Figure 1. Patient flow algorithm.

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Course of BKV-viremia and therapeutic interventions

Comparing the three patient groups, we found no differences regarding gender, recipient age, HLA-matching, initial immunosuppression, frequency of rejection episodes prior to BKV-viremia and immunosuppression at first BKV-viremia (Table 2). Also, median time to first and peak BK-viremia were statistically not different (2.1 and 4.0 months in the definitive PyVAN group, 2.6 and 3.0 months in the presumptive PyVAN group and 4.3 and 4.7 months in the low BK-viremia group). Peak BKV-loads were significantly different among the three groups and highest in the definitive PyVAN group (p < 0.0001; Table 2).

Table 2.  Immunosuppression and viral dynamics among the three categories of active BKV-infection
 Definitive PyVAN (n = 13)Presumptive PyVAN (n = 17)Low BKV-viremia (n = 8)p-level
Induction therapy
 None, n (%)1 (6)0.15
 Simulect, n (%)9 (69)16 (94)6 (75) 
 ATG/IvIg, n (%)4 (31)2 (25) 
Initial immunosuppression
 Tac-MMF-P, n (%)10 (77)11 (65)5 (63)0.71
 Tac-MMF-Sir, n (%)3 (23)6 (35)3 (37) 
Rejection prior to BKV-viremia, n (%)3 (23)4 (24)3 (38)0.72
Immunosuppression at first BKV-viremia
 Tac-MMF-P, n (%)101340.57
 Tac-MMF-Sir, n (%)211 
 Tac-Sir-P, n (%)11 
 Tac-MMF, n (%)11 
 Tac-P, n (%)11 
 Sir-MMF, n (%)1 
Course of BKV-infection (months)
 Transplant to first decoy cells1.8 (0.7–2.5)1.6 (0.6–8.4)3.9 (1.2–6.6)0.07
 Transplant to first BKV-viremia2.1 (1.2–3.8)2.6 (0.9–13.1)4.3 (1.8–7.0)0.05
 Transplant to peak BKV-viremia4.0 (1.6–7.7)3.0 (1.2–13.1)4.7 (1.8–7.3)0.70
Peak BKV-viremia (log10 copies/ml)6.5 (5.3–7.3)5.0 (4.3–6.7)3.9 (3.7–4.0)<0.0001

In 17/38 patients (45%) Tac trough levels were reduced by one step, in 13/38 patients (34%) Tac trough levels had to be reduced by two steps. In 3/38 patients (8%) there was no additional reduction of immunosuppression required than predefined in the standard procedure. In 5/38 patients (13%) other interventions were made: in two patients on Tac-Sir, Tac trough levels were reduced by one step and Sir either stopped or switched to MMF; in two patients on Tac-Sir and Tac-MMF, Tac trough levels were not changed but Sir was stopped and MMF reduced, respectively; in one patient on Sir-MMF, MMF was switched to azathioprine.

In 7/38 patients (18%) the treating physicians deviated from the standard procedure of exclusive reduction of immunosuppression because concurrent rejection was assumed. One patient with pretransplant HLA-DSA received rituximab and IvIg for concurrent antibody-mediated rejection during BKV-viremia; another patient was treated with steroid pulses for concurrent endothelialitis; five patients were treated with steroid pulses for presumed concurrent tubulointerstitial rejection. Treatment of tubulointerstitial rejection was driven by the judgment of the treating physician and was not based on any additional histological or clinical parameters. The frequency of rejection therapy was not different across the three groups (2/13 in the definitive PyVAN, 4/17 in the presumptive PyVAN, 1/8 in the low BK-viremia group; p = 0.76). Following rejection treatment, a transient increase and persistence of BKV-viremia was observed in 1/7 patients, while BKV-viremia further decreased and cleared in the other 6/7 patients.

Clearance of BKV-viremia

Clearance of BKV-viremia was observed in 35/38 (92%) patients. The frequency of clearance was not different across the three groups (p = 0.60; Table 3). The median time from first BKV-viremia to clearance of BKV-viremia was 8.8 months in the definitive PyVAN, 4.6 months in the presumptive PyVAN and 2.9 months in the low BKV-viremia group (p = 0.001; Table 3). Reduction of immunosuppression to achieve clearance of BKV-viremia was most extensive in patients with definitive PyVAN (Table 3).

Table 3.  Clearance of BKV-viremia and subsequent acute rejection episodes
 Definitive PyVAN (n = 13)Presumptive PyVAN (n = 17)LowBKV-viremia (n = 8)p-level
  1. 1Tubulitis Ia (n = 1) and tubulitis Ib (n = 2). All treated with steroid pulses.

  2. 2Borderline tubulitis (n = 3; one treated with steroid pulses), tubulitis Ia (n = 2; one treated with steroid pulses), tubulitis Ib (n = 1; treated with steroid pulses), antibody-mediated rejection in a patient with pretransplant HLA-DSA (n = 1; treated with steroid pulses, IvIg, and rituximab).

Clearance of BKV-viremia, n (%)12 (92)15 (88)8 (100)0.60
Months from first BKV-viremia to BKV-clearance8.8 (2.8–18.5)4.6 (1.2–23.3)2.9 (0.9–4.6)0.001
Reduction of immunosuppression to achieve
 BKV-clearance, n (%)
   Step 12 (17)8 (53)8 (100)0.001
   Step 26 (50)7 (47) 
   Step 34 (33) 
Rejection after BKV-clearance, n (%)
 Clinical rejection (clinical bx)11 (8)1 (7)1 (12)0.67
 Subclinical rejection (surveillance bx)24 (33)3 (20) 
 No rejection (surveillance bx)2 (17)3 (20)3 (38) 
 Stable creatinine (no bx)5 (42)8 (53)4 (50) 
Immunosuppression at last follow-up
 Tac-MMF-P, n4620.72
 Tac-MMF, n796 
 Tac-Aza, n1 
 Tac-P, n1 
 Sir-Aza, n1 
 Sir-P, n1 

In three patients BKV-viremia significantly decreased (>2 log10 copies/ml) but did not fully clear at the end of follow up. One patient with definitive PyVAN and a peak BKV-viremia of 6.11 log10 copies/ml still had 3.55 log10 copies/ml 35 months after first BKV-viremia on a low-dose Tac-MMF-P therapy (Tac trough levels of 4–6 ng/ml). Triple therapy was continued due to ongoing concurrent antibody-mediated rejection. Another patient with presumptive PyVAN and a peak BKV-viremia of 6.02 log10 copies/ml still had 3.37 log10 copies/ml 27 months after first BKV-viremia on a low-dose Tac-MMF dual therapy (Tac trough levels 4–6 ng/ml; MMF dose 250 mg bid). The last patient also had presumptive PyVAN with a peak BKV-viremia of 6.66 log10 copies/ml. Thirty-two months after first BKV-viremia, he still had 4.35 log10 copies/ml on a dual therapy with Sir-P (Sir trough levels <3 ng/ml and 7.5 mg prednisone) which was given after diagnosis and treatment for a posttransplant lymphoma.

Rejection episodes after clearance of BKV-viremia

At least one allograft biopsy was performed in 18/35 patients (51%) clearing BKV-viremia. Three of them were clinically indicated and revealed tubulitis Ia (n = 1) or Ib (n = 2) accounting for a biopsy-proven clinical rejection rate of 8.6%. In the other 15 patients with stable renal function, eight had no rejection in surveillance biopsies, three demonstrated subclinical borderline tubulitis, two subclinical tubulitis Ia, one a subclinical tubulitis Ib and one patient with pretransplant HLA-DSA had subclinical antibody-mediated rejection. Clinical and subclinical rejection episodes were not different among the three groups of active BKV-infection. All three patients with clinical rejection and 4/7 patients with subclinical rejection were treated (Table 3). Following rejection treatment, a short-term rebound of low BKV-viremia (i.e. <4 log10 copies/ml) with re-clearance occurred in two patients. Immunosuppression at last follow-up was also not different across the three groups (Table 3).

Evolution of allograft function

At baseline, serum creatinine was not different among the three groups (definitive PyVAN: median 1.5 mg/dl (range 1.1–2.9); presumptive PyVAN: median 1.5 mg/dl (range 0.8–4.6); low BKV-viremia: median 1.3 mg/dl (range 1.2–2.6); p = 0.39). Three of 13 patients (23%) with definitive PyVAN, 2/17 patients (12%) with presumptive PyVAN and no patient in the low BKV-viremia group had elevated serum creatinine levels during the course of BKV-viremia. At last follow-up serum creatinine was also not different between the three groups (definitive PyVAN: median 1.6 mg/dl (range 0.8–2.7); presumptive PyVAN: median 1.6 mg/dl (range 1.0–3.2); low BKV-viremia: median 1.3 mg/dl (range 0.9–2.3); p = 0.10). Serum creatinine values at last follow-up and at baseline were not different in the definitive and presumptive PyVAN groups (p = 0.77 and p = 0.60), while they were slightly lower in the low BKV-viremia group (p = 0.02; Figure 2).

image

Figure 2. Evolution of serum creatinine in the three categories of active BKV-infection. Serum creatinine values were recorded at five specific time points (i.e. at baseline 1st month, at first BKV-viremia, at peak BKV-viremia, at clearance of BKV-viremia and at last follow-up). Serum creatinine values at baseline 1st month and at last follow-up were compared by paired t-tests and the p-values are given for each category of active BKV-infection. The time scale is given as months posttransplant in the bottom line.

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When patients were grouped according to peak BKV-viremia, eight had less than 4 log10 copies/ml (low BKV-viremia), 17 had 4 to 5.99 log10 copies/ml (medium BKV-viremia) and 13 had ≥6 log10 copies/ml (high BKV-viremia). Serum creatinine levels within these three groups were not different at the above mentioned time points (p ≥ 0.10). Serum creatinine values at last follow-up and at baseline were not different in the medium and high BKV-viremia groups (p = 0.37 and p = 0.29), while they were slightly lower in the low BKV-viremia group (p = 0.02; data not shown).

Tubular expression of HLA-DR and evolution of allograft function

In the reported 38 patients, 57 biopsies were obtained during BKV-viremia. HLA-DR expression on tubular epithelial cells was positive in 20 biopsies (35%), negative in 34 biopsies (60%) and not available in three cases (5%). Six of 7 biopsies (86%) that were treated with additional steroids demonstrated positive HLA-DR staining (all focal positive), while 14/50 biopsies (28%) not treated as rejection were HLA-DR positive (2 diffuse positive, 12 focal positive) (p = 0.01). In patients treated with steroids, creatinine values remained stable in the six patients with positive HLA-DR staining (mean 1.9 mg/dl at biopsy and 1.8 mg/dl at clearance of BKV-viremia; paired t-test p = 0.64) and also in the one patient with negative HLA-DR staining. In patients not treated with steroids, creatinine values remained stable in patients with positive HLA-DR staining (mean 2.4 mg/dl at biopsy and 1.8 mg/dl at clearance of BKV-viremia; p = 0.14) and in patients with negative HLA-DR staining (mean 1.7 mg/dl at biopsy and 1.6 mg/dl at clearance of BKV-viremia; p = 0.54).

Patient and allograft survival

After a median follow-up time of 34 months (range 18–60), there was no graft loss in the 38 patients with BKV-viremia. Three of 38 patients died with functioning allograft and after clearance of BKV-viremia (two males suffered cardiac death and one female died due to urothelial carcinoma). All deceased patients were in the definitive PyVAN group.

Patient survival at one and three years posttransplant were 100% and 92% in the BKV-viremia group and 99% and 97% in the no BKV-viremia group, respectively (p = 0.13). Death-censored allograft survival at one and three years posttransplant were both 100% in the BKV-viremia group and 97% and 96% in the no BKV-viremia group, respectively (p = 0.20).

Discussion

  1. Top of page
  2. Abstract
  3. Introduction
  4. Material and Methods
  5. Results
  6. Discussion
  7. Acknowledgment
  8. Disclosure
  9. References

The main observation in this study was that screening for active BKV-infection along with reduction of immunosuppression is an effective strategy to preserve allograft function in all degrees of active BKV-infection. While this could have been inferred by two previous studies (7,19), our data provide the histopathological evidence due to systematic performance of surveillance allograft biopsies. Brennan et al. and Ginevri et al. have suggested that preemptive reduction of immunosuppression in patients with BKV-viremia prevented development of definitive PyVAN (7,19). However, our data rather indicate that definitive PyVAN can still occur, but mainly with stable allograft function and only detectable by surveillance biopsies.

Active BKV-infection mostly occurs within the first 6 months posttransplant and can be easily detected by noninvasive assays in the urine or plasma (5,7,8,19). In our cohort of 203 consecutive patients, baseline characteristics—except for a male predominance—and immunosuppressive regimens were neither associated for the occurrence of sustained BKV-viremia nor for the degree of BKV-infection. This is consistent with two prospective studies (7,19) and a large retrospective matched control cohort study (20). By contrast, several risk factors for development of definitive PyVAN such as higher Tac levels, previous treated rejection episodes, induction therapy with ATG and poor HLA-matching have been described (21–26). Given the heterogeneity of risk factors of varying predictive power, screening for active BKV-infection should include all patients in the first 1–2 years posttransplant and ideally with a higher sampling density during the more intense immunosuppressive phase in the first 6 months posttransplant.

Early detection of sustained BKV-viremia is very likely the key step for the success of immunosuppression reduction strategies. Indeed, all studies—including ours—that report favorable outcomes of immunosuppression reduction, detected BKV-viremia early posttransplant allowing for timely therapeutic intervention (7,8,19). However, if the diagnosis of definitive PyVAN is made at a later time point, reduction of immunosuppression is less successful, most likely due to more advanced irreversible damage of the allograft (2,20).

Clearance of BKV-viremia in our study was 92% and independent of the degree of BKV-infection. In a few patients BKV-viremia persisted for more than 27 months after first appearance despite reduction of immunosuppression to presumably the lowest possible levels. One patient with ongoing subclinical antibody-mediated rejection and BKV-viremia was especially challenging, because immunosuppression had to be adapted carefully according to the leading process. Such patients with sustained BKV-viremia and a significant risk for aggravation of the concurrent rejection process upon reduction of immunosuppression might be candidates for an adjunct antiviral therapy such as cidofovir (27,28) or leflunomide (29). Unfortunately, these drugs can have severe adverse effects and their efficacy to clear active BKV-infection has not been well studied (18). Another option for such ‘difficult-to-treat’ patients is the acceptance of low level BKV-viremia, because low BKV-replication within the allograft might not dramatically shorten its lifespan (30).

Our immunosuppression reduction strategy was primarily based on lowering Tac levels. In vitro data indicate that Tac is the strongest inhibitor of BKV-specific T cells and thus reducing calcineurin inhibitor levels is a reasonable first step to lower overall immunosuppression in patients with active BKV-infection (10,31). Another group first reduced or discontinued the anti-proliferative drug (i.e. MMF or azathioprin) and then the calcineurin-inhibitor with similar results compared to our study (19). Therefore, reducing the overall immunosuppressive burden is likely more important than lowering or discontinuing a specific drug.

While the overall strategy of reducing immunosuppression seems to be operating well, the presence of acute rejection concurrent with PyVAN remains a significant challenge. Seven of 38 patients (18%) in our study were treated for presumed concurrent rejection while BKV-viremia was present. Histopathological features of antibody-mediated rejection or the presence of endothelialitis are regarded as strong indicators for concurrent rejection. However, tubulointerstitial inflammation is a common observation in patients with BKV-viremia or PyVAN and it is currently impossible to distinguish a BKV-specific infiltrate from an allo-specific infiltrate (32,33). In previous studies, tubular HLA-DR expression was proposed as a potential marker to separate these two different entities (34). Our current data did not show an association of mostly focal HLA-DR expression and allograft function in response to reduction of immunosuppression alone or with added steroids. Clearly, distinction between BKV-specific and allo-reactive infiltrates in patients with BKV-viremia is a challenging and unresolved problem (35–37), which will require larger investigations with longitudinal histological and viral dynamic data as well as a standardized treatment protocol. In such a context, predictive markers for concurrent tubulointerstitial rejection could be evaluated with much better accuracy.

The rate of clinical allograft rejection after clearance of BKV-viremia was 8.6% and independent of the degree of BKV-infection. The observed clinical rejection rate is within the range (0–23%) reported in the literature (7–9,19). In addition, subclinical tubulitis was detected in 6 of 14 patients (43%), who had surveillance biopsies with stable allograft function after clearance of BKV-viremia. It is currently unknown whether these subclinical infiltrates are allo-specific (i.e. rejection) or represent ‘residual’ BKV-specific inflammation that requires more time to completely resolve. Three of these six patients were treated with steroid pulses for presumed rejection. Interestingly, all six patients maintained stable allograft function and none experienced recurrent BKV-viremia. Nevertheless, the risk of rejection after clearance of BKV-viremia is not negligible and patients need to be followed carefully around the clearance of BKV-viremia. Eventually, the dynamic of BKV clearance and monitoring of BKV-specific cellular immune responses might help to guide immunosuppression during this vulnerable phase in order to minimize rejection episodes after clearance of BKV-viremia (7,38).

Patients with definitive PyVAN had higher peak BKV-viremia and required more time to clear it than patients with presumptive PyVAN and low BKV-viremia. Together this indicates a markedly increased overall viral burden, which might induce more allograft damage due to direct and indirect effects (39). Although allograft function remained stable in patients with definitive PyVAN, these favorable results need to be confirmed in a larger population with longer follow-up.

In conclusion, our study indicates that early detection of active BKV-infection along with reduction of immunosuppression is an effective strategy to preserve medium-term allograft function independent of the severity of BKV-infection and even in patients with definitive PyVAN. However, patients with definitive PyVAN have a longer and more extensive period of BKV-viremia, which might induce allograft damage that could affect long-term survival.

Acknowledgment

  1. Top of page
  2. Abstract
  3. Introduction
  4. Material and Methods
  5. Results
  6. Discussion
  7. Acknowledgment
  8. Disclosure
  9. References

SS and HHH are supported by the Swiss National Foundation (grant number 32473B_125482 and 3200B0–110040).

Disclosure

  1. Top of page
  2. Abstract
  3. Introduction
  4. Material and Methods
  5. Results
  6. Discussion
  7. Acknowledgment
  8. Disclosure
  9. References

The authors of this manuscript have no conflicts of interest to disclose as described by the American Journal of Transplantation.

References

  1. Top of page
  2. Abstract
  3. Introduction
  4. Material and Methods
  5. Results
  6. Discussion
  7. Acknowledgment
  8. Disclosure
  9. References