Transplantation and Cellular Therapy 27 (2021) 642649 Transplantation and Cellular Therapy journal homepage: www.tctjournal.org Guideline Standardizing Definitions of Hematopoietic Recovery, Graft Rejection, Graft Failure, Poor Graft Function, and Donor Chimerism in Allogeneic Hematopoietic Cell Transplantation: A Report on Behalf of the American Society for Transplantation and Cellular Therapy Mohamed A. Kharfan-Dabaja1,*, Ambuj Kumar2, Ernesto Ayala1, Mahmoud Aljurf3, Taiga Nishihori4, Rebecca Marsh5, Lauri M. Burroughs6, Navneet Majhail7, A. Samer Al-Homsi8, Zaid S. Al-Kadhimi9, Merav Bar6, Alice Bertaina10, Jaap J. Boelens11, Richard Champlin12, Sonali Chaudhury13, Zachariah DeFilipp14, Bhagirathbhai Dholaria15, Areej El-Jawahri14, Suzanne Fanning16, Ellen Fraint11, Usama Gergis17, Sergio Giralt18, Betty K. Hamilton7, Shahrukh K. Hashmi19,20, Biljana Horn21, Yoshihiro Inamoto22, David A Jacobsohn23, Tania Jain24, Laura Johnston25, Abraham S. Kanate26, Ankit Kansagra27, Adetola Kassim15, Leslie S. Kean28, Carrie L. Kitko29, Jessica Knight-Perry30, Joanne Kurtzberg31, Hien Liu4, Margaret L. MacMillan32, Zahra Mahmoudjafari33, Marco Mielcarek6, Mohamad Mohty34, Arnon Nagler35, Eneida Nemecek36, Timothy S. Olson37, Betul Oran12, Miguel-Angel Perales18, Susan E. Prockop11, Michael A. Pulsipher38, Iskra Pusic39, Marcie L. Riches40, Cesar Rodriguez41, Rizwan Romee42, Gabriela Rondon12, Ayman Saad43, Nina Shah44, Peter J. Shaw45, Shalini Shenoy46, Jorge Sierra47, Julie Talano48, Michael R. Verneris30, Paul Veys49, John E. Wagner32, Bipin N. Savani15, Mehdi Hamadani50, Paul A. Carpenter6 1 Division of Hematology-Oncology and Blood and Marrow Transplantation and Cellular Therapies Program, Mayo Clinic, Jacksonville, Florida 2 Program for Comparative Effectiveness Research, Morsani College of Medicine, University of South Florida, Tampa, Florida 3 Department of Adult Hematology and Stem Cell Transplantation, King Faisal Specialist Hospital and Research Centre, Riyadh, Saudi Arabia 4 Department of Blood and Marrow Transplantation and Cellular Immunotherapy, Moffitt Cancer Center, Tampa, Florida 5 Cincinnati Children's Hospital Medical Center, Cincinnati, Ohio 6 Fred Hutchinson Cancer Research Center, Seattle, Washington 7 Blood and Marrow Transplant Program, Taussig Cancer Institute, Cleveland Clinic, Cleveland, Ohio 8 NYU Langone Health, New York, New York 9 Division of Oncology and Hematology, Department of Internal Medicine, University of Nebraska Medical Center, Omaha, Nebraska 10 Division of Stem Cell Transplant and Regenerative Medicine, Department of Pediatrics, Stanford University, Stanford, California 11 Stem Cell Transplantation and Cellular Therapies Program, Department Pediatrics, Memorial Sloan Kettering Cancer Center, New York, New York 12 Department of Stem Cell Transplantation and Cellular Therapy, The University of Texas MD Anderson Cancer Center, Houston, Texas 13 Ann and Robert H. Lurie Children's Hospital of Chicago, Chicago, Illinois 14 Department of Hematology-Oncology and Hematopoietic Cell Transplant and Cellular Therapy Program, Massachusetts General Hospital, Boston, Massachusetts 15 Department of Hematology-Oncology, Vanderbilt University Medical Center, Nashville, Tennessee 16 Blood and Marrow Transplant Program, University of South Carolina School of Medicine, Greenville, South Carolina 17 Bone Marrow Transplant and Immune Cellular Therapy, Thomas Jefferson University Hospital, Philadelphia, Pennsylvania 18 Department of Medicine, Division of Hematologic Malignancies, Memorial Sloan Kettering Cancer Center Weill Cornell Medical College, New York, New York 19 Division of Hematology, Department of Internal Medicine, Mayo Clinic, Rochester, Minnesota 20 Department of Medicine, Sheikh Shakhbout Medical City, Abu Dhabi, United Arab Emirates 21 Department of Pediatrics, Division of Hematology/Oncology, University of Florida, UF Health Shands Children's Hospital, Gainesville, Florida 22 Department of Hematopoietic Stem Cell Transplantation, National Cancer Center Hospital, Tokyo, Japan 23 Division of Blood and Marrow Transplantation Center for Cancer and Blood Disorders, Children's National Medical Center, Washington, DC 24 Hematologic Malignancies and Bone Marrow Transplantation Program, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, Maryland 25 Division of Blood and Marrow Transplantation, Department of Medicine, Stanford University School of Medicine, Stanford, California 26 West Virginia University, Morgantown, West Virginia 27 UT Southwestern Medical Center, Dallas, Texas Financial disclosure: See Acknowledgments on page 648. *Correspondence and reprint requests: Mohamed A. Kharfan-Dabaja, MD, MBA, FACP, Mayo Clinic Florida, 4500 San Pablo Road, Jacksonville, FL 32224. E-mail address: KharfanDabaja.Mohamed@mayo.edu (M.A. Kharfan-Dabaja). https://doi.org/10.1016/j.jtct.2021.04.007 2666-6367/© 2021 The American Society for Transplantation and Cellular Therapy. Published by Elsevier Inc. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/) M.A. Kharfan-Dabaja et al. / Transplantation and Cellular Therapy 27 (2021) 642649 643 28 Boston Children's Hospital, Dana-Farber Cancer Institute, Boston, Massachusetts 29 Division of Pediatric Hematology/Oncology, Department of Pediatrics, Vanderbilt University Medical Center, Nashville, Tennessee 30 Department of Pediatrics, Division of Hematology/Oncology/BMT, University of Colorado School of Medicine, Aurora, Colorado 31 Pediatric Blood and Marrow Transplant Program, Duke University School of Medicine, Durham, North Carolina 32 Blood and Marrow Transplant Program, Masonic Cancer Center, University of Minnesota Medical School, Minneapolis, Minneapolis 33 Division of Pharmacy, University of Kansas Cancer Center, University of Kansas Health System, Lawrence, Kansas 34 Sorbonne Universite, INSERM, Centre de Recherche Saint-Antoine and Ho ^pital Saint-Antoine, Service d'Hematologie Clinique et Therapie Cellulaire, Paris, France 35 Chaim Sheba Medical Center, Tel Hashomer, Israel 36 Knight Cancer Institute, Oregon Health & Science University, Portland, Oregon 37 Blood and Marrow Transplant Section, Division of Oncology, Department of Pediatrics, Children's Hospital of Philadelphia, Pennsylvania 38 Children's Hospital Los Angeles Cancer and Blood Disease Institute, USC Keck School of Medicine, Los Angeles, California 39 Division of Oncology, Department of Medicine, Washington University School of Medicine, St Louis, Missouri 40 Division of Hematology, University of North Carolina at Chapel Hill, North Carolina 41 Department of Hematology and Oncology, Wake Forest University School of Medicine, Winston-Salem, North Carolina 42 Cellular Therapy and Stem Cell Transplant Program, Dana Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 43 Division of Hematology, The Ohio State University, Columbus, Ohio 44 Division of Hematology-Oncology, Department of Medicine, University of California San Francisco, San Francisco, California 45 The Children's Hospital at Westmead, Sydney, Australia 46 Department of Pediatrics, Washington University School of Medicine, St. Louis, Missouri 47 Department of Hematology, Hospital de la Santa Creu i Sant Pau, Josep Carreras Leukemia Research Institute, Barcelona, Spain 48 Department of Pediatric Hematology/Oncology, Children's Hospital of Wisconsin, Medical College of Wisconsin, Milwaukee, Wisconsin 49 Blood & Marrow Transplant Unit, Great Ormond Street Hospital, University College London, London, United Kingdom 50 Division of Hematology and Oncology, Department of Medicine, Medical College of Wisconsin, Milwaukee, Wisconsin Article history: A B S T R A C T Received 11 April 2021 Allogeneic hematopoietic cell transplantation (allo-HCT) is potentially curative for certain hematologic malignan- Accepted 11 April 2021 cies and nonmalignant diseases. The field of allo-HCT has witnessed significant advances, including broadening indications for transplantation, availability of alternative donor sources, less toxic preparative regimens, new cell Key Words: manipulation techniques, and novel GVHD prevention methods, all of which have expanded the applicability of Allogeneic hematopoietic cell the procedure. These advances have led to clinical practice conundrums when applying traditional definitions of transplantation hematopoietic recovery, graft rejection, graft failure, poor graft function, and donor chimerism, because these Hematopoietic recovery may vary based on donor type, cell source, cell dose, primary disease, graft-versus-host disease (GVHD) prophy- Graft rejection laxis, and conditioning intensity, among other variables. To address these contemporary challenges, we surveyed Graft failure a panel of allo-HCT experts in an attempt to standardize these definitions. We analyzed survey responses from Donor chimerism adult and pediatric transplantation physicians separately. Consensus was achieved for definitions of neutrophil and platelet recovery, graft rejection, graft failure, poor graft function, and donor chimerism, but not for delayed engraftment. Here we highlight the complexities associated with the management of mixed donor chimerism in malignant and nonmalignant hematologic diseases, which remains an area for future research. We recognize that there are multiple other specific, and at times complex, clinical scenarios for which clinical management must be individualized. © 2021 The American Society for Transplantation and Cellular Therapy. Published by Elsevier Inc. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/) INTRODUCTION not be true in the case of primary graft failure with autologous Allogeneic hematopoietic cell transplantation (allo-HCT) is neutrophil recovery. Moreover, in NMA conditioning, espe- potentially curative for certain hematologic malignancies and cially for nonmalignant diseases, engraftment definitions nonmalignant diseases [1,2] and is now more broadly applica- based only on neutrophil recovery but agnostic of chimerism ble owing to improved outcomes using not just HLA-matched are insufficient to address late graft failure, because neutrophil sibling or unrelated donors, but also umbilical cord blood and recovery might occur with complete loss of donor chimerism. haploidentical related donors. Increased options of reduced- Contemporary allo-HCT practice measures the level of intensity (RIC) or nonmyeloablative (NMA) conditioning can donor chimerism by quantifying the percentage of unsorted mitigate transplantation-related mortality in high-risk allo- marrow and/or microfluorimetrically sorted peripheral blood HCT and further expand feasibility [3-7]. More than 95% of leukocytes. Leukocyte compartments most commonly evalu- patients needing an allo-HCT now have access to a suitable ated include granulocytes (via CD33 surface expression) and T donor. These advances have led to clinical practice conun- cells (CD3), sometimes extending to B cells (CD19) and natural drums when applying traditional definitions of hematopoietic killer cells (CD56), depending on the disease indication for recovery, graft rejection, and graft failure, because hematopoi- allo-HCT. Full donor chimerism is most commonly defined etic recovery and engraftment kinetics may vary based on empirically using a cutoff of >90%, >95%, or >97.5%. Partial, donor type, cell source, cell manipulation and dose, primary more commonly known as mixed, donor chimerism has been disease, graft-versus-host disease (GVHD) prophylaxis, and defined empirically using lower-end cutoffs such as >2.5% up conditioning intensity, among other factors [8-12]. In daily to >10% and upper-end cutoffs such as 90% up to 97.5%. clinical practice definitions have been used interchangeably, Absence of donor chimerism has been empirically defined by albeit inconsistently or incorrectly. For instance, recovery of cutoffs of <2.5%, <5%, and <10% [13]. RIC or NMA conditioning neutrophils, operationally defined as achievement of an abso- regimens are more likely to result in mixed donor chimerism lute neutrophil count (ANC) of 500 cells/mL, is at times also compared with myeloablative conditioning (MAC) regimens used to define hematopoietic engraftment. The latter implies [14]. In the 2001 workshop, the authors recommended avoid- that donor neutrophils have reached 500 cells/mL, which might ing using a specific threshold to define mixed chimerism, 644 M.A. Kharfan-Dabaja et al. / Transplantation and Cellular Therapy 27 (2021) 642649 owing to multiple factors that may potentially affect results, Table 1 including assay sensitivity, underlying disease, and cellular Panel Member Demographics compartment(s) involved; accordingly, mixed chimerism was Characteristic Adult Transplantation Pediatric Transplantation simply defined as any reliable detection of host hematopoietic Physicians (n = 20) Physicians (n = 19) cells, hence the wide range still used in current practice [15]. Age, yr, n (%) Although this is conceptually factual, certain levels of mixed 5-45 6 (30) 4 (21) chimerism within this wide range might be considered clini- 46-55 10 (50) 8 (42) cally acceptable or not, depending on the underlying allo-HCT 56-65 3 (15) 6 (32) indication. For instance, in patients who undergo allo-HCT for >65 1 (5) 1 (5) sickle cell disease, a certain level of stable mixed donor mye- Sex, n (%) loid chimerism is considered an acceptable endpoint for dis- Female 3 (15) 10 (53) ease correction because it tracks with donor erythrocyte Male 17 (85) 9 (47) chimerism. Similarly, marrow failure syndromes, like Fanconi Years in practice, anemia or Diamond Blackfan anemia, are correctable with sta- n (%) ble mixed chimerism, but anything less than full donor mye- <5 1 (5) 1 (5) loid chimerism does not erase concerns about future (host) 5-10 3 (15) 2 (11) myelodysplasia or acute myelogenous leukemia. After allo- 11-15 7 (35) 2 (11) HCT for hematologic malignancies, it is intuitive that early, 16-20 5 (25) 6 (32) durable full donor chimerism is desirable, because a low level >20 4 (20) 8 (42) of donor chimerism is associated with an elevated risk for dis- ease relapse [16,17]. To address these contemporary challenges, we organized a panel of pediatric and adult allo-HCT experts to attempt to RESULTS standardize definitions of hematopoietic (neutrophil and Demographics platelet) recovery, graft rejection, primary and secondary graft During the initial round, 20 of the 25 invited identified failure, poor graft function, and donor chimerism. We con- themselves as adult-treating and 19 of 23 invited identified ducted a survey of adult and pediatric transplantation physi- themselves as pediatric-treating transplantation physicians. In cians separately. Whenever applicable, we provide clinical the adult-treating group, the majority were male (85%), and scenarios to highlight the application of these consensus defi- 80% of participants had more than 10 years of experience. In nitions in clinical practice. the pediatric-treating group, female physicians composed 53% of the total participants, and 85% had more than 10 years of experience (Table 1). METHODS Panel Composition Practice Characteristics We assembled a steering committee of 14 physicians with expertise in The majority (55%) in the adult-treating group reported prac- allo-HCT and 1 independent methodology expert with expertise in evidence ticing in transplant centers that perform >100 allo-HCT proce- synthesis and the RAND-modified Delphi methods to develop the survey dures annually, mostly from HLA-matched unrelated donors questions. The steering committee drafted the protocol and consensus state- ments based on systematic review of the literature and clinical practice con- (MUDs) using peripheral blood stem cells as the preferred cell siderations, and set up the expert panel. The first round of the survey source. On the other hand, the majority in the pediatric-treating comprised 3 questions on panel demographics, 4 questions on practice set- group (63%) reported practicing in centers that perform >50 allo- ting, and 26 questions on definitions and clinical management. Considering HCT procedures annually, mostly from HLA-MUDs using bone the differences in transplantation practices across age groups, we assembled separate final panels for adult (n = 25) and pediatric (n = 23) HCT physicians marrow (BM) as the preferred cell source (Table 2). with steering committee representative(s) from either panel. Panel physi- cians included diverse geographical representation and expertise in the field, Definitions and Management Recommendations as demonstrated by a track record of peer-reviewed publications, leadership of clinical trials, and involvement in national and international transplanta- Neutrophil and Platelet Recovery tion societies. The methodology expert assisted with designing, developing, As summarized in Table 3, both the adult and pediatric trans- and administering the surveys along with data collection and analysis, but plantation panels endorsed existing working definitions of neutro- did not participate in the voting process at any stage. phil and platelet recovery set forth by the Center for International Blood and Marrow Transplant Research (CIBMTR). Methodology Delayed Engraftment, Graft Rejection, Graft Failure, Poor Graft Before voting on recommendations, a formal guidance document on the Function, and Secondary Graft Failure RAND-modified Delphi method was shared with all participants; a statement Neither panel could reach consensus on the definition of with 70% vote in favor/against was considered a consensus. A formal evalu- ation of patient values and preferences and cost was not conducted; how- delayed engraftment (Table 3). However, there was consensus ever, an overall assumption was made that recommendations would be on differentiating graft rejection from graft failure, defining feasible in the setting across panel member practices and would not add any poor graft function, and secondary graft failure. Both panels additional burden on existing resources. The questions covered the broad defined graft rejection as an immune-mediated process, domains of definitions and clinical practice recommendations. Consistent with the RAND-modified Delphi methodology, participants rated all state- whereas graft failure was considered to represent a wider ments anonymously for all rounds of voting. Furthermore, all but one rounds array of possibilities, including cell dosing, disease, infections, of voting were administered online via Qualtrics survey management soft- drugs, and an immune-mediated event. ware (Qualtrics, Provo, UT). One round of the survey was administered anon- There was consensus on defining primary graft failure ymously online via Poll Everywhere (pollev.com) during the virtual meeting aimed at reaching consensus, in lieu of an in-person meeting, owing to the based on different cell sources. For instance, both panels COVID-19 pandemic. Descriptive statistics was used to analyze the results of defined graft failure when using peripheral blood stem cells or the survey, which are summarized as proportions. unstimulated BM as lack of achievement of an ANC 500/mL M.A. Kharfan-Dabaja et al. / Transplantation and Cellular Therapy 27 (2021) 642649 645 Table 2 regimen or whether T cell-replete or T cell-depleted grafts Transplant Center and Practice Characteristics were prescribed (Table 4). Center Characteristics Adult Pediatric Transplantation Transplantation Managing Mixed Donor Chimerism in Allo-HCT Physicians Physicians Although both panels endorsed the current definition of Number of allo-HCTs/yr, n (%) mixed donor chimerism as 5% to 95% for both myeloid and <50 2 (10) 7 (37) lymphoid lineages, they also recognized the practical impor- 51-100 7 (35) 9 (47) tance of using the actual percentage of donor cells for clinical 101-200 6 (30) 1 (5) management. In this case, the panels recommended that indi- >200 5 (25) 2 (11) vidual patient management be based on a consideration of the Cell source, n (%)* actual (downward or upward) trajectories of percent donor PBSCs 20 (74) 5 (21) myeloid and lymphoid chimerism, together with complete Unstimulated BM 6 (22) 13 (54) blood counts and clinical status. Because trajectories can help G-CSF-stimulated BM 1 (4) 0 inform medical decision making, the panel recommends mea- UCB 0 4 (17) suring donor chimerism at specific time points during the first year post-allo-HCT (eg, days +30, +90, +180, and +365). Other 0 2 (8)y Pertaining to specific management of downward trajecto- Donor type, n (%)* ries in donor CD3 and CD33 chimerism in the malignant dis- MRD 10 (29) 7 (17) ease setting, Figure 1 highlights the complex decision-making MUD 16 (47) 15 (36) in real-world clinical practice. Although withdrawal (or taper- MMRD/haploidentical 7 (21) 7 (17) ing) of immune suppression appeared to be the most fre- MMUD 0 6 (14) quently chosen approach for declining donor chimerism, the UCB 1 (3) 7 (17) median response was to select 2 possible therapeutic options PBSCs indicates peripheral blood stem cells; UCB, umbilical cord blood; MRD, (range, 1 to 5). Decision making is further complicated by HLA-matched related donor; MUD, HLA-matched unrelated donor; MMRD: other variables, for example, did the downward donor chime- HLA-mismatched related donor; MMUD: HLA-mismatched unrelated donor. * More than one option may apply/the number of responses may exceed rism trajectory occur in the setting of disease relapse/progres- the number of participants. sion or during a continuous remission? Was GVHD present or y Alpha-beta-depleted haploidentical transplantation (n = 1), ex vivo T cell- not? What was the ablative intensity of the conditioning regi- depleted PBSCs (n = 1). men? Was T cell depletion part of conditioning? These are just a few examples. by day +30 with associated pancytopenia. When using cord Figure 2 also highlights the complex clinical decision pro- blood, both panels defined graft failure as lack of achievement cess associated with managing similar downtrends in donor of an ANC 500/mL by day +42 with associated pancytopenia. chimerism in the context of nonmalignant disease. Interest- This assumes that donor chimerism testing is also done to con- ingly, fewer adult transplantation physicians responded to this firm the suspicion of graft failure. There was no consensus by question relative to pediatric transplantation physicians either panel to define graft failure when using G-CSF-stimu- (n = 15 versus n = 17; Figure 2) compared with responses to lated BM (Table 3). the same questions for malignant disease context (n = 19 ver- Poor graft function was defined as frequent dependence on sus n = 16; Figure 1). This may highlight the broader experi- blood and/or platelet transfusions and/or growth factor sup- ence with allo-HCT for nonmalignant diseases in the pediatric port in the absence of other explanations, such as disease transplantation group. Both panels recognized that the level of relapse, drugs, or infection (Table 3). donor chimerism required for nonmalignant disease correc- Secondary graft failure was defined as a decline in hemato- tion depends on the specific underlying disease indication for poietic function (possibly involving hemoglobin and/or plate- transplantation; however, specific levels of donor chimerism lets and/or neutrophils) necessitating blood products or were not discussed for particular disease indications. growth factor support, after having met the standard defini- Finally, the panel could not identify a single best approach tion of hematopoietic (neutrophil and platelet) recovery to the management of down-trending donor chimerism in (Table 3). Both panels recommended assessing secondary graft either the malignant or the nonmalignant disease setting. failure by evaluating hematopoietic function (based on periph- eral blood counts), BM cellularity, and donor chimerism DISCUSSION (Table 4). This work represents an effort to standardize definitions of hematopoietic recovery, graft rejection, primary and second- ary graft failure, poor graft function, and donor chimerism in Donor Chimerism the setting of allo-HCT, and also to provide broad guidance on Both panels endorsed current definitions of full donor chi- the clinical management of mixed donor chimerism. We rec- merism as >95%, mixed donor chimerism as 5% to 95%, and ognize that there are multiple other specific and at times com- absent donor chimerism as <5%, for both myeloid and lym- plex clinical scenarios for which clinical management must be phoid lineages (Table 3). individualized. Pertaining to neutrophil and platelet recovery, both adult Measuring Donor Chimerism and pediatric transplantation physician panels endorsed the Both panels recommended routine measurement of donor existing CIBMTR working definitions. Here, using the word chimerism using CD3 or similar for lymphoid cells and CD33 “recovery” instead of “engraftment” is more appropriate, or similar for myeloid cells. During the first year after allo- because confirmation of donor source ideally requires also HCT, both panels recommended routine measurement of proof of at least mixed/partial donor chimerism, which gener- donor chimerism on approximately days +30, +90, and +180 ally occurs later in the course of transplantation. Neither panel and at 1 year regardless of the intensity of the preparative reached a consensus in defining a specific time point for 646 M.A. Kharfan-Dabaja et al. / Transplantation and Cellular Therapy 27 (2021) 642649 Table 3 Consensus Definitions from the Adult and Pediatric Transplantation Physician Panels Term Definition Neutrophil recovery Both panels endorsed the existing definition of neutrophil recovery as the first of 3 successive days with an absolute neutrophil count of 500/mL after post-transplantation nadir. Platelet recovery Both panels endorsed the definition of platelet recovery as the first of 3 consecutive days with a platelet count of 20,000/ mL or higher in the absence of platelet transfusion for 7 consecutive days. Delayed engraftment No consensus was reached by either panel. Graft rejection versus graft failure Both panels defined graft rejection as an immune-mediated process, whereas graft failure represents a wider array of possibilities, including cell dosing, disease, infection, drugs, and an immune-mediated event. Graft failure (primary)* PBSCs: Both panels defined graft failure as lack of achievement of an ANC 500/mL by day +30 with associated (according to cell source) pancytopenia. Unstimulated BM: Both panels defined graft failure as lack of achievement of an ANC 500/mL by day +30 with associ- ated pancytopenia. G-CSF-stimulated BM: No consensus was reached by either panel. UCB: Both panels defined graft failure as lack of achievement of an ANC  500/mL by day +42 with associated pancytopenia. Poor graft function** Both panels defined poor graft function as frequent dependence on blood and/or platelet transfusions and/or growth factor support in the absence of other explanations, such as disease relapse, drugs, or infections. Secondary graft failure* Both panels defined secondary graft failure as a decline in hematopoietic function (may involve hemoglobin and/or pla- telets and/or neutrophils) necessitating blood products or growth factor support, after having met the standard defini- tion of hematopoietic (neutrophils and platelets) recovery. Donor chimerism Full: Both panels endorsed the existing definition of full donor chimerism as >95% for both myeloid and lymphoid lineages. Mixed or partial: Both panels endorsed the existing definition of mixed donor chimerism as 5% to 95% for both myeloid and lymphoid lineages. Absent: Both panels endorsed the existing definition of absent donor chimerism as <5% for both myeloid and lymphoid lineages. PBSC, peripheral blood stem cells; ANC, absolute neutrophil count; BM, bone marrow cells; G-CSF, granulocyte colony stimulating factor. * Donor chimerism testing is also done to confirm the suspicion of graft failure. ** Assumes that donor myeloid and lymphoid chimerism are within a desirable target level. Table 4 Recommendations of the Adult and Pediatric Transplantation Physician Panels Category Recommendations Assessment of secondary graft failure Evaluating hematopoietic function (based on peripheral blood counts), bone marrow cellularity, and donor chime- rism(s) when assessing secondary graft failure Measuring donor chimerism Routine practice includes measurement of donor chimerism using CD3 or similar for lymphoid cells and CD33 or similar for myeloid cells. During the first year after allo-HCT, measure donor chimerism routinely on days +30, +90, and +180 and at 1 year during the first year post- allo-HCT. Measure sorted (CD3 and/or CD33) donor chimerism routinely in patients receiving MAC, RIC, or NMA conditioning. Measure sorted (CD3 and/or CD33) donor chimerism routinely when using T cell-depleted grafts (in vivo or ex vivo). Managing mixed donor The panel recognizes the impact of the actual percentage of donor cells for clinical management. chimerism (malignant diseases) Consider using the actual percentages of donor myeloid cells and lymphoid cells, blood counts and the clinical sta- tus of the patient for a change in management. Use the actual percentage of mixed donor chimerism at specific time points post-allografting (ie, days +30, +90, +180, and +365) for a change in management. Consider the decline from a prior chimerism level to inform clinical management. Managing mixed donor chimerism The panel recognizes that the level of donor chimerism required for disease correction depends on the disease. (nonmalignant diseases/bone marrow Consider the decline from a prior chimerism level to inform clinical management. failure syndromes) delayed engraftment. This is not surprising, because in theory, administering a CD34-selected cell boost to address poor graft delayed engraftment represents a continuum from the time of function with or without mixed/absent chimerism in the myeloid absence of hematopoietic recovery until objective confirma- compartment and prescribing donor lymphocyte infusion(s) to tion of graft failure. Moreover, from a treatment standpoint, treat low mixed chimerism, but do require at least 5% donor lym- these patients generally continue to receive supportive inter- phoid chimerism. Other interventions to treat graft failure include ventions while cytopenic, such as antimicrobial prophylaxis, considering a second allo-HCT, treatment of infections, and with- blood products, and growth factors, among others. holding suspected myelotoxic drugs, among others, as clinically Guidelines differentiate between graft rejection and graft fail- indicated. ure because they have different therapeutic implications. For graft Different from graft failure, poor graft function assumes rejection, interventions concentrate primarily on overcoming the that donor myeloid and lymphoid chimerism are within a HLA disparity barrier. However, interventions for graft failure are desirable target level and that there are other causes that more varied based on specific situations and might include explain the continuous need for blood products or growth M.A. Kharfan-Dabaja et al. / Transplantation and Cellular Therapy 27 (2021) 642649 647 Options for down-trending Pediatric-Transplant Panel member number A Adult-Transplant Panel member number B CD3 donor chimerism 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 Withdraw immune suppression Increase immune suppression Donor Lymphocyte infusion CD34 cell boost Second allogeneic HCT Options for down-trending Pediatric-Transplant Panel member number C Adult-Transplant Panel member number D 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 CD33 donor chimerism Withdraw immune suppression Increase immune suppression Donor Lymphocyte infusion CD34 cell boost Second allogeneic HCT Figure 1. Transplantation physician responses to options for managing down-trending chimerism in malignant diseases. Management responses are shown for pedi- atric transplantation panel members to declines in CD3 (A) and CD33 donor chimerism (C). Similar responses are shown for adult transplant panel members in (B) and (D), respectively. Pediatric-Transplant Panel member number Adult-Transplant Panel member number Options for down-trending A B 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 CD3 donor chimerism Withdraw immune suppression Increase immune suppression Donor Lymphocyte infusion CD34 cell boost Second allogeneic HCT Pediatric-Transplant Panel member number Adult-Transplant Panel member number Options for down-trending 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 C 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 D CD33 donor chimerism Withdraw immune suppression Increase immune suppression Donor Lymphocyte infusion CD34 cell boost Second allogeneic HCT Figure 2. Transplantation physician responses to options for managing down-trending chimerism in nonmalignant diseases. Management responses are shown for pediatric transplantation panel members to declines in CD3 (A) and CD33 donor chimerism (C). Similar responses are shown for adult transplantation panel members in (B) and (D), respectively. factor support. For instance, diseases such as myelofibrosis practice limitations related to this definition. Intuitively, could cause poor graft function post-transplantation, requiring patients with lower levels of mixed donor chimerism are likely frequent transfusion of blood products during the first few to be treated differently than those with higher levels. More- months, even in the presence of full donor myeloid chimerism. over, mixed chimerism is a dynamic state in which a progres- For primary graft failure, both panels endorsed using a sive decline in donor chimerism might portend or confirm practical definition based on lack of recovery of ANC to 500/ graft loss or disease recurrence, depending on the cell com- mL in the presence of associated cytopenias at a particular time partment analyzed. For instance, a 10% level of CD3 donor chi- point based on the prescribed stem cell source (Table 3). We merism might prompt consideration for donor lymphocyte acknowledge that additional workup is certainly required, infusion or even prescription of antineoplastic therapy in the which should include assessing marrow cellularity and quanti- case of a T cell malignancy. On the other hand, an 80% donor fying donor chimerism levels to confirm graft failure (Table 3), CD3 level, which still meets the definition of mixed donor chi- and that future interventions could vary depending on several merism, is more likely to be followed closely until a clearer factors, such as patient functional status and disease-specific trend is established. Furthermore, management could be dif- reasons, among others. In cases of secondary graft failure, ferent if mixed donor chimerism is observed in only one line- patients have a decline in hematopoietic function and confir- age versus both lineages. We acknowledge that in the setting mation of absent donor chimerism levels after previously of T cell depletion, ex vivo or in vivo, CD3 chimerism might be documented full or mixed donor chimerism levels. low or absent for many months after transplantation. Notwithstanding the different sensitivities of donor chime- When managing declining donor chimerism levels, rism methodologies, both panels endorsed current working approaches tend to consider not only the specific lineage(s) definitions of full donor chimerism in myeloid and lymphoid involved, but also the percentage decline (trajectory) over lineages as >95%, mixed or partial donor chimerism as 5% to time from a known peak level. Repeating donor chimerism lev- 95%, and absence of donor chimerism as <5% (Table 3). els should be considered to confirm the results. When both Although both panels endorsed the definition of mixed donor panels were asked about the immediate next step in managing chimerism as 5% to 95%, they acknowledged several clinical declines in donor CD3 and CD33 chimerism in the setting of 648 M.A. Kharfan-Dabaja et al. / Transplantation and Cellular Therapy 27 (2021) 642649 allo-HCT for malignant diseases, the most frequent response for Cidara Therapeutics, Servier, and Medigene and on the sci- was to withdraw (or taper) immune suppression (Figure 1). In entific advisory boards of MolMed and NexImmune. He has addition, more than one-half of respondents chose more than received research support for clinical trials from Incyte, Kite/ one possible option, underscoring the complexities associated Gilead, and Miltenyi Biotec. He serves in a volunteer capacity with managing such clinical situations. Caveats must certainly as a member of the Board of Directors of Be The Match be applied, because in certain scenarios, such as the presence (National Marrow Donor Program), as well as on the CIBMTR of active GVHD, withdrawing (or tapering) immune suppres- Cellular Immunotherapy Data Resource Executive Committee. sion might be inappropriate. S.E.P. receives support for the conduct of sponsored trials from In the case of declining CD3 and CD33 donor chimerism in the Atara Biotherapeutics, Mesoblast, and Jasper and is an inventor setting of nonmalignant disorders, both panels also chose with- of IP licensed to Atara Biotherapeutics by Memorial Sloan Ket- drawing immune suppression as the next immediate step in man- tering Cancer Center (MSKCC) (assigned all rights to MSKCC aging a downward trend in CD3 and CD33 chimerism. Again, and has no financial interest in Atara Biotherapeutics). I.P. more than one-half of the participants chose more than one serves on advisory boards for Kadmon, Incyte, and Syndex. M. option. Such decisions need to consider several factors, including L.R. serves on an advisory board for BioIntellect. C.R. serves as but not limited to the patient's underlying disease indication for a consultant for Amgen, Bristol Myers Squibb, Takeda, and allo-HCT, whether the downtrend occurred in the context of mini- Sanofi. R.R. has received research funding from Crispr Thera- mal versus heavy pretransfusion, whether the decreases in chime- peutics and serves on a scientific advisory board for Glycostem. rism and blood counts were observed during steady-state versus A.S. serves as a consultant for Magenta, Incyte Pharmaceuti- tapered immune suppression, and the intensity of the preparative cals, and CareDx and receives research support from Amgen, regimen (MAC versus RIC). We acknowledge that optimal man- Kadmon, and OrcaBio. N.S. has received research funding from agement of declining donor chimerism, particularly in patients Celgene/Bristol Myers Squibb, Janssen, bluebird bio, Sutro Bio- with nonmalignant disorders, remains an area in which future pharma, Teneobio, Poseida, Nektar, and has served as an advi- research is definitely needed. sor for GSK, Amgen, Indapta Therapeutics, Sanofi, CareDx, Kite The main goal in developing these consensus definitions Pharma, Karyopharm, Oncopeptides, and CSL Behring. J.S. and recommendations is to harmonize allo-HCT clinical prac- reports personal fees from Astellas, Jazz Pharmaceuticals, Abb- tice. These guidelines are not intended to replace clinical judg- vie, Daiichi Sankyo, Pfizer, and grants and personal fees from ment, as there are several situations were decisions need to be Novartis. M.R.V. reports advisory board participation for Jazz individualized. Pharmaceuticals and Novartis and consultancy with Fate Ther- apeutics and Terumo. J.E.W. serves as a clinical advisor to Magenta Therapeutics and Rocket Pharmaceuticals. M.H. ACKNOWLEDGMENTS reports receiving research support/funding from Takeda Phar- Financial disclosure: Please refer to conflicts of interest state- maceutical, Spectrum Pharmaceuticals, and Astellas Pharma; ment which lists financial disclodures whenever applicable. serving as a consultant for Janssen, Incyte, ADC Therapeutics, Conflict of interest statement: M.A.K-D. reports consultancy Celgene, Omeros, Verastem, and MorphoSys; and serving on for Daiichi Sankyo and Pharmacyclics. L.M.B. has received sup- the speakers bureau for Sanofi Genzyme, AstraZeneca, and Bei- port for a clinical trial conducted through the Fred Hutchinson Gene. E.A., A. Kumar, M.A., T.N., R.M., A.S.A., Z.S.A., A.B., R.C., A. Cancer Research Center by Medac, including supply of the E-J., E.F., B. Hamilton, S.K.H., B. Horn, D.A.J., L.J., A.S.K., A. Kansa- study drug treosulfan; is a member of the data safety and mon- gra, A. Kassim, L.S.K., C.L.K., J.K-P., J.K., M.L.M., Z.M., M. Miel- itoring boards (DSMB) for a clinical trial with Rocket Pharma- carek, M. Mohty, A.N., E.N., T.S.O., M.A.P., G.R., P.J.S., S.S., J.T., P. ceuticals and another clinical trial with Jasper Therapeutics. N. V., B.N.S., and P.A.C. declare no conflicts of interest. M. has served as a consultant for Anthem and has received honoraria from Incyte and Nkarta, M.B. reports employment by Bristol Myers Squibb. J.J.B. reports consultancy for Race Oncology, Takeda, Avrobio, Bluerock, Omeros, and Advanced REFERENCES Clinical. S.C. serves on an advisory board for bluebird bio. Z.D. 1. Jenq RR, van den Brink MRM. Allogeneic haematopoietic stem cell trans- has received research support from Inctye and RegImmune- plantation: individualized stem cell and immune therapy of cancer. Nat Rev Cancer. 2010;10:213–221. Corp and has served as a consultant for Syndax Pharmaceuti- 2. Gooley TA, Chien JW, Pergam SA, et al. 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