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Keywords:

  • haploidentical;
  • stem cell transplantation;
  • paediatrics

Abstract

  1. Top of page
  2. Abstract
  3. Introduction
  4. Graft rejection
  5. Graft versus host disease
  6. Graft versus leukaemia
  7. Immune reconstitution
  8. Clinical paediatric perspective
  9. References

The broader application of stem cell transplantation (SCT) for paediatric diseases has been limited by a lack of human leucocyte antigen (HLA)-matched donors. Virtually all children, however have at least one haploidentical parent who could serve as a donor. Such a donor is immediately available and the considerable costs of additional HLA typing, registry and banking expenditures that are necessary to procure an unrelated donor, could be reduced. Recent technological advances appear to have overcome the historical problems of graft rejection and severe graft versus host disease in the haploidentical setting, and in the latest studies the overall survival for children undergoing haploidentical SCT for leukaemia is now comparable with that following unrelated donor bone marrow or cord blood transplantation. Post-transplant infectious complications and leukaemia relapse remain the most important barriers yet to overcome, and new directions in the use of adoptive cellular immunity appear to be promising in this respect. Haploidentical SCT is now a viable option for those children who do not have an HLA compatible sibling or fully matched unrelated donor. The relative merits of a haploidentical family donor versus mismatched unrelated bone marrow or cord blood donation needs to be assessed in prospective, randomized clinical trials.


Introduction

  1. Top of page
  2. Abstract
  3. Introduction
  4. Graft rejection
  5. Graft versus host disease
  6. Graft versus leukaemia
  7. Immune reconstitution
  8. Clinical paediatric perspective
  9. References

Although stem cell transplantation (SCT) has been successfully used to treat a wide range of paediatric diseases over the past three decades, broader application of SCT has been limited by a lack of human leucocyte antigen (HLA)-matched donors (Horowitz, 1999). As a HLA genotypically identical sibling donor is only available in 20–30% of cases, attention has switched during the last decade to alternative donors, i.e. closely matched unrelated donors from panels of adult volunteers (Hansen et al, 1997; Davies et al, 2000), cord blood banks (Kurtzberg et al, 1996; Wagner et al, 1996; Gluckman et al, 1997), and partially mismatched related donors (Aversa et al, 1994; Henslee-Downey et al, 1997).

The chance of finding a phenotypically matched unrelated donor from the Worldwide Registry varies with the patient's race, ranging from 60–70% for caucasians to under 10% for ethnic minorities (Beatty et al, 1995). The probability increases, if single antigen mismatched bone marrow donors (Casper et al, 1995; Oakhill et al, 1996) or up to two antigen mismatched cord blood donations (Rubinstein et al, 1998) are acceptable as permissible mismatches. A major drawback for a child, who urgently needs a SCT procedure, is the time interval from initiating a search to procuring a donor from the volunteer panels; this may take 2–4 months for a bone marrow donor, although this may be reduced to only a few weeks for a cord blood donation (Barker et al, 2002). Additionally, the cost incurred in procuring such a donor may be substantial. However, virtually all children have at least one haploidentical parent who could serve as a donor. Such a donor is immediately available and the availability is not dependent on racial or ethnic group and the considerable costs of additional HLA typing, registry and banking expenditures can be avoided.

Haploidentical transplants have been attempted sporadically over the last two decades, however, initial efforts were largely unsuccessful and the associated mortality was prohibitively high (Rowe & Lazarus, 2001). The major problems were those of intractable graft versus host disease (GVHD) (Beatty et al, 1985; Anasetti et al, 1989), and graft rejection (Soiffer et al, 1991). Experience in children with severe combined immunodeficiency (SCID) had clearly demonstrated that GVHD could be prevented by extensive T-cell depletion of the bone marrow graft with no need for post-transplant immunosuppression (Reisner et al, 1983), but unfortunately, until the early 1990s, the benefit of preventing GVHD was largely offset in patients with leukaemia by the high incidence of graft rejection (Kernan et al, 1987).

Several different approaches have been adopted in an attempt to overcome the inherent difficulties of haploidentical transplantation. In South Carolina, Henslee-Downey et al (1997) exploited sequential immunomodulation pre- and post-SCT using ex vivo T-cell depletion with the T10B9 monoclonal antibody and in vivo T-cell lysis with the immunotoxin H65-RTA, after an intensive conditioning regimen. Amongst 72 paediatric patients, they reported an 88% probability of sustained engraftment and a 16% and 51% probability of acute and chronic GVHD respectively. The Perugia group (Aversa et al, 1994) employed stem cell dose escalation to overcome rejection of heavily T-cell depleted mismatched bone marrow, by supplementing the graft with T-cell depleted granulocyte colony stimulating factor (G-CSF) mobilized peripheral blood progenitor cells (PBPCs). In so doing they achieved a ‘megadose’ SCT, increasing the number of CD34+ cells 10-fold. Amongst 43 mostly adult patients, 41 of 43 (95%) achieved primary sustained engraftment and only one of 41 developed acute GVHD grades II–IV despite no postgrafting immunosuppression.

A factor that has confounded the interpretation of outcome data in the haploidentical setting is the inherent advanced disease status amongst recipients of such a procedure (Aversa et al, 1998); however, as a result of the pioneering studies mentioned above over the past few years, data is emerging to suggest that haploidentical transplant procedures may have a role in patients other than those in desperate straits. Although the problems of graft rejection and GVHD may have largely been overcome, further major obstacles remain, primarily the slow immune reconstitution post haploidentical SCT which puts the patients at risk of infection for a significantly longer period of time compared with matched donor SCT (Ochs et al, 1995; Kook et al, 1996) and high relapse rates. The current strategies that now enable haploidentical transplants to be conducted more safely, the recent outcome of such transplants in the paediatric setting, and the ongoing developments which are attempting to address some of the remaining problems, will now be reviewed.

Graft rejection

  1. Top of page
  2. Abstract
  3. Introduction
  4. Graft rejection
  5. Graft versus host disease
  6. Graft versus leukaemia
  7. Immune reconstitution
  8. Clinical paediatric perspective
  9. References

Engraftment failure can be due to an immune rejection mediated by anti-donor cytotoxic T-lymphocyte precursors (CTL-p) in the host (Reisner et al, 1986) or to competition for ‘niches’ from residual host stem cells (Lapidot et al, 1989), either of which having survived the conditioning. In unmanipulated grafts, the presence of donor T cells usually suppresses residual host immunity, facilitating engraftment, but such suppression is abolished by profound T-cell depletion of the graft in the haploidentical setting. It was originally believed that this barrier to engraftment could be overcome by a global increase in the intensity of the conditioning regimen (Bozdech et al, 1985; Champlin et al, 1987), however, such intensive regimens led to unacceptable toxicity, particularly in heavily pretreated patients with advanced disease. On the other hand, more targeted approaches have led to an improved outcome: a number of groups tried to enhance myeloablation by the addition of agents specifically targeting stem cells, such as melphalan (Lapidot et al, 1989), busulphan or thiotepa (Terenzi et al, 1990), in combination with total body irradiation (TBI); others have utilized more subtle refinements of immunosuppression to facilitate engraftment without increasing toxicity, including the use of high-dose methylprednisolone (Kernan et al, 1989), total nodal irradiation (Soiffer et al, 1991), anti-T-cell antibodies (Fischer et al, 1986; Henslee-Downey et al, 1996), antithymocyte globulin (ATG) (Malilay et al, 1989; Aversa et al, 1998), and fludarabine, a purine analogue which inhibits adenosine deaminase (Aversa et al, 1998). Indeed, as in the HLA-matched setting with highly immunosuppressive conditioning regimens, it may become possible to obviate the need for myeloablation, particularly in patients with marrow aplasia. Investigators at Baylor College (Texas) have recently achieved durable engraftment from a haploidentical donor in a child with Fanconi anaemia using a regimen consisting of fludarabine, Campath 1H and two anti-CD45 antibodies (M. K. Brenner, personal communication). Similarly, based on their animal studies (Luznik et al, 2003), the Johns Hopkins group have reported sustained engraftment in eight of 10 adult patients with haematological malignancies receiving apartially HLA-mismatched bone marrow transplant using a fludarabine 150 mg/m2/cyclophosphamide 80 mg/kg/TBI 2 Gy regimen with tacrolimus/mycophenolate mofetil (FK506/MMF) post-transplant immunosuppression (O'Donnell et al, 2002). However, it should be emphasized that data on such approach are extremely limited and patients should only be transplanted on such protocols in the context of well-designed clinical trials.

Donor natural killer (NK)-cell alloreactivity may also play a role in engraftment (Ruggeri et al, 1999). This is discussed more fully below, but essentially occurs when HLA-mismatched recipient target cells do not express the specific class I alleles, which block inhibitory receptors (killer immunoglobulin-like receptors, KIRs) on donor NK cells. In the Perugia experience, seven of 40 patients with no KIR incompatibility in the GVH direction rejected, whereas none of 20 KIR-mismatched patients rejected (Ruggeri et al, 1999).

Probably the most important development in preventing rejection has been the concept that escalation of the stem cell dose directly contributes to the likelihood of establishing donor type chimaerism. Increasing the donor stem cell dose may facilitate competition with residual host stem cells for available niches. Additionally, it was realized that cells within the CD34+ compartment possessed potent veto activity, which neutralize host alloreactive CTL, and the greater the number of CD34+ cells, the greater the induction of tolerance (Uharek et al, 1992; Bachar-Lusting et al, 1995; Reisner & Martelli, 1995). When haematopoietic growth factors became available, the concept of a megadose SCT could be applied in humans; large numbers of CD34+ cells (e.g. 10 × 106/kg) could be transplanted into patients by collecting PBPCs from donors following mobilization with G-CSF (Aversa et al, 1994). The Perugia group (Aversa et al, 1998), have successfully developed a megadose haploidentical SCT procedure using a relatively non-toxic conditioning regimen consisting of thiotepa (a single TBI dose of 800 cGy), fludarabine and ATG, which achieves over 90% sustained engraftment with a 30-d treatment-related mortality (TRM) of only 10%.

In addition to cells within the CD34+ fraction, several other donor cells possessing veto activity have been described, amongst which the most potent would appear to be CTLs (Sambhara & Miller, 1991). As these cells also possess marked GVH reactivity, investigators have developed new approaches to deplete alloreactive clones directed against the host by stimulating the donor T cells against third party stimulators in the absence of interleukin 2 (IL-2); only the activated anti-third party CTLs survive IL-2 starvation in the primary culture, so that alloreactive CTLs directed against the host are depleted. In mouse models, these non-alloreactive CTLs are endowed with a very potent veto activity which appears mediated through both FasL and CD8-mediated apoptosis (Reich-Zeliger et al, 2000).

If the tolerizing effects of CD34+ and/or other veto cells could be increasingly exploited then this would further facilitate transplantation across the haploidentical barrier under safer conditions, without the need for supra-lethal conditioning. This would reduce the risks in patients with existing co-morbidities, and facilitate transplantation in children with genetic diseases where such conditioning is harder to justify.

Graft versus host disease

  1. Top of page
  2. Abstract
  3. Introduction
  4. Graft rejection
  5. Graft versus host disease
  6. Graft versus leukaemia
  7. Immune reconstitution
  8. Clinical paediatric perspective
  9. References

Historically, GVHD was one of the overwhelming problems associated with haploidentical transplantation, making the procedure almost prohibitive. Initially, it was shown that the risk of acute GVHD was significantly increased in patients receiving marrow grafts from donors incompatible for one, two, or three HLA loci (Beatty et al, 1985; Anasetti et al, 1989). GVHD in this setting is largely mediated by alloreactive donor T cells recognizing mismatched HLA molecule–peptide complexes, and methods to prevent the high rate of GVHD have relied largely upon the depletion of most or all of the donor T cells. It is known from early studies of donor lymphocytes in the haploidentical setting that cell doses as low as 3 × 104/kg can mediate lethal GVHD (Aversa et al, 1998). A variety of negative selection strategies using bone marrow as a stem cell source have been utilized, such as soyabean lectin agglutination, monoclonal or polyclonal antibodies either as single agents or in combination (reviewed by Ho & Soiffer, 2001). With the introduction of PBPCs as a preferred stem cell source, the large number of PBPCs that needed to be processed required the introduction of alternative T-cell depletion strategies and methods such as combinations of positive selection of peripheral CD34+ progenitors followed by an additional negative depletion step have been described (Collins et al, 1997; Aversa et al, 1998). In the last few years a single-step positive selection of CD34+ progenitors using a method of high-gradient magnetic-activated cell sorting (MACS) has been increasingly used as an indirect method for T-cell depletion (Schumm et al, 1999). The T-cell depletion obtained with this method is in the range of 4–5 logs, achieving a threshold of 1–5 × 104/kg CD3+ cells/kg, below which GVHD prophylaxis is not required (Reisner & Martelli, 2000). Using this approach the problem of GVHD has been largely overcome, with very low rates of GVHD in the largest series (Aversa et al, 1998; Handgretinger et al, 2001). Additionally, this positive selection method is associated with an indirect depletion of B-lymphocytes of >3 logs, which reduces the incidence of Epstein-Barr virus (EBV)-associated lymphoproliferative disease, otherwise associated with selective T-cell depletion (Shapiro et al, 1988). However, the rigorous T-cell depletion attained by CD34 selection occurs at the cost of loss of the graft versus leukaemia (GVL) effect and delayed immune reconstitution, resulting in high relapse rates and a high morbidity/mortality from infection. Novel strategies designed for the selective depletion/tolerization of alloreactive T cells causing GVHD are discussed below. Recent reports have questioned whether the expression of CD34 is sufficient to characterize the most pluripotent stem cell (Zanjani et al, 1998), and investigators are looking at the selection of more primitive cells using antigens expressed at an earlier developmental stage (e.g. CD133) (Yin et al, 1997). Further, the possibility still exists that by positive selection of stem cells based on their antigenic expression, useful accessory cells may be discarded, so that investigators are again revisiting the possibility of large-scale negative T-cell (and B-cell) depletion strategies of PBPCs to address this issue (Handgretinger et al, 2002). In particular, regulatory T cells, which play a critical role in the maintenance of immunological tolerance, may also have the potential to prevent GVHD. Recently, much attention has focussed on CD4+/CD25+ T cells. In a murine model (Taylor et al, 2002), depletion of CD4+ CD25+ cells from the graft resulted in increased GVHD and infusion of ex vivo expanded CD4+ CD25+ cells significantly inhibited GVHD. The major issue preventing clinical application of such an approach is that suppression by CD4+ CD25+ cells is not specific for alloantigens, so that loss of anti-viral and anti-leukaemic responses would be predicted. Novel approaches to generate alloantigen-specific regulatory T cells are needed.

In the haploidentical setting, the mismatched haplotype of the donor can originate from either of the parents, accordingly referred to as non-inherited maternal antigens (NIMA haplotype) or non-inherited paternal antigens (NIPA haplotype). Exposure to maternal antigens in utero appears to reduce the risk of GVHD, as non-T-cell depleted bone marrow transplants donated by 1–2 antigen mismatched haploidentical sibling donors to recipients mismatched for NIPA caused more acute and chronic GVHD and TRM than transplants donated by haploidentical siblings mismatched for NIMA (Van Rood et al, 2002).

Graft versus leukaemia

  1. Top of page
  2. Abstract
  3. Introduction
  4. Graft rejection
  5. Graft versus host disease
  6. Graft versus leukaemia
  7. Immune reconstitution
  8. Clinical paediatric perspective
  9. References

In the HLA-matched bone marrow transplant setting, patients with high risk leukaemia who do not develop GVHD after an extensively T-cell depleted graft are at an increased risk of relapse (Marmont et al, 1991). Paradoxically, clinical reports of patients with advanced acute myeloid leukaemia (AML) undergoing haploidentical SCT, suggest a relatively low incidence of GVHD and a low relapse rate (Aversa et al, 1998; Ruggeri et al, 2002). Recent work has suggested that donor NK alloreactivity, a biological phenomenon which is unique to mismatched transplants, may play a significant role in this GVL effect in mismatched transplants (Ruggeri et al, 1999).

Natural killer function is regulated by a complex array of interactions between receptors on their cell surface and target cell ligands that inhibit or activate NK-mediated lysis (reviewed in Farag et al, 2002). In particular, NK cells are normally blocked from killing self or HLA-matched targets by the expression of KIRs, which recognize self major hisotcompatibility complex class I ligands, and which (upon engagement with their cognate class I ligands) deliver inhibitory signals to block activation of NK-cell lysis (Moretta & Moretta, 1997; Farag et al, 2002). After a KIR-mismatched haploidentical SCT, the engrafted stem cells appear to give rise to a transient (1–3 months) wave of reconstituting NK cells with a high frequency of donor versus recipient alloreactive NK clones. In vitro assays have demonstrated that alloreactive NK clones have potent cytotoxic activity against leukaemic targets from patients with chronic myeloid leukaemia (CML) and AML, but not against blasts from most patients with acute lymphoblastic leukaemia (ALL) (Ruggeri et al, 1999). This may reflect the lack of expression of lymphocyte function-associated antigen 1 (LFA1), an adhesion molecule which is necessary for NK binding to target, on ALL blasts (Ruggeri et al, 1999).

In the clinical setting, long-term follow-up of 57 high risk AML patients who underwent haploidentical SCT showed an impressive effect of NK alloreactivity on subsequent relapse. No relapses occurred amongst 20 patients transplanted from haploidentical donors with KIR mismatch in the GVL direction. In contrast, 28 of 37 patients relapsed when transplanted with grafts with no potential for NK alloreactivity (P < 0·001) (Ruggeri et al, 2002). This apparent GVL effect was not associated with GVHD; indeed KIR-mismatched patients had a significantly lower incidence of significant GVHD. No difference in relapse rate was observed in patients transplanted for ALL. This data strongly support the hypothesis that in HLA-mismatched transplants, GVL may be mediated by NK alloreactivity when a KIR mismatch occurs in the GVH direction. In contrast to the T-cell mediated GVL effect in HLA-matched transplants, this appears to be independent of GVHD and murine studies suggest this may be due to the elimination of recipent antigen presenting cells, which are critical for initiating GVHD, by alloreactive donor NK cells (Shlomchik et al, 1999; Ruggeri et al, 2002). Larger, prospective studies are needed to substantiate these results. If these data are confirmed, KIR mismatching between donor and recipient should become a major criteria for donor selection in haploidentical SCT for myeloid malignancies.

The GVL effect of donor CTL recognizing mismatched minor histocompatibility, overexpressed myeloid and leukaemic antigens is largely lost in haploidentical SCT, because of the rigorous T-cell depletion employed. However, strategies aimed at generation of allorestricted CTL recognizing haematopoetic antigens such as WT-1, HA-1 and CD45 (Gao et al, 2000; Mutis et al, 2002; Amrolia et al, 2003a) or selective depletion of alloreactive donor T cells with preservation of CTL responses to myeloid tumour antigens (Amrolia et al, 2003b) may offer the prospect of restoring a CTL-mediated GVL effect in the haploidentical setting.

Immune reconstitution

  1. Top of page
  2. Abstract
  3. Introduction
  4. Graft rejection
  5. Graft versus host disease
  6. Graft versus leukaemia
  7. Immune reconstitution
  8. Clinical paediatric perspective
  9. References

Having largely overcome the problems of engraftment and GVHD, much of the focus at the present time is centred on the delayed immune reconstitution that follows haploidentical SCT. Several mechanisms underlie slow immunological reconstitution, which include (i) the profound T- and B-cell depletion associated with CD34 selection [generally the number of CD3+ cells infused with the graft is <5 × 104/kg (Schumm et al, 1999)], (ii) ATG in the conditioning regimen which may antagonize expansion of these residual cells, (iii) the degree of HLA disparity between the host and donor, and (iv) decaying thymic function in adults (Roux et al, 2000; Weinberg et al, 2001). This slow recovery places patients at significant risk from post-transplant viral, fungal and other opportunistic infections (Kook et al, 1996; Small et al, 1999), and is the most important cause of mortality in adults undergoing haploidentical SCT, reaching as high as 40% in some studies (Aversa et al, 1998). This risk would appear to be considerably reduced in the paediatric population, where immune reconstitution is somewhat more rapid (Handgretinger et al, 1999; Ortín et al, 2002). A prospective study of immune reconstitution post-CD34-selected haplo-identical SCT in children showed the median time to normal T- and B-cell numbers and proliferative responses was 7–8 months (Eyrich et al, 2001). T-cell reconstitution appeared biphasic, with an initial phase characterized by an oligoclonal repertoire of memory T cells (presumably because of peripheral expansion of mature T cells from the graft) followed at 6 months by a wave of naïve T cells, presumably derived from donor stem cells emigrating from the thymus, which broadens the T-cell repertoire. Immune reconstitution may be more rapid in the paediatric setting because higher CD34+ cell doses are feasible and because of the presence of intact thymic function. CD34+ cell megadoses of >20 × 106/kg appear to enhance the recovery of CD3 and CD4 lymphocyte populations (Handgretinger et al, 1999; Ortín et al, 2002).

Interleukin 12 is known to be a major cytokine in the initiation of protective T-helper 1 (Th-1) immunity against opportunistic infections. The observation that G-CSF, used to hasten neutrophil recovery post-SCT, blocks IL-12 production by antigen-presenting cells prompted the discontinuation of this cytokine post-transplant. As the early stopping of G-CSF, clinical data suggest the restoration of IL-12 levels to normal values much sooner, with CD4+ cell numbers and function markedly improving (Volpi et al, 2001).

Much attention at present is being focused on the restoration of specific immunity after transplant, in order to prevent infective complications and perhaps also relapse. Donor lymphocyte infusions (DLI) are frequently used for the correction of immunodeficiency in HLA-matched transplants, but this approach is unlikely to be feasible in the haploidentical setting, because the frequency of alloreactive T cells is estimated to be 2–3 logs greater than that of viral-specific T cells. The use of low dose DLI from haploidentical donors was not shown to improve immune reconstitution (Eyrich et al, 2001) and is frequently associated with severe GVHD (Aversa et al, 1998; Kawano et al, 1998; Eyrich et al, 2001). Several groups have explored the possibility of using genetically modified DLI transduced with an herpes simplex virus thymidine kinase (HSV-TK) ‘suicide’ gene to allow their selective elimination by activating the prodrug gancyclovir, in the event of serious GVHD (Bonini et al, 1997; Tiberghien et al, 2001). However, the polyclonal stimulation needed for efficient retroviral transduction of T cells may compromise their anti-viral activity (Sauce et al, 2002) and mutations in the HSV-TK gene may lead to gancyclovir resistance, so that it may not be possible to switch off GVHD (Garin et al, 2001). Rather than use bulk DLI, other groups have attempted to restore T-cell immunity to specific pathogens using adoptive immunotherapy with specific T-cell lines. This approach has been used extremely successfully for the prevention of cytomegalovirus (CMV) and EBV-related disease in the HLA-matched setting (Walter et al, 1995; Rooney et al, 1998). However, in the haploidentical setting, in order to avoid GVHD, T-cell cloning is required. Perruccio et al (2001) generated large numbers of donor T-cell clones against Aspergillus fumigatus and CMV antigens, screened them for cross-reactivity to host alloantigens and infused pooled non-alloreactive clones into adult recipients at a dose of 5 × 105/kg on day 15 after transplant. All patients developed Aspergillus- and CMV-specific responses within 3 weeks compared with untreated patients who developed the corresponding pathogen-specific T-cell responses more than 9 months post-transplant (Perruccio et al, 2001). This approach, while elegant, will only restore immunity to individual target pathogens and is far too labour-intensive to be used routinely. A number of strategies to tolerize or delete alloreactive T cells responsible for GVHD, but preserve anti-viral and anti-leukaemic T-cell responses from bulk populations of donor lymphocytes are, therefore, being investigated.

A Boston group has studied the selective induction of anergy in alloreactive T cells by culturing the graft ex vivo with irradiated recipient peripheral blood mononuclear cells (PBMC) in the presence of cytotoxic T-lymphocyte associated antigen 4 (CTLA-4) immunoglobulin to block B7:CD28 mediated co-stimulation (Boussiotis et al, 1994; Gribben et al, 1996; Guinan et al, 1999). An initial clinical study in 11 haploidentical transplants, in which T-replete alloanergized grafts were given to adult patients with acute leukaemia, showed impressive results with remarkably low rates of GVHD (Guinan et al, 1999), however, it was not demonstrated that this improved immune reconstitution. Unfortunately this antibody is no longer clinically available. The Boston group are initiating clinical trials with similar antibodies such as anti-B7-1 and Quesenberry et al (2001) have shown a similar approach is feasible in a murine model using blockade of CD40 ligand-mediated co-stimulation. The problems with this approach are that anergy may not be complete or permanent, resulting in GVHD, and that the anergized cells may have a negative effect on bystander T cells with anti-viral and anti-leukaemic activity. It will therefore be critical to assess the reconstitution of viral-specific immunity in further clinical trials using this approach.

An alternative strategy is to selectively deplete donor T cells, specifically of alloreactive T cells responsible for GVHD, by deleting T cells that are activated in response to recipient antigen presenting cells. A number of groups have targeted surface markers expressed on activated T cells, e.g. CD25 and CD69 (Montagna et al, 1999; Koh et al, 2002), using magnetic microbeads to remove, or an immunotoxin to kill, donor T cells expressing activation markers after co-culture with recipient PBMC. This approach has the advantages that alloreactive T cells are permanently removed and cannot influence the function of the remaining T cells. Andre-Schmutz et al (2002) have recently published data from a Phase I/II clinical study of addback of 105 – 8 × 105/kg allodepleted donor lymphocytes, generated using an anti-CD25 (IL-2 receptor α) immunotoxin, in 15 patients undergoing HLA-mismatched SCT. The incidence of significant GVHD was impressively low (two of 15), but it is too early to be sure whether the infused T cells improved anti-viral immune reconstitution. We have investigated a similar approach using recipient EBV-transformed B cells as stimulators, in order to more reliably deplete alloreactive T cells and to preserve responses to potential tumour antigens in myeloid malignancies (Amrolia et al, 2003b). We have demonstrated that this approach preserves in vitro T-cell responses to CMV, adenovirus and EBV (through the retention of T cells recognizing EBV antigens in the context of the non-shared HLA haplotype). An alternative approach to allodepletion relies on the principle that activated T cells selectively retain a photo-active rhodamine derivative and hence host-reactive T cells can be deleted by photodynamic purging (Chen et al, 2002; Guimond et al, 2002). These approaches show considerable promise, but it remains to be demonstrated that a sufficient level of allodepletion can be achieved to enable the addback of enough T cells to restore anti-viral and anti-leukaemic activity without causing GVHD. This issue can only be addressed in carefully designed clinical studies.

Clinical paediatric perspective

  1. Top of page
  2. Abstract
  3. Introduction
  4. Graft rejection
  5. Graft versus host disease
  6. Graft versus leukaemia
  7. Immune reconstitution
  8. Clinical paediatric perspective
  9. References

The role of haploidentical SCT for the treatment of genetic diseases is firmly established. Due to the immediate availability of a donor, haploidentical SCT has been extensively utilized in SCID (Fischer et al, 1990; Buckley et al, 1999), when a matched family donor is not available. The outcome in such patients has improved dramatically in recent years with >75% of patients surviving for a year (Antoine et al, 2003). For non-SCID immunodeficiencies, where timing is less critical, a closely matched unrelated donor is often preferred to a haploidentical donor as the former approach gives superior engraftment with an improved 3-year survival approaching that of genotypically HLA identical SCT (Antoine et al, 2003). Nevertheless, for a significant number of children with genetic diseases who come from ethnic minority groups no closely matched donor can be found; for this reason haploidentical SCT has been extensively used in hemophagocytic lymphohistiocytosis (Arico et al, 1996; Jabado et al, 1997), and although the results are improving they are not as yet equivalent to matched donor SCT.

Most recent studies reporting the use of haploidentical SCT in leukaemia involve adult patients (Aversa et al, 1998). There have been eight studies in the last 5 years which have reported a group of 10 or more children undergoing haploidentical SCT, and these are listed in Table I. The most recent report, and that with the most favourable outcome, comes from Birmingham Children's Hospital in the UK (Ortín et al, 2002). Children with malignant conditions received TBI (14·4 Gy in eight fractions) and cyclophosphamide (60 mg/kg/d for two consecutive days) according to standard UK practice for matched related and unrelated donor transplants (Lawson et al, 2000). In addition, patients received 5 d of fludarabine (25 mg/m2/d) prior to the above and equine anti-lymphocyte globulin (ALG) 12·5 mg/kg/d from day −2 to day +2. All patients received PBPCs after CD34 selection with CliniMACS (Miltenyi Biotech, Bergisch-Gladbach, Germany). The mean CD34 cell dose administered was 11 × 106/kg, with a mean CD3+ T-cell dose of 7 × 104/kg. Due to the relatively high T-cell dose, cyclosporin A was given for the first 30 d then tailed. Among 16 patients with malignant diseases, all engrafted, there was no TRM, three patients have relapsed and died, and 13 of 16 (81·3%) remained alive, well and in complete remission (CR) after a median follow-up of 480 ± 255 d.

Table I.  Studies within the last 5 years reporting haploidentical stem cell transplantation in 10 or more paediatric patients.
ReferenceNo. of patients 2–3 Ag. mmConditions CD 34 dose (×106/kg)CD three dose (×104/kg)ConditioningSerotherapy/ immunosuppresionEngraftment (%)TRM (%)GVHD (>grade II) (%)Outcome and median follow-up
  1. Ag. mm, antigen mismatch; TRM, transplant-related mortality; GVHD, graft versus host disease; M, malignant; CML, chronic myeloid leukaemia; AA, aplastic anaemia; MDS, myelodysplastic syndrome; N/A, not available; BM, bone marrow; NC, nucleated cells; TBI, total body irradiation; TLI, total lymphoid radiation; ATG, antithymocyte globulin; ALG, antilymphocyte globulin; VP16, etoposide; Thio, thiotepa; Cy, cyclophosphamide; CsA, cyclosporin  A; BM, bone marrow; Flu, fludarabine; Bu, busulphan; MTX, methotrexate; MP, methylprednisolone; ext, extensive; AW, alive and well.

  2. *Two patients experienced de novo graft versus host disease (GVHD). Four patients rejected the graft and received donor lymphocyte infusions (DLI). Three of these patients had GVHD.

  3. †Ten patients failed to engraft initially and received a second graft. Six of these engrafted.

  4. ‡Four patients failed to engraft and two rejections were seen. Two of these patients engrafted after a second transplant.

  5. §The only significant cases of GVHD were in three of eight patients receiving DLI.

  6. ¶Two patients failed to engraft and three rejections were seen. Four of these patients engrafted after a second transplant.

  7. **GVHD only seen after donor lymphocyte infusions.

Speiser et al (1997)10349M (CML)N/AN/ATBI/CySerotherapy/CsA ± MTX80545637% survival at 24 months
Passweg et al (2000) 1010M12·90·04TBI/VP16/CyATG pregraft OKT3 postgraft70403030% AW at 24 months
Kawano et al (1998) 13 812 M 1 genetic7·70·1TBI/TLI: 8MP/CsA + MTX693815*38% AW at 24 months
Henslee-Downey et al (1997) 725966 M 6 AA/MDSBM 1·5 × 108 NC/kg7·5TBI/VP16/Cy/ AraCATG postgraft MP pre- and post-CsA87†3316 Ext. chronic: 835% AW at 24 months
Peters et al (1999) 14149 M 5 genetic21·54·7TBI/TLI: 6 Bu/Thio/Cy: 8ATG pre- and post-CsA in genetic7936757% AW at 15 months
Handgretinger et al (1999) 232018 M 5 genetic14·21·4TBI/Thio/Cy Bu/Thio/Cy No conditioning: 1ALG: 17 ALG, OKT3, steroids: 3 ATG: 1 ATG, OKT3, steroids: 183‡1313§48% AW (median follow-up N/A)
Klingebiel et al (2000) 393431 M 8 genetic19·61·08N/ACsA: 797¶1810**46% AW at 6 months
Ortín et al (2002) 212116 M 5 genetic11·137·01Flu/TBI/Cy Flu/Bu/CyALG day −2 to +2 CsA100 55 Chronic: 1976% AW at 16 months

The Birmingham study is notable for the high, sustained engraftment rate and the lack of infectious deaths. As noted previously, successful engraftment after haploidentical SCT is determined by a complex relationship between stem cell dose, extent of T-cell depletion, intensity of pretransplant conditioning regimen, and pre- and post-transplant immunosuppression (Passweg et al, 2000). In the series reported by Peters et al (1999), the incidence of graft failure was 29% after a mean CD34 dose of 21·5 × 106/kg. Handgretinger et al (1999) reported a 20% failure rate with grafts containing an average number of 14·2 × 106 CD34 cells/kg. Conversely, in Ortín et al (2002), all patients with malignant disease engrafted, despite a modest mean CD34 cell dose of 11·13 × 106/kg. This may reflect the routine use of TBI in conditioning, as well as a significantly higher donor T-cell dose.

Henslee-Downey et al (1997) first stressed the importance of TBI in the conditioning regimen to reduce the rejection risk. Whilst avoiding TBI in children may reduce long-term side-effects (Leiper, 2002) and several studies (Handgretinger et al, 1999; Peters et al, 1999) include a number of children who achieved successful engraftment without TBI, in all the reports patients who received TBI were more likely to achieve sustained engraftment and maintain complete donor chimaerism. This was particularly evident in one study (Kawano et al, 1998), where the median dose of CD34 cells (7·7 × 106) and T cells (1 × 105) was relatively low and engraftment failed in most of those who did not receive TBI. A number of children, who have experienced rejection, have been successfully engrafted with a second infusion of CD34 cells following further immunosuppression with methylprednisolone, anti-CD3 antibody OKT3, with or without fludarabine (Klingebiel et al, 2000). It may be possible to predict impending rejection by the early detection of recipient T cells post-SCT (Peters et al, 1999).

In the paediatric studies by Ortín et al (2002) and Handgretinger et al (1999), the number of infectious deaths were low. Ortín et al (2002) reported that <50% (eight of 21) patients experienced viral infections, with only three cases of CMV reactivation amongst 14 CMV-positive donor-recipient pairs and no deaths attributable to infection. Handgretinger et al (1999) reported two deaths caused by viral infections (one adenovirus, one CMV) out of 23 patients. As a comparison, Bunjes et al (2000) reported a series of 10 haploidentical transplants in adults where all six patients with positive pretransplant serology for CMV developed viraemia, and five developed CMV disease despite prophylactic gancyclovir. Likewise, infections accounted for 24 of 34 non-leukaemic deaths in 76 mainly adult patients transplanted in a recent Perugia series (Aversa et al, 2001). These differences may reflect the relatively preserved thymic function in children compared with adults, together with the higher stem cell doses attainable. High CD34 cell doses appear to hasten immune recovery (Handgretinger et al, 1999; Ortín et al, 2002), but in the Birmingham study (Ortín et al, 2002) the mean CD34 dose (11·13 × 106/kg) was lower than other paediatric studies (see Table I). It is possible that the relatively high T-cell dose infused in this study may have been responsible for the reduced incidence of viral complications.

As mentioned previously, a factor that has confounded the interpretation of outcome data in the haploidentical setting is the inherent advanced disease amongst the recipients of such a procedure (Aversa et al, 1998), and it is notable in the study by Ortín et al (2002) that all patients were in CR at the time of transplant.

The overall survival rate for children undergoing haploidentical SCT from the eight studies listed in Table I ranged from 27% to 71%, with a mean of 44% at a median follow-up of 18 months. This is very comparable with the results achieved in a large group of children with acute leukaemia (Rocha et al, 2001) or leukaemia and genetic diseases (Barker et al, 2001) who underwent unrelated donor bone marrow or cord blood transplantation. Very few studies have directly compared haploidentical SCT with other forms of allogeneic SCT. The relative merits of choosing an unrelated bone marrow or peripheral blood donor versus a cord blood donation versus a haploidentical family member will be dictated to some degree by the disease and urgency of transplant. In situations where both approaches are feasible, prospective randomized studies to determine the best approach are now appropriate. The potential advantages and disadvantages of each are given in Table II. A study of haploidentical SCT in chronic myeloid leukaemia that included 21 children showed a superior survival of 47% with donors mismatched for 0–1 antigens compared with 27% for donors mismatched for 2–3 antigens (Speiser et al, 1997). Drobyski et al (2002) reported superior survival for adults and children with haematological malignancies undergoing matched unrelated donor SCT (58%) versus either mismatched unrelated (34%, P = 0·01), or haploidentical (21%, P = 0·002) SCT. Relapse was lower in matched unrelated patients (25%, P < 0·01) and mismatched unrelated patients (26%, P < 0·014) than in haploidentical patients (42%). TRM was significantly higher in recipients of mismatched unrelated grafts (45%, P < 0·01) and haploidentical grafts (42%, P < 0·001) compared with recipients of matched unrelated marrow grafts (23%). However, this study was not randomized and the outcome of the haploidentical group, which included relatively young patients (median age of 22 years), was poor in comparison with paediatric studies (see Table I); this was presumably due to a significant proportion of high-risk patients. Nevertheless, in the absence of a sibling donor, the majority of paediatric transplant centres would choose a fully matched unrelated donor as the next choice, time permitting, and then centres differ in their subsequent choice between a cord blood donation (matched for ≥4/6 antigens), a mismatched bone marrow donor (≥9/10 antigens including HLA-A, -B, -C, DRB1, DQB1), or a haploidentical family member. The United Kingdom's Children's Cancer Study Group (UKCCSG), which collectively performs 250 paediatric allogeneic transplants each year, recently achieved a consensus document detailing indications for paediatric SCT and donor selection guidelines. Proposed indications for allogeneic SCT including haploidentical SCT are given in Table III (modified from Cornish et al, 1998) and the proposed HLA hierarchy and optimum stem cell source and choice of donor are outlined in Table IV (A. Vora & A. Green, personal communication).

Table II.  Potential advantages and disadvantages of the different types of stem cell donor.
DonorAvailabilityAccess (re-access)CostRejection riskEngraftmentGVHD riskGVLImmune reconstitution
  1. GVHD, graft versus host disease; GVL, graft versus leukaemia; T, T cell; NK, natural killer cell.

Unrelated bone marrow10/10 = 50% >9/10 = 80% Ethnic minority = 20%Slow (possible)HighLowModerateModerateTModerate
Unrelated cord blood≥5/6 antigens = 45% ≥4/6 antigens = 90%Fast (no)ModerateHighSlowLowTSlow
Haploidentical family>90%Immediate (yes)LowModerateFastLowNKVery slow
Table III.  Proposed indications for transplant procedures in children.
DiseaseDisease statusAllogeneic matched related* Allogeneic unrelated†Haploidentical relatedAutologous blood or marrow
  1. *Also includes five of six matched related donors in certain cases.

  2. †Includes 9/10, 10/10 from HLA-A, -B, -C, DRB1, DQB1.

  3. ‡Not t(15, 17); t(8, 21); inv16; Down's syndrome.

  4. §If relapse <1 year from diagnosis.

  5. ¶If relapse >1 year from diagnosis.

  6. **Worth considering, if killer immunoglobulin-like receptor (KIR)-mismatched donor available because of graft versus leukaemia effect of natural killer cells.

  7. ††Philadelphia positive t(9, 22), near haploid, >25% blasts in bone marrow (BM) at day 28.

  8. ‡‡For high-risk patients by the Berlin–Frankfurt–Munster classification (Uderzo et al, 2001), i.e. T-cell disease or BM relapse <18 months from diagnosis, any standard or intermediate risk patient with minimal residue disease (MRD) >104 at week 5 of re-induction will be offered blood and marrow transplantation (BMT), if closely matched related/unrelated donor available.

  9. §§If no haematological response to Imatinib of interferon, or haematological relapse on Imatinib.

  10. ¶¶If cytogenetics abnormal or blood product dependent.

  11. ***RAEBT, sAML eligible for AML trials, except Down's syndrome, where BMT is not necessary.

  12. †††Alternative donor SCT only considered in patients with acquired AA, who have failed immunosuppressive therapy.

  13. R, in routine use for selected patients; CRP, to be undertaken in approved Clinical Research Protocols; D, developmental or pilot studies can be approved in specialist units; NR, not generally recommended; CMML, chronic myelomonocytic leukaemia; JMML, juvenile myelomonocytic leukaemia; RA, refractory anaemia; RARS, RA with ringed sideroblasts; RAEBT, RA with excess blasts in transformation; AML, acute myeloid leukaemia; sAML, secondary AML, ALL, acute lymphoblastic leukaemia; CML, chronic myeloid leukaemia; T-NHL, T cell non-Hodgkins lymphoma.

  14. Developed through the UKCCSG BMT group (modified from Cornish et al, 1998). Indications for allogeneic SCT in children devised by the UKCCSG BMT group (modified from Cornish et al, 1998).

AMLCR1R‡NRNRNR
CR2RCRPCRP§
Relapse/refractoryDDCRP**NR
ALLHigh-risk CR1R††R††NRNR
CR2‡‡R‡‡R‡‡CRP‡‡CRP
CR3RRRNR
Relapse/refractoryNRNRNRNR
CMLChronic phaseRRCRP§§CRP
Advanced phaseRRCRP§§CRP
Blast crisisDNRCRP§§NR
T-NHLAs ALL    
Hodgkin's diseaseCR1NRNRNRNR
CR2NRNRNRCRP
CR3DDNRD
RefractoryNRNRNRD
MyelodyspasiaJMMLRRCRPNR
RA¶¶, RARS, CMML,RCRPCRPNR
RAEBT, sAML***    
Immunodeficiency and inborn errorsRRR
ThalassaemiaRNRNR
Sickle cell diseaseRNRNR
Aplastic anaemia and inherited monocytopeniaRCRP†††CRP†††
Table IV.  A proposed HLA hierarchy and preferred stem cell source to guide choice of the most optimal SCT donor.
  1. HLA, human leucocyte antigen; SCT, stem cell transplantation; MSD, matched sibling donor; MUD/MMUD, matched/mismatched unrelated donor; MFD/MMFD, matched/mismatched family donor; SCID, severe combined immunodeficiency; KIR, killer immunoglobulin-like receptor; BM, bone marrow; PBSC, peripheral blood stem cell.

  2. Devised by UKCCSG BMT group 2002 (A. Vora & A. Green, personal communication).

HLA hierarchy
 1. MSD= phenotypic MFD = fully matched related cord blood.*
 2. 10/10 molecular match UD = 6/6 matched unrelated cord blood.*
 3. 1 allelic MMUD = 5/6 MFD = 2/6 MM unrelated cord blood.*
 4. 2 allelic MMUD = ≥2 MMFD.†
*With nucleated cell dose >3·7 × 107/kg recipient weight. Options 3 and 4 should only be considered in poor risk patients for whom alternative treatment is unavailable, or has failed. Option 4 usually requires in vitro T-cell depletion. MMFD may be preferred over unrelated donors for certain disorders (e.g. severe combined immunodeficiency).
†Most ≥2 MMFD for children are parents, as the use of GCSF in young sibling donors is not encouraged. Choice of parent depends on factors given below and KIR mismatch in myeloid diseases.
Stem cell source
 1. MSD‡ − BM > PBSC.
 2. MUD‡ − BM = PBSC.
 3. MMUD/MMFD − PBSC > BM.
‡Adult sibling and unrelated donors should be allowed a choice after non-directive counselling by a doctor, independent of the transplant programme.
CMV statusGender
PatientDonorPatientDonor
PosPos > NegMaleMale > Female
NegNeg > PosFemaleMale > Female
Age: Young > Old
ABO blood groups: Aim for a match in heavily pretransfused patients

There is a small amount of experience with haploidentical SCT in the non-malignant haematological diseases. In Pesaro, 28 children with homozygous β-thalassaemia received unmodified marrow from a haploidentical family donor mismatched at zero HLA antigens (n = 6), one HLA locus (n = 15), two loci (n = 5), and three loci (n = 2) (Gaziev et al, 2000). Overall survival and thalassaemia free survival reached a plateau at 65% and 21% respectively. As might be expected in a heavily transfused cohort, the rejection rate was high, occurring in 16 of 29 patients, but rather surprisingly, HLA disparity did not have a significant effect on survival. Transplant-related mortality was 34%, GVHD was a major contributing cause of death (50%), followed by infections (30%). In children with thalassaemia who do not have a matched family donor and who require SCT because they cannot receive adequate conventional treatment, more encouraging data are coming from the use of very closely matched unrelated donors using extended haplotype matching (La Nasa et al, 2002).

For patients with severe aplastic anaemia refractory to immunosuppressive therapy who do not have a matched sibling donor, major advances have recently been made with closely matched unrelated donors (Kojima et al, 2000; Vassiliou et al, 2001). In the absence of an unrelated donor, there have been only a few anecdotal reports of haploidentical SCT in such patients, with no long-term survivors so far (Handgretinger et al, 1999; Kremens et al, 2001). There has been a more favourable outcome in Fanconi anaemia (Boulad et al, 2000; Elhasid et al, 2000), and it is likely that with the ongoing improvements in haploidentical SCT more successes will be achieved in bone marrow failure syndromes.

Most children undergoing haploidentical SCT receive preparation with myeloablative chemoradiotherapy. Occasional patients will not tolerate this intensity of conditioning because of pre-existing co-morbidities. Given that highly purified CD34 cells possess potent veto activity, there has been some limited success with the use of reduced intensity fludarabine/melphalan/ATG-based regimens with the addition busulphan 8 mg (Veys et al, 2002) or total lymphoid irradiation (Matthes-Martin et al, 2002) to facilitate engraftment in the haploidentical setting. Further work with additional veto cells (Aversa et al, 2001), targeted radiotherapy (Pagel et al, 2002) and monoclonal antibodies (Wulf et al, 2003) is likely to improve this approach in the future.

In conclusion, there has been much progress in both the clinical and scientific approaches to haploidentical transplants. Recent experience now shows that a high rate of engraftment can be achieved without severe GVHD and with low regimen-related mortality. Post-transplant infectious complications and relapse remain the most important barriers yet to overcome, and new directions in the use of adoptive cellular immunity appear to be promising. Haploidentical SCT is a viable option for those children who do not have an HLA compatible sibling donor or fully matched unrelated donor. The majority of children requiring such a procedure will have ALL in second CR (CR2) at high risk of relapse or AML in CR2 where relapse has occurred within the first year from diagnosis. Haploidentical SCT should not be pursued for children with ALL not in remission but because of the potential GVL effect of NK cells in AML, studies of haploidentical SCT in refractory AML may be justified. The relative merits of a haploidentical family donor versus mismatched bone marrow or cord blood donation need to be assessed in prospective, multi-centre randomized clinical trials. In the meantime, ongoing translational research is very likely to improve this modality of treatment further.

References

  1. Top of page
  2. Abstract
  3. Introduction
  4. Graft rejection
  5. Graft versus host disease
  6. Graft versus leukaemia
  7. Immune reconstitution
  8. Clinical paediatric perspective
  9. References
  • Amrolia, P.J., Reid, S.D., Gao, L., Schultheis, B., Dotti, G., Brenner, M.K., Melo, J.V., Goldman, J.M. & Stauss, H.J. (2003a) Allorestricted cytotoxic T cells specific for human CD45 show potent antileukemic activity. Blood, 101, 10071014.
  • Amrolia, P.J., Muccioli-Casadei, G., Yvon, E., Huls, H., Bollard, C., Weider, E., Molldrem, J.J., Rooney, C., Heslop, H., Kuehnle, I. & Brenner, M.K. (2003b) Selective depletion of donor allo-reactive T-cells without loss of anti-viral and anti-leukemic responses using an anti-CD25 immunotoxin. Blood, 102, 22922299.
  • Anasetti, C., Amos, D., Beatty, P.G., Appelbaum, F.R., Bensinger, W., Buckner, C.D., Clift, R., Doney, K., Martin, P.J. & Mickelson, E. (1989) Effect of HLA compatibility on engraftment of bone marrow transplants in patients with leukemia or lymphoma. The New England Journal of Medicine, 320, 842847.
  • Andre-Schmutz, I., Le Deist, F., Hacein-Bey-Abina, S., Vitetta, E., Schindler, J., Chedeville, G., Vilmer, E., Fischer, A. & Cavazzana-Calvo, M. (2002) Immune reconstitution without graft-versus-host disease after haemopoietic stem-cell transplantation: a phase 1/2 study. Lancet, 360, 130137.
  • Antoine, C., Muller, S., Cant, A., Cavazzana-Calvo, M., Veys, P., Vossen, J., Fasth, A., Heilmann, C., Wulffraat, N., Seger, R., Blanche, S., Friedrich, W., Abinun, M., Davies, G., Bredius, R., Schulz, A., Landais, P., Fischer, A., European Group for Blood and Marrow Transplantation & The European Society for Immunodeficiency (2003) Long-term survival and transplantation of haemopoietic stem cells for immunodeficiencies: report of the European experience 1968–99. Lancet, 361, 553560.
  • Arico, M., Janka, G., Fischer, A., Henter, J.I., Blanche, S., Elinder, G., Marinetti, M. & Rusca, M.P. (1996) Hemophagocytic lymphohistiocytosis. Report of 122 children from the International Registry. FHL Study Group of the Histiocyte Society. Leukemia, 10, 197203.
  • Aversa, F., Tabilio, A., Terenzi, A., Velardi, A., Falzetti, F., Giannoni, C., Lacucci, R., Zei, T., Martelli, M.P. & Gambelunghe, C. (1994) Successful engraftment of T-cell-depleted haploidentical ‘three-loci’ incompatible transplants in leukemia patients by addition of recombinant human granulocyte colony-stimulating factor-mobilized peripheral blood progenitor cells to bone marrow inoculm. Blood, 84, 39483955.
  • Aversa, F., Tabilio, A., Velardi, A., Cunningham, I., Terenzi, A., Falzetti, F., Ruggeri, L., Barbabietola, G., Aristei, C., Latini, P., Reisner, Y. & Martelli, M.F. (1998) Treatment of high-risk acute leukemia with T-cell-depleted stem cells from related donors with one fully mismatched HLA haplotype. The New England Journal of Medicine, 339, 11861193.
  • Aversa, F., Velardi, A., Tabilio, A., Reisner, Y. & Martelli, M.F. (2001) Haploidentical stem cell transplantation in leukemia. Blood Reviews, 15, 111119.
  • Bachar-Lusting, E., Rachmim, N., Li, H.W., Lan, F. & Reisner, Y. (1995) Megadose of T cell-depleted bone marrow overcomes MHC barriers in sublethally irradiated mice. Nature Medicine, 1, 12681273.
  • Barker, J.N., Davies, S.M., DeFor, T., Ramsay, N.K., Weisdorf, D.J. & Wagner, J.E. (2001) Survival after transplantation of unrelated donor umbilical cord blood is comparable to that of human leukocyted antigen-matched unrelated donor bone marrow: results of a matched-pair analysis. Blood, 97, 29572961.
  • Barker, J.N., Krepski, T.P., DeFor, T.E., Davies, S.M., Wagner, J.E. & Weisdorf, D.J. (2002) Searching for unrelated donor hematopoietic stem cells: availability and speed of umbilical cord blood versus bone marrow. Biology of Blood and Marrow Transplantation, 8, 257260.
  • Beatty, P.G., Clift, R.A., Mickelson, E.M., Nisperos, B.B., Flournoy, N., Martin, P.J., Sanders, J.E., Stewart, P., Buckner, C.D. & Storb, R. (1985) Marrow transplantation from related donors other than HLA-identical siblings. The New England Journal of Medicine, 313, 765771.
  • Beatty, P.G., Mori, M. & Milford, E. (1995) Impact of racial genetic polymorphism on the probability of finding an HLA-matched donor. Transplantation, 60, 778783.
  • Bonini, C., Ferrari, G., Verzeletti, S., Servida, P., Zappone, E., Ruggieri, L., Ponzoni, M., Rossini, S., Mavilio, F., Traversari, C. & Bordignon, C. (1997) HSV-RK gene transfer into donor lymphocytes for control of allogeneic graft-versus-leukemia. Science, 276, 17191724.
  • Boulad, F., Gillio, A., Small, T.N., George, D., Prasad, V., Torok-Castanza, J., Regan, A.D., Collins, N., Auerbach, A.D., Kernan, N.A. & O'Reilly, R.J. (2000) Stem cell transplantation for the treatment of Fanconi anaemia using a Fludarabine-based cytoreductive regimen and T-cell-depleted related HLA-mismatched peripheral blood stem cell grafts. British Journal of Haematology, 111, 11531157.
  • Boussiotis, V.A., Gribben, J.G., Freeman, E.J. & Nadler, L.M. (1994) Blockade of the CD 28 co-stimulatory pathway: a means to induce tolerance. Current Opinion in Immunology, 6, 797807.
  • Bozdech, M.J., Sondel, P.M., Trigg, M.E., Longo, W., Kohler, P.C., Flynn, B., Billing, R., Anderson, S.A., Hank, J.A. & Hong, R. (1985) Transplantation of HLA-haploidentical T-cell-depleted marrow for leukemia: addition of cytosine arabinoside to the pretransplant conditioning prevents rejection. Experimental Hematology, 13, 12011210.
  • Buckley, R.H., Schiff, S.E., Schiff, R.I., Markert, L., Williams, L.W., Roberts, J.L., Myers, L.A. & Ward, F.E. (1999) Hematopoietic stem-cell transplantation for the treatment of severe combined immunodeficiency. The New England Journal of Medicine, 340, 508516.
  • Bunjes, D., Duncker, C., Wiesneth, M., Stefanic, M., Dohr, D., Harsdorf, S., Karakas, T., Reske, S.N. & Bergmann, L. (2000) CD34+ selected cells in mismatched stem cell transplantation: a single center experience of haploidentical peripheral blood stem cell transplantation. Bone Marrow Transplantation, 25(Suppl. 2), S9S11.
  • Casper, J., Camitta, B., Truitt, R., Baxter-Lowe, L.A., Bunin, N., Lawton, C., Murray, K., Hunter, J., Pietryga, D. & Garbrecht, F. (1995) Unrelated bone marrow donor transplants for children with leukaemia or myelodysplasia. Blood, 85, 23542363.
  • Champlin, R.E., Ho, W.G., Mitsuyasu, R., Burnison, M., Greenberg, P., Holly, G., Winston, D.W., Feig, S.A. & Gale, R.P. (1987) Graft failure and leukemia relapse following T lymphocyte-depleted bone marrow transplants: effect of intensification of immunosuppressive conditioning. Transplant Procedures, 19, 26162619.
  • Chen, B.J., Cui, X., Liu, C. & Chao, N.J. (2002) Prevention of graft-versus-host disease while preserving graft-versus-leukemia effect after selective depletion of host reactive T cells by photodynamic cell purging process. Blood, 99, 30833088.
  • Collins, P., Watts, M., Brocklesby, M., Gerritsen, B. & Veys, P. (1997) Sucessful engraftment of haploidentical stem cell transplant for familial haemophagocytic lymphohistiocytosis using both bone marrow and peripheral blood stem cells. British Journal of Haematology, 96, 644646.
  • Cornish, J., Goulden, N. & Potter, M. (1998) Unrelated donor bone-marrow transplantation. In: The Clinical Practice of Stem Cell Transplantation (ed. by J.G.Barrett & J.C. Treleaven), pp. 363391. Isis Medical Media Ltd, Oxford.
  • Davies, S.M., Kollman, C., Anasetti, C., Antin, J.H., Gajewski, J., Casper, J.T., Nademanee, A., Noreen, H., King, R., Confer, D. & Kernan, N.A. (2000) Engraftment and survival after unrelated-donor bone marrow transplantation: a report from the national marrow donor program. Blood, 96, 40964102.
  • Drobyski, W.R., Klein, J., Flomenberg, N., Pietryga, D., Vesole, D.H., Margolis, D.A. & Keever-Taylor, C.A. (2002) Superior survival associated with transplantation of matched unrelated versus one-antigen-mismatched unrelated or highly human leukocyte antigen-disparate haploidentical family donor marrow grafts for the treatment of hematologic malignancies: establishing a treatment algorithm for recipients of alternative donor grafts. Blood, 99, 806814.
  • Elhasid, R., Weyl, M., Arush, B., Katz, T., Gan, Y., Shechter, Y., Sami, I., Postovsky, S., Reisner, Y. & Rowe, J.M. (2000) Successful haploidentical bone marrow transplantation in Fanconi anemia. Bone Marrow Transplantation, 26, 12211223.
  • Eyrich, M., Lang, P., Lal, S., Bader, P., Handgretinger, R., Klingebiel, T., Niethammer, D. & Schlegel, P.G. (2001) A prospective analysis of the pattern of immune reconstitution in a paediatric cohort following transplantation of positively selected human leucocyte antigen-disparate haematopoietic stem cells from parental donors. British Journal of Haematology, 114, 422432.
  • Farag, S.S., Fehniger, T.A., Ruggeri, L., Velardi, A. & Caligiuri, M.A. (2002) Natural killer cell receptors: new biology and insights into the graft-versus-leukemia effect. Blood, 100, 19351947.
  • Fischer, A., Griscelli, C., Blanche, S., Le Deist, F., Veber, F., Lopez, M., Delaage, M., Olive, D., Mawas, C. & Janossy, G. (1986) Prevention of graft failure by an anti-HLFA-1 monoclonal antibody in HLA-mismatched bone marrow transplantation. Lancet, 2, 10581061.
  • Fischer, A., Landais, P., Friedrich, W., Morgan, G., Gerritsen, B., Fasth, A., Porta, F., Griscelli, C., Goldman, S.F., Levinsky, R. & Vossen, J. (1990) European experience of bone marrow transplantation for severe combined immunodeficiency. Lancet, 2, 850854.
  • Gao, L., Bellantuono, I., Elsasser, A., Marley, S.B., Gordon, M.Y., Goldman, J.M. & Stauss, H.J. (2000) Selective elimination of leukemic CD34(+) progenitor cells by cytotoxic T lymphocytes specific for WT1. Blood, 95, 21982203.
  • Garin, M.I., Garrett, E., Tiberghien, P., Apperley, J.F., Chalmers, D., Melo, J.V. & Ferrand, C. (2001) Molecular mechanism for ganciclovir resistance in human T lymphocytes transduced with retroviral vectors carrying herpes simplex virus thymidine kinase gene. Blood, 97, 122129.
  • Gaziev, D., Galimberti, M., Lucarelli, G., Polchi, P., Giardini, C., Angelucci, E., Baronciani, D., Sodani, P., Erer, B., Biagi, M.D., Andreani, M., Agostinel, F., Donati, M., Nesci, S. & Talevi, N. (2000) Bone marrow transplantation from alternative donors for thalassemia: HLA-phenotypically identical relative and HLA nonidentical sibling or parent transplants. Bone Marrow Transplantation, 25, 815821.
  • Gluckman, E., Rocha, V., Boyer-Chammard, A., Locatelli, F., Arcese, W., Pasquini, R., Ortega, J., Souillet, G., Ferreira, E., Laporte, J.P., Fernandez, M. & Chastang, C. (1997) Outcome of cord-blood transplantation from related and unrelated donors. The New England Journal of Medicine, 337, 373381.
  • Gribben, J.G., Guinan, E.C., Boussiotis, V.A., Ke, X.Y., Linsley, L., Sieff, C., Gray, G.S., Freeman, G.J. & Nadler, L.M. (1996) Complete blockade of B7 family-mediated costimulation is necessary to induce human alloantigen-specific anergy: a method to ameliorate graft-versus-host disease and extend the donor pool. Blood, 87, 48874893.
  • Guimond, M., Balassy, A., Barrette, M., Brochu, S., Perreault, C. & Roy, D.C. (2002) P-glycoprotein targeting: a unique strategy to selectively eliminate immunoreactive T cells. Blood, 100, 375382.
  • Guinan, E.C., Boussiotis, V.A., Neuberg, D., Brennan, L.L., Hirano, N., Nadler, L.M. & Gribben, J.G. (1999) Transplantation of anergic histoincompatible bone marrow allografts. The New England Journal of Medicine, 340, 17041714.
  • Handgretinger, R., Schumm, M., Lang, P., Greil, J., Reiter, A., Bader, P., Niethammer, D. & Klingebiel, T. (1999) Transplantation of megadoses of purified haploidentical stem cells. Annual New York Academy of Science, 872, 351361.
  • Handgretinger, R., Klingebiel, T., Lang, P., Schumm, M., Neu, S., Geiselhart, A., Bader, P., Schlegel, P.G., Greil, J., Stachel, D., Herzog, R.J. & Niethammer, D. (2001) Megadose transplantation of purified peripheral blood CD34(+) progenitor cells from HLA-mismatched parental donors in children. Bone Marrow Transplantation, 27, 777783.
  • Handgretinger, R., Leimig, T., Babarin-Dorner, A., Holladay, M.A., Houston, J. & Gordon, P. (2002) The potential role of graft engineering strategies in haploidentical stem cell transplantation. Bone Marrow Transplantation, 30, S11.
  • Hansen, J.A., Petersdorf, E., Martin, P.J. & Anasetti, C. (1997) Hematopoietic stem cell transplants from unrelated donors. Immunology Review, 157, 141151.
  • Henslee-Downey, P.J., Parrish, R.S., MacDonald, J.S., Romond, E.H., Marciniak, E., Coffey, C., Ciocci, G. & Thompson, J.S. (1996) Combined in vitro and in vivo T lymphocyte depletion for the control of graft-versus-host disease following haploidentical marrow transplant. Transplantation, 61, 738745.
  • Henslee-Downey, P.J., Abhyankar, S.H., Parrish, R.S., Pati, A.R., Godder, K.T., Neglia, W.J., Goon-Johnson, K.S., Geier, S.S., Lee, C.G. & Gee, A.P. (1997) Use of partially mismatched related donors extends access to allogeneic marrow transplant. Blood, 89, 38643872.
  • Ho, V.T. & Soiffer, R.J. (2001) The history and future of T-cell depletion as graft-versus-host disease prophylaxis for allogeneic hematopoietic stem cell transplantation. Blood, 98, 31923204.
  • Horowitz, M.M. (1999) Uses and growth of hematopoietic cell transplantation, Hematopoietic cell transplantation. Blackwell Science, pp. 1218.
  • Jabado, N., de Graeff-Meeder, E.R., Cavazzana-Calvo, M., Haddad, E., Le Deist, F., Benkerrou, M., Dufourcq, R., Caillat, S., Blanche, S. & Fischer, A. (1997) Treatment of familial hemophagocytic lymphohistiocytosis with bone marrow transplantation from HLA genetically nonidentical donors. Blood, 90, 47434748.
  • Kawano, Y., Takaue, Y., Watanabe, A., Takeda, O., Arai, K., Itoh, E., Ohno, Y., Teshima, T., Harada, M., Watanabe, T., Okamoto, Y., Abe, T., Kajiume, T., Matsushita, T., Ikeda, K., Endo, M., Kuroda, Y., Asano, S., Tanosaki, R., Yamaguchi, K., Law, P. & McMannis, J.D. (1998) Partially mismatched paediatric transplants with allogeneic CD34(+) blood cells from a related donor. Blood, 92, 31233872.
  • Kernan, N.A., Flomenberg, N., Dupont, B. & O'Reilly, R.J. (1987) Graft rejection in recipients of T-cell-depleted HLA-nonidentical marrow transplants for leukemia. Identification of host-derived antidonor allocytotoxic lymphocytes. Transplantation, 43, 842847.
  • Kernan, N.A, Emanuel, D. & Castro-Malaspina, H. (1989) Post transplant immunosuppression with antithymocyte globulin and methylprednisolone prevents immunulogically mediated graft failure following a T-cell depleted marrow transplant. Blood, 74, 123a.
  • Klingebiel, T.E., Lang, P., Schumm, M., Bader, P., Schiegel, P.G., Eyrich, M., Greil, D., Niethammer, D. & Handgretinger, R. (2000) Current results in haploidentical stem cell transplantation in children. Bone Marrow Transplantation, 25(Suppl. 1), S1.
  • Koh, M.B., Prentice, H.G., Corbo, M., Morgan, M., Cotter, F.E. & Lowdell, M.W. (2002) Alloantigen-specific T-cell depletion in a major histocompatibility complete fully mismatched murine model provides effective graft-versus-host disease prophylaxis in the presence of lymphoid engraftment. British Journal of Haematology, 118, 108116.
  • Kojima, S., Horibe, K., Inaba, J., Yoshimi, A., Takahashi, Y., Kudo, K., Kato, K. & Matsuyama, T. (2000) Long-term outcome of acquired aplastic anaemia in children: comparison between immunosuppressive therapy and bone marrow transplantation. British Journal of Haematology, 111, 321328.
  • Kook, H., Goldman, F., Padley, D., Giller, R., Rumelhart, S., Holida, M., Lee, N., Peters, C., Comito, M., Huling, D. & Trigg, M. (1996) Reconstruction of the immune system after unrelated or partially matched T-cell-depleted bone marrow transplantation in children: immunophenotypic analysis and factors affecting the speed of recovery. Blood, 88, 10891097.
  • Kremens, B., Basu, O., Grosse-Wilde, H., Sauerwien, W., Schaefer, U.W. & Havers, W. (2001) Transplantation of CD34-enriched peripheral stem cells from an HLA-haplotype mismatched donor to a patient with severe aplastic anemia. Bone Marrow Transplantation, 27, 111113.
  • Kurtzberg, J., Laughlin, M., Graham, M.L., Smith, C., Olson, J.F., Halperin, E.C., Ciocci, G., Carrier, C., Stevens, C.E. & Rubinstein, P. (1996) Placental blood as a source of hematopoietic stem cells for transplantation into unrelated recipients. The New England Journal of Medicine, 335, 157166.
  • La Nasa, G., Giardini, C., Ariolu, F., Locatelli, F., Arras, M., De Stefano, P., Ledda, A., Pizzati, A., Sanna, M.A., Vacca, A., Lucarelli, G. & Contu, L. (2002) Unrelated donor bone marrow transplantation for thalassemia: the effect of extended haplotypes. Blood, 99, 43504356.
  • Lapidot, T., Terenzi, A., Singer, T.S., Salomon, O. & Reisner, Y. (1989) Enhancement by dimenthyl myleran of donor type chimerism in murine recipients of bone marrow allografts. Blood, 73, 20252032.
  • Lawson, S.E., Harrison, G., Richards, S., Oakhill, A., Stevens, R., Eden, O.B. & Darbyshire, P.J. (2000) The UK experience in treating relapsed childhood acute lymphoblastic leukaemia: a report on the medical research council UKALLR1 study. British Journal of Haematology, 108, 531543.
  • Leiper, A.D. (2002) Non-endocrine late complications of bone marrow transplantation in childhood: Part II. British Journal of Haematology, 118, 2343.
  • Luznik, L., Slansky, J.E., Jalla, S., Borrello, I., Levitsky, H.I., Pardoll, D.M. & Fuchs, E.J. (2003) Successful therapy of metastatic cancer using tumour vaccines in mixed a llogeneic bone marrow chimeras. Blood, 101, 16451652.
  • Malilay, G.P., Sevenich, E.A., Condie, R.M. & Filipovich, A.H. (1989) Prevention of graft rejection in allogeneic bone marrow transplantation: I. Preclinical studies with antithymocyte globulins. Bone Marrow Transplantation, 4, 107112.
  • Marmont, A.M., Horowitz, M.M., Gale, R.P., Sobocinski, K., Ash, R.C., van Bekkum, D.W., Champlin, R.E., Dicke, K.A., Goldman, J.M. & Good, R.A. (1991) T-cell depletion of HLA-identical transplants in leukemia. Blood, 78, 21202130.
  • Matthes-Martin, S., Peters, C., Lawatschka, A., Fritsch, G., Lion, T., Haas, O.A. & Gadner, H. (2002) Reduced intensity conditioning in paediatric stem cell transplantation. Bone Marrow Transplantation, 30(Suppl. 1), S12S14.
  • Montagna, D., Yvon, E., Calcaterra, V., Comoli, P., Locatelli, F., Maccario, R., Fisher, A. & Cavazzana-Calvo, M. (1999) Depletion of alloreactive T cells by a specific anti-interleukin-2 receptor p55 chain immunotoxin does not impair in vitro antileukemia and antiviral activity. Blood, 93, 35503557.
  • Moretta, A. & Moretta, L. (1997) HLA class I specific inhibitory receptors. Current Opinions in Immunology, 9, 694701.
  • Mutis, T., Blokland, E., Kester, M., Schrama, E. & Goulmy, E. (2002) Generation of minor histocompatibility antigen HA-1-specific cytotoxic T cells restricted by nonself HLA molecules: a potential strategy to treat relapsed leukemia after HLA-mismatched stem cell transplantation. Blood, 100, 547552.
  • O'Donnell, P.V., Luznik, L., Jones, R.J., Vogelsang, G.B., Leffell, M.S., Phelps, M., Rhubart, P., Cowan, K., Piantados, S. & Fuchs, E.J. (2002) Nonmyeloablative bone marrow transplantation from partial HLA-mismatched related donors using posttransplantation cyclophosphamide. Bone Marrow Transplantation, 8, 377386.
  • Oakhill, A., Pamphilon, D.H., Potter, M.N., Steward, C.G., Goodman, S., Green, A., Goulden, P., Goulden, N.J., Hale, G., Waldmann, H. & Cornish, J.M. (1996) Unrelated donor bone marrow transplantation for children with relapsed acute lymphoblastic leukaemia in second complete remission. British Journal of Haematology, 94, 574578.
  • Ochs, L., Shu, X.O., Miller, J., Enright, H., Wagner, J., Filipovich, A., Miller, W. & Weisdorf, D. (1995) Late infections after allogeneic bone marrow transplantation: comparison of incidence in related and unrelated donor transplant recipients. Blood, 10, 39793986.
  • Ortín, M., Raj, R., Kinning, E., Williams, M. & Darbyshire, P.J. (2002) Partially matched related donor peripheral blood progenitor cell transplantation in paediatric patients adding Fludarabine and anti-lymphocyte gamma-globulin. Bone Marrow Transplantation, 30, 359366.
  • Pagel, J.M., Matthews, D.C., Appelbaum, F.R., Bernstein, I.D. & Press, O.W. (2002) The use of radioimmunoconjucates in stem cell transplantation: mini review. Bone Marrow Transplantation, 29, 807816.
  • Passweg, J.R., Kuhne, T., Gregor, M., Favre, G., Avoledo, P., Tichelli, A. & Gratwohl, A. (2000) Increased stem cell dose, as obtained using currently available technology, may not be sufficient for engraftment of haploidentical stem cell transplants. Bone Marrow Transplantation, 26, 10331036.
  • Perruccio, K., Tosti, A., Posati, S., Carotti, A., Burchielli, E., Aversa, F., Martelli, M.F. & Velardi, A. (2001) Transfer of functional immune responses to Aspergillus and CMV after haploidentical hematopoietic transplantation. Bone Marrow Transplantation, 27, OS102 (abstract).
  • Peters, C., Matthes-Martin, S., Fritsch, G., Holter, W., Lion, T., Witt, V., Hocker, P., Fischer, G., Dieckmann, K., Handgretinger, R., Klingebiel, T. & Gadner, H. (1999) Transplantation of highly purified peripheral blood CD34+ cells from HLA-mismatched parental donors in 14 children: evaluation of early monitoring of engraftment. Leukemia, 13, 20702078.
  • Quesenberry, P.J., Zhong, S., Wang, H. & Stewart, M. (2001) Allogeneic chimerism with low-dose irradiation, antigen presensitization, and costimulator blockade in H-2 mismatched mice. Blood, 97, 557564.
  • Reich-Zeliger, S., Zhao, Y., Krauthgamer, R., Bachar-Lustig, E. & Reisner, Y. (2000) Anti-third party CD8+ CTLs as potent veto cells: coexpression of CD8 and FasL is a prerequisite. Immunity, 13, 507515.
  • Reisner, Y. & Martelli, M.F. (1995) Bone marrow transplantation across HLA barriers by increasing the number of transplanted cells. Immunology Today, 16, 437440.
  • Reisner, Y. & Martelli, M.F. (2000) Tolerance induction by ‘megadose’ transplants of CD34+ stem cells: a new option for leukemia patients without an HLA-matched donor. Current Opinions in Immunology, 12, 536541.
  • Reisner, Y., Kapoor, N., Kirkpatrick, D., Pollack, M.S., Cunningham-Rundles, S., Dupont, B., Hodes, M.Z., Good, R.A. & O'Reilly, R.J. (1983) Transplantation for severe combined immunodeficiency with HLA-A, B, D, DR incompatible parental marrow cells fractionated by soybean agglutinin and sheep red blood cells. Blood, 61, 341348.
  • Reisner, Y., Ben-Bassat, I., Douer, D., Kaploon, A., Schwartz, E. & Ramot, B. (1986) Demonstration of clonable alloreactive host T cells in a primate model for bone marrow transplantation. Proceedings of the National Accademy of Sciences of the United States of America, 83, 40124015.
  • Rocha, V., Cornish, J., Sievers, E.L., Filipovich, A., Locatelli, F., Peters, C., Remberger, M., Michel, G., Arcese, W., Dallorso, S., Tiedemann, K., Busca, A., Chan, K.W., Kato, S., Ortega, J., Vowels, M., Zander, A., Souillet, G., Oakill, A., Woolfrey, A., Pay, A.L., Green, A., Garnier, F., Ionescu, I., Werne, P., Sirchia, G., Rubinstein, P., Chevret, S. & Gluckman, E. (2001) Comparison of outcomes of unrelated bone marrow and umbilical cord blood transplants in children with acute leukemia. Blood, 97, 29622971.
  • Rooney, C.M., Smith, C.A., Ng, C.Y., Loftin, S.K., Sixbey, J.W., Gan, Y., Srivastava, D.K., Bowman, L.C., Krance, R.A., Brenner, M.K. & Heslop, H.E. (1998) Infusion of cytotoxic T cells for the prevention and treatment of Epstein-Barr virus-induced lymphoma in allogeneic transplant recipients. Blood, 92, 15491555.
  • Roux, E., Dumont-Girard, F., Starobinski, M., Siegrist, C.A., Helg, C., Chapuis, B. & Roosnek, E. (2000) Recovery of immune reactivity after T-cell-depleted bone marrow transplantation depends on thymic activity. Blood, 96, 22992303.
  • Rowe, J.M. & Lazarus, H.M. (2001) Genetically haploidentical stem cell transplantation for acute leukemia. Bone Marrow Transplantation, 27, 669676.
  • Rubinstein, P., Carrier, C., Scaradavou, A., Kurtzberg, J., Adamson, J., Migliaccio, A.R., Berkowitz, R.L., Cabbad, M., Dobrila, N.L., Taylor, P.E., Rosenfield, R.E. & Stevens, C.E. (1998) Outcomes among 562 recipients of placental-blood transplants from unrelated donors. The New England Journal of Medicine, 339, 15651577.
  • Ruggeri, L., Capanni, M., Casucci, M., Volpi, I., Tosti, A., Perruccio, K., Urbana, E., Negrin, R.S., Martelli, M.F. & Velardi, A. (1999) Role of natural killer cell alloreactivity in HLA-mismatched hematopoietic stem cell transplantation. Blood, 94, 333339.
  • Ruggeri, L., Capanni, M., Urbani, E., Perruccio, K., Shlomchik, W.D., Tosti, A., Posati, S., Rogaia, D., Frassoni, F., Aversa, F., Martelli, M.F. & Velardi, A. (2002) Effectiveness of donor natural killer cell alloreactivity in mismatched hematopoietic transplants. Science, 295, 20972100.
  • Sambhara, S.R. & Miller, R.G. (1991) Programmed cell death of T cells signaled by the T cell receptor and the α 3 domain of class I MHC. Science, 252, 14241427.
  • Sauce, D., Bodinier, M., Garin, M., Petracca, B., Tonnelier, N., Duperrier, A., Melo, J.V., Apperley, J.F., Ferrand, C., Herve, P., Lang, F., Tiberghien, P. & Robinet, E. (2002) Retrovirus-mediated gene transfer in primary T lymphocytes impairs their anti-Epstein-Barr virus potential through both culture-dependent and selection process-dependent mechanisms. Blood, 99, 11651173.
  • Schumm, M., Lang, P., Taylor, G., Duci, S., Klingebiel, T., Buhring, H.J., Geiselhart, A., Niethammer, D. & Handgretinger, R. (1999) Isolation of highly purified autologous and allogeneic peripheral CD34+ cells using the CliniMACS device. Journal of Hematotherapy, 8, 209218.
  • Shapiro, R.S., McClain, K., Frizzera, G., Gajl-Peczalska, K.J., Kersey, J.H., Blazar, B.R., Arthur, D.C., Patton, D.F., Greenberg, J.S. & Burke, B. (1988) Epstein-Barr virus associated B cell lymphoproliferative disorders following bone marrow transplantation. Blood, 71, 12341243.
  • Shlomchik, W.D., Couzens, M.S., Tang, C.B., McNiff, J., Robert, M.E., Liu, J., Shlomchik, M.J. & Emerson, S.G. (1999) Prevention of graft versus host disease by inactivation of host antigen-presenting cells. Science, 285, 412415.
  • Small, T.N., Papadopoulos, E.B., Boulad, F., Black, P., Castro-Malaspina, H., Childs, B.H., Collins, N., Gillio, A., George, D., Jakubowski, A., Heller, G., Fazzari, M., Kernan, N., MacKinnon, S., Szabolcs, P., Young, J.W. & O'Reilly, R.J. (1999) Comparison of immune reconstitution after unrelated and related T-cell-depleted bone marrow transplantation: effect of patient age and donr leukocyte infusions. Blood, 93, 467480.
  • Soiffer, R.J., Mauch, P., Tarbell, N.J., Anderson, K.C., Freedman, A.S., Rabinowe, S.N., Takvorian, T., Murrey, C.I., Coral, F., Bosserman, L. Dear, K., Nadler, L.M. & Ritz, J. (1991) Total lymphoid irradiation to prevent graft rejection in recipients of HLA non-identical T cell-depleted allogeneic marrow. Bone Marrow Transplantation, 7, 2333.
  • Speiser, D.E., Hermans, J., Van Biezen, A., Starobinski, M., Jeannet, M., Jacobsen, N., Niederwieser, D., Gratwohl, A. & Apperley, J.F. (1997) Haploidentical family member transplants for patients with chronic myeloid leukaemia: a report of the Chronic Leukaemia Working Party of European Group for Blood and Marrow Transplantation (EBMT). Bone Marrow Transplantation, 19, 11971203.
  • Taylor, P.A., Friedman, T.M., Korngold, R., Noelle, R.J. & Blazar, B.R. (2002) Tolerance induction of alloreactive T cells via ex vivo blockade of the CD40:CD40L costimulatory pathway results in the generation of a potent immune regulatory cell. Blood, 99, 46014609.
  • Terenzi, A., Lubin, I., Lapidot, T., Salomon, O., Faktorowich, Y., Rabi, I., Martelli, M.F. & Reisner, Y. (1990) Enhancement of T-cell depleted bone marrow allografts in mice by thiotepa. Transplantation, 50, 717720.
  • Tiberghien, P., Ferrand, C., Lioure, B., Milpied, N., Angonin, R., Deconinck, E., Certoux, J.M., Robinet, E., Saas, P., Petracca, B., Juttner, C., Reynolds, C.W., Longo, D.L., Herve, P. & Cahn, J.Y. (2001) Administration of herpes simplex-thymidine kinase-expressing donor T cells with a T-cell-depleted allogeneic marrow graft. Blood, 97, 6372.
  • Uderzo, C., Conter, V., Dini, G., Locatelli, F., Miniero, R. & Tamaro, P. (2001) Treatment of childhood acute lymphoblastic leukaemia after the first relapse: curative strategies. Haematologica, 86, 17.
  • Uharek, L., Glass, B., Gassmann, W., Eckstein, V., Steinmann, J., Loeffler, H. & Mueller-Ruchholtz, W. (1992) Engraftment of allogeneic bone marrow cells: experimental investigations on the role of cell dose, graft-versus-host reactive T cells and pretransplant immunosuppression. Transplant Proceedings, 24, 30233025.
  • Van Rood, J.J., Loberiza, Jr, F.R., Zhang, M.J., Oudshoorn, M., Claas, F., Ca, M.S., Champlin, R.E., Gale, R.P., Ringden, O., Hows, J.M. & Horowitz, M.H. (2002) Effect of tolerance to noninherited maternal antigens on the occurrence of graft-versus-host disease after bone marrow transplantation from a parent or an HLA-haploidentical sibling. Blood, 99, 15721577.
  • Vassiliou, G.S., Webb, D.K.H., Pamphilon, D. & Veys, P.A. (2001) Improved outcome of alternative donor bone marrow transplantation in children with sever aplastic anaemia using a conditioning regimen containing low-dose total body irradiation, Cyclophosphamide and Campath. British Journal of Haematology, 114, 701705.
  • Veys, P., Gaspar, H., Amrolia, P., Rao, K., Davies, G., Jones, A., Wynn, R., Will, A. & Goulden, N. (2002) Alternative donor stem cell transplantation using reduced intensity conditioning. Bone Marrow Transplantation, 30(Suppl. 1), S14S15.
  • Volpi, I., Peruccio, K., Tosti, A., Capanni, M., Ruggeri, L., Posati, S., Aversa, F., Tabilio, A., Romani, L., Martelli, M.F. & Velardi, A. (2001) Post-grafting granulocyte colony-stimulating factor administration impairs functional immune recovery in recipients of HLA haplotype-mismatched hematopoietic transplants. Blood, 97, 25142521.
  • Wagner, J.E., Rosenthal, J., Sweetman, R., Shu, X.O., Davies, S.M., Ramsay, N.K., McGlave, P.B., Sender, L. & Cairo, M.S. (1996) Successful transplantation of HLA-matched and HLA-mismatched umbilical cord blood from unrelated donors: analysis of engraftment and acute graft-versus-host disease. Blood, 88, 795802.
  • Walter, E.A., Greenberg, P.D., Gilbert, M.J., Finch, R.J., Watanabe, K.S., Thomas, E.D. & Riddell, S.R. (1995) Reconstitution of cellular immunity against cytomegalovirus in recipients of allogeneic bone marrow by transfer of T-cell clones from the donor. New England Journal of Medicine, 333, 10381044.
  • Weinberg, K., Blazar, B.R., Wagner, J.E., Agura, E., Hill, B.J., Smogorzewska, M., Koup, R.A., Betts, M.R., Collins, R.H. & Douek, D.C. (2001) Factors affecting thymic function after allogeneic hematopoietic stem cell transplantation. Blood, 97, 14581466.
  • Wulf, G.G., Luo, K.L., Goodell, M.A. & Brenner, M.K. (2003) Anti-CD45-mediated cytoreduction to facilitate allogeneic stem cell transplantation. Blood, 101, 24342439.
  • Yin, A.H., Miraglia, S., Zanjani, E.D., Almeida-Porada, G., Ogawa, M., Leary, A.G., Olweus, J., Kearney, J. & Buck, D.W. (1997) AC133, a novel marker for human hematopoietic stem and progenitor cells. Blood, 90, 50025012.
  • Zanjani, E.D., Almeida-Porada, G., Livingston, A.G., Flake, A.W. & Ogawa, M. (1998) Human bone marrow CD34-cells engraft in vivo and undergo multilineage expression that includes giving rise to CD34+ cells. Experimental Hematology, 26, 353360.