Viral specific T cell therapy in kidney transplant recipients – A single‐center experience

Viral infections such as adenovirus (ADV), BK virus (BKV), and cytomegalovirus (CMV) after kidney transplantation negatively impact outcomes in transplant recipients despite advancements in screening and antiviral therapy. We describe our experience of using the virus‐specific T cell therapy (VSTs) in kidney transplant recipients (KTR) at our transplant center.


INTRODUCTION
Adenovirus (ADV), BK virus (BKV), and cytomegalovirus (CMV) infections after kidney transplantation are a common cause of increased morbidity leading to hospitalizations, graft loss, and patient mortality. 1sseminated ADV infection may present with hemorrhagic cystitis, nephritis, hepatitis, gastroenteritis, or pneumonia leading to significant morbidity and mortality. 2,3Valganciclovir (VGCV) is used for the prophylaxis and treatment of CMV infection after a kidney transplant.
However, it can cause significant myelosuppression or may not be an option in the presence of resistant CMV strain. 4Use of antivirals such as cidofovir or foscarnet has been limited by nephrotoxicity. 4BKVassociated nephropathy (BKVAN) can be present in up to 10% of KTR with as many as 50% progressing to allograft failure. 5Currently, there is no approved antiviral therapy, and immunosuppression (IS) reduction forms the mainstay of management.[8] There is emerging evidence in hematopoietic stem cell transplant (HSCT) recipients that virus-specific T cell therapy (VSTs) is a safe and effective treatment option to overcome these barriers. 9Unfortunately, the use of VSTs in solid organ transplant recipients has not been well studied.Herein we describe our initial experience of VSTs in ADV, BKV, and CMV infections at our transplant center.Alpha for all tests was a two-tailed p = .05,unadjusted for multiple tests, and significance tests were conducted using SAS 9.4 software.

Patient characteristics
The overall median age of the cohort was 60 years, 65% were men and 52% were of Caucasian race (Table S1).Native kidney disease was due to diabetes and hypertension in 52% of these patients.There were 17 deceased donor kidney transplants, five live donor kidney transplants, and one simultaneous liver-kidney (SLK) transplant.Anti-thymocyte globulin (ATG) was used for induction in 21 (92%) patients.Seventeen (74%) patients had more than five HLA ABDR mismatches between the kidney donor and the recipient.Maintenance IS was composed of tacrolimus and mycophenolate mofetil (MMF) in 14 (61 %), belatacept and MMF in two (9%) recipients, and tacrolimus, MMF, and prednisone in seven (30%) of transplant recipients.The median time to infection after transplant surgery was 3 months.

Clinical and laboratory characteristics pre-VSTs
In this cohort of 23 KTR; two patients had acute rejection 1-3 months prior to the development of infections (Tables S2 and S3).One patient had Banff acute cellular rejection IIB treated with steroids and ATG.
The second patient received four sessions of plasma exchange and bortezomib for active AMR.Out of 23 patients, 17 had BKV, four CMV, and two ADV infections.All patients had a reduction in maintenance IS after diagnosis of infection.MMF was stopped in 70% KTR and substituted with prednisone as per the discretion of the treating nephrologist.The remaining patients on MMF had a 50%-75% dose reduction.Fever was not a common presenting symptom and was present in only 13% of the patients.Nausea, vomiting, or diarrhea were seen in patients with CMV infections (13%).Two patients with BK viremia had evidence of moderate to severe hydronephrosis requiring percutaneous nephrostomy.Disseminated ADV infection presented with fever (>101 • F), nosebleed, and gross hematuria.AKI was present in 91% of the patients.Renal allograft biopsy was performed in 13 out of 23 patients (57%).The median time to BK viremia post-KT was  14 weeks.CMV resistance panel testing was performed in patients who didn't respond to antiviral therapy.Antiviral resistance to either UL97 or UL54 mutation was noted in 100% of the CMV infections.Prior to VSTs, one patient with BKV and two patients with ADV received intravenous immunoglobulin (IVIG, total dose 2 g/kg in BKV and 0.5 g/kg in ADV).No patient in our cohort received Cidofovir due to risk of nephrotoxicity.

Response to VSTs and kidney allograft outcome
The median time to initiation of VSTs after reduction in IS was 2 months (range 0.5-5) (Figures 1 and 2, and Table S4).The median number of VSTs infusion was 2 (range 1-4).The median follow-up period after receiving VSTs was 8 months (range 3-14 months).VSTs were initiated sooner in ADV infection due to a lack of improvement with IS reduction and IVIG.KTR infected with ADV received one infusion of VSTs leading to resolution of viremia and renal allograft recovery by 8 weeks post-therapy.In all four cases of resistant CMV infections, VSTs were associated with more than 50% reduction in VL as described in Figure 1.It was noted that BK VL decreased from the last pre-VSTs level to 12 weeks post-VSTs assessment in all 17 BK cases (p < .001).VSTs use was associated with improvement in serum creatinine and proteinuria (p < .001)(Figure 2).One patient was diagnosed with immune complex membranoproliferative glomerulonephritis (MPGN) post VSTs, and another KTR developed TCMR (grade >1B) leading to chronic allograft nephropathy.We did not notice any immediate infusion reaction after VST administration.Fever persisted for 2-3 days in patients with disseminated ADV infection.Of note, we did not see any case of GVHD, or cytokine release syndrome which are reported as rare potential complications of VSTs. 11,12

DISCUSSION
We report our findings using third-party VSTs in ADV, BKV, and CMV infections.To our knowledge, this is the largest case series in the literature reporting the use of VSTs in KTR.
Adoptive T cell-based therapy is a potential therapy to treat viral infection post-transplant.It has been used in HSCT patients for various opportunistic infections such as ADV, BKV, CMV, Epstein-Barr virus, and human herpesvirus 6. 1,[9][10]13 ADV-specific T cells and antibodies are the highest in children and young adults. ADVpecific CD4 and CD8 T cells have been correlated with resolution of viremia.14,15 In early BKV circulating CD4 T helper, T cytotoxic cells and specific polyfunctional CD8 positive T cells are detectable.
BKV-specific T cells have been shown to be associated with viral clearance. 14,7Following primary infection, CMV-specific CD4 and CD8 T cells, and CMV serology are associated with individual immunocompetence to CMV.Patients with CMV-specific cellular immunity are less likely to develop viremia compared to those without detectable immunity. 14,16st patients in our cohort responded to VST therapy as shown in Figure 1.In addition, there was a significant improvement in AKI except in two patients described below.This seems encouraging given the almost 60% risk of deteriorating renal allograft function secondary to acute rejection from reduced IS or progression in interstitial fibrosis and tubular atrophy. 7,8e patient developed an immune complex MPGN 3 weeks after VSTs for BKV infection.While there is no report of such a presenta- June 2021 and December 2022, 23 adults (age >18 years) KTR with ADV, BKV, or CMV infection received third-party (offthe-shelf) VSTs through a clinical trial at a tertiary center.Patient demographics, clinical presentation, IS management, antiviral treatment, and outcomes were characterized.The general approach to treat these infections at our center involved lowering the overall amount of IS.The initial step is dose reduction or discontinuation of antimetabolite followed by a reduction in the trough goal of tacrolimus.Based on immunologic risk, many patients were placed on prednisone after discontinuation of anti-metabolite.All KTR received VGCV for CMV prophylaxis as per our institution's protocol.Breakthrough CMV infections were treated with VGCV, ganciclovir (GCV), maribavir, or foscarnet as recommended by the transplant infectious disease physician.Intravenous immunoglobulin (IVIG) was used as adjunctive therapy in ADV infections.A patient who did not respond to the above interventions as indicated by rising viral load (VL), antiviral resistance, acute kidney injury (AKI defined as an increase in serum creatinine >0.3 mg/dL) or development of tissue invasive disease was considered for VSTs.VSTs are manufactured from the peripheral blood mononuclear cells (PBMCs) isolated from unrelated healthy volunteers.The PBMCs are incubated with specific viral antigens to generate interferon-gamma (IFN-y) by memory lymphoid cells with peptide specificity.These cells are then cultured in a media containing cytokines following which the final VST product constituted by IFN-y positive CD4 and CD8 T cells is harvested.1,9,10High-resolution human leukocyte antigen (HLA) typing between recipient and third-party VST donor was done to match HLA types with specificity for individual viruses.Criteria for VST infusion included any of the following: ADV PCR greater than 1000 copies/ml, BK viremia or nephropathy with BK PCR greater than 1000 copies/ml, CMV PCR greater than 500 IU/ml, CMV with concurrent therapy resistance or severe neutropenia due to antiviral.Key exclusion criteria were the use of T cell depleting therapy within 2 weeks of VSTs infusion, steroids >0.5 mg/kg/day, active malignancy, active graft versus host disease (GVHD) grades II-IV, and uncontrolled bacterial or fungal infection.Repeat VSTs infusions were allowed every 4 weeks if GVHD or other adverse effects had not occurred.Antiviral and IS management before and after VSTs was at the discretion of the clinical team.Selected pre-VSTs to post-VSTs changes in lab values were tested using two-tailed sign-rank tests applied to the post-minus-pre values.

2 1
months (range 1-55 months).Seven (41%) of the patients with BK viremia developed biopsy-proven BKVAN.Biopsy-proven acute granulomatous interstitial nephritis due to ADV was diagnosed in one patient.The median time to CMV infection was 2.5 months (range 1-5 months) after transplant surgery.Antiviral therapy treatment included GCV, VGCV, maribavir, and foscarnet.AKI led to the discontinuation of foscarnet in two patients.The median duration of antiviral therapy was * BK VL decreased from 0-2 wk.pre-VSTs level to 12 weeks post-VSTs (p<0.001)Trend of (Median) viral load pre-and post-virus-specific T cell therapy (VSTs).

2 (
Median) Serum creatinine and proteinuria pre-and post-virus-specific T cell therapy (VSTs).* VSTs use was associated with improvement in serum creatinine and proteinuria (p < .001).
tion after VSTs in the published literature thus far, the presentation of de novo MPGN chronologically post VSTs argues for further investigation.The second patient developed TCMR 4 weeks after the second VST infusion for BKV infection.Prior to VSTs, this patient's MMF was stopped due to infection.Our study did have certain limitations.First, it was a single-center study with a lack of external validity.Second, it was a retrospective study with limited numbers.Studying larger populations would offer more conclusive results.Third, with the simultaneous use of multiple strategies in the treatment of viral infections, it is difficult to fully delineate the impact of each individual treatment regimen.However, the trend in improvement of the VL post VSTs suggests the efficacy of VSTs which was not seen with reduction in IS alone.Fourth, the degree of