Antilymphoid antibody preconditioning and tacrolimus monotherapy for pediatric kidney transplantation

Objective: Heavy post-transplant immunosuppression may contribute to long-term immunosuppression dependence by subverting tolerogenic mechanisms; thus, we sought to determine if this undesirable consequence could be mitigated by pretransplant lymphoid depletion and minimalistic post-transplant monotherapy. Study design: Lymphoid depletion in 17 unselected pediatric recipients of live (n = 14) or deceased donor kidneys (n = 3) was accomplished with antithymocyte globulin (ATG) (n = 8) or alemtuzumab (n = 9). Tacrolimus was begun post-transplantation with subsequent lengthening of intervals between doses (spaced weaning). Maintenance immunosuppression, morbidity, graft function, and patient/graft survival were collated. Results: Steroids were added temporarily to treat rejection in two patients (both ATG subgroup) or to treat hemolytic anemia in two others. After 16 to 31 months (mean 22), patient and graft survival was 100% and 94%, respectively. The only graft loss was in a nonweaned noncompliant recipient. In the other 16, serum creatinine was 0.85 ± 0.35 mg/dL and creatinine clearance was 90.8 ± 22.1 mL/1.73 m2. All 16 patients are on monotherapy (15 tacrolimus, one sirolimus), and 14 receive every other day or 3 times per week doses. There were no wound or other infections. Two patients developed insulin-dependent diabetes. Conclusion: The strategy of lymphoid depletion and minimum post-transplant immunosuppression appears safe and effective for pediatric kidney recipients. © 2006 Elsevier Inc. All rights reserved

Renal transplantation in children managed with lymphocyte depleting agents and low-dose maintenance tacrolimus monotherapy

OBJECTIVE. Describe the safety and efficacy of antithymocyte globulin or alemtuzumab preconditioning, steroid avoidance and reduced calcineurin inhibitor (CNI) immunosuppression in 34 children undergoing renal transplantation. METHODS. ATG (n=8) or alemtuzumab (n=26) were infused at the time of transplantation. This was followed by low-dose twice a day tacrolimus monotherapy with consolidation to once daily dosing by 6 months and once every other day dosing by 12 months. Follow-up ranged from 0.5-2.9 years (mean 1.33 years), with a minimum of 6 months. RESULTS. Both ATG and alemtuzumab were well tolerated. Lymphopenia occurred routinely and resolved after 3-6 months. Acute cellular rejection occurred in 9%; it was related to medical nonadherence in two patients and resulted in one graft loss at 1.5 years. Important adverse events included transient neutropenia in 10 children (none with serious infection), and autoimmune hemolytic anemia in two (resolved with a steroid course in both and conversion to sirolimus in one). Estimated glomerular filtration rate (e-GFR) was stable and averaged 88 mL/min/1.73 m at latest follow-up. Fifteen preadolescents had a greater increase in height Z-score at 1 year (1.3 vs. 0.5, P=0.001), and a higher e-GFR (94.8±21 vs. 76.6±20 ml/min/1.73 m, P<0.05), when compared to case-matched historical controls who were weaned off steroids by 6 months after transplantation and received twice daily tacrolimus monotherapy. CONCLUSION. This simple regimen appears safe, has a low risk for acute cellular rejection or other adverse effects, and is associated with excellent growth and renal function. Such a regimen may also improve compliance and limit CNI nephrotoxicity. © 2007 Lippincott Williams & Wilkins, Inc

Immunoglobulin G, A, and M Responses to BK Virus in Renal Transplantation

Immunoglobulin G (IgG), IgA, and IgM antibodies were measured in serum samples from 71 organ donors, 81 kidney transplant recipients at transplantation, and 67 patients during the posttransplant period by using a virus-like particle-based enzyme-linked immunosorbent assay (ELISA). BK virus (BKV) and JC virus DNA were detected in urine and plasma by real-time PCR. IgG antibodies to BKV were demonstrated in the majority (80.3 to 100%) of patients irrespective of clinical category, but titers were highest in patients with active viral replication. IgA antibodies were present with greater frequency (72.7 to 81.3% versus 0 to 23.6%; P < 0.001) and higher titer (mean optical density, 0.11 to 0.15 versus 0.05 to 0.08; P < 0.001) in patients who were BKV DNA positive than those who were BKV DNA negative. IgM antibodies showed a similar pattern of reactivity but lower frequency in the setting of active viral replication (9.1 to 43.7% versus 0 to 1.4%; P < 0.001). A rise in IgG level of >0.577 optical density (OD) units or a rise in IgA or IgM level of >0.041 OD units was strongly associated with active viral replication. Urine viral load showed a positive correlation with IgM titer (r = 0.22) but a negative correlation with IgG titer (r = −0.28) and IgA titer (r = −0.1). Chronic dialysis patients typically did not have serologic or virologic evidence of active BKV infection. Anti-BKV titers did not rise in patients with JC viruria. In conclusion, measurement of anti-BKV antibody titer and class response can be used to detect the onset of viral replication. ELISAs can be quite specific despite considerable sequence homology between BK virus and JC virus

Phylogenetic Analysis of Polyomavirus BK Sequences

Polyomavirus BK (BKV) has emerged as an important pathogen in kidney transplant patients. Existing taxonomic classifications of BKV come from conventional DNA sequence alignments based on limited data derived from the VP1 gene. We have used a phylogenetic whole-genome approach to examine the pattern of diversity and evolutionary relationships between 45 BKV strains isolated from multiple clinical settings. This analysis supports the classification of BKV into six genotypes, of which types V and VI have not been previously recognized. BKV strains hitherto classified as type I are, in fact, quite heterogeneous, and several cluster with our newly defined genotypes V and VI. The sequence information needed for assigning genotypes can be captured by VP1, VP2, VP3, or large T-gene sequencing. The most polymorphic coding region in the viral genome is VP1, but significant variation is also present in the large T-antigen gene, wherein polymorphisms are found in 11.39% of all nucleotide sites, 46.22% of which are cluster specific. Type-specific amino acid changes within the VP1 region are predicted to map to the BC and DE loops. The number of taxonomically informative amino acid changes in the large T antigen exceeds even that of the VP1 region. Viral strains isolated from healthy subjects and from patients with human immunodeficiency virus infection, Wiskott-Aldrich syndrome, and vasculopathy with capillary leak syndrome formed distinct subclusters. However, within the kidney transplant population, BKV strains derived from patients with asymptomatic viruria did not show complete separation from strains associated with allograft nephropathy

Correlates of Quantitative Measurement of BK Polyomavirus (BKV) DNA with Clinical Course of BKV Infection in Renal Transplant Patients

BK virus-allograft nephropathy (BKVAN) is an increasingly recognized complication after kidney transplantation. Quantitative tests have been advocated to monitor patients, but data demonstrating their efficacy are relatively limited. We developed a real-time PCR assay to quantitate BK virus loads in the setting of renal transplantation, and we correlated the BK virus load with clinical course and with the presence of BK virus in renal biopsy specimens. BK virus loads were measured in urine, plasma, and kidney biopsy samples in three clinical settings: (i) patients with asymptomatic BK viruria, (ii) patients with active BKVAN, and (iii) patients with resolved BKVAN. Active BKVAN was associated with BK viremia greater than 5 × 103 copies/ml and with BK viruria greater than 107 copies/ml in all cases. Resolution of nephropathy led to resolution of viremia, decreased viruria levels, and disappearance of viral inclusions, but low-level viral DNA persisted in biopsy specimens even for patients whose viruria was cleared. All but one patient in the resolved BKVAN group carried a urinary viral load below 107 copies/ml. Viral loads in patients with asymptomatic viruria were generally lower but in some cases overlapped with levels more typical of BKVAN. One patient with asymptomatic viruria and with a viral load overlapping values seen in BKVAN had developed nephropathy by the time of follow-up. In conclusion, serial measurement of viral loads by quantitative PCR is a useful tool in monitoring the course of BK virus infection. The results should be interpreted in conjunction with the clinical picture and biopsy findings

Renal transplantation in children managed with lymphocyte depleting agents and low-dose maintenance tacrolimus monotherapy

Objective. Describe the safety and efficacy of antithymocyte globulin or alemtuzumab preconditioning, steroid avoidance and reduced calcineurin inhibitor (CN!) immunosuppression in 34 children undergoing renal transplantation. Methods. ATG (n=8) or alemtuzumab (n=26) were infused at the time of transplantation. This was followed by low-dose twice a day tacrolimus monotherapy with consolidation to once daily dosing by 6 months and once every other day dosing by 12 months. Follow-up ranged from 0.5-2.9 years (mean 1.33 years), with a minimum of6 months. Results. Both ATG and alemtuzumab were well tolerated. Lymphopenia occurred routinely and resolved after 3-6 months. Acute cellular rejection occurred in 9%; it was related to medical nonadherence in two patients and resulted in one graft loss at 1.5 years. Important adverse events included transient neutropenia in 10 children (none with serious infection), and autoimmune hemolytic anemia in two (resolved with a steroid course in both and conversion to sirolimus in one). Estimated glomerular filtration rate (e-GFR) was stable and averaged 88 mLimin/I.73 m 2 at latest follow-up. Fifteen preadolescents had a greater increase in height Z-score at 1 year (1.3 vs. 0.5, P=O.OOI), and a higher e-GFR (94.8:!:21 vs. 76.6:!:20 mllmin/1.73 m 2 , P&lt;0.05), when compared to case-matched historical controls who were weaned off steroids by 6 months after transplantation and received twice daily tacrolimus mono therapy. Conclusion. This simple regimen appears safe, has a low risk for acute cellular rejection or other adverse effects, and is associated with excellent growth and renal function. Such a regimen may also improve compliance and limit CNI nephrotoxicity

Development of a Loop-Mediated Isothermal Amplification Assay for Rapid Detection of BK Virus

Loop-mediated isothermal amplification (LAMP) is a novel method for rapid amplification of DNA. Its advantages include rapidity and minimal equipment requirement. The LAMP assay was developed for BK virus (BKV), which is a leading cause of morbidity in renal transplant recipients. The characteristics of the assay, including its specificity and sensitivity, were evaluated. BKV LAMP was performed using various incubation times with a variety of specimens, including unprocessed urine and plasma samples. A ladder pattern on gel electrophoresis, typical of successful LAMP reactions, was observed specifically only for BKV and not for other viruses. The sensitivity of the assay with 1 h of incubation was 100 copies/tube of a cloned BKV fragment. Additionally, a positive reaction was visually ascertained by a simple color reaction using SYBR green dye. BKV LAMP was also successful for urine and plasma specimens without the need for DNA extraction. Due to its simplicity and specificity, the LAMP assay can potentially be developed for “point of care” screening of BKV