Allogeneic Bone Marrow Infusion Research Articles

Posttransplant cyclophosphamide in allogeneic bone marrow transplantation for the treatment of nonmalignant hematological diseases.

We present a single-center retrospective series of allogeneic bone marrow transplantation (BMT) with the use of posttransplant cyclophosphamide (PTCy) in the setting of nonmalignant hematological conditions. Nine patients were treated between 2013 and 2019. Nonmyeloablative conditioning consisted of antithymocyte globulin, fludarabine, low-dose cyclophosphamide, and total body irradiation (200cGy) followed by allogeneic bone marrow infusion. Post-BMT GVHD prophylaxis was with PTCy, tacrolimus, and mycophenolate mofetil. At a median follow-up of 24 months (range 4, 63), all patients are alive, with donor-derived hematopoiesis and free of significant acute or chronic GVHD. Donors were haploidentical (n = 6), fully matched unrelated (n = 2), and fully matched sibling (n = 1). Neutrophil and platelet engraftment occurred at a median of 21 days and 33 days, respectively, after transplantation. Three patients (3/9, 33%) experienced stage 1-2 acute skin GVHD. The only cases of chronic GVHD are in three patients (3/9, 33%) with ocular disease (two mild, one moderate). No patient has required systemic immunosuppression beyond 12 months after BMT. PTCy-based nonmyeloablative allogeneic BMT is safe and effective for nonmalignant hematologic conditions and should be prospectively compared with historical regimens.

Safety of hematopoietic cell infusion in children with malignant and non-malignant diseases.

HPC infusions have been associated with a variety of adverse events related to either patient or HPC product-related factors. Studies documenting infusion-related AEs in children are limited. We reviewed HPC infusion records in 354 children. Infusion-related adverse events were classified as follows: grade 0-absent, grade I-mild, grade II-moderate, grade III-severe, grade IV-life-threatening, and grade V-death. The percentage of patients with grade 0, I, and II-IV AEs was as follows: 0=67%, I=23.4%, and II-V=9.6% (one patient had fatal anaphylactic reaction to dimethyl sulfoxide). The incidence of grade II-IV hypertension was 7.1%. There was a higher incidence of AEs with infusion of allogeneic bone marrow versus allogeneic PBSCs (47.4% vs 25.3%, P=.001). Cryopreserved products had a lower incidence of infusion-associated AEs compared with fresh HPC products (24% vs 39.4%, P=.003). Allogeneic HPC infusion volume (>100mL) was a significant risk factor for infusion-associated AEs (P<.001). Patients >10years who received autologous HPC infusions had higher risk of AEs when compared to patients <10years (P=.01). Our study demonstrated that despite a high incidence of infusion-associated hypertension, HPC infusion is relatively safe in children. Investigating strategies to optimize management of hypertension in the setting of HPC infusion is warranted.

(92) - Results of a First-in-Man Study of Mesenchymal Stem Cell Therapy for Bronchiolitis Obliterans Syndrome

s S43 Recipients of larger volumes of MSC (2 and 4 million cells/kg) have shown no improvement in FEV1 but have only 2 months follow at the time of this report (See Figure 2). Conclusion: Preliminary results from this study suggest a beneficial effect on lung function derived from low volume infusion of allogeneic bone-marrow derived MSC. This effect may not occur on recipients of larger volumes of stem cells but longer follow up is needed.

The absence of Peyer’s patches or host TNF-A does not affect GVHD progression in an allogeneic BMT model using extensive conditioning regimens

Peyer’s patches (PP) have been previously shown to play a critical role in the initiation of acute lethal GVHD in a non-irradiated murine model (Murai et al., 2003). This report utilized IL-7 receptor antibody treatment of pregnant mice to inhibit the development of PPs in the fetus. We wanted to assess GVHD progression in mice congenitally deficient in PP presence and function. The absence of various members of the TNF and TNF-R families also affects secondary lymphoid structure and organization. The presence of PP has varied between different independently generated TNF knockout strains which may be related to the close linkages of tnf with lta and ltb genes. A new TNF deficient strain has been developed that lacks PPs, displays the defects characteristic of TNF ablation but not the LT associated defects characterized by lack of lymph nodes and defects in splenic microarchitecture. To examine the role of host-derived TNF family members in acute lethal GVHD, we also had to assess the role of secondary lymphoid structures. We utilized a full MHC mismatched mode of BALB/c (H2d) cells into myeloablated C57BL/6 (H2b) recipients. We lethally irradiated wild type (WT) mice, TNF-α deficient mice with Peyer’s patches (PP+) and TNF-α deficient mice without Peyer’s patches (PP-) followed by infusion of allogeneic bone marrow and spleen cells. We observed no difference in the survival of the three groups of mice (median day of death ranged from 26 to 27). In addition, no significant differences were observed in GVHD associated histopathological lesions in the small intestine. Hisopathological lesions observed in the colon were similar in both the PP+ and PP- TNF-α deficient recipients. Based on our observations, we conclude that (1) Peyer’s patches and (2) host TNF-α are not required for the development of acute lethal GVHD in mice that received extensive conditioning and allogeneic bone marrow transplantation.

The contribution of cytotoxic and noncytotoxic function by donor T-cells that support engraftment after allogeneic bone marrow transplantation

The present studies were designed for investigation of the requirements for cytotoxic function in donor T-cells transplanted to support engraftment after infusion of allogeneic bone marrow. The experiments examined the capacity of donor CD8 T-cells lacking Fas ligand and/or perforin function to facilitate donor B6 congenic (B6-Ly5.1) BM engraftment across major histocompatibility complex class I/II barriers after transplantation. T-cell-depleted BM cells from B6-Ly5.1 donors were transplanted into sublethally irradiated (5.5 Gy) BALB/c recipients together with different lymphocyte populations from wild-type B6 (B6-wt) donors or donors lacking functional cytotoxic pathways. Early presence of lineage-committed donor progenitor cells was assessed by the presence of day 5 splenic colony-forming units-granulocyte-macrophage (CFU-GM). Recipients of BMT without donor T-cells did not demonstrate significant CFU-GM activity 5 days post-BMT. Lineage-committed progenitor cells in recipient spleens could be supported by addition to the BM of wild-type (B6-wt) and cytotoxically single- (perforin, B6-pko or FasL, B6-gld) or double-deficient (B6-cdd) CD8 T-cells. However, B220+-enriched B-cells could not support the presence of day 5 donor CFU-GM. For further assessment of the capacity of cytotoxically impaired T-cells to participate in the engraftment process, the ability of these and normal CD8 cells to support the homing of donor cells to the BM was examined after infusion of carboxyfluorescein diacetete succinimidyl ester-labeled progenitors. In a syngeneic model lacking resistance, cytotoxically impaired donor T-cells supported increased numbers of progenitor cells in the marrow equivalent to the support provided by wild-type donor T-cells. Examination of peripheral chimerism indicated that during the first month after B6-->BALB/c BMT, donor chimerism was detected in BMT recipients receiving unfractionated T-cells or CD8+ T-cells from B6-wt donors, and chimerism was maintained at least 80 days after BMT. In contrast, B6-cdd unfractionated or CD8+ T-cells failed to maintain long-term B6 donor chimerism in the host. Experiments with highly enriched populations of positively selected CD8+ T-cells from B6-pko, B6-gld, or B6-cdd donors demonstrated that although each of these T-cell populations could promote the initial presence of donor CFU-GM early post-BMT, B6-pko and B6-cdd CD8+ T-cell populations were not able to support long-term peripheral chimerism. These results demonstrate that donor T-cells lacking major cytotoxic effector pathways have functions that support initial donor progenitor cell presence in the host hematopoietic compartment after BMT. They also demonstrate that support of long-term donor BM engraftment requires CD8+ T-cells with intact cytotoxic, that is, perforin, function. Finally, syngeneic B6-->B6 BMT suggests activation of CD8+ T-cells posttransplantation apparently is required to support enhanced progenitor cell activity. This study provides new findings concerning the role of cytotoxic function in the process of facilitating allogeneic donor BM engraftment.

Predominance of NK1.1+TCR alpha beta+ or DX5+TCR alpha beta+ T cells in mice conditioned with fractionated lymphoid irradiation protects against graft-versus-host disease: "natural suppressor" cells.

We developed a nonmyeloablative host conditioning regimen in a mouse model of MHC-mismatched bone marrow transplantation that not only reduces radiation toxicity, but also protects against graft-vs-host disease. The regimen of fractionated irradiation directed to the lymphoid tissues and depletive anti-T cell Abs results in a marked change in the residual host T cells, such that NK1.1+ or DX5+asialo-GM1+ T cells become the predominant T cell subset in the lymphoid tissues of C57BL/6 and BALB/c mice, respectively. The latter "natural suppressor" T cells protect hosts from graft-vs-host disease after the infusion of allogeneic bone marrow and peripheral blood cells that ordinarily kill hosts conditioned with sublethal or lethal total body irradiation. Protected hosts become stable mixed chimeras, but fail to show the early expansion and infiltration of donor T cells in the gut, liver, and blood associated with host tissue injury. Cytokine secretion and adoptive transfer studies using wild-type and IL-4(-/-) mice showed that protection afforded by NK1.1+ and DX5+asialo-GM1+ T cells derived from either donors or hosts conditioned with lymphoid irradiation is dependent on their secretion of high levels of IL-4.

The history of bone-marrow transplantation

As it influenced so much else this century, so the atomic bomb essentially marked the beginnings of interest in bone-marrow transplantation. The first detonation of a nuclear device in New Mexico in 1945, and the subsequent effects of the atomic bomb on the populations of Hiroslaima and Nagasaki provoked great and urgent interest in the effects of irradiation. While studying the effects of lethal irradiation in mice, Jacobson (1949)’ discovered a protective effect of splenic shielding: the spleen in mice contains haemopoietic tissue. He went on to demonstrate a similar protective effect by shielding the femur (1951).2 A year later, Lorenz showed that this protection could also be conferred by intravenous bonemarrow infusion.3 At first, it was thought that the radiation protection was due to humoral elements in the syngeneic bone marrow. Over the next five years, however, a number of investigators demonstrated that, not only was the protection due to cellular elements, but that these donor cells effectively repopulated the irradiated marrow.cw.“9 An animal whose haemopoietic system was derived from another animal was termed a ‘radiation chimera’.’ In 1958, a radiation accident occurred in Vinca, Yugoslavia, and six physicists developed radiationinduced aplasia. Although one died, Mathe” described the successful infusion of allogeneic bone marrow into the remaining five, and subsequently

Allogeneic hematolymphoid microchimerism and prevention of autoimmune disease in the rat. A relationship between allo- and autoimmunity.

Conventional allogeneic bone marrow transplantation after myeloablation can prevent experimental autoimmunity and has been proposed as treatment for humans. However, trace populations of donor hematolymphoid cells persisting in solid organ allograft recipients have been associated in some circumstances with therapeutic effects similar to replacement of the entire bone marrow. We therefore examined whether inducing hematolymphoid microchimerism without myeloablation could confer the ability to resist mercuric chloride (HgCl2)-induced autoimmunity. Brown-Norway (BN) rats were pretreated with a syngeneic or allogeneic bone marrow infusion under transient FK506 immunosuppression before receiving HgCl2. They were compared with BN rats receiving either no pretreatment (naive) or FK506 alone. Administration of HgCl2 to naive BN rats induced marked autoantibody production, systemic vasculitis and lymphocytic infiltration of the kidneys, liver and skin in all of the animals and a 47% mortality. In contrast, BN rats pretreated with HgCl2-resistant allogeneic Lewis bone marrow and transient FK506 showed less clinical disease and were completely protected from mortality. More specifically, IgG anti-laminin autoantibody production was decreased by 40% (P < 0.05), and there was less histopathological tissue injury (P < 0.005), less in vitro autoreactivity (P < 0.05), less of an increase in class II MHC expression on B cells (P < 0.01), and 22% less weight loss (P < 0.01), compared with controls. Protection from the experimental autoimmunity was associated with signs of low grade activation of the BN immune system, which included: increased numbers of circulating B and activated T cells before administration of HgCl2, and less autoreactivity and spontaneous proliferation in vitro after HgCl2.

Clinical use of hematopoietic growth factors in allogeneic bone marrow transplantation

The use of the recombinant hematopoietic growth factors G-CSF and GM-CSF have shortened the period of neutropenia, or avoided this problem, in many cancer patients who have received cytotoxic therapy. Although these benefits have been particularly striking in the autologous bone marrow and/or autologous peripheral blood progenitor cell transplant setting, most data suggest that the use of G-CSF and GM-CSF only marginally enhance recovery of the neutrophil count when administered after allogeneic bone marrow infusion. Furthermore, in the allograft setting these expensive agents have not provided benefit in the form of enhanced platelet count recovery, lessening the incidence of graft-versus-host disease, or improvement in overall survival. These data do not justify routine widespread use of G-CSF and GM-CSF and suggest that these agents should be reversed for patients who experience delay in engraftment after allogeneic bone marrow infusion. Administration of erythropoietin, on the other hand, may reduce the need for homologous red blood cell transfusions, and may increase the safety margin for both the allogeneic bone marrow recipient and as well as the donor. Recombinant hematopoeitic growth factors targetted specifically to enhance platelet recovery after transplantation (such as interleukin-3, interleukin-6, and interleukin-11) have shown promise after autotransplantation and after conventional dose chemotherapy, and likely will be evaluated in the allogeneic transplant patient.

PATHOLOGY OF EXPERIMENTAL PULMONARY BONE MARROW EMBOLISM

In order to investigate the initial lesions of pulmonary bone marrow embolism and its pathogenesis, the author studied the pulmonary changes of 70 rabbits during a 24‐hour period after the Infusion of 500 mg of fresh allogeneic bone marrow into the marginal ear veins. After 30 minutes, leukostasis was observed in the non‐embolized small arteries. After 2 hours, leukostasis increased and by 5 to 10 hours, it reached a maximum, still decreased considerably after 24 hours. In the perivascular connective tissue, edema and inflammatory cell infiltration had occurred as a result of Increased vascular permeability due to leukostasis. Fifteen minutes after intravenous administration of a single shot of indomethacln (5 mg/kg), the Infusion of allogeneic bone marrow was performed. Five hours after the infusion, the suppression of pulmonary vascular leukostasis and interstitial edema were observed. The effect of drugs on morphological changes, however, is extremely small in the group pre‐treated with diphenhydramine hydrochloride (3 mg/kg). The author concluded that the mechanical Injury of vascular endothelial cells by emboli and the accumulation of leukocytes in the pulmonary vessels may play an important role in the pathogenesis of the initial change of pulmonary bone marrow embolism. It is also suggested that the embolized bone marrow in the small arteries and vasculitis may lead to arteriosclerosis in the future.

Allogeneic bone marrow transplantation in mice after total lymphoid irradiation: influence of breeding conditions and strain of recipient mice.

Different groups of C57BL/ka or BALB/c mice received a dose of 34 Gy or 42 Gy of fractionated total lymphoid irradiation (TLI) before bone marrow transplantation with 30 X 10(6) BALB/c or C57BL nucleated bone marrow cells, respectively. BALB/c mice that were not bred in specific pathogen-free conditions before TLI showed a high morbidity and mortality rate after 34 Gy of TLI and allogeneic bone marrow transplantation as compared with BALB/c or C57BL that were bred in pathogen-free conditions before irradiation. Many of the conventionally bred BALB/c mice had clinical and histologic signs of graft-vs-host disease after TLI and allogeneic bone marrow infusion. Although leucocytosis and lymphocytosis and the immunologic competence as measured with in vitro tests were equally depressed after 34 Gy TLI in BALB/c and C57BL mice, chimerism was nevertheless significantly easier to obtain in BALB/c mice. The incidence of chimerism after TLI could be enhanced in C57BL mice by increasing the total radiation dose from 34 to 42 Gy. This augmentation of chimerism was paralleled by the induction of more suppressor cells after 42 Gy of TLI in C57BL mice.

12 - Treatment of Aplastic Anaemia by Antilymphocyte Globulin with or without Marrow Infusion

Forty-one patients, suffering from severe aplastic anaemia were treated either with ALG alone (27 patients) or ALG followed by infusion of allogeneic bone marrow (14 patients). Eighteen patients (67 per cent) are presently alive after ALG alone at over 100 to over 550 days. Fourteen (52 per cent) showed sustained improvement of haematopoiesis, two are alive without change, one recovered autologous haematopoiesis after cyclophosphamide conditioning and transfusion of HLA identical marrow and one is lost to follow-up. Eight patients (57 per cent) are currently alive after ALG and transfusion of haplotype identical marrow with self-sustaining autologous haematopoiesis at over 200 days to over four and a half years. No lethal complications occurred and none of the bone marrow infusions led to permanent engraftment or graft-versus-host disease. The mechanism of action is not known, but our results support the hypothesis that unspecified autoimmune reactions block the normal outgrowth of haematopoietic precursor cells in a substantial number of patients with aplastic anaemia. This therapeutic approach seems to offer good chances of survival, especially for those patients who do not have an HLA identical sibling. Its value should be further investigated.

TREATMENT OF APLASTIC ANÆMIA BY ANTILYMPHOCYTE GLOBULIN WITH AND WITHOUT ALLOGENEIC BONE-MARROW INFUSIONS

29 patients with severe aplastic anæmia were treated with either antilymphocyte globulin (A.L.G.) alone (15 patients) or A.L.G. followed by infusion of allogeneic bone-marrow (14 patients). The overall response to both forms of treatment in terms of 1-year survival was 55%; 12 of the 29 patients showed a sustained hæmatological improvement, during a period of observation of up to 4 1/2 years. No potentially fatal complications were observed. None of the bone-marrow infusions led to a permanent "take" or graft-versus-host disease. How A.L.G. acts is unknown, but our findings accord with the hypothesis that, in a substantial proportion of cases of aplastic anæmia, unspecified autoimmune reactions block the development of residual stem cells. A.L.G. seems to offer a good chance of survival, especially for those patients who do not have HLA-matched siblings. Its value should be further established.

Acquired Aplastic Anaemia in Adults

A follow-up study of 10 patients suffering from acquired aplastic anaemia, comprising methocrylate-embedded bone marrow biopsies and CFU cultures, is presented. The haematopoietic recovery patterns and changes in the inflammatory infiltration after permanent engraftment could be distinguished from those in non-transplanted patients. After anti-thymocyte globulin treatment followed by allogeneic bone marrow infusion, the recovery pattern resembled that in non-transplanted patients. The persistently low colony-forming capacity in some patients could be explained by the existence of lymphoid inhibitory cells, which suggests an immunologic auto-destructive mechanism.