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Hematopoietic stem cell transplantation from alternative sources in adults with high-risk acute leukemia
Blood Cells, Molecules, and Diseases 33 (2004) 294 – 302 www.elsevier.com/locate/ybcmd
Hematopoietic stem cell transplantation from alternative sources in adults $ with high-risk acute leukemia Franco Aversaa,*, Yair Reisnerb, Massimo F. Martellia a
Department of Hematology, University of Perugia, Perugia, Italy b Department of Immunology, Weizmann Institute, Israel Submitted 9 August 2004 Available online 2 October 2004 (Communicated by M. Lichtman, M.D., 12 August 2004)
Abstract Since 75% of patients with high-risk acute leukemia do not have a human leukocyte antigen (HLA)-identical sibling, alternative sources for hematopoietic stem cell transplantation (HSCT) are matched unrelated donors (MUD), unrelated umbilical cord blood (UD-UCB) and one HLA haplotype mismatched family members (haploidentical). The chance of finding a suitable donor in the international voluntary donor registries is limited by frequency of the HLA phenotype and the time required to identify the right donor from a potential panel, to establish eligibility and to harvest the cells. In adult MUD recipients, eventfree survival ranges up to 50% and refers only to patients who undergo transplant, without taking into account those who do not find a donor. Umbilical cord blood offers the advantages of easy procurement, the absence of risks to donors, the reduced risk of transmitting infections, immediate availability of cryopreserved samples and acceptance of mismatches at two of the six antigens. Although UD-UCB transplantation is a viable option for children, it is seldom considered for adults. The great divergency between body weight and the number of hematopoietic cells in a standard cord blood unit, particularly if associated with a two-antigen mismatch, increases the risk of graft failure and delays hematopoietic reconstitution. Work on full-haplotype mismatched transplants has been proceeding for over 20 years. Originally, outcome in leukemia patients was disappointing because of high incidence of severe graft-vs.-host disease in T-replete transplants and high rejection rates in T-cell-depleted transplants. The breakthrough came with the use of a megadose of T-cell-depleted progenitor cells after a high-intensity conditioning regimen. Treating end-stage patients inevitably confounded clinical outcome in the early pilot studies. Today, high-risk acute leukemia patients are treated at less advanced stages of disease, receive a reasonably well tolerated conditioning regimen, and benefit from advances in post-transplant immunological reconstitution. All these factors contribute to markedly reduce transplant-related mortality. Overall, event-free survival and transplant-related mortality compare favorably with reports from unrelated matched transplants. T-cell-depleted megadose stem cell transplant from a mismatched family member, who is immediately available, can be offered as a viable option to candidates with highrisk acute leukemias. D 2004 Elsevier Inc. All rights reserved. Keywords: Haploidentical transplant; High-risk acute leukemia; T cell depletion; Megadose of CD34+ cells
Introduction
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This paper is based on a presentation at a Focused Workshop on Haploidentical Stem Cell Transplantation sponsored by The Leukemia and Lymphoma Society held in Naples, Italy, from July 8 to 10, 2004. * Corresponding author. Istituto di Ematologia, Policlinico Monteluce, Via Brunamonti, 51, Perugia 06100, Italy. Fax: +39 75 578 3691. E-mail address: [email protected] (F. Aversa). 1079-9796/$ - see front matter D 2004 Elsevier Inc. All rights reserved. doi:10.1016/j.bcmd.2004.08.021
The data bases of unrelated donors, which now include more than eight million potential volunteers, have markedly increased the chance of finding a complete phenotypic match or a dreasonably toleratedT HLA mismatch. The odds depend on the ethnic group—ranging 60–70% for Caucasians to under 10% for ethnic minorities—and
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frequency of the HLA phenotype [9,10]. The chance of finding a suitable donor is limited even more by the extremely stringent age restrictions, because of the rise in morbidity and mortality with age. Furthermore, the enormous polymorphism of human HLA genes is reflected by new DNA-based techniques for HLA typing, which reveal an impressive number of new alleles within the antigens that were previously defined by serology. Matching by these methods reduces the risk of graft rejection and GvHD but the chance of finding a suitable donor is also reduced [9–13]. The National Marrow Donor Programme study showed that disparities for HLA-A, -B, -C or -DRB1 alleles were independent risk factors for acute GvHD [14]. HLA-A or -B allele mismatch was a significant risk factor for the occurrence of chronic GvHD [15]. A recent randomized study showed severe acute and extensive chronic GvHD were both significantly reduced when antithymocyte globulin was given, but transplant related mortality and overall survival remained unmodified [16]. In MUD transplants a 0.53 transplant-related mortality (TRM) in T-cell-replete transplants and 0.64–0.71 TRM in Tcell-depleted transplants have been reported [17]. Age and disease status are major factors in TRM in adults. In partially T-cell-depleted MUD transplants, Drobyski et al. [18] reported a 0.23 probability of TRM at 2 years, with patients over 20 years of age at significantly higher risk. In T-cellreplete MUD transplants, Cornelissen et al. [19] observed 0.61 TRM in 127 adults with ALL. After transplantation from an identical, or partially matched related, donor or MUD, only patient’s age and disease status markedly impacted on TRM (25% vs. 48% at a cutoff at 37 years old; 38% vs. 60% early vs. advanced disease) [20]. Reports on large series from many transplant centers indicate that transplantation from MUD is feasible but is considerably more difficult than the use of HLA-identical sibling donors. A large International Bone Marrow Transplantation Registry (IBMTR) study of 2055 allogeneic bone marrow transplants showed leukemia-free survival was better in HLA-identical sibling recipients than in MUD recipients, particularly when patients were transplanted in early stage disease [21]. Even for transplants performed
Fig. 1. The probability of having a matched-related stem cell donor is about 75%.
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between 1994 and 2000, IBMTR data continue to show increased risk associated with the use of MUD compared to HLA-identical sibling donors in patients otherwise comparable for age, diagnosis and stage disease at transplant [12]. The Seattle report [22] shows only 7% of the 81 patients in relapse, 19% of the 16 who never achieved remission, 28% for the 40 transplanted in second complete remission and 50% of the 16 in first complete remission survive leukemiafree at 5 years. Age also influences outcome with only 14% of the adult surviving leukemia-free as compared to 32% of patients under 18 years of age. Similar data are reported by the National Marrow Donor Program in 756 AML patients transplanted from matched unrelated donors [23,24]. Eventfree survival correlates with the age of patients (25% and 35% in adults and children, respectively) and stage of disease, with the difference being even more marked in patients in advanced stages of the disease at transplant (7% event-free survival for adults and 23% for children). So, in adult MUD recipients, event-free survival ranges up to 50% and refers only to patients who undergo transplant, without taking into account those who do not find a donor. Furthermore, the time required to identify the right donor from a potential panel, to establish eligibility and to harvest the cells may, in patients who urgently need a transplant, lead to relapse and failure of intention to treat. Consequently, for patients who do not have a matched donor or who urgently need a transplant, attention has focussed on unrelated cord blood and the haploidentical three loci mismatched family member.
Unrelated donor umbilical cord blood transplant Compared with matched unrelated hematopoietic stem cell transplant, umbilical cord blood (UCB) offers the advantages of easy procurement, the absence of risks to donors, the reduced risk of transmitting infections, immediate availability of cryopreserved samples and acceptance of mismatches at two of the six antigens. Several reports show unrelated UCB transplant is feasible for children with leukemia [25–30]. Disease status at transplant is a risk factor. Up to 50% of patients transplanted in first or second complete remission but only 8% of those transplanted in more advanced stages of disease survive disease-free. Gluckman and Rocha [30] have recently reported results of unrelated UCB transplantation in 196 children (median age 7 years) with ALL (35 CR I, 85 CR II and 75 in more advanced stage) transplanted between 1994 and 2001. The probability of neutrophil recovery at day 60 was 84%, the estimated probability of acute GvHD grade II–IV was 38%. Estimated 2-year event-free survival was 40%, 34% and 29% for patients transplanted in CR I, CR II or more advanced stage, respectively. A recent study [30] compared outcomes in 99 children with acute leukemia transplanted from UCB units with those of 442 children transplanted from unrelated matched bone marrow donors.
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UCB transplant recipients had delayed hematopoietic recovery, decreased incidence of acute GvHD and higher TRM at 100 days after transplant. Overall, UCB transplant was associated with an 80% probability of granulocyte engraftment at day 60, suggesting engraftment is a major concern particularly when the cord blood unit contains less than 3.5 107 CD34+ cells/Kg and the degree of HLA disparity increases. Risk factors were age over 15 and HLA disparity. Event-free survival at two years was 31% in the UCB group and 43% in the MUD-BMT. Although UCB transplantation is a viable option for children, it is seldom considered for adults [30–32]. The great divergency between body weight and the number of hematopoietic cells in a standard cord blood unit, particularly if associated with a two-antigen mismatch, increases the risk of graft failure and delays hematopoietic reconstitution. Gluckman and Rocha [30] analyzed 108 adults (median age 26 years; median weight 60 kg) receiving UCUCB transplants. Neutrophil recovery at day 60 was observed in 81% of patients, 15 did not engraft. TRM at day 100 was 54%, 1 year EFS was 21%. Favorable factors for survival were disease status at transplant and the number of nucleated cells in the unit (z1 107/kg recipient b.w.). These data indicate that in adult leukemia patients, outcomes after UC-UCB transplant might be improved by selecting units that contain a large number of nucleated cells and patients who are in early stage disease.
Haploidentical transplantation Many patients with acute leukemia at high risk of relapse who could benefit from an allogeneic stem cell transplant fail to find a matched donor, whether related or unrelated, and too many patients with high-risk acute leukemia who urgently need a transplant, still today relapse and die during the search for an unrelated graft. Nearly all of them have at least one haploidentical three loci mismatched family member who is promptly available as donor. Haploidentical transplantation offers an immediate source of hematopoietic stem cells to almost all patients and the few patients who reject the haploidentical transplant have the advantage of another, immediately available donor within the family circle. Until the 1990s, haploidentical transplantation for acute leukemia was usually unsuccessful because of the high incidence of severe GvHD in unmanipulated transplants [33] and graft rejection in extensively T-cell-depleted transplants [34]. It has been known since the early 1980s that effective T cell depletion can completely prevent both acute and chronic GVHD even when using haploidentical parental bone marrow differing at the three major HLA loci [35–37]. This remarkable potential of T cell depletion, based on numerous murine studies in the 1970s, was first demonstrated in the treatment of patients with severe combined immune deficiency (SCID) [38]. The problem of GvHD,
which is almost uniformly lethal in such haploidentical transplants, was completely prevented by 3 log T cell depletion using soybean lectin and E-rosetting. Since this initial study, more than 300 transplants from haploidentical donors have been carried out worldwide for SCID patients with a high rate of long-term partial or complete immune reconstitution [39]. Following the encouraging results in SCID patients, it was reasonable to assume that in leukemia patients who were pretreated by supralethal radiochemotherapy, residual immunity at the day of the transplant would be dramatically reduced and, therefore, graft rejection would not be a major problem. Unfortunately, results in patients with acute leukemia, who received a T-cell-depleted mismatched transplant after supralethal radiotherapy-based conditioning regimen, showed graft the incidence of rejection, which is a rare occurrence in unmanipulated transplants, rose dramatically [34,40]. Resistance to engraftment is mediated by recipient anti-donor CTLps, which survive the conditioning regimen [41]. Enhancing immunosuppression and myeloablation by adding different agents such as anti-T antibodies, cytosine arabinoside and thiotepa to the standard conditioning still did not ensure engraftment of T-cell-depleted mismatched bone marrow cells [42–46]. The turning point in the history of the T-cell-depleted haplotransplant came in 1993 [47] with the clinical application of an extensively T-cell-depleted megadose of stem cells, a concept which Yair Reisner successfully pioneered in animal models in the late 1980s [48]. bMegadoseQ stem cell transplants consisted initially of G-CSF mobilized peripheral blood stem cells and bone marrow cells, both ex vivo depleted of T cells by soybean agglutination and E-rosetting [47,49]. The conditioning protocol included TBI, cyclophosphamide, thiotepa and ATG. Eighty percent of the 36 high-risk adult leukemia patients we transplanted between 1993 and 1995 achieved primary sustained engraftment. Although no post-transplant immunosuppressive therapy was used as prophylaxis, the incidence of GVHD was significantly lower than in T-cell-replete mismatched transplants. Subsequently, changes were made in the transplant protocol (Table 1). In an attempt to reduce extrahematological toxicity, we set out to develop new conditioning regimens. After we had observed in a murine model that the immunosuppressive effect of TBI + cyclophosphamide, a potential factor in extra-hematological toxicity, could be achieved with TBI + fludarabine, a drug that was widely used in lymphoproliferative disorders, we substituted fludarabine for cyclophosphamide in our conditioning protocol in October 1995 (Fig. 2) [50]. Furthermore, the total lung dose radiation was decreased from 6 to 4 Gy. At the same time, to eliminate GvHD, the number of CD3+ cells in the graft was further reduced by changing the method of T cell depletion. Positive immunoselection of peripheral blood CD34+ cells was used instead of soybean agglutination and E rosetting. Initially, one-round E-rosetting was followed
F. Aversa et al. / Blood Cells, Molecules, and Diseases 33 (2004) 294–302 Table 1 Full-haplotype mismatched HSCT for high-risk acute leukaemia (March 1993–May 2004) Years Patients
1993–1995 (n = 36)
Conditioning Methods
G-CSF post
1995–1997 (n = 44)
1999–2004 (n = 104)
TBI, TT, CY, ATG TBI, TT, F, ATG TBI, TT, F, ATG SBA + E-rosette ( 1) + CD34 selection E-rosette ( 2) CD34 selection (CliniMACS) (Ceprate SC) Yes Yes No
Graft content (median) CD34+ (106/kg) 10.8 CD3+ (104/kg) 22.4
10.5 2.0
13.6 1.2
by positive immuno-selection of the CD34+ cells using the Ceprate-SC system [51]. Since January 1999, we have started selecting CD34+ cells in a one-step procedure using the Clinimacs device or, in some cases, in a two-step (positive/ negative selection-antiCD34+/antiCD2+) procedure using the Isolex instrument (Table 1). Automated peripheral blood CD34+ cell immunoselection for graft processing is time and labor saving and ensures a high CD34+ cell recovery rate. Besides providing 4.5 log T-cell depletion of the graft, it guarantees a 3.5 log B-cell depletion, which helps prevent EBV-related lymphoproliferative disorders [52]. The use of bone marrow as source of stem cells was abandoned in 1996 as sufficient numbers of CD34+ cells could be collected from G-CSF-mobilized peripheral blood. Furthermore, since 1999, post-transplant G-CSF administration has been suspended because experimental data suggested it induced immunosuppression in haploidentical transplant recipients [53,54]. In a large series of high-risk acute leukemia, patients these modifications to our original protocol ensured, sustained full donor type engraftment in over 95%, rapid hematopoietic recovery and a very low incidence of acute GvHD grade II–IV without the need for any post-transplant
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immune suppression as prophylaxis [51,55]. Handgretinger et al. [56] reported similar engraftment rates and no GVHD in children with acute leukemia after a chemotherapy-alone based conditioning regimen. In the Perugia and Tqbingen studies, GVHD was prevented by an ex vivo T-cell depletion alone. Another factor that may have contributed to T cell depletion in vivo, thereby reducing the incidence of GvHD, was the ATG in the conditioning regimen, which persisted in plasma for several days [57]. The same principle applies to the Tqbingen trial in which OKT3 was administered during the conditioning and after the transplant. Another reflection emerging from these two reports is that whether TBI is part of the conditioning or not does not seem to influence the engraftment rate when a stem cell megadose is given. Several principles emerge from these clinical results. First, the threshold dose of T lymphocytes (3 104/kg) for GvHD as established in haploidentical transplants in SCID patients, was seen to be valid for leukemia patients [39,58]. It should be noted that in the latter category of patients, this threshold dose is defined in the context of substantial levels of anti-T cell antibodies (ATG or OKT3) administered as part of the conditioning regimen. Secondly, a megadose of purified CD34+ cells is a crucial factor together with a highly immuno-myeloablative conditioning regimen, in overcoming the barrier of residual anti-donor cytotoxic T-lymphocyte precursors (CTL-p) in Tcell-depleted mismatched transplants [59–61]. A feasible hypothesis is that it reduces the frequencies of anti-donor CTL-ps in vivo. In vitro studies show that cells within the CD34+ cell population exhibit bvetoQ activity, i.e., in bulk mixed lymphocyte reactions they are able to neutralize specific CTL-ps directed against their antigens but not against a third party [62]. Furthermore, early myeloid CD33+ cells ( CD34+ and CD34- ), harvested 7–12 days after ex vivo expansion of CD34+ cells, are also endowed with marked veto activity, which is not found in late myeloid cells expressing CD14 or CD11b [63]. Therefore,
Fig. 2. Conditioning regimens during two time periods. Beginning in October 1995, fludarabine was substituted for cyclophosamide.
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soon after transplantation, infused CD34+ cells and their CD33+ progeny, which expand exponentially, could inhibit residual anti-donor CTL-ps in recipients. It is also worth noting that although the Perugia conditioning regimen was highly immunosuppressive and myeloablative, the extra-hematological toxicity was minimal. In particular, there was no case of veno-occlusive disease of the liver and no patient has died of lung toxicity since the total radiation dose to the lungs was reduced to 4 Gy. One concern in T-cell-depleted transplants is that the risk of acute leukemia relapse is higher because of the lack of GvHD-related GvL effect [35,64]. Allogeneic hematopoietic transplantation eradicates leukemia through the anti-leukemic effect of the conditioning regimen, and of the immune cells in the graft that recognize and eliminate the leukemia cells that survive the conditioning regimen. In T-replete transplants, this graft-vs.-leukemia (GvL) effect is mediated by T cell alloreactions directed against host MHC antigens, which have a broad tissue distribution. In the mismatched transplant. extensive T-cell depletion is essential because otherwise alloreactive donor T cells are associated with a high incidence of severe GvHD. The intensely myeloablative conditioning regimen could have compensated for the lack of T-cell-mediated GvL effect. In fact, a TBI- and thiotepa-based conditioning regimen and no post-transplant immunosuppression seems to be associated with a low incidence of leukemia relapse whether the T-cell-depleted transplant is matched sibling or mismatched relative. In the matched sibling setting, we observed a 0.12 and 0.28 probability of relapse, respectively, in patients with AML and ALL in CRI-II [36] and similar results were reported in AML patients by the SloanKettering group [37]. In our T-cell-depleted haploidentical transplants, we did not observe any increase in the probability of leukemia relapse in ALL and AML patients who were transplanted in first or later remission at high risk of relapse. As expected, this is significantly different to the relapse rate in patients transplanted in relapse. Disease status at transplant, in the haploidentical, like all other forms of transplant, is the major risk factor for leukemia relapse. Although relapse rates in matched and mismatched transplants overlap when patients are transplanted in first complete remission, it is worth noting that only patients at very high risk of relapse were inserted in the haploidentical transplant program, indicating that time should not be lost when patients with unfavorable prognostic features do not have a matched donor. Another finding which emerged from all our experience with haploidentical transplants is the role of donor vs. recipient natural killer cell alloreactivity, a biological phenomenon that is unique to the mismatched transplant, and that plays a role in the control of leukemia relapse in AML patients [65,66]. In many donor–recipient pairs, donor NK inhibitory receptors (KIRs) do not recognize as self the recipient’s Class I alleles. Consequently, the donor NK cells are not blocked and they are constantly activated to lyse the
recipient’s lymphohematopoietic cells. In these cases, the repertoire of the regenerating NK cells is characterized by significant numbers of alloreactive NK cells. This occurs in the absence of GvHD. The NK alloreactive clones lyse cryopreserved AML and CML cells but not most of adult ALL cells perhaps because the lymphoblasts lack LFA-1, an adhesion molecule that is essential in the formation of target-effector conjugates. In a retrospective analysis of patients transplanted in Perugia since the mismatched transplant program was started in 1993, multivariate analysis showed lack of an NK alloreactive donor was the strongest independent risk factor for AML relapse (hazard ratio 4.24; 95 percent CI 1.34–13.45; P = 0.014) after adjustment for disease status at transplant (relapse vs. remission: hazard ratio = 2.76; 95% CI 1.06–7.21; P = 0.038) [67]. Therefore the potential for donor vs. recipient NK cell alloreactivity, which can be predicted by standard HLA typing, is recommended when selecting the donor of choice from among the mismatched family members. Transplant-related mortality (TRM), a major problem in all transplants from alternative sources, mainly depends on slow immunological recovery that increases susceptibility to life-threatening infections. Several mechanisms are responsible for the post-transplant immune deficiency. In adults for up to a year after transplant, because of their decayed thymic function, early immune recovery stems from expansion of the mature T cells in the graft, and months later, from de novo production of naRve T cells [68–71]. Unfortunately, tissue damage by conditioning regimens prevents T cell homing to peripheral lymphoid tissues, where generation and maintenance of T cell memory take place. In unrelated donor and cord blood transplants GvHD prophylaxis, GvHD itself and its therapy antagonize T cell expansion and function and slow neutrophil count recovery after cord blood transplants is an additional factor. In haploidentical transplants, extensive T-cell depletion is crucial in preventing GvHD, so the T cell repertoire is limited and the ATG in the conditioning could delay T cell homeostatic expansion. Another aspect of the post-transplant immune deficiency, which has emerged from recent studies, is the impact of GCSF in transplant recipients. G-CSF is generally given to hasten neutrophil recovery. As we noticed that G-CSF blocks IL-12 production in antigen-presenting cells (APCs) and decreases pathogen-specific T cell responses in donor cells in vitro and in vivo, in January 1999, we decided to stop giving G-CSF to recipients [54]. The engraftment rate remained unchanged, and IL-12 production by APCs was restored to normal much sooner, with CD4+ cell numbers and function markedly improving. Most of the posttransplant CD4+ T cell clones exhibited protective Th1/Th0 cytokine production features, i.e., all clones expressed functional IL-12 receptors and few produced IL 4 and IL 10. These behavioral patterns were very different to what had been observed in recipients who had received G CSF, whose CD4+ clones had clearly exhibited nonprotective type 2 functional features.
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Finally, the majority of haploidentical transplant recipients were in advanced-stage disease, had been heavily pretreated and had undergone several courses of chemotherapy that left them vulnerable to infection. Many had already suffered life-threatening infections and their long history of failed remissions had led to bacterial and fungal colonization before transplant. Interestingly, no fatal infectious events occurred after the first year post-transplant, showing that with no significant chronic GvHD and no immune suppressive therapy, the risk of infection-related mortality disappears, unlike what is observed in matched unrelated transplants [71,72]. Any further reduction in TRM will only be achieved by hastening post-transplant immune recovery and several strategies are under investigation. One is the adoptive transfer of non-alloreactive T cells generated by purging IL-2 receptor (CD25) mixed lymphocyte response (MLR)reactive T cells [73]. Another involves the infusion of lymphocytes co-cultured with irradiated cells from the recipients in the presence of CTLA4-Ig—an agent that inhibits B7/CD28-mediated costimulation [74–76]. Preliminary studies report donor T lymphocytes specifically directed against pathogens (CMV, Aspergillus) are being infused at doses of up to 1 106/kg body weight safely across the HLA barrier [77]. Thymic output of naive T cells could be enhanced by administering keratinocyte growth factor (KGF) to protect thymic stroma from radiation injury [78] or IL7 to promote T cell differentiation [79]. In murine models, stromal cells are being engineered with IL7 to support T cell recovery and function [80]. In mismatched transplants, as in matched unrelated transplants, survival correlates with disease status at transplant. Encouragingly, at present, probability of long-term survival after haploidentical transplant overlaps with that of adults at the same stage of disease who received matched unrelated transplants [17–24,51,56,81]. One major factor
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that has undoubtedly confounded interpretation of the outcome in full-haplotype mismatched transplant is that most recipients had advanced stage disease at transplant. In the 65 patients with AML, who were transplanted in Perugia after March 1993, 47 were in relapse at transplant [82]. For these, the probability of survival is a poor 0.05 for ALL and 0.18 for AML. Nevertheless, for the AML patients who had no option other than a haploidentical transplant, the result is not altogether displeasing. Outcomes improve when patients were in second, third or even fourth CR at transplant being about 30% for both ALL and AML and confirming once again the survival advantage of transplant in remission. For patients who were in high risk CRI at transplant, EFS is, respectively, 28% and 53% for the 19 with ALL and the 22 with AML (Fig. 3). Similarly, in the Tubingen, series of 37 children with ALL who received a full-haplotype mismatched transplant disease status impacted on outcomes. The 21 patients in remission at transplant have a 0.39 probability of EFS as compared to the 0.13 in relapse [83].
Conclusions Much progress has been made over the past 10 years in the clinical, biological and technical aspects of the T-celldepleted full-haplotype mismatched transplants for acute leukemia. The high rejection rate and incidence of severe GvHD, which prevented the haploidentical transplant from being clinically feasible, have been overcome by combining high-intensity conditioning and a megadose of stem cells. The highly immunosuppressive and myeloablative conditioning regimen is well tolerated even by advanced stage, heavily pretreated patients. acute leukemia patients. To
Fig. 3. The event-free survival for 84 cases of ALL (left panel) and for 100 cases of AML after haploidentical stem cell transplantation. Three groups are shown for each type of leukaemia: patients transplanted in first remission (CR I), in a second or subsequent remission (CR z II), or in relapse.
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avoid GvHD, the threshold dose of T lymphocytes (V4 104/kg) as established in SCID patients, emerged as valid for leukemia patients receiving mismatched transplants. No post-transplant immunosuppression is required. When a stem cell megadose is given, the engraftment rate does not seem to differ whether TBI is part of the conditioning or not. The megadose of stem cells overcomes the barrier of the residual antitoxic CTL precursors through a veto activity exerted by CD34+ cells. Outcomes in haploidentical transplants depend more on disease status at transplant and on the patient’s history than on HLA-incompatibility. Ten years’ experience with haploidentical transplantation show that patients with high-risk ALL should be transplanted in early-stage disease in order to reduce the risk of relapse as much as possible and that for patients with AML, donor search criteria should include the family member who has the potential for donor-vs.-recipient NK cell alloreactivity, in an attempt to maximize the antileukemic effect of the haploidentical transplant. These results encourage extending the haploidentical transplant to patients with an indication to transplant. Haploidentical donors are found within the family for almost all patients with no undue delay between decisionmaking and transplantation, which is a crucial factor in urgent cases. Experimental studies that are currently under way all aim at bringing post-transplant infections under control. Once this aim is achieved, it is to be expected that transplant physicians will be less hesitant to treat patients in better condition and earlier disease stage and the mismatched transplant will be offered, not as a last resort, but as a routine option in the early stages of the disease to high-risk acute leukemia patients who need an allogeneic hematopoietic stem cell transplant.
[4]
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[15]
Acknowledgments The authors would like to thank all the laboratory and technical staff, the attending physicians and nurses of the Bone Marrow Transplant Unit at the University of Perugia, Italy, for their dedication and professional skills. Special thanks to Dr. Geraldine Anne Boyd for help in writing this paper.
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Hematopoietic stem cell transplantation from alternative sources in adults $ with high-risk acute leukemia Franco Aversaa,*, Yair Reisnerb, Massimo F. Martellia a
Department of Hematology, University of Perugia, Perugia, Italy b Department of Immunology, Weizmann Institute, Israel Submitted 9 August 2004 Available online 2 October 2004 (Communicated by M. Lichtman, M.D., 12 August 2004)
Abstract Since 75% of patients with high-risk acute leukemia do not have a human leukocyte antigen (HLA)-identical sibling, alternative sources for hematopoietic stem cell transplantation (HSCT) are matched unrelated donors (MUD), unrelated umbilical cord blood (UD-UCB) and one HLA haplotype mismatched family members (haploidentical). The chance of finding a suitable donor in the international voluntary donor registries is limited by frequency of the HLA phenotype and the time required to identify the right donor from a potential panel, to establish eligibility and to harvest the cells. In adult MUD recipients, eventfree survival ranges up to 50% and refers only to patients who undergo transplant, without taking into account those who do not find a donor. Umbilical cord blood offers the advantages of easy procurement, the absence of risks to donors, the reduced risk of transmitting infections, immediate availability of cryopreserved samples and acceptance of mismatches at two of the six antigens. Although UD-UCB transplantation is a viable option for children, it is seldom considered for adults. The great divergency between body weight and the number of hematopoietic cells in a standard cord blood unit, particularly if associated with a two-antigen mismatch, increases the risk of graft failure and delays hematopoietic reconstitution. Work on full-haplotype mismatched transplants has been proceeding for over 20 years. Originally, outcome in leukemia patients was disappointing because of high incidence of severe graft-vs.-host disease in T-replete transplants and high rejection rates in T-cell-depleted transplants. The breakthrough came with the use of a megadose of T-cell-depleted progenitor cells after a high-intensity conditioning regimen. Treating end-stage patients inevitably confounded clinical outcome in the early pilot studies. Today, high-risk acute leukemia patients are treated at less advanced stages of disease, receive a reasonably well tolerated conditioning regimen, and benefit from advances in post-transplant immunological reconstitution. All these factors contribute to markedly reduce transplant-related mortality. Overall, event-free survival and transplant-related mortality compare favorably with reports from unrelated matched transplants. T-cell-depleted megadose stem cell transplant from a mismatched family member, who is immediately available, can be offered as a viable option to candidates with highrisk acute leukemias. D 2004 Elsevier Inc. All rights reserved. Keywords: Haploidentical transplant; High-risk acute leukemia; T cell depletion; Megadose of CD34+ cells
Introduction
$
This paper is based on a presentation at a Focused Workshop on Haploidentical Stem Cell Transplantation sponsored by The Leukemia and Lymphoma Society held in Naples, Italy, from July 8 to 10, 2004. * Corresponding author. Istituto di Ematologia, Policlinico Monteluce, Via Brunamonti, 51, Perugia 06100, Italy. Fax: +39 75 578 3691. E-mail address: [email protected] (F. Aversa). 1079-9796/$ - see front matter D 2004 Elsevier Inc. All rights reserved. doi:10.1016/j.bcmd.2004.08.021
The data bases of unrelated donors, which now include more than eight million potential volunteers, have markedly increased the chance of finding a complete phenotypic match or a dreasonably toleratedT HLA mismatch. The odds depend on the ethnic group—ranging 60–70% for Caucasians to under 10% for ethnic minorities—and
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frequency of the HLA phenotype [9,10]. The chance of finding a suitable donor is limited even more by the extremely stringent age restrictions, because of the rise in morbidity and mortality with age. Furthermore, the enormous polymorphism of human HLA genes is reflected by new DNA-based techniques for HLA typing, which reveal an impressive number of new alleles within the antigens that were previously defined by serology. Matching by these methods reduces the risk of graft rejection and GvHD but the chance of finding a suitable donor is also reduced [9–13]. The National Marrow Donor Programme study showed that disparities for HLA-A, -B, -C or -DRB1 alleles were independent risk factors for acute GvHD [14]. HLA-A or -B allele mismatch was a significant risk factor for the occurrence of chronic GvHD [15]. A recent randomized study showed severe acute and extensive chronic GvHD were both significantly reduced when antithymocyte globulin was given, but transplant related mortality and overall survival remained unmodified [16]. In MUD transplants a 0.53 transplant-related mortality (TRM) in T-cell-replete transplants and 0.64–0.71 TRM in Tcell-depleted transplants have been reported [17]. Age and disease status are major factors in TRM in adults. In partially T-cell-depleted MUD transplants, Drobyski et al. [18] reported a 0.23 probability of TRM at 2 years, with patients over 20 years of age at significantly higher risk. In T-cellreplete MUD transplants, Cornelissen et al. [19] observed 0.61 TRM in 127 adults with ALL. After transplantation from an identical, or partially matched related, donor or MUD, only patient’s age and disease status markedly impacted on TRM (25% vs. 48% at a cutoff at 37 years old; 38% vs. 60% early vs. advanced disease) [20]. Reports on large series from many transplant centers indicate that transplantation from MUD is feasible but is considerably more difficult than the use of HLA-identical sibling donors. A large International Bone Marrow Transplantation Registry (IBMTR) study of 2055 allogeneic bone marrow transplants showed leukemia-free survival was better in HLA-identical sibling recipients than in MUD recipients, particularly when patients were transplanted in early stage disease [21]. Even for transplants performed
Fig. 1. The probability of having a matched-related stem cell donor is about 75%.
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between 1994 and 2000, IBMTR data continue to show increased risk associated with the use of MUD compared to HLA-identical sibling donors in patients otherwise comparable for age, diagnosis and stage disease at transplant [12]. The Seattle report [22] shows only 7% of the 81 patients in relapse, 19% of the 16 who never achieved remission, 28% for the 40 transplanted in second complete remission and 50% of the 16 in first complete remission survive leukemiafree at 5 years. Age also influences outcome with only 14% of the adult surviving leukemia-free as compared to 32% of patients under 18 years of age. Similar data are reported by the National Marrow Donor Program in 756 AML patients transplanted from matched unrelated donors [23,24]. Eventfree survival correlates with the age of patients (25% and 35% in adults and children, respectively) and stage of disease, with the difference being even more marked in patients in advanced stages of the disease at transplant (7% event-free survival for adults and 23% for children). So, in adult MUD recipients, event-free survival ranges up to 50% and refers only to patients who undergo transplant, without taking into account those who do not find a donor. Furthermore, the time required to identify the right donor from a potential panel, to establish eligibility and to harvest the cells may, in patients who urgently need a transplant, lead to relapse and failure of intention to treat. Consequently, for patients who do not have a matched donor or who urgently need a transplant, attention has focussed on unrelated cord blood and the haploidentical three loci mismatched family member.
Unrelated donor umbilical cord blood transplant Compared with matched unrelated hematopoietic stem cell transplant, umbilical cord blood (UCB) offers the advantages of easy procurement, the absence of risks to donors, the reduced risk of transmitting infections, immediate availability of cryopreserved samples and acceptance of mismatches at two of the six antigens. Several reports show unrelated UCB transplant is feasible for children with leukemia [25–30]. Disease status at transplant is a risk factor. Up to 50% of patients transplanted in first or second complete remission but only 8% of those transplanted in more advanced stages of disease survive disease-free. Gluckman and Rocha [30] have recently reported results of unrelated UCB transplantation in 196 children (median age 7 years) with ALL (35 CR I, 85 CR II and 75 in more advanced stage) transplanted between 1994 and 2001. The probability of neutrophil recovery at day 60 was 84%, the estimated probability of acute GvHD grade II–IV was 38%. Estimated 2-year event-free survival was 40%, 34% and 29% for patients transplanted in CR I, CR II or more advanced stage, respectively. A recent study [30] compared outcomes in 99 children with acute leukemia transplanted from UCB units with those of 442 children transplanted from unrelated matched bone marrow donors.
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UCB transplant recipients had delayed hematopoietic recovery, decreased incidence of acute GvHD and higher TRM at 100 days after transplant. Overall, UCB transplant was associated with an 80% probability of granulocyte engraftment at day 60, suggesting engraftment is a major concern particularly when the cord blood unit contains less than 3.5 107 CD34+ cells/Kg and the degree of HLA disparity increases. Risk factors were age over 15 and HLA disparity. Event-free survival at two years was 31% in the UCB group and 43% in the MUD-BMT. Although UCB transplantation is a viable option for children, it is seldom considered for adults [30–32]. The great divergency between body weight and the number of hematopoietic cells in a standard cord blood unit, particularly if associated with a two-antigen mismatch, increases the risk of graft failure and delays hematopoietic reconstitution. Gluckman and Rocha [30] analyzed 108 adults (median age 26 years; median weight 60 kg) receiving UCUCB transplants. Neutrophil recovery at day 60 was observed in 81% of patients, 15 did not engraft. TRM at day 100 was 54%, 1 year EFS was 21%. Favorable factors for survival were disease status at transplant and the number of nucleated cells in the unit (z1 107/kg recipient b.w.). These data indicate that in adult leukemia patients, outcomes after UC-UCB transplant might be improved by selecting units that contain a large number of nucleated cells and patients who are in early stage disease.
Haploidentical transplantation Many patients with acute leukemia at high risk of relapse who could benefit from an allogeneic stem cell transplant fail to find a matched donor, whether related or unrelated, and too many patients with high-risk acute leukemia who urgently need a transplant, still today relapse and die during the search for an unrelated graft. Nearly all of them have at least one haploidentical three loci mismatched family member who is promptly available as donor. Haploidentical transplantation offers an immediate source of hematopoietic stem cells to almost all patients and the few patients who reject the haploidentical transplant have the advantage of another, immediately available donor within the family circle. Until the 1990s, haploidentical transplantation for acute leukemia was usually unsuccessful because of the high incidence of severe GvHD in unmanipulated transplants [33] and graft rejection in extensively T-cell-depleted transplants [34]. It has been known since the early 1980s that effective T cell depletion can completely prevent both acute and chronic GVHD even when using haploidentical parental bone marrow differing at the three major HLA loci [35–37]. This remarkable potential of T cell depletion, based on numerous murine studies in the 1970s, was first demonstrated in the treatment of patients with severe combined immune deficiency (SCID) [38]. The problem of GvHD,
which is almost uniformly lethal in such haploidentical transplants, was completely prevented by 3 log T cell depletion using soybean lectin and E-rosetting. Since this initial study, more than 300 transplants from haploidentical donors have been carried out worldwide for SCID patients with a high rate of long-term partial or complete immune reconstitution [39]. Following the encouraging results in SCID patients, it was reasonable to assume that in leukemia patients who were pretreated by supralethal radiochemotherapy, residual immunity at the day of the transplant would be dramatically reduced and, therefore, graft rejection would not be a major problem. Unfortunately, results in patients with acute leukemia, who received a T-cell-depleted mismatched transplant after supralethal radiotherapy-based conditioning regimen, showed graft the incidence of rejection, which is a rare occurrence in unmanipulated transplants, rose dramatically [34,40]. Resistance to engraftment is mediated by recipient anti-donor CTLps, which survive the conditioning regimen [41]. Enhancing immunosuppression and myeloablation by adding different agents such as anti-T antibodies, cytosine arabinoside and thiotepa to the standard conditioning still did not ensure engraftment of T-cell-depleted mismatched bone marrow cells [42–46]. The turning point in the history of the T-cell-depleted haplotransplant came in 1993 [47] with the clinical application of an extensively T-cell-depleted megadose of stem cells, a concept which Yair Reisner successfully pioneered in animal models in the late 1980s [48]. bMegadoseQ stem cell transplants consisted initially of G-CSF mobilized peripheral blood stem cells and bone marrow cells, both ex vivo depleted of T cells by soybean agglutination and E-rosetting [47,49]. The conditioning protocol included TBI, cyclophosphamide, thiotepa and ATG. Eighty percent of the 36 high-risk adult leukemia patients we transplanted between 1993 and 1995 achieved primary sustained engraftment. Although no post-transplant immunosuppressive therapy was used as prophylaxis, the incidence of GVHD was significantly lower than in T-cell-replete mismatched transplants. Subsequently, changes were made in the transplant protocol (Table 1). In an attempt to reduce extrahematological toxicity, we set out to develop new conditioning regimens. After we had observed in a murine model that the immunosuppressive effect of TBI + cyclophosphamide, a potential factor in extra-hematological toxicity, could be achieved with TBI + fludarabine, a drug that was widely used in lymphoproliferative disorders, we substituted fludarabine for cyclophosphamide in our conditioning protocol in October 1995 (Fig. 2) [50]. Furthermore, the total lung dose radiation was decreased from 6 to 4 Gy. At the same time, to eliminate GvHD, the number of CD3+ cells in the graft was further reduced by changing the method of T cell depletion. Positive immunoselection of peripheral blood CD34+ cells was used instead of soybean agglutination and E rosetting. Initially, one-round E-rosetting was followed
F. Aversa et al. / Blood Cells, Molecules, and Diseases 33 (2004) 294–302 Table 1 Full-haplotype mismatched HSCT for high-risk acute leukaemia (March 1993–May 2004) Years Patients
1993–1995 (n = 36)
Conditioning Methods
G-CSF post
1995–1997 (n = 44)
1999–2004 (n = 104)
TBI, TT, CY, ATG TBI, TT, F, ATG TBI, TT, F, ATG SBA + E-rosette ( 1) + CD34 selection E-rosette ( 2) CD34 selection (CliniMACS) (Ceprate SC) Yes Yes No
Graft content (median) CD34+ (106/kg) 10.8 CD3+ (104/kg) 22.4
10.5 2.0
13.6 1.2
by positive immuno-selection of the CD34+ cells using the Ceprate-SC system [51]. Since January 1999, we have started selecting CD34+ cells in a one-step procedure using the Clinimacs device or, in some cases, in a two-step (positive/ negative selection-antiCD34+/antiCD2+) procedure using the Isolex instrument (Table 1). Automated peripheral blood CD34+ cell immunoselection for graft processing is time and labor saving and ensures a high CD34+ cell recovery rate. Besides providing 4.5 log T-cell depletion of the graft, it guarantees a 3.5 log B-cell depletion, which helps prevent EBV-related lymphoproliferative disorders [52]. The use of bone marrow as source of stem cells was abandoned in 1996 as sufficient numbers of CD34+ cells could be collected from G-CSF-mobilized peripheral blood. Furthermore, since 1999, post-transplant G-CSF administration has been suspended because experimental data suggested it induced immunosuppression in haploidentical transplant recipients [53,54]. In a large series of high-risk acute leukemia, patients these modifications to our original protocol ensured, sustained full donor type engraftment in over 95%, rapid hematopoietic recovery and a very low incidence of acute GvHD grade II–IV without the need for any post-transplant
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immune suppression as prophylaxis [51,55]. Handgretinger et al. [56] reported similar engraftment rates and no GVHD in children with acute leukemia after a chemotherapy-alone based conditioning regimen. In the Perugia and Tqbingen studies, GVHD was prevented by an ex vivo T-cell depletion alone. Another factor that may have contributed to T cell depletion in vivo, thereby reducing the incidence of GvHD, was the ATG in the conditioning regimen, which persisted in plasma for several days [57]. The same principle applies to the Tqbingen trial in which OKT3 was administered during the conditioning and after the transplant. Another reflection emerging from these two reports is that whether TBI is part of the conditioning or not does not seem to influence the engraftment rate when a stem cell megadose is given. Several principles emerge from these clinical results. First, the threshold dose of T lymphocytes (3 104/kg) for GvHD as established in haploidentical transplants in SCID patients, was seen to be valid for leukemia patients [39,58]. It should be noted that in the latter category of patients, this threshold dose is defined in the context of substantial levels of anti-T cell antibodies (ATG or OKT3) administered as part of the conditioning regimen. Secondly, a megadose of purified CD34+ cells is a crucial factor together with a highly immuno-myeloablative conditioning regimen, in overcoming the barrier of residual anti-donor cytotoxic T-lymphocyte precursors (CTL-p) in Tcell-depleted mismatched transplants [59–61]. A feasible hypothesis is that it reduces the frequencies of anti-donor CTL-ps in vivo. In vitro studies show that cells within the CD34+ cell population exhibit bvetoQ activity, i.e., in bulk mixed lymphocyte reactions they are able to neutralize specific CTL-ps directed against their antigens but not against a third party [62]. Furthermore, early myeloid CD33+ cells ( CD34+ and CD34- ), harvested 7–12 days after ex vivo expansion of CD34+ cells, are also endowed with marked veto activity, which is not found in late myeloid cells expressing CD14 or CD11b [63]. Therefore,
Fig. 2. Conditioning regimens during two time periods. Beginning in October 1995, fludarabine was substituted for cyclophosamide.
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soon after transplantation, infused CD34+ cells and their CD33+ progeny, which expand exponentially, could inhibit residual anti-donor CTL-ps in recipients. It is also worth noting that although the Perugia conditioning regimen was highly immunosuppressive and myeloablative, the extra-hematological toxicity was minimal. In particular, there was no case of veno-occlusive disease of the liver and no patient has died of lung toxicity since the total radiation dose to the lungs was reduced to 4 Gy. One concern in T-cell-depleted transplants is that the risk of acute leukemia relapse is higher because of the lack of GvHD-related GvL effect [35,64]. Allogeneic hematopoietic transplantation eradicates leukemia through the anti-leukemic effect of the conditioning regimen, and of the immune cells in the graft that recognize and eliminate the leukemia cells that survive the conditioning regimen. In T-replete transplants, this graft-vs.-leukemia (GvL) effect is mediated by T cell alloreactions directed against host MHC antigens, which have a broad tissue distribution. In the mismatched transplant. extensive T-cell depletion is essential because otherwise alloreactive donor T cells are associated with a high incidence of severe GvHD. The intensely myeloablative conditioning regimen could have compensated for the lack of T-cell-mediated GvL effect. In fact, a TBI- and thiotepa-based conditioning regimen and no post-transplant immunosuppression seems to be associated with a low incidence of leukemia relapse whether the T-cell-depleted transplant is matched sibling or mismatched relative. In the matched sibling setting, we observed a 0.12 and 0.28 probability of relapse, respectively, in patients with AML and ALL in CRI-II [36] and similar results were reported in AML patients by the SloanKettering group [37]. In our T-cell-depleted haploidentical transplants, we did not observe any increase in the probability of leukemia relapse in ALL and AML patients who were transplanted in first or later remission at high risk of relapse. As expected, this is significantly different to the relapse rate in patients transplanted in relapse. Disease status at transplant, in the haploidentical, like all other forms of transplant, is the major risk factor for leukemia relapse. Although relapse rates in matched and mismatched transplants overlap when patients are transplanted in first complete remission, it is worth noting that only patients at very high risk of relapse were inserted in the haploidentical transplant program, indicating that time should not be lost when patients with unfavorable prognostic features do not have a matched donor. Another finding which emerged from all our experience with haploidentical transplants is the role of donor vs. recipient natural killer cell alloreactivity, a biological phenomenon that is unique to the mismatched transplant, and that plays a role in the control of leukemia relapse in AML patients [65,66]. In many donor–recipient pairs, donor NK inhibitory receptors (KIRs) do not recognize as self the recipient’s Class I alleles. Consequently, the donor NK cells are not blocked and they are constantly activated to lyse the
recipient’s lymphohematopoietic cells. In these cases, the repertoire of the regenerating NK cells is characterized by significant numbers of alloreactive NK cells. This occurs in the absence of GvHD. The NK alloreactive clones lyse cryopreserved AML and CML cells but not most of adult ALL cells perhaps because the lymphoblasts lack LFA-1, an adhesion molecule that is essential in the formation of target-effector conjugates. In a retrospective analysis of patients transplanted in Perugia since the mismatched transplant program was started in 1993, multivariate analysis showed lack of an NK alloreactive donor was the strongest independent risk factor for AML relapse (hazard ratio 4.24; 95 percent CI 1.34–13.45; P = 0.014) after adjustment for disease status at transplant (relapse vs. remission: hazard ratio = 2.76; 95% CI 1.06–7.21; P = 0.038) [67]. Therefore the potential for donor vs. recipient NK cell alloreactivity, which can be predicted by standard HLA typing, is recommended when selecting the donor of choice from among the mismatched family members. Transplant-related mortality (TRM), a major problem in all transplants from alternative sources, mainly depends on slow immunological recovery that increases susceptibility to life-threatening infections. Several mechanisms are responsible for the post-transplant immune deficiency. In adults for up to a year after transplant, because of their decayed thymic function, early immune recovery stems from expansion of the mature T cells in the graft, and months later, from de novo production of naRve T cells [68–71]. Unfortunately, tissue damage by conditioning regimens prevents T cell homing to peripheral lymphoid tissues, where generation and maintenance of T cell memory take place. In unrelated donor and cord blood transplants GvHD prophylaxis, GvHD itself and its therapy antagonize T cell expansion and function and slow neutrophil count recovery after cord blood transplants is an additional factor. In haploidentical transplants, extensive T-cell depletion is crucial in preventing GvHD, so the T cell repertoire is limited and the ATG in the conditioning could delay T cell homeostatic expansion. Another aspect of the post-transplant immune deficiency, which has emerged from recent studies, is the impact of GCSF in transplant recipients. G-CSF is generally given to hasten neutrophil recovery. As we noticed that G-CSF blocks IL-12 production in antigen-presenting cells (APCs) and decreases pathogen-specific T cell responses in donor cells in vitro and in vivo, in January 1999, we decided to stop giving G-CSF to recipients [54]. The engraftment rate remained unchanged, and IL-12 production by APCs was restored to normal much sooner, with CD4+ cell numbers and function markedly improving. Most of the posttransplant CD4+ T cell clones exhibited protective Th1/Th0 cytokine production features, i.e., all clones expressed functional IL-12 receptors and few produced IL 4 and IL 10. These behavioral patterns were very different to what had been observed in recipients who had received G CSF, whose CD4+ clones had clearly exhibited nonprotective type 2 functional features.
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Finally, the majority of haploidentical transplant recipients were in advanced-stage disease, had been heavily pretreated and had undergone several courses of chemotherapy that left them vulnerable to infection. Many had already suffered life-threatening infections and their long history of failed remissions had led to bacterial and fungal colonization before transplant. Interestingly, no fatal infectious events occurred after the first year post-transplant, showing that with no significant chronic GvHD and no immune suppressive therapy, the risk of infection-related mortality disappears, unlike what is observed in matched unrelated transplants [71,72]. Any further reduction in TRM will only be achieved by hastening post-transplant immune recovery and several strategies are under investigation. One is the adoptive transfer of non-alloreactive T cells generated by purging IL-2 receptor (CD25) mixed lymphocyte response (MLR)reactive T cells [73]. Another involves the infusion of lymphocytes co-cultured with irradiated cells from the recipients in the presence of CTLA4-Ig—an agent that inhibits B7/CD28-mediated costimulation [74–76]. Preliminary studies report donor T lymphocytes specifically directed against pathogens (CMV, Aspergillus) are being infused at doses of up to 1 106/kg body weight safely across the HLA barrier [77]. Thymic output of naive T cells could be enhanced by administering keratinocyte growth factor (KGF) to protect thymic stroma from radiation injury [78] or IL7 to promote T cell differentiation [79]. In murine models, stromal cells are being engineered with IL7 to support T cell recovery and function [80]. In mismatched transplants, as in matched unrelated transplants, survival correlates with disease status at transplant. Encouragingly, at present, probability of long-term survival after haploidentical transplant overlaps with that of adults at the same stage of disease who received matched unrelated transplants [17–24,51,56,81]. One major factor
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that has undoubtedly confounded interpretation of the outcome in full-haplotype mismatched transplant is that most recipients had advanced stage disease at transplant. In the 65 patients with AML, who were transplanted in Perugia after March 1993, 47 were in relapse at transplant [82]. For these, the probability of survival is a poor 0.05 for ALL and 0.18 for AML. Nevertheless, for the AML patients who had no option other than a haploidentical transplant, the result is not altogether displeasing. Outcomes improve when patients were in second, third or even fourth CR at transplant being about 30% for both ALL and AML and confirming once again the survival advantage of transplant in remission. For patients who were in high risk CRI at transplant, EFS is, respectively, 28% and 53% for the 19 with ALL and the 22 with AML (Fig. 3). Similarly, in the Tubingen, series of 37 children with ALL who received a full-haplotype mismatched transplant disease status impacted on outcomes. The 21 patients in remission at transplant have a 0.39 probability of EFS as compared to the 0.13 in relapse [83].
Conclusions Much progress has been made over the past 10 years in the clinical, biological and technical aspects of the T-celldepleted full-haplotype mismatched transplants for acute leukemia. The high rejection rate and incidence of severe GvHD, which prevented the haploidentical transplant from being clinically feasible, have been overcome by combining high-intensity conditioning and a megadose of stem cells. The highly immunosuppressive and myeloablative conditioning regimen is well tolerated even by advanced stage, heavily pretreated patients. acute leukemia patients. To
Fig. 3. The event-free survival for 84 cases of ALL (left panel) and for 100 cases of AML after haploidentical stem cell transplantation. Three groups are shown for each type of leukaemia: patients transplanted in first remission (CR I), in a second or subsequent remission (CR z II), or in relapse.
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avoid GvHD, the threshold dose of T lymphocytes (V4 104/kg) as established in SCID patients, emerged as valid for leukemia patients receiving mismatched transplants. No post-transplant immunosuppression is required. When a stem cell megadose is given, the engraftment rate does not seem to differ whether TBI is part of the conditioning or not. The megadose of stem cells overcomes the barrier of the residual antitoxic CTL precursors through a veto activity exerted by CD34+ cells. Outcomes in haploidentical transplants depend more on disease status at transplant and on the patient’s history than on HLA-incompatibility. Ten years’ experience with haploidentical transplantation show that patients with high-risk ALL should be transplanted in early-stage disease in order to reduce the risk of relapse as much as possible and that for patients with AML, donor search criteria should include the family member who has the potential for donor-vs.-recipient NK cell alloreactivity, in an attempt to maximize the antileukemic effect of the haploidentical transplant. These results encourage extending the haploidentical transplant to patients with an indication to transplant. Haploidentical donors are found within the family for almost all patients with no undue delay between decisionmaking and transplantation, which is a crucial factor in urgent cases. Experimental studies that are currently under way all aim at bringing post-transplant infections under control. Once this aim is achieved, it is to be expected that transplant physicians will be less hesitant to treat patients in better condition and earlier disease stage and the mismatched transplant will be offered, not as a last resort, but as a routine option in the early stages of the disease to high-risk acute leukemia patients who need an allogeneic hematopoietic stem cell transplant.
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Acknowledgments The authors would like to thank all the laboratory and technical staff, the attending physicians and nurses of the Bone Marrow Transplant Unit at the University of Perugia, Italy, for their dedication and professional skills. Special thanks to Dr. Geraldine Anne Boyd for help in writing this paper.
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