Introduction

 

Post-transplant lymphoproliferative diseases (PTLD) are heteroge- neous lymphoid disorders ranging from  indolent polyclonal prolifer- ations to  aggressive lymphomas, which may  complicate solid  organ transplantation (SOT) or  haematopoietic stem cell  transplantation [1]. The highest risk of SOT-related PTLD occurs  in heart or lung  (up to 20 %) transplant recipients, while the  risk is lower (1-5%) in liver and  renal ones,  where inferior and  shorter doses of immunosuppres- sors are used  [1,2].

 

Most  PTLD cases  are  of B cell origin  (> 85-90%) and  Epstein Barr Virus (EBV) is associated in two  thirds of these. While  EBV-negative PTLD tends to occur  in older  patients, with a longer latency after  SOT (> 5 years  versus 6−12 months in EBV+ patients) [3,4], no differences in response or survival have  been described [5]. The standard of care for PTLD has  traditionally involved reduction of immunosuppressive treatment and  rituximab-based chemotherapy regimens [6]. Patients refractory to rituximab have  dire prognosis and  other approaches are justified [7].  Specific   cytotoxic T-lymphocytes infusions have  been * Corresponding author.

 

E-mail address: hernani_raf@gva.es (R. Hernani).

 

successfully used  in EBV-related PTLD, although their availability is limited [4].

 

CD19 chimeric antigen receptor T-cell (CAR-T) therapy has shown remarkable  efficacy in  patients with relapsed or  refractory diffuse large  B-cell lymphoma (DLBCL) [8,9]. However, data  on outcomes of CAR-T in the  setting of PTLD are  scarce  [10−15].  Moreover, limited information is available on the influence of concomitant immunosup- pressive drugs on CAR-T function [16,17].

 

Herein we  describe management of a patient treated with CAR-T therapy for PTLD after  renal transplantation. Additionally, we suggest a strategy for minimizing immunosuppressive treatment in this clini- cal scenario. This report also  reviews the  limited published data  in the setting of SOT-related PTLD treated with CAR-T.

 

Methods

 

Both cytokine release syndrome (CRS) and  immune effector cell- associated neurotoxicity syndrome (ICANS) were graded according to the  American Society  for Transplantation and  Cellular  Therapy con- sensus [18]. Assessment of response to CAR-T therapy was  based on the  LYRIC criteria [19]. Infectious prophylaxis followed Spanish con- sensus  guidelines [20].  Cytopenias were  graded  according to  the Common  Terminology  Criteria  for  Adverse   Events   v5.0  scale.   To access   data   for  the   review  process,  a  search  was   performed  in PubMed using  the  search terms “post-transplant lymphoproliferative disease”, “PTLD”, “chimeric antigen receptor T-cell” and “CAR-T”. Oral presentations  from    Haematology  conferences   since    2017    (not Pubmed-indexed)  were   also    included.  Informed  consent   was obtained from  the  patient to use his clinical  information for research purposes.

 

Case report

 

A 56-year-old male  was diagnosed with DLBCL (EBV negative, ger- minal center phenotype, no c-Myc  rearrangement) in 2017.  In 2000, he  had  undergone cadaveric renal transplantation due  to  end-stage kidney disease of unknown etiology. At the  time of PTLD diagnosis, his  immunosuppression regimen included tacrolimus, mycopheno- late  mofetil (MMF) and  prednisone (5 mg/day). Both tacrolimus and MMF were switched to  everolimus (1.25  mg/12 h)  after  diagnosis (target levels: 3.5−5 ng/ml) and prednisone was maintained. Protein- uria  (1.92  g/24  h) was  detected two  years  after  everolimus conver- sion,  which responded well  to angiotensin converting enzyme inhibitors (ACEI). He had  stage  IV disease with peritoneal infiltration and  an international prognostic index (IPI) of 3. The patient received six courses of rituximab, cyclophosphamide, doxorubicin, vincristine and  prednisone (R-CHOP), and  two  additional doses of rituximab, achieving complete response (CR), as assessed by positron emission tomography (PET). Five months later,  biopsy  of an enlarged iliac adenopathy confirmed disease recurrence, for which the  patient was given  two  cycles  of rituximab, ifosfamide, carboplatin and  etoposide (R-ICE) salvage therapy, achieving CR2. Despite the  absence of mor- phologic  bone    marrow  infiltration  by   lymphoma,  the    patient remained with persistent cytopenias that precluded the  mobilization of autologous stem cells. At that time,  the  patient was  not  eligible  for allotransplant due  to lack of a suitable donor. Four  courses of dose- adjusted rituximab, gemcitabine and  oxaliplatin (R-GEMOX) were therefore prescribed as  consolidation therapy.  Despite this,  a  PET scan  performed two  months later showed supra- and  infradiaphrag- matic progression that was confirmed by tissue biopsy.  Polatuzumab, bendamustine and   rituximab (PBR) were administered (only   five cycles  due   to  haematological  toxicity), reaching  CR3. Six  months later,  an  isolated inguinal relapse was  again  confirmed by PET scan. He underwent radiation therapy (30 Gy), but  posttreatment evalua- tion showed infradiaphragmatic progression (stage II, IPI 1).

 

We  decided to proceed to CAR-T therapy, selecting axicabtagene ciloleucel (Axi-cel) for two  reasons: firstly, because despite a shorter duration,  CD28  costimulatory  domain  seems  to  produce  a  more intense signalling than 4-1BB [21], and secondly, since reintroduction of immunosuppressive agents might be required after  CAR-T if graft rejection was  suspected. Everolimus was  temporally discontinued before apheresis (following Kite@  instructions) and  then definitively stopped before CAR-T infusion. Prednisone was  maintained at physi- ological  dose  during the  entire process avoid  adrenal insufficiency. Bridging  therapy with 2 cycles of cyclophosphamide (500  mg/m2 day 1)  and  prednisone (60  mg/m2  day  1−5) was  prescribed.  PET-scan

after  bridging therapy demonstrated  peritoneal progression (Deau- ville score  5), despite which it was  decided to procced to CAR-T ther- apy.  At the  time of CAR-T infusion, creatinine, glomerular filtration rate  (Cockcroft-Gault  Equation)  and  24-h  urine  protein  were 1.35 mg/dL, 56.7 mL/min and 0.06 g, respectively.

 

Clinical course after CAR-T therapy

 

Time  from  apheresis to CAR-T infusion was  64 days.  The patient received the  standard lymphodepleting chemotherapy regimen con- sisting of three days  of FluCy (dose-adjusted fludarabine 24 mg/m2/d and  cyclophosphamide  500 mg/m2/d).  Axi-cel was finally administered in  August  2020.  The  patient developed grade I CRS, from  day +1 to +10, which resolved after  one  dose  of tocilizumab. He also developed grade II ICANS on day +7, which resolved immediately after  initiating dexamethasone (10 mg with subsequent tapering over seven days,  with a cumulative dose  of 70 mg).  Kidney  function was closely  monitored, as were inflammatory markers including c-reac- tive  protein, procalcitonin, ferritin and  interleukin-6, reaching peak levels  of 114 mg/L, 0.72 ng/mL, 1428  ng/mL and  1715  pg/mL, respec- tively. No impairment of renal function was detected, and  the  patient was  finally discharged on day +14. However, clinical  course was  sud- denly complicated by a septic shock  and  right lower lobe pneumonia due  to Pseudomonas aeruginosa, that required  intensive  care  unit admission for vasopressor support. On day +90 cytomegalovirus infection was  detected, which resolved spontaneously without pre- emptive therapy. During   the  following weeks, the  patient experi- enced grade 3 anaemia and grade 4 thrombocytopenia and neutrope- nia,  which were managed with cytokine combination therapy with darbopoetin alfa (150 mg/week), eltrombopag (50 mg/day), and gran-

ulocyte colony-stimulating factor  (480 mg 2−3 times per week). Bone marrow was hypocellular without dysplasia or signs of viral infection or   hemophagocytic lymphohistiocytosis. Creatinine and  glomerular filtration  rate   have   remained  relatively  stable  (1.31   mg/dL   and 60   mL/min  at   last   follow-up). Significant 24-h  proteinuria  was detected three months after  CAR-T infusion (0.6 g), reaching a peak one   month later (1.25   g).  It  is  progressively responding  to  ACEI (0.84   g  after   two   months).  If  worsening,  a  graft-biopsy  will   be planned to discard graft  rejection. At one  month after  CART infusion, the  patient achieved CR disease status, and  this  response was  sus- tained up to the last follow-up (10 months).

Tailoring immunosuppressive treatment and renal monitoring during

 

CAR-T therapy

 

Everolimus was  not  reintroduced after  CAR-T therapy. Both clini- cal  and  analytical parameters were closely  monitored for  signs  of graft  rejection (typically manifesting as  hypertension and  protein- uria)  or indirect markers of increased risk of graft  rejection (develop- ment of anti-HLA  antibodies, recovery of B lymphocytes or  plasma cells). Prednisone at physiological dose was discontinued two months after   CAR-T infusion, without  subsequent signs  of  adrenal insuffi- ciency.  CAR-T lymphocytes were detected by flow  cytometry up  to three months after  infusion, in keeping with previous data  [21,22]. At last  follow-up, no  clear  signs  of graft  rejection had  developed, and the patient continues without immunosuppressive treatment.

 

Literature review

 

Table 1 summarizes available data  on  the  use  of CAR-T for PTLD after  SOT (n = 12) [10−15]. As can  be seen,  EBV was  only  associated in  2  cases  (17%), which appeared soon  after  transplant (4  and  21 months). Kidney  and  liver transplants were the  most frequent (7 and 3, respectively) and  DLBCL was  the  predominant  histology (n  = 11 [92%]). Notably,  five  patients completely discontinued immunosup- pressive drugs before or soon  after  CAR-T infusion. Despite the  short observation period, one  case  of graft  rejection was  documented. The patient remained in CR and  it was  decided not  to reintroduce immu- nosuppressive therapy  in  order  to   reduce  the   relapse  risk.   Ten patients developed CRS (grade I - II [n = 9]; grade III [n = 1]), and four patients developed ICANS (grade III [n = 3];  grade IV [n = 1]). One patient with disease persistence died  at day  +115  of unclarified encephalopathy, whereas the  other two  other patients died  of infec- tion  and  progressive disease, respectively. Complete remission was achieved in  seven patients (58%), all of them still  alive  and  five  in ongoing response.

 

Discussion

 

Very  few  cases  have  been reported on  CAR-T therapy for  PTLD after  solid organ transplantation. Here, we have  described the  clinical outcome of one  further patient and  propose a strategy for tailoring immunosuppressive treatment  and   organ  monitoring  in  patients with kidney allografts after   CAR-T infusion, to  add  to  the  limited body of evidence-based medical literature.

 

CAR-T therapy was  decided in our  patient after  refractoriness to several lines  of immunochemotherapy and  non-transplant-eligible status. We were concerned that concomitant use of immunosuppres- sors  prior   to  apheresis or  after   CAR-T infusion could  theoretically suppress expansion of CAR-T lymphocytes [17], an thus increase the relapse rate  [16]. Nonetheless, partial reduction or total discontinua- tion of immunosuppressive therapy would pose  the risk of enhancing the  allo-immune response and  leading to chronic-graft rejection [23,24],  mainly due  to  humoral mechanisms, typified by  hyperten- sion  and  renal dysfunction (detected by a serum creatinine increase or the  appearance of proteinuria).

 

After  discussing the  abovementioned issues within a multidisci- plinary team of nephrologists and  clinical  and  laboratory haematolo- gists,   it   was   decided  to   discontinue  immunosuppressive  agents before CAR-T infusion in order to enhance expansion and  capability to reduce disease relapse. It is worth noting that no antiHLA antigens, plasma cells  or  B lymphocytes were detected  before infusion. We hypothesized that the  immune pathways that could  induce allo-graft rejection would be heavily suppressed, presumably as a result of pre- vious  treatments [25].  In light  of this,  immunosuppressors were no longer used  and  close  monitoring was  carried out  (Fig. 1) to detect preliminary  signs   of  rejection.  Seven   months  after   infusion the patient remains in CR without immunosuppressive drugs.  Proteinuria has  been detected, but  it  is progressively decreasing after  starting ACEI.

The  present data,   together with previous reports, suggest that CAR-T is an effective treatment for PTLD, without severe toxicity and capable of inducing sustained responses. Specific  further research in this  patient subset is currently underway [26,27].  Although Luttwak et  al found no  alterations in  CAR-T expansion despite concomitant immunosuppressive therapy [11], reduction or complete discontinu- ation of immunosuppressive treatment remains the  first step  to treat PTLD. According to published data  and our own  experience, it may be feasible to  completely discontinue immunosuppressive therapy, although more data   and  longer follow-up are  required to  confirm this  hypothesis. A close  monitoring of graft  function and  immune reconstitution might herald an  indication to reintroduce the  immu- nosuppressive treatment.

 

Parallel to the  growing use  of CAR-T therapy, the  number of kid- ney  transplants is also  rising  annually (http://www.transplant-obser vatory.org), which might increase the  number of CAR-T procedures offered to PTLD. The final  decision on whether to initiate CAR-T ther- apy  should hinge on  multidisciplinary team evaluation and  be  tai- lored  to  each  patient on  a  case-by-case basis.  Patients with lower dose  of immunosuppressive treatment and  without recent history of graft rejection are probably the best  candidates.