Surgery for Renal Hyperparathyroidism in the Era of Cinacalcet: A Single-Center Experience.

BACKGROUND AND AIMS: There are only few data on the influence of cinacalcet on the outcome of parathyroidectomy in patients with renal hyperparathyroidism. Indication and timing of surgery have changed since its introduction, especially with regard to kidney transplantation. Therefore, we retrospectively analyzed patients undergoing parathyroidectomy for renal hyperparathyroidism in our institution. MATERIAL AND METHODS: Between 2008 and 2015, 196 consecutive operations in 191 patients were analyzed. About 80 operations (41%) were performed in patients receiving cinacalcet compared with 116 operations (59%) in patients without cinacalcet. Clinical data, preoperative medication, pre- and postoperative laboratory values, type and details of surgery including complications, as well as cardiovascular complications and kidney transplantation with graft function were recorded. RESULTS: Demographical data were similar in patients with or without cinacalcet treatment. A total of 54% of patients received a kidney graft before or after parathyroidectomy. Pre- and postoperative parathormone levels were similar in both groups (preoperatively 755 vs 742 ng/L, postoperatively 50 vs 46 ng/L, p > 0.10), whereas patients with cinacalcet showed significantly lower calcium levels preoperatively (2.28 vs 2.41 mmol/L, p = 0.0002). There was no difference in recurrence or persistence of hyperparathyroidism, duration of surgery, hospital stay, or complication rate. Creatinine levels in patients with tertiary hyperparathyroidism were similar after 1-year follow-up. CONCLUSION: Cinacalcet did not influence outcome of patients with parathyroidectomy for renal hyperparathyroidism and can be safely offered to patients not responding to medical treatment.

The effect of induction therapy on established CMV specific T cell immunity in living donor kidney transplantation.

Cytomegalovirus (CMV) infection influences both short and long term outcomes in immunosuppressed organ transplant recipients. The aim of this study was to evaluate the effect of different induction immunosuppression regimens on CMV specific T cell response in patients with already established CMV immunity. In 24 seropositive living donor kidney recipients, the frequency of CMV specific T cells was determined by ELISPOT (Enzyme-Linked ImmunoSpot) assay prior and 6 months after transplantation. Recipients' peripheral blood mononuclear cells were stimulated with immediate-early (IE1) and phosphoprotein 65 (pp65) CMV-derived peptide pools and the number of cells producing interferon gamma (IFN-gamma) was assessed. Patients received quadruple immunosuppression based either on depletive rabbit antithymocyte globulin (rATG) or non-depletive basiliximab induction and tacrolimus/mycophenolate mofetil/steroids. Patients with rATG induction received valgancyclovir prophylaxis. No effects of different induction agents on CMV specific T cell immunity were found at sixth month after kidney transplantation. There were no associations among dialysis vintage, pretransplant CMV specific T cell immunity, and later CMV DNAemia. Similarly, no effect of CMV prophylaxis on CMV specific T cell immunity was revealed. This study shows no effect of posttransplant immunosuppression on CMV specific T cell immunity in living donor kidney transplant recipients with CMV immunity already established, regardless of lymphocyte depletion and CMV prophylaxis.

Pink-beam serial crystallography.

Serial X-ray crystallography allows macromolecular structure determination at both X-ray free electron lasers (XFELs) and, more recently, synchrotron sources. The time resolution for serial synchrotron crystallography experiments has been limited to millisecond timescales with monochromatic beams. The polychromatic, "pink", beam provides a more than two orders of magnitude increased photon flux and hence allows accessing much shorter timescales in diffraction experiments at synchrotron sources. Here we report the structure determination of two different protein samples by merging pink-beam diffraction patterns from many crystals, each collected with a single 100 ps X-ray pulse exposure per crystal using a setup optimized for very low scattering background. In contrast to experiments with monochromatic radiation, data from only 50 crystals were required to obtain complete datasets. The high quality of the diffraction data highlights the potential of this method for studying irreversible reactions at sub-microsecond timescales using high-brightness X-ray facilities.

Estimated Nephron Number of the Donor Kidney: Impact on Allograft Kidney Outcomes.

BACKGROUND: Low birth weights have been associated with a reduction in nephron number with compensatory hypertrophy of existing glomeruli. The impact of donor birth weight as an estimate of nephron number on allograft function, however, has not been examined. METHODS: We collected donor birth weight, kidney weight, and volume from 91 living kidney donor-recipient pairs before nephrectomy and after 12, 36, and 60 months. Nephron number was calculated from donor birth weight and age. RESULTS: Donor birth weight, kidney weight/body surface area (BSA), and kidney volume showed a moderate positive correlation with allograft estimated glomerular filtration rate (eGFR) at 12 months (P < .05). Donor age showed a negative moderate correlation with allograft eGFR at 12 months (P = .015). The strongest correlation with allograft eGFR was observed for calculated donor kidney nephron number at 12, 36, and 60 months (R, 0.340, 0.305, and 0.476, respectively; P < .05). No impact was observed on allograft daily proteinuria of any investigated marker (P > .05). Recipients of donors with birth weight <2.5 kg had need of a significantly greater number of antihypertensive drugs (P < .05). CONCLUSIONS: Calculated nephron number from donor birth weight and age is suggested to be superior to donor kidney weight/BSA and volume regarding allograft function. Calculated nephron number could estimate expected eGFR and guide decision making in cases of impaired allograft function.

Donor-Recipient Matching Based on Predicted Indirectly Recognizable HLA Epitopes Independently Predicts the Incidence of De Novo Donor-Specific HLA Antibodies Following Renal Transplantation.

De novo donor-specific HLA antibodies (dnDSA) are recognized as a risk factor for premature allograft failure. Determinants of DSA specificity are generated via the indirect allorecognition pathway. Here, we present supportive data for the relevance of predicted indirectly recognizable HLA epitopes (PIRCHE) to predict dnDSA following kidney transplantation. A total of 2787 consecutive kidney transplants performed between 1995 and 2015 without preformed DSA have been analyzed. De novo DSA were detected by single antigen bead assay. HLA epitope mismatches were determined by the HLAMatchmaker and PIRCHE approach and correlated in uni- and multivariate analyses with 10-year allograft survival and incidence of dnDSA. The PIRCHE-II score moderately predicted allograft survival. However, the predictive value of elevated PIRCHE-II scores >9 for the incidence of dnDSA was statistically significant (p < 0.001). In a multivariate Cox regression analysis adjusted for antigen mismatch and HLAMatchmaker epitopes, the PIRCHE-II score could be identified as an independent risk factor for dnDSA. The PIRCHE-II score independently from the antigen mismatch and HLAMatchmaker epitopes could be revealed as being a strong predictor for dnDSA. PIRCHE may help to identify acceptable mismatches with decreased risk of dnDSA and thus improve long-term renal allograft survival.

Clinical Development of Cell Therapies: Setting the Stage for Academic Success.

Cellular therapies have potential to treat a wide range of diseases with autologous immunotherapies showing unprecedented therapeutic promise in clinical trials. Such therapies are mainly developed by academic researchers applying small-scale production, targeting rare and unmet medical needs. Here, we highlight the clinical translation of immunotherapy product in an academic setting, which may serve as a success model for early academic development of cell-based therapeutics.

Strain Lattice Imprinting in Graphene by C60 Intercalation at the Graphene/Cu Interface.

Intercalation of C60 molecules at the graphene-substrate interface by annealing leads to amorphous and crystalline structures. A comparison of topography and electronic structure with wrinkles and moiré patterns confirms intercalation. The intercalated molecules imprint a local strain/deformation on the graphene layer whose magnitude is controlled by the intermolecular distance. The crystalline intercalated structure exhibits a superlattice peak in the local density of states. This work provides control of local strain in graphene.

Comparing Humoral and Cellular Immune Response Against HBV Vaccine in Kidney Transplant Patients.

Host protection upon vaccination usually results from the complex interplay of humoral and cellular components of the immune system. Exploring hepatitis B surface antigen (HBsAg)-specific T cell responses and their correlation with humoral responses under immunosuppression, we analyzed 51 renal transplant recipients, differing in HBV vaccine-specific antibody titers (non [NRs]-, low [LRs]-, and high responders [HRs]) and in 22 healthy controls (HCs) in a cross-sectional study. HBsAg-specific T cells were analyzed by flow cytometry according to expression of activation markers CD40L and/or CD69, and the cytokines IFNγ, IL-2, TNFα, and IL-17. No significant differences in responder rate and magnitude of HBsAg-specific T cell responses were found between HCs and HRs. Interestingly, HBsAg-specific Th-cells were also observed in 50% of humoral NRs. Frequencies of HBsAg-specific CD40L+ Th-cells were significantly higher in HRs compared to LRs (p = 0.009) and in LRs in comparison to NRs (p = 0.043). All but NRs showed a predominance of multi-potent HBsAg-specific TNFα+IL-2+ Th-cells. As expected, HBsAg-specific CD8(+) T cells were rarely found. In conclusion, mounting of hepatitis B vaccine-specific T cell responses is possible in kidney transplant recipients despite immunosuppression. Detection of HBV-specific Th-cells in a significant proportion of humoral NRs contributes to the current discussion on conferring immune protection by cellular memory in such patients.

Mesenchymal Stromal Cells Prevent Allostimulation In Vivo and Control Checkpoints of Th1 Priming: Migration of Human DC to Lymph Nodes and NK Cell Activation.

Although the immunomodulatory potency of mesenchymal stromal cells (MSC) is well established, the mechanisms behind are still not clear. The crosstalk between myeloid dendritic cells (mDC) and natural killer (NK) cells and especially NK cell-derived interferon-gamma (IFN-γ) play a pivotal role in the development of type 1 helper (Th1) cell immune responses. While many studies explored the isolated impact of MSC on either in vitro generated DC, NK, or T cells, there are only few data available on the complex interplay between these cells. Here, we investigated the impact of MSC on the functionality of human mDC and the consequences for NK cell and Th1 priming in vitro and in vivo. In critical limb ischemia patients, who have been treated with allogeneic placenta-derived mesenchymal-like stromal cells (PLX-PAD), no in vivo priming of Th1 responses toward the major histocompatibility complex (MHC) mismatches could be detected. Further in vitro studies revealed that mDC reprogramming could play a central role for these effects. Following crosstalk with MSC, activated mDC acquired a tolerogenic phenotype characterized by reduced migration toward CCR7 ligand and impaired ability to stimulate NK cell-derived IFN-γ production. These effects, which were strongly related to an altered interleukin (IL)-12/IL-10 production by mDC, were accompanied by an effective prevention of Th1 priming in vivo. Our findings provide novel evidence for the regulation of Th1 priming by MSC via modulation of mDC and NK cell crosstalk and show that off-the-shelf produced MHC-mismatched PLX-PAD can be used in patients without any sign of immunogenicity.

Human CD45RA(-) FoxP3(hi) Memory-Type Regulatory T Cells Show Distinct TCR Repertoires With Conventional T Cells and Play an Important Role in Controlling Early Immune Activation.

Adoptive immunotherapy with regulatory T cells (Treg) is a new option to promote immune tolerance following solid organ transplantation (SOT). However, Treg from elderly patients awaiting transplantation are dominated by the CD45RA(-) CD62L(+) central memory type Treg subset (TregCM), and the yield of well-characterized and stable naïve Treg (TregN) is low. It is, therefore, important to determine whether these TregCM are derived from the thymus and express high stability, suppressive capacity and a broad antigen repertoire like TregN. In this study, we showed that TregCM use a different T cell receptor (TCR) repertoire from conventional T cells (Tconv), using next-generation sequencing of all 24 Vß families, with an average depth of 534 677 sequences. This showed almost no contamination with induced Treg. Furthermore, TregCM showed enhanced suppressive activity on Tconv at early checkpoints of immune activation controlling activation markers expression and cytokine secretion, but comparable inhibition of proliferation. Following in vitro expansion under mTOR inhibition, TregCM expanded equally as well as TregN without losing their function. Despite relatively limited TCR repertoire, TregCM also showed specific alloresponse, although slightly reduced compared to TregN. These results support the therapeutic usefulness of manufacturing Treg products from CD45RA(-) CD62L(+) Treg-enriched starting material to be applied for adoptive Treg therapy.

The Loss of BKV-specific Immunity From Pretransplantation to Posttransplantation Identifies Kidney Transplant Recipients at Increased Risk of BKV Replication.

Quantification of BKV-load and BKV-specific immunity have been evaluated to monitor BKV-replication and outcomes in kidney transplant recipients (KTRs) with BKV-infection. However, it remains crucial to better understand how immune markers can predict the risk for later infection. We studied all KTRs between 2008 and 2011. Twenty-four KTRs were diagnosed with BKV-replication and a control group of 127 KTRs was used for comparison. Samples were collected before at +1, +2, and +3 months posttransplantation. BKV-specific and alloreactive T cells were measured using an interferon-γ Elispot assay. The extent of immunosuppression was quantified by lymphocyte subpopulations and interferon-gamma levels. KTRs with a loss of BKV-specific T cells directed to Large T-antigen from pretransplantation to posttransplantation were at increased risk of BKV-replication (p < 0.001). In contrast, KTRs with stable/rising BKV-specific T cells were more likely not to develop BKV-replication (p < 0.05). KTRs developing BKV-replication showed significantly lower CD3+, CD4+, CD8+ T cells and interferon-γ levels posttransplantation, but significantly higher alloreactive T cells (p < 0.05). Monitoring pretransplant and posttransplant BKV-specific T cells is suggested a sensitive marker to identify KTRs at increased risk of BKV-replication. Increased susceptibility to immunosuppression predisposes KTRs to a loss of protective BKV-specific immunity that results in impaired virus control and BKV-replication.

International prognostic index, type of transplant and response to rituximab are key parameters to tailor treatment in adults with CD20-positive B cell PTLD: clues from the PTLD-1 trial.

Tailoring treatment by patient strata based on the risk of disease progression and treatment toxicity might improve outcomes of patients with posttransplant lymphoproliferative disorder (PTLD). We analysed the cohort of 70 patients treated in the international, multicenter phase II PTLD-1 trial (NCT01458548) to identify such factors. Of the previously published scoring systems in PTLD, the international prognostic index (IPI), the PTLD prognostic index and the Ghobrial score were predictive for overall survival. None of the scoring systems had a considerable effect on the risk for disease progression. Age and ECOG performance status were the baseline variables with the highest prognostic impact in the different scoring systems. Baseline variables not included in the scoring systems that had an impact on overall survival and disease progression were the type of transplant and the response to rituximab at interim staging. Thoracic organ transplant recipients who did not respond to rituximab monotherapy were at particularly high risk for death from disease progression with subsequent CHOP-based chemotherapy. Patients in complete remission after four courses of rituximab and patients in partial remission with low-risk IPI had a low risk of disease progression. We speculate that chemotherapy might not be necessary in this patient cohort.

Atomic scale surface structure and morphology of InAs nanowire crystal superlattices: the effect of epitaxial overgrowth.

While shell growth engineering to the atomic scale is important for tailoring semiconductor nanowires with superior properties, a precise knowledge of the surface structure and morphology at different stages of this type of overgrowth has been lacking. We present a systematic scanning tunneling microscopy (STM) study of homoepitaxial shell growth of twinned superlattices in zinc blende InAs nanowires that transforms {111}A/B-type facets to the nonpolar {110}-type. STM imaging along the nanowires provides information on different stages of the shell growth revealing distinct differences in growth dynamics of the crystal facets and surface structures not found in the bulk. While growth of a new surface layer is initiated simultaneously (at the twin plane interface) on the {111}A and {111}B nanofacets, the step flow growth proceeds much faster on {111}A compared to {111}B leading to significant differences in roughness. Further, we observe that the atomic scale structures on the {111}B facet is different from its bulk counterpart and that shell growth on this facet occurs via steps perpendicular to the ⟨112⟩B-type directions.

Five-year outcomes in kidney transplant patients converted from cyclosporine to everolimus: the randomized ZEUS study.

ZEUS study was an open-label, 12-month, multicenter study in which 300 de novo kidney transplant recipients were randomized to continue receiving cyclosporine (CsA) or convert to everolimus at 4.5 months posttransplant. Five-year follow-up data were available for 245/269 patients (91.1%) who completed the core 12-month study (123 everolimus, 109 CsA). At 5 years, adjusted estimated GFR was 66.2 mL/min/1.73 m(2) with everolimus versus 60.9 mL/min/1.73 m(2) with CsA; the mean difference was 5.3 mL/min/1.73 m(2) in favor of everolimus (95% CI 2.4, 8.3; p < 0.001 [intent-to-treat population]). In a post hoc analysis of patients remaining on study drug at 5 years (everolimus 77, CsA 86), mean difference was 8.2 mL/min/1.73 m(2) (95% CI 4.3, 12.1; p < 0.001) in favor of everolimus. The cumulative incidence of biopsy-proven acute rejection postrandomization was 13.6% with everolimus versus 7.5% with CsA (p = 0.095), largely accounted for by grade I rejection (16/21 patients and 7/11 patients, respectively). Postrandomization, graft loss, mortality, serious adverse events and neoplasms were similar in both arms. In conclusion, conversion of kidney transplant patients to everolimus at 4.5 months posttransplant is associated with a significant improvement in renal function that is maintained to at least 5 years. The increase in early mild acute rejection did not affect long-term graft function.

Clindamycin-primaquine for pneumocystis jiroveci pneumonia in renal transplant patients.

BACKGROUND: Trimethoprim/sulfamethoxazole (TMP/SMX) is considered first-line therapy for pneumocystis jiroveci pneumonia (PCP) in renal transplant patients. Alternatives have not been formally studied. Clindamycin-primaquine (C-P) is effective in HIV-associated PCP, but data in renal transplant patients are lacking. PATIENTS AND METHODS: Retrospective cohort study of 57 consecutive renal transplant patients who developed PCP and were treated with C-P (n = 23) or TMP/SMX (n = 34). RESULTS: A non-significantly higher failure rate was observed in patients on C-P due to lack of efficacy (30.4 versus 20.6%, p = 0.545). The difference was more pronounced in severe PCP (60 versus 37.5%, p = 0.611) and a significantly lower efficacy of C-P was seen when used as salvage therapy. The two patients who had received C-P after not responding to TMP/SMX failed this regimen, but all seven patients who had failed initial treatment with C-P and had been switched to TMP/SMX were cured (p = 0.028). No treatment-limiting adverse reactions were reported for patients on C-P while six patients (17.6%) on TMP/SMX developed possibly related treatment-limiting toxicity (p = 0.071). However, in only two patients adverse events were definitely related to TMP/SMX (5.9%). CONCLUSIONS: Clindamycin-primaquine appears to be safe and well tolerated for treating PCP in renal transplant patients but is probably less effective than TMP/SMX, the standard regimen. However, our data indicates that C-P represents an acceptable alternative for patients with contraindications or treatment emergent toxicities during TMP/SMX use. Notably, TMP/SMX was also acceptably tolerated in most patients. TMP/SMX remains an effective salvage regimen in case of C-P failure.

The role of CD4(+) T cells in BKV-specific T cell immunity.

Reactivation of polyomavirus BK (BKV) infection represents a severe complication in kidney transplant (KTX) patients. We previously reported an association between a declining BK viral load and the reconstitution of CD4(+) T cell BKV-specific immunity in patients following kidney transplantation. However, the specific contribution of CD4(+) T cells in the regulation of BKV-replication is unknown. Nevertheless, in vitro enrichment of BKV-specific T cells and subsequent adoptive T cell transfer may improve the restoration of immune competence in KTX patients with BKV infection. To date, strategies to capture human BKV-specific T cells with the ensuing expansion to clinically useful numbers are lacking. Here, we demonstrated a comprehensive flow cytometric analysis of the BKV-specific T cell response that permits access to the majority of T cells specific for immunodominant BKV antigens. A full-spectrum evaluation of the BKV-specific T cell response was performed by stimulating peripheral blood mononuclear cells (PBMC) with a mixture of BKV immunodominant peptide pools at varying concentrations and measuring activation marker expression and cytokine secretion. We also examined the effects of co-stimulation and PBMC resting time prior to activation. We defined the narrow range of stimulation conditions that permit the capture and expansion of functional BKV-specific T cell lines. The generated BKV-specific T cell lines showed the highest specificity and functionality when the T cells were captured according to IFNγ-secretion. This study highlights the multifunctional and cytolytic BKV-specific CD4(+) T cells as a dominant population within the generated T cell product. This method offers a novel approach for the generation of BKV-specific T cell lines for adoptive immunotherapy and underscores the critical role of CD4(+) T cells in the clearance of BKV.

Current characteristics and outcome of cytomegalovirus infections after kidney transplantation.

INTRODUCTION: The clinical course of cytomegalovirus (CMV) infections in the current era is poorly described. We characterized the symptoms and outcome of all CMV infections in a large cohort of kidney transplant recipients. Among 1129 kidney transplant recipients transplanted between 2004 and 2011 in Charité Universitätsmedizin Berlin and Helsinki University Hospital, 297 patients with CMV infection were characterized. RESULTS: CMV disease occurred in 217/1129 patients (19.2%), and CMV infection in 297/1129 (26.3%). Gastrointestinal symptoms were recorded in 58% and fever in 47% patients with primary CMV disease, compared to 46% and 27% patients with symptomatic CMV reactivation, whereas leukopenia or thrombocytopenia were seen in only 17-28% patients, and malaise in 9-10%. Tissue-invasive CMV gastroenteritis was confirmed in 11% and CMV pneumonia in only 1% of patients with CMV disease. Only 1 patient died because of CMV infection (mortality 0.3%). Virus-related factors or the use of secondary prophylaxis did not predict the risk of recurrence, which occurred in 33% patients. CONCLUSION: In conclusion, CMV disease remains a common problem after kidney transplantation. Gastrointestinal symptoms were common, especially in patients with primary CMV infection, whereas bone marrow suppression, hepatopathy, or malaise were seen less frequently.

Percutaneous computer tomography-guided ethanol sympathicolysis for the treatment of resistant arterial hypertension.

PURPOSE: As an alternative to catheter-based radiofrequency (RF) ablation, renal sympathicolysis can also be achieved by image-guided percutaneous injection of ethanol around the renal artery. MATERIALS AND METHODS: We report the case of a 50-year-old man with refractory hypertension and end-stage renal failure of unclear etiology who was treated with computed tomography-guided percutaneous periarterial ethanol sympathicolysis. RESULTS: The procedure was painless. The patient's BP decreased within 6 days from a baseline value of 172/84 mm Hg (1 week before treatment) to a sustained decreased value of 143/70 mm Hg 1 month after intervention, i.e., a decrease by 29/14 mm Hg. The patient's hypertension-related headache resolved. CONCLUSION: Image-guided periarterial ethanol injection for renal sympathetic denervation in a patient with drug-resistant hypertension is feasible. We provide a detailed description of this new interventional procedure and discuss its potential advantages compared with catheter-based RF ablation.

Novel GMP-compatible protocol employing an allogeneic B cell bank for clonal expansion of allospecific natural regulatory T cells.

The adoptive transfer of natural regulatory T cells (nTreg) is a new option to reshape undesired immune reactivity in autoimmunity and transplantation toward "tolerance." The first clinical trials using adoptive transfer of polyclonal nTreg demonstrated safety and hints of efficacy. However, the low frequencies of antigen-specific cells among the pool of polyclonal nTreg and their broad antigen nonspecific suppression are limitations of this approach regarding efficacy and safety. Recently, the isolation and expansion of (allo)antigen-specific nTreg have successfully been achieved by using Treg-specific activation markers but the yield is relatively low. Here, we describe a novel good manufacturing practice (GMP)-compatible expansion protocol of alloantigen-specific nTreg based on the stimulation of nTreg by allogeneic activated B cells. Their functionality and specificity are superior compared to polyclonal nTreg both in vitro and in vivo. Employing an allogeneic B cell bank, designed to cover the majority of HLA types, allows fast GMP-compliant manufacturing for donor-specific nTreg for clinical application in organ and stem cell transplantation. TCR repertoire analyses by next generation sequencing revealed impressive expansion by several log-steps of even very low-abundance alloantigen-specific nTreg clones. This novel method offers a simple approach for expanding antigen-specific nTreg and is characterized by high replicability and easy transferability to full GMP standards.

TCR repertoire analysis by next generation sequencing allows complex differential diagnosis of T cell-related pathology.

Clonotype analysis is essential for complete characterization of antigen-specific T cells. Moreover, knowledge on clonal identity allows tracking of antigen-specific T cells in whole blood and tissue infiltrates and can provide information on antigenic specificity. Here, we developed a next generation sequencing (NGS)-based platform for the highly quantitative clonotype characterization of T cells and determined requirements for the unbiased characterization of the input material (DNA, RNA, ex vivo derived or cell culture expanded T cells). Thereafter we performed T cell receptor (TCR) repertoire analysis of various specimens in clinical settings including cytomegalovirus (CMV), polyomavirus BK (BKV) reactivation and acute cellular allograft rejection. Our results revealed dynamic nature of virus-specific T cell clonotypes; CMV reactivation was linked to appearance of new highly abundant antigen-specific clonalities. Moreover, analysis of clonotype overlap between BKV-, alloantigen-specific T cell-, kidney allograft- and urine-derived lymphocytes provided hints for the differential diagnosis of allograft dysfunction and enabled appropriate therapy adjustment. We believe that the established approach will provide insights into the regulation of virus-specific/anti-tumor immunity and has high diagnostic potential in the clinical routine.