Hans Hägglund

Low incidence of acute graft-versus-host disease, using unrelated HLA-A-, HLA-B-, and HLA-DR-compatible donors and conditioning, including anti-T-cell antibodies.

BACKGROUND Using unrelated bone marrow, there is an increased risk of graft-versus-host disease (GVHD). METHODS HLA-A-, HLA-B-, and HLA-DR-compatible unrelated bone marrow was given to 132 patients. The diagnoses included chronic myeloid leukemia (n=43), acute lymphoblastic leukemia (n=29), acute myeloid leukemia (n=27), myelodysplastic syndrome (n=4), lymphoma (n=3), myeloma (n=1), myelofibrosis (n=1), severe aplastic anemia (n=12), and metabolic disorders (n=12). The median age was 25 years (range 1-55 years). HLA class I was typed serologically, and class II was typed by polymerase chain reaction using sequence-specific primer pairs. Immunosuppression consisted of antithymocyte globulin or OKT3 for 5 days before transplantation and methotrexate combined with cyclosporine. RESULTS Engraftment was seen in 127 of 132 patients (96%). Bacteremia occurred in 47%, cytomegalovirus (CMV) infection in 49%, and CMV disease in 8%. The cumulative incidences of acute GVHD > or = grade II and of chronic GVHD were 23% and 50%, respectively. The 5-year transplant-related mortality rate was 39%. The overall 5-year patient survival rate was 49%; in patients with metabolic disorders and severe aplastic anemia, it was 61% and 48%, respectively. The disease-free survival rate was 47% in patients with hematological malignancies in first remission or first chronic phase and 38% in patients with more advanced disease (P=0.04). Acute GVHD was associated with early engraftment of white blood count (P=0.02). Poor outcome in multivariate analysis was associated with acute myeloid leukemia (P=0.01) and CMV disease (P=0.04). CONCLUSION Using HLA-A-, HLA-B-, and HLA-DR-compatible unrelated bone marrow and immunosuppression with antithymocyte globulin, methotrexate, and cyclosporine, the probability of GVHD was low and survival was favorable.

Predictive factors for outcome of allogeneic hematopoietic cell transplantation for adult acute lymphoblastic leukemia

Between January 1990 and December 1997, 182 adults with acute lymphoblastic leukemia (ALL) received allogeneic hematopoietic cell transplants according to Fred Hutchinson Cancer Research Center protocols. Patients eligible for transplantation included those in first remission, especially those at high risk of relapse (n = 41), and any patient in second or later remissions (n = 46) or in relapse (n = 95). The median patient age was 29.4 years (range, 18.0-57.6 years), and the median duration of disease was 13.3 months (range, 2.4-221.9 months). Fifty-six patients had Philadelphia chromosome-positive ALL. Most patients (n = 169) received a conditioning regimen of cyclophosphamide 120 mg/kg plus 12.0 to 15.75 Gy of total body irradiation and a combination of cyclosporine and methotrexate as graft-versus-host disease (GVHD) prophylaxis. One hundred twenty-one patients received stem cells from HLA-identical donors (88 related donors and 33 unrelated donors), and 61 received stem cells from HLA-mismatched donors (26 related donors and 35 unrelated donors). Actuarial disease-free survival at 5 years was 21% for all patients, 43% for patients in first remission, 24% for patients in second or later remissions, and 9% for patients in relapse. Univariate and multivariate Cox regression analyses were performed to identify factors associated with survival, relapse, nonrelapse mortality, and disease-free survival. Factors significantly associated (P <.01) with improved survival and disease-free survival included younger age and being in first remission. Lower disease-free survival was associated with receiving cyclosporine alone as GVHD prophylaxis (P <.01). Risk of relapse correlated only with disease status at transplantation: patients who underwent transplantation in relapse had a 9-fold increased risk compared with patients who underwent transplantation in first remission. Acute or chronic GVHD had no significant effect on relapse. Increased nonrelapse mortality was associated with HLA-mismatched donors, a positive cytomegalovirus serology before transplantation, and GVHD prophylaxis with only cyclosporine. Patients with Philadelphia chromosome-positive ALL had survival and relapse rates similar to patients with normal cytogenetics.

Does high-dose intravenous immune globulin treatment after bone marrow transplantation increase mortality in veno-occlusive disease of the liver?

Forty-five recipients of bone marrow from HLA-identical siblings were given intravenous immune globulin (IVIG) 0.5 g/kg once a week during the first 3 months after transplantation. Fifty-three consecutive previously transplanted HLA-identical siblings were included as controls. Only patients who were cytomegalovirus (CMV) seropositive or had a CMV-seropositive donor were included. There were no major differences in patient characteristics between the two groups. However, more patients in the IVIG group received individualized graft-versus-host disease (GVHD) prophylaxis with less cyclosporine (P<0.01), more controls received liposomal amphotericin B (P=0.01), and more patients in the IVIG group received low-dose heparin as prophylaxis against veno-occlusive disease of the liver (P<0.001). Median follow-up was 21 months in the IVIG group and 47 months in the control group. There were no differences between the groups with regard to time to engraftment, hospitalization time, or days with fever. No differences between the IVIG group and control group were detected in the incidence of acute GVHD grade II-IV (17% vs. 23%) or chronic GVHD (30% vs. 42%). The incidence of bacterial septicemias (53% vs. 63%) and invasive fungal infections (9% vs. 6%) was unaffected by IVIG treatment. The incidence of symptomatic CMV infection was the same in the two groups (14% vs. 16%). One control patient died of CMV interstitial pneumonitis, and 1 patient from each group died from viral interstitial pneumonitis of other origin. The incidence of veno-occlusive disease (VOD) was 16% in the IVIG group versus 6% in the controls (P=NS). Fatal VOD occurred in 11% of the IVIG group compared with none of the controls (P=0.02). Other transplant-related complications did not differ between the two groups. Two-year survival was 62% in the IVIG group and 60% in the controls (P=NS). No significant beneficial effect was seen with IVIG, which may increase mortality in VOD. The use of high dose IVIG as prophylaxis in marrow transplant recipients is questioned.

Treatment of hepatic venoocclusive disease with recombinant human tissue plasminogen activator or orthotopic liver transplantation after allogeneic bone marrow transplantation.

Ten allogeneic bone marrow transplant (BMT) recipients with hepatic venoocclusive disease (VOD) were treated with recombinant human tissue plasminogen activator (rt-PA). Two of them subsequently underwent orthotopic liver transplantation (OLT). One additional patient with VOD underwent OLT without prior rt-PA treatment. Treatment with rt-PA was started a median of 14 (1--35) days after BMT. The dose of rt-PA given to adults was 10-50 mg i.v. and that given to children was 3-10 mg i.v. Treatment was given for 2-4 days. In three patients, the dose was administered over a longer period or it was repeated. Four patients responded to rt-PA therapy and six did not. Eight patients suffered from hemorrhages, one intracranial and three gastrointestinal. Four patients required blood transfusions. Four had minor subcutaneous hemorrhages and/or epistaxis. One patient died of intracranial hemorrhage and five from hepatic and/or multiorgan failure. Two patients treated with rt-PA, 10 mg/day for 4 days, are alive; one is alive and well 3 months after BMT, the other has relapsed after 7 months. The three patients undergoing OLT died of chronic hepatic failure, cerebral edema, and pneumonia. Our experience suggests that rt-PA should not be administered in high doses and that the treatment should not be given over a longer period, because of the risk of severe hemorrhages.

Ex vivo expansion of canine dendritic cells from CD34+ bone marrow progenitor cells.

BACKGROUND The aims of this study were to ex vivo expand canine dendritic cells and determine their phenotype and functional characteristics. METHODS CD34+-selected cells and CD34+-depleted canine bone marrow (BM) cells were cultured in Iscoves modified medium for 14 days. Cytokines added to the cultures included human granylocyte/macrophage colony-stimulating factor 5 ng/ml, hFlt3 ligand 200 ng/ml, and human tumor necrosis factor-alpha 10 ng/ml. Cultured cells and purified subpopulations were assessed for cell surface antigen expression, morphology, and function by flow cytometric analysis, electron microscopy, and an allogeneic mixed lymphocyte reaction at day 14. RESULTS Two main cell populations were identified, DR++(bright)/CD14- and DR+(dim)/CD14+. Ex vivo expanded CD34+-selected cells showed increased allostimulatory activity compared to both cultured CD34+-depleted cells and mononuclear cells. In contrast, ex vivo expansion from CD34+-depleted cells was unsuccessful. After sorting cells from the ex vivo expanded CD34+-selected bone marrow to enrich for DR++/CD14- cells, a 42-fold increase (median) of allostimulatory activity was observed as compared with sorted DR+/CD14+ cells (P=0.02). CONCLUSIONS Cells with dentric cell-like phenotypes and functions can be cultured from canine CD34+-selected bone marrow cells. Future studies will address the roles of these cells in engraftment, graft versus host reactions and graft-host tolerance in a canine hematogoietic stem cell transplantaton model.

Transplantation of Autologous and Allogeneic Bone Marrow With Liver From A Cadaveric Donor for Primary Liver Cancer1

BACKGROUND In histocompatibility mismatched experimental animals, a combination of T-cell-depleted autologous and allogeneic marrow may induce mixed chimerism and tolerance. Patients with large primary liver tumors have a poor outcome. We investigated whether it were possible to induce mixed chimerism and obtain an antitumor effect in a patient with a large primary liver cancer after combined liver and bone marrow transplantation (BMT). METHODS A 46-year-old female with a primary non resectable liver cancer received a liver transplant from a cadaveric donor. Subsequently, she was conditioned with 4x2 Gy of total lymphoid irradiation, 120 mg/kg cyclophosphamide, and 7.5 Gy total body irradiation. Twelve days after liver transplantation, she received T-cell-depleted autologous:cadaveric 5/6 antigen HLA-mismatched marrow in a proportion of CD34+ cells of 0.5:3.0x10(6)/kg. Chimerism status was determined with polymerase chain reaction amplification of variable number tandem repeats from DNA obtained from CD3+, CD19+, and CD45+ magnetic-bead-separated cells. RESULTS The early posttransplant period was uneventful; liver function was normal and the hematopoietic engraftment of donor and recipient origin was prompt. Alpha-fetoprotein levels dropped from 440 to 35 microg/l. One month after marrow transplantation, donor T-cells decreased markedly. Monoclonal antibody OKT-3 and 10(5)/kg donor T-cells were given. One month later, the patient developed diarrhea and abdominal pain. A colonoscopy showed moderate gastrointestinal acute graft-versus-host disease and a Cryptosporidium infection. Three months after BMT, she became a complete donor chimera. Chimera cells showed little, if any, reactivity in mixed lymphocyte cultures to recipient and donor cells, but reacted to third party. Five months after BMT, she developed progressive Aspergillus fumigatus pneumonia and died. No tumor was found at the autopsy. CONCLUSION We obtained mixed donor-recipient hematopoietic chimerism without severe acute graft-versus-host-disease, after combined T-cell depleted autologous and allogeneic BMT and a transplantation of a liver from an HLA-mismatched cadaveric donor. Additional donor T-cells enhanced donor bone marrow engraftment, but rejected the autograft. On the basis of this first attempt, further clinical studies are warranted.

Flt3 ligand promotes engraftment of allogeneic hematopoietic stem cells without significant graft-versus-host disease

Background. Graft-versus-host (GVH) reactions contribute to stable engraftment of allogeneic hematopoietic stem cell transplants. It was hypothesized that the in vivo expansion of recipient dendritic cells (DC) with the administration of ligand for Flt3 (FL) could promote allogeneic engraftment after reduced-intensity conditioning by enhancing the GVH effect. Methods. FL was first administered to three nonirradiated healthy dogs for 13 days at a dosage of 100 &mgr;g/kg/day. Next, nine dogs received 4.5 Gy total-body irradiation (TBI) and unmodified marrow grafts from dog leukocyte antigen (DLA)-identical littermates without posttransplant immunosuppression. FL was administered to the recipients at a dosage of 100 &mgr;g/kg/day from day −7 until day +5. Results. In normal dogs, FL produced significant increases in monocytes (CD14+) and neutrophils in the peripheral blood, a marked increase in CD1c+ cells with DC-type morphology in lymph nodes, and increased alloreactivity of third-party responders to peripheral blood mononuclear cells in mixed lymphocyte reactions (P <0.001). Sustained engraftment was observed in eight of nine (89%) FL-treated dogs compared with 14 of 37 (38%) controls (P =0.02, logistic regression). All engrafted FL-treated dogs became stable complete (n=2) or mixed (n=6) hematopoietic chimeras without significant graft-versus-host disease (GVHD). Recipient chimeric dogs (n=4) were tolerant to skin transplants from their marrow donors but rejected skin grafts from unrelated dogs within 7 to 9 days (median, 8 days). Conclusions. In this study, the authors showed that FL administered to recipients promotes stable engraftment of allogeneic marrow from DLA-identical littermates after 4.5 Gy TBI without significant GVHD.

Successful allogeneic bone marrow transplantation in a 2.5-year-old boy with ongoing cytomegalovirus viremia and severe aplastic anemia after orthotopic liver transplantation for non-A, non-B, non-C hepatitis.

BACKGROUND A 2.5-year-old boy received a cadaveric orthotopic liver transplant for acute liver failure due to non-A, non-B, non-C hepatitis. After transplantation, he developed thrombocytopenia and neutropenia and subsequently severe aplastic anemia. The patient also suffered from recurrent cytomegalovirus (CMV) viremia, treated with foscarnet and ganciclovir. METHODS For treatment of his aplastic anemia, the patient underwent an allogeneic bone marrow transplantation from his HLA-identical sister after conditioning with cyclophosphamide at 200 mg/kg and antithymocyte globulin at 3 mg/kg for 5 days. Prophylactic acyclovir was given because of ongoing CMV viremia at the time of bone marrow transplantation. RESULTS The transplant course was uneventful, with rapid engraftment. There were no signs of liver dysfunction, graft-versus-host disease, or reactivation of CMV. The patient is in excellent health, with normal liver and bone marrow function 3 years after bone marrow transplantation. CONCLUSION This case report shows that allogeneic bone marrow transplantation is feasible and well tolerated in a patient with severe aplastic anemia after liver transplantation for acute fulminant viral hepatitis.

Improved survival after bone marrow transplantation for early leukemia using busulfan–cyclophosphamide and individualized prophylaxis against graft-versus-host disease: a long-term follow-up

To minimize immunosuppression, allow a graft‐versus‐leukemia (GVL) effect, and reduce relapse incidence, 73 leukemic recipients of human leukocyte antigens‐identical sibling marrow were given graft‐versus‐host disease (GVHD) prophylaxis based on the estimated risk of GVHD development. Methotrexate (MTX) monotherapy was given to patients with an estimated low risk of developing GVHD, whereas MTX+cyclosporine (CsA) was given to ‘high‐risk’ patients. After engraftment, CsA was discontinued, and weekly MTX was reinstituted and given until 3 months post‐bone marrow transplant. Conditioning consisted of busulfan (BU)+cyclophosphamide (CY) (n=35) or CY+total body irradiation (TBI) (n=38). Retrospective controls were given CY+TBI and MTX+CsA (n=39). The median observation time was 5 yr 11 months. Chronic GVHD increased to 53% in the individual BU+CY group and 46% in the individual CY+TBI group, compared to 25% in the control group (p=0.05). This increase was restricted to the limited form. The actuarial relapse incidence decreased to 20% in the individual BU+CY group, compared to 52% in the control group, p=0.03. In the individual CY+TBI group, the relapse incidence was 44% (n.s. versus controls, p=0.04 versus individual BU+CY). The 5‐yr relapse‐free survival (RFS) in the individual BU+CY group was 66%, in the control group, 41% (p=0.07), and in the individual CY+TBI group, 45% (p=0.1 versus individual BU+CY). Patients with early leukemia in the individual BU+CY group had a RFS of 83%, compared to 44% in the control group (p=0.02) and 42% in the individual CY+TBI group (p=0.01). In the multivariate analysis, advanced leukemia beyond first complete remission and first chronic phase and conditioning with CY+TBI were correlated to poor RFS. In summary, the individualized prophylaxis itself did not reduce the relapse incidence. However, in patients with early leukemia conditioning with BU+CY, our method of individualizing the GVHD prophylaxis might be of value, since this group had the best RFS in this study.

Improved survival with ursodeoxycholic acid prophylaxis in allogeneic stem cell transplantation: long-term follow-up of a randomized study.

We report the long-term results of a prospective randomized study on the use of ursodeoxycholic acid (UDCA) for prevention of hepatic complications after allogeneic stem cell transplantation. Two hundred forty-two patients, 232 with malignant disease, were randomized to receive (n = 123) or not to receive (n = 119) UDCA from the beginning of the conditioning until 90 days post-transplantation. The results were reported after 1-year follow-up. UDCA administration reduced significantly the proportion of patients developing high serum bilirubin levels as well as the incidence of severe acute graft-versus-host disease (GVHD), liver GVHD, and intestinal GVHD. In the UDCA prophylaxis group, nonrelapse mortality (NRM) was lower and overall survival better than in the control group. After a 10-year follow-up, the difference in the survival and NRM in favor of the UDCA-treated group, seen at 1 year, was maintained (survival 48% versus 38%, P = .037; NRM 28% versus 41%, P = .01). A landmark analysis in patients surviving at 1 year post-transplantation showed no significant differences between the study groups in the long-term follow-up in chronic GVHD, relapse rate, NRM, disease-free survival, or overall survival. These long-term results continue to support the useful role of UDCA in the prevention of transplant-related complications in allogeneic transplantation.