Does Post-Transplantation Cyclophosphamide Inhibit Graft-versus-Leukemia? (2020)

Allogeneic blood or marrow transplantation, or alloBMT, is a potentially curative therapy for a wide variety of hematologic malignancies and nonmalignant hematologic disorders. In the past, eligibility for alloBMT was limited to patients with an HLA-matched sibling or unrelated donor because partially HLA-mismatched transplants were associated with unacceptably high incidences of graft-versus-host disease (GVHD) and nonrelapse mortality, resulting in poor survival [1Szydlo R. Goldman J.M. Klein J.P. et al.Results of allogeneic bone marrow transplants for leukemia using donors other than HLA-identical siblings.J Clin Oncol. 1997; 15: 1767-1777Crossref PubMed Scopus (381) Google Scholar]. Fortunately, modern methods of GVHD prophylaxis, especially high-dose post-transplantation cyclophosphamide (PTCy), have effectively dismantled the HLA barrier to successful allogeneic hematopoietic cell transplantation (HCT) [2Kanakry C.G. Fuchs E.J. Luznik L. Modern approaches to HLA-haploidentical blood or marrow transplantation.Nat Rev Clin Oncol. 2016; 13: 10-24Crossref PubMed Scopus (157) Google Scholar]. The theory behind PTCy is that the drug is selectively toxic to proliferating alloreactive T cells while sparing the nonalloreactive population responsible for protection against infection. Thus, prevention of GVHD by PTCy requires 2 steps: (1) activation of alloreactive donor and host T cells upon infusion of the haploidentical stem cell graft and (2) administration of cyclophosphamide when the alloreactive T cells are most susceptible to being killed by the drug [3Mayumi H. Umesue M. Nomoto K. Cyclophosphamide-induced immunological tolerance: an overview.Immunobiology. 1996; 195: 129-139Crossref PubMed Scopus (96) Google Scholar]. A recent meta-analysis found results of haploidentical blood or marrow transplantation (haploBMT) plus PTCy were as good or better than results of transplants from HLA-matched unrelated donors, although still not as good as transplants from HLA-matched siblings [4Gagelmann N. Bacigalupo A. Rambaldi A. et al.Haploidentical stem cell transplantation with posttransplant cyclophosphamide therapy vs other donor transplantations in adults with hematologic cancers: a systematic review and meta-analysis.JAMA Oncol. 2019; 5: 1739-1748Crossref PubMed Scopus (28) Google Scholar]. These studies have established haploidentical HCT with PTCy as a viable option for patients lacking a matched sibling donor. The initial PTCy regimen developed at Johns Hopkins comprised a single dose of cyclophosphamide 50 mg/kg i.v. on day 3 followed by tacrolimus and mycophenolate mofetil (MMF) commencing on day 4 [5O'Donnell P.V. Luznik L. Jones R.J. et al.Nonmyeloablative bone marrow transplantation from partially HLA-mismatched related donors using posttransplantation cyclophosphamide.Biol Blood Marrow Transplant. 2002; 8: 377-386Abstract Full Text Full Text PDF PubMed Scopus (294) Google Scholar]. Unacceptably high incidences of graft failure and severe GVHD led us to add a second dose of cyclophosphamide on day 4 and to delay initiation of tacrolimus and MMF to day 5, and this produced acceptable incidences of graft failure and GVHD [6Luznik L. O'Donnell P.V. Symons H.J. et al.HLA-haploidentical bone marrow transplantation for hematologic malignancies using nonmyeloablative conditioning and high-dose, posttransplantation cyclophosphamide.Biol Blood Marrow Transplant. 2008; 14: 641-650Abstract Full Text Full Text PDF PubMed Scopus (1024) Google Scholar]. However, concerns were expressed that the incidence of relapse was too high, leading to the concern that PTCy was suppressing the graft-versus-leukemia effect of haploidentical HCT. Chiusolo et al. [7Chiusolo P. Bug G. Olivieri A. et al.A modified post-transplant cyclophosphamide regimen, for unmanipulated haploidentical marrow transplantation, in acute myeloid leukemia: a multicenter study.Biol Blood Marrow Transplant. 2018; 24: 1243-1249Abstract Full Text Full Text PDF PubMed Scopus (34) Google Scholar] modified the regimen by starting cyclosporine on day 0, MMF on day 1, and administering cyclophosphamide on days 3 and 5 after transplantation. The idea behind this modification is that early initiation of the calcineurin inhibitor (CNI) would prevent some of the T cell activation that is required for cyclophosphamide to induce tolerance, and so the graft-versus-leukemia effect would be better preserved. To determine the optimal timing and sequence of CNI and PTCy, Ruggeri et al. [8Ruggeri A. Labopin M. Battipaglia G. et al.Timing of post-transplantation cyclophosphamide administration in haploidentical transplantation: a comparative study on behalf of the Acute Leukemia Working Party of the European Society for Blood and Marrow Transplantation.Biol Blood Marrow Transplant. 2020; 26: 1915-1922Abstract Full Text Full Text PDF Scopus (5) Google Scholar] analyzed outcomes of 509 patients with leukemia in Europe receiving haploBMT incorporating different PTCy-based regimens of GVHD prophylaxis. In total, 385 patients received high-dose cyclophosphamide on days 3 and 4 and then started MMF plus a CNI (tacrolimus in 215 and cyclosporine in 170) on day 5, whereas 124 started cyclosporine and MMF on days 0 and 1, respectively, followed by high-dose cyclophosphamide on days 3 and 5. Results are summarized Table 1. Based upon these results, the authors conclude that an early start of immunosuppression combined with PTCy on days 3 and 5 improves leukemia-free survival and refined GVHD-free, relapse-free survival due primarily to a lower incidence of relapse.Table 1Outcomes of HaploBMT with Different PTCy Regimens in EuropeGVHD Prophylaxis, %OutcomePTCy Days 3, 4Tacrolimus/MMF Day ≥5PTCy Days 3, 4CsA/MMF Day ≥5CsA Day –1 or 0, MMF Day 1PTCy Days 3, 5P Valuen215170124Acute GVHD II-IV253918≤.01Chronic GVHD2521240.5Relapse363726.02Nonrelapse mortality262021.35Overall survival (2 years)444859.15Leukemia-free survival (2 years)434653.05Refined GVHD-free, relapse-free survival333946.02Notable outcomes are bolded.CsA indicates Cyclosporine A. Open table in a new tab Notable outcomes are bolded. CsA indicates Cyclosporine A. The study was limited by its retrospective nature and by differences between groups in key characteristics that are also associated with outcomes. For example, patients starting CNI before PTCy were significantly younger and more frequently received myeloablative conditioning and bone marrow grafts compared with patients starting CNI after PTCy. Their donors had a higher prevalence of cytomegalovirus seronegativity. These differences can confound the interpretation of the effects of GVHD prophylaxis on major outcomes. A valid comparison of the different PTCy-containing GVHD prophylaxis regimens would require a prospective, randomized trial in which the conditioning regimen and graft source are fixed or at least specified for each patient prior to randomization. The key question being addressed by this study is, when is the best time to initiate CNI, before or after PTCy? When our group translated PTCy to the clinic, 1 study in particular guided us to start CNI after the completion of high-dose cyclophosphamide. Nomoto et al. [9Nomoto K. Eto M. Yanaga K. Nishimura Y. Maeda T. Interference with cyclophosphamide-induced skin allograft tolerance by cyclosporin A.J Immunol. 1992; 149: 2668-2674PubMed Google Scholar] found that cyclosporine given from day –5 to day –1 blocked the induction of tolerance by the infusion of MHC-compatible allogeneic cells on day –2 and cyclophosphamide administration on day 0. Thus, while C3H strain mice, treated with major histocompatibility complex-compatible AKR strain spleen cells plus cyclophosphamide, accepted AKR skin grafts for over 100 days, the addition of the cyclosporine prior to tolerance induction reduced skin graft survival to a median of 18.4 days. The logical conclusion of this study was that T cell activation by antigen is required for cyclophosphamide to induce tolerance. Since the major limitation of T cell replete haploBMT is excessive bidirectional alloreactivity resulting in unacceptably high incidences of graft failure and severe GVHD, we did not want to diminish the tolerizing effect of cyclophosphamide by blocking T cell activation with early CNI administration. While it is certainly plausible that giving CNI before PTCy could decrease leukemia relapse by preventing the induction of tolerance in leukemia-specific T cells, it is hard to explain why the incidence of graft failure and of GVHD would not increase due to the blockade of tolerance induction in host and donor alloreactive T cells, respectively. It is possible that starting cyclosporine on day 0, as developed by Chiusolo et al., [7Chiusolo P. Bug G. Olivieri A. et al.A modified post-transplant cyclophosphamide regimen, for unmanipulated haploidentical marrow transplantation, in acute myeloid leukemia: a multicenter study.Biol Blood Marrow Transplant. 2018; 24: 1243-1249Abstract Full Text Full Text PDF PubMed Scopus (34) Google Scholar], is not so different from starting the drug on day 5, after PTCy administration. Alloreactive donor T cells home to secondary lymphoid organs and become activated almost immediately following infusion into allogeneic recipients [10Ford W.L. Atkins R.C. Specific unresponsiveness of recirculating lymphocytes after exposure to histocompatibility antigen in F 1 hybrid rats.Nat New Biol. 1971; 234: 178-180Crossref PubMed Scopus (94) Google Scholar]. Even if a CNI is given before the cells on day 0, it is likely that the concentration of the drug is too low to prevent the activation of most alloreactive host and donor T cells. Thus, it is likely that PTCy will have some or most of its tolerizing effect even when CNI is started on day 0. For this reason, it may be interesting to test the effect of starting CNI earlier, perhaps 3 or 4 days prior to the haplo allograft infusion. The study by Ruggeri et al. [8Ruggeri A. Labopin M. Battipaglia G. et al.Timing of post-transplantation cyclophosphamide administration in haploidentical transplantation: a comparative study on behalf of the Acute Leukemia Working Party of the European Society for Blood and Marrow Transplantation.Biol Blood Marrow Transplant. 2020; 26: 1915-1922Abstract Full Text Full Text PDF Scopus (5) Google Scholar] addresses the important question of the proper timing and sequence of CNI and PTCy administration for optimizing outcomes of haploBMT. While results suggest that CNI initiation before PTCy improves outcomes, this possibility needs to be tested in a prospective, randomized trial to eliminate imbalances in treatment groups. Financial disclosure: Supported by P01 CA225618 from the US National Cancer Institute ( E.J.F. ). Conflict of interest statement: There are no conflicts of interest to report.