BK Virus Nephritis Research Articles

Familial CD45RA- T cells to treat severe refractory infections in immunocompromised patients.

Immunocompromised patients are susceptible to high-risk opportunistic infections and malignant diseases. Most antiviral and antifungal drugs are quite toxic, relatively ineffective, and induce resistance in the long term. The transfer of pathogen-specific Cytotoxic T-Lymphocytes has shown a minimal toxicity profile and effectiveness in treating Cytomegalovirus, Adenovirus, Epstein - Barr virus, BK Virus and Aspergillus infections, but this therapy have the main limitations of regulatory issues, high cost, and absence of public cell banks. However, CD45RA- cells containing pathogen-specific memory T-cells involve a less complex manufacturing and regulatory process and are cheaper, feasible, safe, and potentially effective. We present preliminary data from six immunocompromised patients: four who had severe infectious diseases and two who had EBV lymphoproliferative disease. All of them underwent multiple safe familial CD45RA- T-cell infusions as adoptive passive cell therapy, containing Cytomegalovirus, Epstein - Barr virus, BK virus, and Aspergillus-specific memory T-cells. We also present the method for selecting the best donors for CD45RA- cells in each case and the procedure to isolate and store these cells. The infusions were safe, there was no case of graft-versus host disease, and they showed a clear clinical benefit. The patients treated for BK virus nephritis, Cytomegalovirus encephalitis, Cytomegalovirus reactivation, and disseminated invasive aspergillosis experienced pathogen clearance, complete resolution of symptoms in 4-6 weeks and a lymphocyte increase in 3 of 4 cases after 3-4 months. Donor T cell transient microchimerism was detected in one patient. The two patients treated for EBV lymphoproliferative disease underwent chemotherapy and several infusions of CD45RA- memory T-cells containing EBV cytotoxic lymphocytes. Donor T-cell microchimerism was observed in both patients. The viremia cleared in one of the patients, and in the other, despite the viremia not clearing, hepatic lymphoproliferative disease remained stable and was ultimately cured with EBV-specific Cytotoxic T-Lymphocytes. The use of familial CD45RA- T-cells containing specific Cytotoxic T-lymphocytes is a feasible, safe and potential effective approach for treating severe pathogen infections in immunocompromised patients through a third party donor. Furthermore, this approach might be of universal use with fewer institutional and regulatory barriers.

POS-794 Conversion to sirolimus in kidney transplant patients: the main indications

The mTOR inhibitors (Sirolimus) has been used in kidney transplantation for very specific indications, essentially in case of neoplastic complications after transplantation secondary to calcineurin inhibitors.Then, the indications have been extended.To describe the different indications of sirolimus conversion in renal transplant patients in Tunisia, we carried out a multicenter study. This is a retrospective multicentric cross-sectional and analytical study performed over a period of 30 years (from 1986 to 2017), which collected all kidney transplant patients who received mTOR inhibitors during any time of their follow-up for a minimum period of 6 months. During study period, 89 patients were collected from the different Tunisian renal transplantation units. The majority of patients came from the SAHLOUL and CHARLES NICOLLES hospitals. The average age of patients at the time of transplantation was 34.71 years. Seventy percent were male. Initial nephropathy was indeterminate in 39 % of patients. All patients received sirolimus during their post-transplant follow-up with a mean of 3.13 +/- 4.73 years. The first indication (49% of cases) of Sirolimus initiation in our study was the diagnosis of neoplastic and pre-neoplastic lesions. Neoplastic lesions were noted in 36 patients (40%) and pre-neoplastic lesions in 08 patients. The most frequent neoplasia was Kaposi's sarcoma in diagnosed in 22.5% of patients. The second indication for switching to sirolimus was calcineurin inhibitors toxicity. In fact, 21 patients in our study had received Sirolimus because of calcineurin inhibitors toxicity. Tacrolimus toxicity was noted in 13 patients (15%) and Ciclosporin toxicity was found in 08 patients (9%). On the other hand, seven patients in our study had received Sirolimus after renal transplantation because of BK virus nephritis. Two patients had presented a thrombotic micro angiopathy related to tacrolimus requiring switching to Sirolimus. One patient presented with severe leuko-neutropenia with white blood cell count of 880/mm³ and neutrophils of 150/mm³. The etiological investigation was negative. In front of persistence of this leuko-neutropenia despite decreasing and then withdrawing MMF, a switch to Sirolimus was decided. In addition, Sirolimus was administered to one patient because of a primary CMV infection that occurred after 04 months of renal transplantation and was resistant to treatment with Foscavir and Ganciclovir. Furthermore, 13 patients had been programmed to be converted to Sirolimus. The conversion was decided, in 04 cases, in front of past history of lesions providing degeneration before the transplantation. These antecedents were: villous atrophy, a colloid thyroid nodule, Bourneville's tuberous sclerosis causing degeneration of skin nodules, and a mixed germ cell tumor. In 09 patients, the systematic conversion to Sirolimus was a part of a national protocol. Conversion from calcineurin inhibitor to sirolimus is one of the major thrusts of the current development of immunosuppression in kidney transplantation. The aim is to minimize the side effects of calcineurin inhibitors and ensure better graft and patient survival. According to our study, neoplastic complications and calcineurin inhibitors toxicity are the main indications of conversion to Sirolimus in Tunisia.

Comparable Outcome of CD3+-/CD19+ Depleted Haploidentical with Matched Sibling Donor Hematopoietic Stem Cell Transplantation for Advanced Stage Sickle Cell Disease: Result of a Pilot Study

IntroductionSickle cell disease (SCD) is an inherited disorder with an estimate of 300.000 affected newborns per year worldwide. Despite significant improvements in preventive measures, supportive care and conventional therapy, substantial morbidity and mortality with a reduced life expectancy persists. Allogeneic hematopoietic stem cell transplantation (HSCT) with a matched sibling donor (MSD) is currently the curative standard of care for SCD patients (pts). But MSD and unrelated matched donor availability is <20% and graft- versus-host disease (GvHD) remains to be associated with significant morbidity and mortality with T cell repleted grafts and standard GvHD prophylaxis, which is unacceptable, particularly in pts with non-malignant diseases. A T cell depleted haploidentical HSCT (T-haplo-SCT) expands donor availability while exhibiting low GvHD rates. In our single center pilot study, we compared T-haplo-SCT with a MSD HSCT in pts with advanced stage SCD, using almost identical transplant conditions except donor source and GvHD prophylaxis.Methods17 pts were transplanted for advanced stage SCD-related complications. Five SCD pts received bone marrow (BM) from a MSD and 12 peripheral blood stem cells (PBSC) from a haploidentical donor. The conditioning regimen for both groups consisted of thiotepa (2x5mg/kg), fludarabine (4x40mg/m2), treosulfan (3x14g/m2) and anti-thymocyte globulin (ATG, Neovii, 3x15mg/kg), given on day -9 to -7 for T-haplo-SCT and on days -3 to -1 for MSD. In addition, Cyclosporine A (CSA) or Tacrolimus and MMF was applied for GvHD prophylaxis. All pts underwent exchange transfusions prior to conditioning and means of fertility preservation. The MSD pts included four females and one male, median age 22 years (range 9 to 25 years). Three suffered from homozygous SCD, two from HbS/ß-thalassemia. The BM graft consisted of a median of 2,2 x 106 CD34+ cells/kg bodyweight, with a A/S in three and a A/A genotype in two donors. In T-haplo-SCT, five were females and seven were males. Median age was 18 yrs (range 3 to 28 yrs). Nine had homozygous SCD, three HbS/ß-thalassemia. The PB grafts consisted of a median of 12,2 x 106 CD34+ cells, 11,2 x 103 CD3+ and 23,3 x 103 CD19+ cells/kg. Haploidentical donors were fully haplotype mismatched relatives (1 mother, 8 fathers, 1 cousin, 1 sister and one step sister). Donor genotype was Hb A/A in one and Hb A/S in all others.ResultsThe median follow-up was 21 months in the MSD group and 26 months in T-haplo-SCT group. The median hospitalization was 38 days for both. The median time to engraftment was faster in T-haplo-SCT versus MSD: 16 versus 23 days for leukocytes, 18 versus 23 days for granulocytes and 11 versus 21 days for thrombocytes. The median T (>500/µl) and B cell recovery (>100/µl) in T-haplo-SCT versus MSD was 115 versus 103 days and 85 versus 140 days, respectively.Two pts in the T-haplo-SCT group developed a stable mixed PB chimerism with no SCD-related complications off immunosuppression. Lineage specific chimerism analyses of red cell precursors (CD235a, glycophorine A) in the BM of these pts showed a complete red cell chimerism. The median HbS level off immunosuppression was comparable in both groups (37-38%). In all 17 pts, the conditioning regimen was well tolerated. Mild veno-occlusive disease (VOD/SOS) was diagnosed in 1 MSD patient and a transient thromboembolic event in one T-haplo-SCT patient. One patient developed a life threatening capillary leak syndrome during ATG treatment and subsequent PRES and was rescued by a switch to everolimus MMF prophylaxis. One patient from the T-haplo-SCT succumbed to a CMV pneumonitis combined with rotavirus gastroenteritis. None of our pts developed a grade III-IV acute GvHD. Five T-haplo-SCT and one MSD patient developed steroid responsive grade I-II acute GvHD. One of five MSD and three of twelve T-haplo-SCT pts developed mild to moderate steroid responsive chronic GvHD, which resolved in the meantime. CMV and EBV reactivations were observed but could be treated with standard of care except one patient mentioned above. One patient after T-haplo-SCT developed BK virus nephritis. All pts are stably engrafted off immunosuppression and free of SCD-related complications.ConclusionsIn our experience T-haplo-SCT with CD3+/CD19+ depleted PBSC is safe and efficient with a very low incidence of GvHD and is at least comparable to MSD HSCT, even for advanced stage SCD pts. DisclosuresCorbacioglu:Jazz Pharmaceuticals: Consultancy, Honoraria.

Challenges of BK Viremia in Heart Transplant Recipient with Multiple Comorbidities

<h3>Introduction</h3> BK viremia is rarely reported in non-kidney solid organ transplant recipients. We present a case of a heart transplant recipient with BK viremia. We highlight the challenges of treating BK viremia in a patient with multiple post-transplant morbidities. <h3>Case Report</h3> Our patient is a 16 year old male with history of complex single ventricle anatomy status post heart transplant due to failing Fontan physiology with protein losing enteropathy (PLE). His early post-transplant course was complicated by continued PLE, post-transplant lymphoproliferative disorder (PTLD), and acute kidney injury requiring temporary hemodialysis for nine months. Three years post-transplant, he presented with worsening renal function and proteinuria in the setting of relapsed PTLD. Serum BK virus was found to be positive at high viral loads. Antiviral therapy with leflunomide was initiated. He underwent renal biopsy that showed 70% glomerulosclerosis with BK inclusions consistent with BK virus nephritis. Adjunct therapy with cidofovir was started, but was discontinued due to its side effect of nephrotoxicity. He is currently undergoing treatment with rituximab and virus specific T-cells to both Epstein-barr and BK viruses with a promising initial response. This patient's history of chronic kidney disease and long-term immunosuppressive therapy including multiple exposures to nephrotoxic agents makes the decision of how to treat his BK viremia a challenge. <h3>Summary</h3> BK viremia in post heart transplant recipients is not commonly reported, but poses a challenge to management given the limited therapeutic options available. Development of standard monitoring protocols as well as refinement of treatment options is necessary in this patient population that is at high risk for acute and chronic kidney injury.

BK Virus Pneumonia with CMV Colitis in a Kidney Transplant Recipient: Successful Treatment with Non-Cidofovir Based Therapy

BK virus (BKV) pneumonia is a rare entity especially seen in immunosuppressed patients, for which cidofovir is the used treatment option. We describe a case of a young female patient who presented for altered kidney function six months following kidney transplantation for focal segmental glomerulosclerosis and was found to have BKV nephritis. Her in-hospital stay was complicated by BKV pneumonia requiring mechanical ventilation, in addition to CMV colitis. She was treated with leflunomide/ciprofloxacin and ganciclovir for her pneumonia and colitis, respectively. The patient improved clinically except that her kidney function deteriorated. Leflunomide/ciprofloxacin combination may constitute an effective and safe alternative to cidofovir for the treatment of BKV pneumonia, in particular when cidofovir is not available. J Microbiol Infect Dis 2021; 11(1):36-41.

Haploidentical CD3 or α/β T-cell depleted HSCT in advanced stage sickle cell disease.

Despite significant improvements in the supportive care, sickle cell disease (SCD) leads to significant morbidity and mortality. Allogeneic hematopoietic stem cell transplantation (HSCT), the only curative option, is limited due to matched donor availability. This could be met with T-cell-depleted haploidentical HSCT. Twenty advanced-stage SCD patients, median age 15 years, and 9 patients, median age 14 years, were transplanted with CD3/CD19- or TCRαβ/CD19-depleted grafts and from matched sibling donors (MSDs). The conditioning consisted of ATG, thiotepa, fludarabine, and treosulfan. The median follow-up in the T-haplo-HSCT and the MSD patients was 21 (9-62) and 25 (7-60) months, respectively. The OS in the T-haplo-HSCT and MSD was 90% and 100%, respectively. In the T-haplo-HSCT group, two patient succumbed to a CMV pneumonitis and a macrophage activation syndrome (MAS). One patient in the T-haplo-HSCT group requires renal replacement therapy because of BK virus nephritis. None developed grade III-IV acute GvHD. In the T-haplo-HSCT and in the MSD, 20% and 22%, respectively, developed a mild or moderate chronic GvHD. These results demonstrate the feasibility, safety, and efficacy of T-haplo-HSCT also for adult advanced stage SCD patients.

Urinary Virome Perturbations in Kidney Transplantation.

The human microbiome is important for health and plays a role in essential metabolic functions and protection from certain pathogens. Conversely, dysbiosis of the microbiome is seen in the context of various diseases. Recent studies have highlighted that a complex microbial community containing hundreds of bacteria colonizes the healthy urinary tract, but little is known about the human urinary viruses in health and disease. To evaluate the human urinary virome in the context of kidney transplantation (tx), variations in the composition of the urinary virome were evaluated in urine samples from normal healthy volunteers as well as patients with kidney disease after they had undergone kidney tx. Liquid chromatography-mass spectrometry/mass spectrometry analysis was undertaken on a selected cohort of 142 kidney tx patients and normal healthy controls, from a larger biobank of 770 kidney biopsy matched urine samples. In addition to analysis of normal healthy control urine, the cohort of kidney tx patients had biopsy confirmed phenotype classification, coincident with the urine sample analyzed, of stable grafts (STA), acute rejection, BK virus nephritis, and chronic allograft nephropathy. We identified 37 unique viruses, 29 of which are being identified for the first time in human urine samples. The composition of the human urinary virome differs in health and kidney injury, and the distribution of viral proteins in the urinary tract may be further impacted by IS exposure, diet and environmental, dietary, or cutaneous exposure to various insecticides and pesticides.

The Detection of BK Virus by Quantitative PCR of Plasma as Screening Test to Identify Renal Transplant Recipients at Risk of BK Virus Nephropathy

Background: BK virus infection and nephritis are increasing problems in renal transplant recipients (RTRs). Increasing prevalence of this infection and nephritis have been correlated with newer immunosuppressive agent in recent years. Routine post-transplant screening for BK viremia prior to the occurrence of nephritis and the reduction in immunosuppressive therapy for subjects with BKV infection appear to be attractive objectives. Objectives: This study performed to screening for BK virus in blood to identify RTRs at risk of development BKV associated nephritis (BKVN) .

Tolerance Induction With Hematopoietic Stem Cell for Kidney Transplantation

Tolerance induction is considered to be a final goal in the field of organ transplantation. We have performed simultaneous bone marrow and kidney transplantation in MHC mismatched patients to induce transient mixed chimerism and donor-specific tolerance. From December 2011 to May 2014, seven MHC mismatched patients received simultaneous bone marrow and kidney transplantation. Median follow-up was 34 months after transplant. The preconditioning regimen consisted of cyclophosphamide, fludarabine, rituximab (375 mg/m2, −7 and −2 days before transplantation), antithymocyte globulin (1.5 mg/mg/day,-2 ~ 0 days) and thymic irradiation. Maintenance immunosuppressions were tacrolimus and steroids. Immunosuppression tapering was usually initiated when the patient sustained stable graft function for at least 12 months post-transplant. Immunosuppression was slowly tapered over 6 ~ 12 months. Four out of the seven patients were successfully tapered off immunosuppression. One patient is currently undergoing immunosuppression tapering. Two patients failed at immunosuppression withdrawal; Because of acute rejection and severe BK virus nephritis. 5 patients experienced BK viremia during their post-transplant period. We analyzed T cell reconstitution after bone marrow and kidney transplantation, showed a major expansion of CD8+ effector memory (CD45RA-CCR7-), end-stage effector (CD45RA + CCR7-) T cells, and late-differentiated CD27-CD28- cells. We screened BMTKT patients for the expression of an inhibitory receptor such as PD-1, 2B4, CD160, BTLA, TIGIT, Tim-3 and LAG-3. They were increased an expression of 2B4 and PD-1. And, we analyzed the functionally distinct subpopulations in regulatory T cells. Patients with experienced acute cellular rejection were more elevated CD45RA-Foxp3low non-suppressive Treg cells (Fr-III) than CD45RA-Foxp3high effector Treg cells (Fr-II). We have performed seven cases of simultaneous bone marrow and kidney transplantation in MHC mismatched patients. Four out of the seven patients achieved immunosuppression withdrawal and revealed increased CD45RA-Foxp3high effector Treg cells (Fr-II).

Mining the human urine proteome for monitoring renal transplant injury

The human urinary proteome provides an assessment of kidney injury with specific biomarkers for different kidney injury phenotypes. In an effort to fully map and decipher changes in the urine proteome and peptidome after kidney transplantation, renal allograft biopsy matched urine samples were collected from 396 kidney transplant recipients. Centralized and blinded histology data from paired graft biopsies was used to classify urine samples into diagnostic categories of acute rejection, chronic allograft nephropathy, BK virus nephritis, and stable graft. A total of 245 urine samples were analyzed by liquid chromatography-mass spectrometry using isobaric Tags for Relative and Absolute Quantitation (iTRAQ) reagents. From a group of over 900 proteins identified in transplant injury, a set of 131 peptides were assessed by selected reaction monitoring for their significance in accurately segregating organ injury causation and pathology in an independent cohort of 151 urine samples. Ultimately, a minimal set of 35 proteins were identified for their ability to segregate the 3 major transplant injury clinical groups, comprising the final panel of 11 urinary peptides for acute rejection (93% area under the curve [AUC]), 12 urinary peptides for chronic allograft nephropathy (99% AUC), and 12 urinary peptides for BK virus nephritis (83% AUC). Thus, urinary proteome discovery and targeted validation can identify urine protein panels for rapid and noninvasive differentiation of different causes of kidney transplant injury, without the requirement of an invasive biopsy.

The Identification of Novel Potential Injury Mechanisms and Candidate Biomarkers in Renal Allograft Rejection by Quantitative Proteomics

Early transplant dysfunction and failure because of immunological and nonimmunological factors still presents a significant clinical problem for transplant recipients. A critical unmet need is the noninvasive detection and prediction of immune injury such that acute injury can be reversed by proactive immunosuppression titration. In this study, we used iTRAQ -based proteomic discovery and targeted ELISA validation to discover and validate candidate urine protein biomarkers from 262 renal allograft recipients with biopsy-confirmed allograft injury. Urine samples were randomly split into a training set of 108 patients and an independent validation set of 154 patients, which comprised the clinical biopsy-confirmed phenotypes of acute rejection (AR) (n = 74), stable graft (STA) (n = 74), chronic allograft injury (CAI) (n = 58), BK virus nephritis (BKVN) (n = 38), nephrotic syndrome (NS) (n = 8), and healthy, normal control (HC) (n = 10). A total of 389 proteins were measured that displayed differential abundances across urine specimens of the injury types (p < 0.05) with a significant finding that SUMO2 (small ubiquitin-related modifier 2) was identified as a "hub" protein for graft injury irrespective of causation. Sixty-nine urine proteins had differences in abundance (p < 0.01) in AR compared with stable graft, of which 12 proteins were up-regulated in AR with a mean fold increase of 2.8. Nine urine proteins were highly specific for AR because of their significant differences (p < 0.01; fold increase >1.5) from all other transplant categories (HLA class II protein HLA-DRB1, KRT14, HIST1H4B, FGG, ACTB, FGB, FGA, KRT7, DPP4). Increased levels of three of these proteins, fibrinogen beta (FGB; p = 0.04), fibrinogen gamma (FGG; p = 0.03), and HLA DRB1 (p = 0.003) were validated by ELISA in AR using an independent sample set. The fibrinogen proteins further segregated AR from BK virus nephritis (FGB p = 0.03, FGG p = 0.02), a finding that supports the utility of monitoring these urinary proteins for the specific and sensitive noninvasive diagnosis of acute renal allograft rejection.

Análisis citológico para la evaluación urinaria en pacientes trasplantados renales con infección del virus BK

IntroductionTubulointerstitial nephritis associated with the BK virus is a complication of kidney transplantation. Markers of BK virus replication include the presence of viral inclusions called «Decoy cells» in urine cytology and high titers of ADN viral load in plasma, urine, and renal tissue. ObjectiveTo evaluate the diagnostic utility of urine cytology for the early identification of BK virus infection by using 3 different types of cytological screening. ResultsUrinary cytology as a diagnostic test showed high sensitivity (81.4%) in identifying renal transplant recipients with BK virus infection. Specificity was only 22.6%. The 3 methods of urine cytology screening appeared to have similar effectiveness in quantifying cell density. ConlusionsUrine cytology is a reliable test for the detection of Decoy cells but is not diagnostic. Irrespective of the screening method used, detection of Decoy cells may be an indirect and sensitive method for estimating the existence of BK viral replication, although urinary polymerase chain reaction and kidney biopsy have greater specificity in the diagnosis of BK virus nephritis.

A pilot study of reduced dose cyclosporine and corticosteroids to reduce new onset diabetes mellitus and acute rejection in kidney transplant recipients

BackgroundNew onset diabetes mellitus (NODM) and acute rejection (AR) are important causes of morbidity and risk factors for allograft failure after kidney transplantation.MethodsIn this multi-center, open label, single-arm pilot study, 49 adult (≥18 years of age), low immunologic risk, non-diabetic recipients of a first deceased or living donor kidney transplant received early steroid reduction to 5 mg/day combined with Thymoglobulin® (Genzyme Transplant, Cambridge, MA, USA) induction, low dose cyclosporine (2-hour post-dose (C2) target of 600 to 800 ng/ml) and mycophenolic acid (MPA) therapy.ResultsSix months after transplantation, two patients (4%) developed NODM and one patient (2%) developed AR. Four patients had impaired fasting glucose tolerance based on 75-g oral glucose tolerance testing (OGTT). There was one patient death. There were no episodes of cytomegalovirus (CMV) infection or BK virus nephritis. In contrast, in a historical cohort of n = 27 patients treated with Thymoglobulin induction, and conventional doses of cyclosporine and corticosteroids, the incidence of NODM and AR was 18% and 15%.ConclusionsThe pilot study results suggest that Thymoglobulin induction combined with early steroid reduction, reduced cyclosporine exposure and MPA, may reduce the incidence of both NODM and AR in low immunological risk patients. A future controlled study enriched for patients at high risk for NODM is under consideration.Trial registrationClinicalTrials.gov: http://NCT00706680

Management and Outcome of BK Viremia in Renal Transplant Recipients

BK viremia can lead to nephritis, which can progress to irreversible kidney transplant failure. Our prospective study provides management and outcome of BK viremia in renal transplant recipients. Two hundred forty de novo kidney-only recipients were enrolled from July 2007 to July 2010 and followed for 1 year. Standard immunosuppression with Thymoglobulin/interleukin 2 receptor blocker and mycophenolate mofetil/tacrolimus (Tac)/prednisone was employed. Quantitative BK virus (BKV) DNA surveillance in plasma/urine was performed at 1, 3, 6, 12, and 24 months after transplantation. Patients with significant viremia (defined as ≥10,000 viral copies/mL) underwent renal biopsy and treated with 30% to 50% reduction in doses of both mycophenolate mofetil and Tac without antiviral therapy. The target 12-hr Tac trough levels were lowered to 4 to 6 ng/mL in the significant viremia group, whereas the target levels remained unchanged at 5 to 8 ng/mL for all other groups. Sixty-five patients (27%) developed BK viremia; 28 (12%) of whom had significant viremia. A total of five (21%) of the 23 (of 28) patients who underwent biopsy presented with subclinical BKV nephritis. The mean plasma BKV DNA declined by 98% (range, 76%-100%) at 1 year after peak viremia. Acute cellular rejection seen in four (14%) of 28 patients, responded to bolus steroids. There was no decline in estimated glomerular filtration rate over time from 1 month after transplantation to 1 year after peak viremia (P=0.57). Reduction in immunosuppression alone resulted in the successful resolution of viremia with preservation of renal function and prevention of clinical BKV nephritis and graft loss.

Lower Prevalence of BK Virus Infection in African American Renal Transplant Recipients

Because the occurrence of BK virus (BKV) nephritis is far less frequent than BK viremia or viruria, analysis of risk factors for BKV nephritis as an endpoint could lead to erroneous findings. We undertook a prospective study to evaluate the risk factors for the occurrence of BKV infections using BK viruria and viremia as endpoints. Two hundred forty renal only transplant recipients were prospectively enrolled into our institutional review board-approved single center study to evaluate various aspects of posttransplant BKV infection. All patients were followed up for a minimum of 6 months posttransplant. Of the 240 subjects, 154 were whites, 61 African Americans, and 25 belonged to other races. A total of 94 developed BKV infection (any degree of BK viruria or viremia) whereas 146 developed no infection. Among these, 33 had BK viruria alone, 61 had BK viremia with viruria and 25 had significant viremia defined as BKV DNA more than 10,000 copies/mL of plasma. Lower proportion of African Americans developed BKV infection, 14 of 61 (23%), as opposed to whites, 67 of 154 (47%). Logistic regression model showed lower risk of any BKV infection in African American recipient race (OR, 0.38; 95% CI, 0.17-0.82; P=0.016) and higher risk of significant BKV infection with occurrence of acute rejection (OR, 3.9; 95% CI, 1.31-11.8; P=0.015). The Kaplan-Meier analysis shows a trend toward greater freedom from BKV infection in African Americans as opposed to other racial groups (P=0.33). Renal transplant recipients of African American race had a lower risk of posttransplant BKV infection compared with whites, independent of other confounding risk factors.

BK Virus (BKV): Infection, Propagation, Quantitation, Purification, Labeling, and Analysis of Cell Entry

BK virus (BKV) can cause BKV nephritis in renal transplant patients and has become a significant reason for graft loss in this decade. BKV is latent in the urogenital tract and most likely is transported with the donor kidney to recipients. BKV replication occurs in the nucleus of human renal proximal tubular cells (HRPTEC) and daughter viruses are delivered to other cells to spread infection. A few in vitro studies have been reported about the mechanism and kinetics of BKV infection. However, there are still a lot of unknown factors regarding BKV infection. This unit describes the handling of BKV, BKV propagation, determination of titer and ability to infect cells, as well as purification and labeling of BKV in order to analyze BKV cell entry.

Repression of BK Virus Infection of Human Renal Proximal Tubular Epithelial Cells by Pravastatin

BK virus (BKV), a human polyomavirus, causes BKV nephritis, which often leads to graft loss after renal transplantation. Currently, the only efficient therapy against BKV nephritis seems to be a reduction or change of immunosuppressive agents, but this may increase the inherent risk of rejection. Here, we report the ability of 3-hydroxy-3-methyl-glutaryl coenzyme A reductase inhibitor (statin), which is routinely used to treat hypercholesterolemia, to repress BKV entry pathways in human renal proximal tubular epithelial cells (HRPTEC) and, correspondently, prevent BKV infection. HRPTEC were co-incubated with BKV and pravastatin. Then the percentage of HRPTEC infected with BKV by immunofluorescent analysis and large T-antigen expression which suggested BKV infection by Western blots was assessed in the absence and presence of pravastatin. The distribution of purified and labeled BKV particles in the presence and absence of pravastatin was also investigated. Both the percentage of BKV infected cells and the large T-antigen expression were significantly decreased in HRPTEC pretreated and co-incubated with pravastatin. However, when pravastatin was added 72 hr after BKV infection it failed to decrease percentage of BKV infected cells. It is likely, that pravastatin's inhibitory effect is explained by depletion of caveolin-1, a critical element of caveolae. BKV enters HRPTEC by caveolar-mediated endocytosis. We provide evidence that pravastatin dramatically decreased caveolin-1 expression in HRPTEC and interfered with internalization of labeled BKV particles. Our data suggest that pravastatin, acting through depletion of caveolin-1, prevented caveolar-dependent BKV internalization and repressed BKV infection of HRPTEC.

Successful Treatment of BK Viremia Using Reduction in Immunosuppression Without Antiviral Therapy

Treatment of BK virus (BKV) infection in renal transplant recipients remains controversial. This retrospective analysis evaluated efficacy and safety of reducing immunosuppression without antiviral therapy. This single center analysis included 24 patients diagnosed with BK viremia between September 2001 and December 2003. Sixteen patients (66%) presented with BKV nephritis and eight patients (34%) presented with viremia without evidence of nephritis on renal biopsy. At time of diagnosis, mean plasma BKV DNA (copies/mL) was 460,409 (range 10,205-1,920,691). Mean doses reduction of mycophenolate mofetil and tacrolimus were 44% and 41%, respectively, from time of diagnosis of BKV infection to complete resolution of viremia. A decline in BK viral load was noticed within 15 to 30 days, with successful elimination of viremia over a mean period of 5.8 months (range, 1-9.5). Mean serum creatinine at time of diagnosis of BK viremia was 1.8 mg/dL (range, 1.2-2.8). Mean follow-up period is 30.9 months postdiagnosis. At the most recent visit, serum creatinine was 2.0 mg/dL (range, 1.0-3.6) (P=0.14). With reduction in immunosuppressive therapy, three patients (13%) developed acute cellular rejection and were treated successfully with intravenous bolus steroids. During follow-up, one patient had a relapse of BKV nephritis during pregnancy and lost her graft. After mean follow-up period of 43.5 months posttransplantation, all 24 patients are alive and 23 have a functioning graft. Seventeen patients (71%) have stable or improved graft function. Our analysis shows that reduction in immunosuppression therapy alone results in clearance of the BK viremia with good long-term outcome.

BK Virus Nephritis after Renal Transplantation

BK virus nephritis is an increasing problem and is posing a threat to improving renal transplant graft survival. The pathogenesis of this condition remains to be investigated. Higher prevalence of BK virus infection in recent years has been correlated with declining acute rejection rates and the use of potent immunosuppressive agents. Patients with this infection usually have asymptomatic viremia and/or nephritis with or without worsening of renal function. The diagnosis of this disease is based on detecting the virus or its effects in urine, blood, and renal tissue. In the past, approximately 30 to 60% of patients with BK virus nephritis developed graft failure. In recent years, the combination of early detection, prompt diagnosis, and therapies including preventive measures have resulted in better outcomes.

Recovery of BK Virus Large T-Antigen–Specific Cellular Immune Response Correlates With Resolution of BK Virus Nephritis

BK virus (BKV) infection of kidney transplant patients is an increasing problem and is thought to be secondary to potent immunosuppressive therapy. BKV infection progresses to BKV nephritis (BKVN) in approximately 8% of transplants and in half of these cases the graft is lost. We used an interferon-gamma enzyme-linked immunosorbent spot (ELISPOT) assay to measure the cellular immune response to peptides encoding BKV large T antigen. Eight kidney transplant patients with BKVN were tested at the time of diagnosis of BKVN and then after resolution of active BKV infection. When total spot counts from all peptide pools were combined, the mean ELISPOT signal per 10,000 cells at the time of BKVN diagnosis was 23.1 (range 3.4-59.7), with a median of 21.8. This increased to 70.2 (range 5.4-189.4) with a median of 37.0 (P=0.1216) after resolution of active BKV infection. To further increase specificity of response, we counted the number of peptide pools with ELISPOT activity of greater than 10 spots per well after subtraction of background. The mean number of pools fitting this criteria at the time of BKVN diagnosis was 2.1 (range 0-8) with a median of 1.5; this increased to 8 (range 1-18) and a median of 6.5 after recovery (P=0.0338). This demonstrates that recovery of cellular immune response to large T antigen corresponds with resolution of active BKV infection. This may prove useful in monitoring patients' cellular immunity and recovery from active BKV infection when treated with reduction in immunosuppressive therapy.