Autologous bone marrow transplantation with 4- hydroperoxycyclophosphamide purged marrows for acute nonlymphocytic leukemia in late remission or early relapse

Twenty-four patients with acute nonlymphocytic leukemia (ANLL) were treated with high-dose chemotherapy or chemoradiotherapy followed by infusion of autologous marrow purged with 100 micrograms/mL of 4- hydroperoxycyclophosphamide (4HC). The marrow harvests were performed when there were less than 5% blasts in the marrow. Seven patients were transplanted in second complete remission (CR), eight in third CR, one in fourth CR, and eight in early relapse. The median time to achieve 500 neutrophils/microL or 1,000 leukocytes/microL was 30 days. A platelet count of 20,000/microL and 50,000/microL was achieved at a median of 67 and 91 days, respectively. One patient failed to engraft by day 58. There were five other transplant-related deaths: sepsis (one), intracerebral hemorrhage (one), veno-occlusive disease (one), and interstitial pneumonia (two). Four of seven evaluable patients transplanted in early relapse obtained a CR lasting 112, 143, 189, and greater than 615 days. Eight of 11 evaluable patients transplanted in CR have relapsed at a median of 153 days (range, 104 to 311). The actuarial survival for all patients was 19%. There was a trend toward improved relapse-free survival for patients transplanted in remission as opposed to those transplanted in relapse (P = .11).     

Autologous bone marrow transplantation in children with acute leukemia

Eight children with acute leukemia were treated with chemoradiotherapy or high-dose chemotherapy followed by infusion of autologous marrow unpurged or purged mainly with monoclonal antibodies. Four were treated in their first remission, three in their second remission, and one in the fourth remission. Patients were isolated in laminar air flow rooms with sterile conditioning. Infused mononuclear cells ranged from 1.0 to 3.8 x 10(7)/kg. Hematologic engraftment was obtained in five patients. The other three patients, who were treated with monoclonal antibody or drug, received unpurged back up marrow because of delayed bone marrow reconstitution. Two recovered, but one died before engraftment. Hematologic recovery was delayed after autologous bone marrow transplantation. The median time to achieve 1,000 leukocytes/microliters, 500 neutrophils/microliters, and last platelet transfusion was 36 days (12-90), 36 days (12-90), and 70 days (39-214), respectively. Moreover, further increases beyond these levels remained unusually slow, especially in platelets. Two of eight patients relapsed at 5 and 15 months and the other patient died of interstitial pneumonitis. Actuarial disease free survival was 62.5%. There was a trend toward improved disease free survival for patients transplanted in the first remission (100%).     

Autologous bone marrow transplantation in acute myelogenous leukemia: in vitro treatment with myeloid cell-specific monoclonal antibodies

Second or third chemotherapy-induced remissions in acute myelogenous leukemia (AML) are limited by early relapse of the leukemia. We developed monoclonal antibodies (MoAbs) that are cytotoxic to myeloid leukemia cells to treat bone marrow from these patients ex vivo for autologous transplantation. In this pilot study, bone marrow was harvested from ten patients with AML in remission, treated with one or two complement-fixing MoAbs, PM-81 and AML-2-23, which react with myeloid differentiation antigens, incubated with rabbit complement, and cryopreserved. These MoAbs were chosen because they have broad reactivity with AML cells but not with pluripotent progenitor cells. At the time of transplant, 6 patients were in second complete remission, 1 each was in third complete or partial remission, and 2 were in early first relapse. The patients were treated with cyclophosphamide (60 mg/kg a day for 2 days) and total body irradiation (200 cGy twice a day for 3 days) and given infusions of MoAb-treated bone marrow. Full bone marrow reconstitution was observed in eight patients; two patients did not recover platelets. Seven of the ten patients are surviving and disease-free at 21.0, 15.0, 13.0, 10.0, 6.0, 3.0, and 2.0 months posttransplant. Treating bone marrow with MoAbs to myeloid differentiation antigens does not interfere with pluripotential stem cell engraftment. Longer follow-up and a controlled study are necessary to prove that the apparent efficacy of this therapeutic approach in some patients is attributable to MoAb-mediated killing of leukemia cells.     

Autologous bone marrow transplantation for acute myeloid leukemia following in vitro treatment with neuraminidase and monoclonal antibodies

Although monoclonal antibodies (MoAbs) to CD15, especially PM-81, react with leukemic blasts from the majority of patients with acute myeloid leukemia (AML), a small subset of patients have cells that are CD15 negative or dim. We determined previously that neuraminidase will increase the reactivity of PM-81 with AML blasts, as well as blasts from many patients with acute lymphoblastic leukemia (ALL). In this report, we describe the laboratory results and clinical course of the first patient with AML whose harvested bone marrow was treated with neuraminidase prior to MoAbs and complement treatment. Neuraminidase increased the percentage of the patient's leukemia cells that reacted with PM-81 from 18% to 90% and more than doubled the percentage of AML blasts that were lysed by PM-81 and complement. The patient suffered no acute toxicity, engrafted rapidly, and was transfusion independent by day 21 post-ABMT. This report demonstrates the probable safety and efficacy of pretreatment of bone marrow with neuraminidase, and increases the number of patients with AML or ALL who may benefit from ABMT using marrow purging with MoAb to CD15.     

Autologous bone marrow transplantation in acute leukemia with marrow purged with alkyl-lysophospholipid.

Alkyl-lysophospholipids are anticancer agents that are selectively toxic to leukemic cells and relatively sparing of normal bone marrow cells. Thus, they would be likely candidates for purging remission marrows before autologous bone marrow transplant. One of the more promising agents is edelfosine, which could be safely used for purging without prolonging marrow recovery. Assays for marrow progenitor cells were performed before and after purging and cryopreservation in 64 patients. There was no significant reduction in colony formation after purging when compared with unpurged cryopreserved marrow, but there was a significant reduction after cryopreservation. Twenty-four patients with acute leukemia in second (16 patients) or third remission (3 patients), early relapse (3 patients), or in first remission with successfully treated extramedullary relapse (2 patients) received marrow-ablative chemotherapy and total body irradiation followed by infusion of marrow purged for 4 hours with 50 to 100 micrograms/mL of edelfosine. There were 9 lymphoblastic and 15 myelogenous leukemia patients. The median time to granulocyte recovery to 500/microL was 26 and 33 days for the 50 and 75 microgram/mL doses, respectively. The patient whose marrow was purged at the dose of 100 micrograms/mL failed to engraft. The median time to platelet recovery to 25,000/microL was 45 and 37 days for the 50 and 75 micrograms/mL doses, respectively. Twenty-nine percent of the patients remain disease free from 131 to 1,291 days, with a median of 356 days. These results have established that purging with 75 micrograms/mL of edelfosine is a safe dose and is recommended for a phase II trial.     

Autologous bone marrow transplantation in acute myelogenous leukemia: in vitro treatment with myeloid-specific monoclonal antibodies and drugs in combination

We report the results of a preclinical study comparing four different purging protocols using a promyelocytic human cell line HL-60 and myeloid leukemic progenitor cells (colony-forming unit-leukemic [CFU-L]) from acute myelogenous leukemia (AML) patients assayed in semisolid culture. We studied the antileukemic effect of (1) Single-cycle complement-mediated lysis by two different monoclonal antibodies (MoAbs) (M195 [CD33] and F23 [CD13] 40 micrograms/mL), reactive with distinct antigens found on early myeloid cells and monocytes, used alone and in combinations; (2) 4-Hydroperoxycyclophosphamide (4-HC) (80 mumol/L or 100 mumol/L) alone; or (3) combined with VP-16 (5 micrograms/mL) and (4) a cocktail of 1 through 3 as above (combined immunochemotherapy). More than 4 logs of HL-60 tumor cell elimination were observed after 1 hour of incubation with both MoAbs plus 4-HC + VP-16 while the single treatment (immunotherapy or chemotherapy) provided 1.5 and 3.5 logs of colony-forming inhibition, respectively. When the same protocols were tested on cryopreserved leukemic cells from eight patients with AML, we observed a mean value of CFU-L inhibition of 92.3% +/- 2.5% SD, 95.5% +/- 1.4% SD, and 99% +/- 0.8% SD after MoAbs and complement lysis, 4-HC, and 4-HC + VP-16 treatment, respectively. The combined treatment of MoAbs and 4-HC + VP-16 produced more than 3-log reduction of CFU-L colony formation. By comparison, the mean recovery of committed normal bone marrow progenitors after incubation with MoAbs and complement was 12% for CFU-granulocyte-macrophage (CFU-GM), 22.9% for burst-forming unit erythroid (BFU-E), and the recovery following 4-HC + VP-16 treatment was 4.4% for CFU-GM and 5.6% BFU-E. In subsequent experiments, highly purified CD34+ blast cells, enriched by positive selection, and stimulated in liquid culture by cytokines (interleukin-1 [IL-1], IL-3, and combination of both) or MO-conditioned medium (MoCM), demonstrated that immunochemotherapy spares hematopoietic colony-forming cells earlier than day 14 CFU-GM, in vitro.     

Autologous bone marrow transplantation for acute myeloid leukemia using monoclonal antibody-purged bone marrow

We report our experience from a clinical trial of autologous bone marrow transplantation (ABMT) in the treatment of 30 patients with acute myeloid leukemia (AML) using monoclonal antibody (MoAb) and complement-treated bone marrow. All patients were in complete remission (CR) at the time of transplant: 6 patients were in first CR, 18 in second CR, and 6 in third CR. The median age of all patients was 42 years (range 11 to 57 years). For marrow ablation, 28 patients were treated with cyclophosphamide and total body irradiation. One patient was treated with busulfan and cyclophosphamide and one was treated with busulfan and VP-16. Each patient was then transfused with autologous bone marrow that had been harvested previously and treated with two MoAbs, PM-81 and AML-2-23, and rabbit complement. Median time to recovery of neutrophils (500/microL) was 30 days, and platelets (20,000/microL) was 45 days. Median time for initial erythrocyte engraftment, assessed by a flow cytometric reticulocyte assay, was 13 days. Median overall and relapse-free survival of first CR patients was at least 17.4 months post-ABMT and the 2- and 3-year actuarial overall and relapse-free survival was 67% (+/- 19%). Median survival for the 24 patients in second or third CR was 6.8 months post-ABMT and 9.3 months since CR; however, six patients survived disease-free from 16 to 61 months post-ABMT. For the second and third CR group it was observed that six patients (5 of the 6 survivors) showed "inversions," when their post-ABMT remission lasted longer than any previous one. Actuarial 2- and 3-year disease-free and overall survival of patients in second and third CR was 25% (+/- 9%) and 18% (+/- 9%), and 29% (+/- 9%) and 23% (+/- 9%), respectively. ABMT avoids the problems of graft-versus-host disease and of finding suitable donors for allogeneic marrow transplantation.     

Autologous bone marrow transplantation using marrow incubated with Asta Z 7557 in adult acute leukemia

The sensitivity of human myeloblastic leukemic (CFU-L) and normal hemopoietic stem cells (CFU-GM and BFU-e) to Asta Z 7557 (INN Mafosfamide) was studied with regard to autologous bone marrow transplantation (ABMT) with cleansed marrow for consolidation therapy in adult patients with acute leukemia (AL) in remission. Establishment of the dose-response curves for CFU-GM (n = 37), BFUe (n = 11), and myeloblastic CFU-L (n = 9) demonstrated a wide range of sensitivity from patient to patient for all three progenitors. Whereas CFU-L, CFU- GM, and BFU-e grown in semisolid cultures disclosed similar sensitivities to Asta Z 7557, long-term culture (LTC) studies (n = 41) indicated a higher resistance of early progenitors. In an effort to achieve a maximum tumor cell kill and yet spare a sufficient amount of normal stem cells to ensure consistent engraftment, we defined the optimal dose for marrow cleansing as the dose sparing 5% CFU-GM (LD95). This dose was established from a preincubation test (PIT) realized on a 10-mL marrow aspirate taken 15 days before marrow collection in each individual patient. Twenty-four adult patients while in remission of AL (20 in complete remission, four in partial remission) were consolidated by cyclophosphamide 60 mg/kg X 2 and total body irradiation at 10 Gy followed by ABMT with marrow cleansed by Asta Z 7557 according to the specification described above. Patients were divided in two groups: group 1, unfavorable prognosis (11 patients); group 2, standard prognosis [13 patients in first complete remission (CR)]. All patients engrafted on leukocytes (median day for recovery to 10(9)/L: day 30), patients with ALL recovered faster than patients with ANL (median day 19 v 34). Similarly, recovery of platelets to 50.10(9)/L occurred sooner in patients with ALL (median day 67, range day 23 through 90) whereas three patients with acute nonlymphoblastic leukemia (ANLL) in group 2 had to be supported with platelet transfusions for more than one year. In group 1, six patients had recurrent tumor within six months; three patients died from toxicity with no evidence of tumor. Two patients are still disease-free with a short follow-up (nine and ten months). In group 2, two patients died from toxicity with no evidence of leukemia three and 16 months post-ABMT. One patient with a M5 ANLL and one patient with ALL relapsed at six and 15 months, respectively. Nine patients have remained in CR or are disease-free with a median follow-up of 22 months.(ABSTRACT TRUNCATED AT 400 WORDS)     

Autologous bone marrow transplantation for acute myelogenous leukemia using 4-hydroperoxycyclophosphamide and VP-16 purged bone marrow.

Thirty adult patients with acute myelogenous leukemia (AML) in remission were treated with hyperfractionated total body irradiation, VP-16, and cyclophosphamide followed by infusion of autologous bone marrow purged with 4-hydroperoxycyclophosphamide and VP-16. Fifteen patients were transplanted in first remission (R1), 13 in second remission (R2), and two in third remission (R3). All patients had hematopoietic engraftment. The median time to achieve a white blood cell count of 1.0 x 10(9)/l and a neutrophil count of 500 x 10(6)/l was 32 days. The median time to achieve an unsupported platelet count of 50 x 10(9)/l was 70 days. There were four transplant-related deaths, all in patients in R2. Ten patients have relapsed, including both patients transplanted in R3. The disease-free survival (DFS) of all patients is 52%, median follow-up not yet reached. Although the number of patients in each group is small and follow-up is limited, there is a trend toward improved actuarial DFS in patients transplanted in R1 compared with patients transplanted in R2 (72 vs 39%; p = 0.081). Use of VP-16 in the conditioning regimen as well as in the purging procedure is feasible in the treatment of patients with AML.     

Autologous bone marrow transplantation in acute lymphoblastic leukemia: biological and clinical aspects

The purpose of this review is to assess the current status of autologous bone marrow transplantation in acute lymphoblastic leukemia. In the first part of the review the biological aspects of minimal residual disease are discussed and in the second part the most recent clinical results analyzed. Only the studies that contained adequate response and survival data were selected for analysis, including peer-reviewed articles, book chapters and proceedings of international meetings. The most current or complete references were used for series reported more than once.     

Autologous bone marrow transplantation in chronic myelogenous leukemia

Introduction of interferon-alpha therapy to chronic myelogenous leukemia (CML) has improved the survival rate of CML patients compared with conventional busulfan therapy. There still, however, are some IFN-resistant cases. To improve the survival rate of these IFN-resistant cases, bone marrow transplantation (BMT) has been tried at the world wide level. In cases without any allogeneic donors, autologous BMT is another choice. We recently have proposed the flow chart therapy system to select the auto-BMT candidates in CML patients. This system, briefly, consists of (1) bone marrow collection as early stage of CML as possible, (2) IFN-alpha treatment with administration of weekly methotrexate or occasional use of hydroxyurea, (3) early detection of accelerated or blastic phase of CML by using scoring system, (4) conditioning regimens of auto-BMT for CML and (5) post-BMT follow-up with IFN-alpha. Following this system, we have initiated the treatment of CML cases. Our tentative results on one case favorable outcome including complete disappearance of Ph1 positive clone. However, there are several questions to be answered in the auto-BMT for CML, namely, (1) do we need to purge Ph1 progenitor cells or not, if yes, how? (2) does the long term use of IFN affect the bone marrow microenvironment resulting in graft failure? Although our preliminary results gave some answers on these questions, further clinical and basic studies are required to obtain higher survival rates in CML treatment.     

Autologous bone marrow transplantation in acute leukaemia

Intensive chemoradiotherapy with autologous bone marrow rescue has been widely explored as treatment for acute myeloblastic and acute lymphoblastic leukaemia. Encouraging preliminary results have been reported but in no situation has this modality of therapy been proved to be superior to conventional chemotherapy. Attempts to remove minimal residual disease from the harvested marrow have been made by both immunological and pharmacological means, but the value of such procedures has not been tested rigorously. Determination of the precise role of autologous bone marrow transplantation in the treatment of acute leukaemia must await the outcome of randomised controlled trials and will take several years.     

In vitro restoration of polyclonal hematopoiesis in a chronic myelogenous leukemia after in vitro treatment with 4- hydroperoxycyclophosphamide

Bone marrow cells of a 45-year-old female with Philadelphia chromosome (Ph1)-positive, early-phase chronic myelogenous leukemia (CML), who was heterozygous for the glucose-6-phosphate dehydrogenase (G6PD) locus, were pretreated in vitro with 4-hydroperoxycyclophosphamide (4-HC) and tested for G6PD activity in several colony formation assays and for karyotypic abnormalities. All cells within the mixed (CFU-GEMM), the erythroid burst (BFU-E), and the granulocyte-macrophage (CFU-GM) colonies expressed type A and type B G6PD activity and a normal karyotype, whereas untreated cells expressed type A G6PD and the Ph1 chromosome. This reversal of G6PD activity type and the disappearance of the Ph1 chromosome in colonies grown from 4-HC-treated cells indicate that this cytotoxic agent spares a residual normal stem cell population in bone marrow cells of early-phase CML patients. This finding, in turn, suggests a therapeutic approach in CML based on in vitro chemotherapy of autologous bone marrow grafts.     

Autologous bone marrow transplantation with 4-hydroperoxycyclophosphamide purged marrows for children with acute non-lymphoblastic leukemia in second remission

Although autologous bone marrow transplantation (ABMT) following purging with 4-hydroperoxycyclophosphamide (4HC) has been effective for some adults with acute non-lymphoblastic leukemia (ANLL), there are few data on ABMT for children. We have performed ABMT for 13 patients (median age, 5 years) with ANLL in second remission. Bone marrow was treated ex vivo with 4HC to kill residual leukemic cells. In order to enhance the anti-leukemic effect of 4HC, exogenous red blood cells were eliminated from the marrow/4HC mixture. All patients were conditioned for transplantation with busulfan (16 mg/kg) and cyclophosphamide (200 mg/kg), followed by infusion of the 4HC treated marrow. Eleven of 13 patients had complete hematologic reconstitution; there were no transplant-related deaths. Five patients relapsed at 2, 4, 5, 6 and 6 months post-BMT. Eight patients are disease-free survivors; the median time for survival is 24 months, with a projected disease-free survival of 61%. ABMT is a safe and efficacious treatment modality for children with ANLL in second remission. Our results suggest that the disease-free survival for pediatric patients may be superior to that seen in adults.     

Allogeneic bone marrow transplantation in acute nonlymphocytic leukemia: a pilot study

The objective of the current study, initiated in 1976, was to improve upon the high relapse rate and subsequent mortality in children and young adults with acute nonlymphocytic leukemia (ANLL). Seventeen patients, ages 6--28, with ANLL in first bone marrow remission, received cyclophosphamide and total body irradiation using a radiation scheme of 750 rad (7.5 Gy) total dose, delivered at a dose rate of 26 rad (26 cGy) per minute. Allogeneic marrow from HLA-matched sibling donors was followed by prophylactic therapy or graft-versus-host disease (GVHD). Median follow-up of the entire group is 20+ mo; survivors have been followed for a minimum of 14+ mo. Interstitial pneumonitis was observed in 6% of patients, and GVHD was observed in 29%. Seventy percent of patients are alive and in complete continuous remission. Two patients have relapsed (at 7 and 24 mo). Actuarial relapse-free survival is 76% at 1 yr and 64% at 5 yr. Quality of life in this disease-free survivors is excellent; all patients are free of active GVHD, receive no maintenance chemotherapy, and have high Karnofsky performances scores. High dose rate total body irradiation plus cyclophosphamide followed by allogeneic BMT may provide an opportunity for long-term complication-free survival in a substantial proportion of children and young adults with ANLL.     

Autologous bone marrow transplantation in high-risk remission T-lineage acute lymphoblastic leukemia using immunotoxins plus 4-hydroperoxycyclophosphamide for marrow purging

Fourteen patients with high-risk T-lineage acute lymphoblastic leukemia (ALL) in complete remission underwent autologous bone marrow transplantation (BMT) in an attempt to eradicate their residual disease burden. A combined immunochemotherapy protocol using a cocktail of two immunotoxins directed against CD5/Tp67 and CD7/Tp41 T-lineage differentiation antigens in combination with the in vitro active cyclophosphamide congener 4-hydroperoxy-cyclophosphamide (4-HC) was used to purge autografts. Despite high dose pretransplant radiochemotherapy and effective purging of autografts, 9 of 14 patients relapsed at a median of 2.5 months (range, 1.2 to 16.8 months) post BMT. Two patients remain alive and disease free at 26 and 28 months post BMT. We used a novel quantitative minimal residual disease (MRD) detection assay, which combines fluorescence activated multiparameter flow cytometry and cell sorting with leukemic progenitor cell (LPC) assays, to analyze remission bone marrow (BM) samples from T-lineage ALL patients for the presence of residual LPCs. Notably, high numbers of residual LPC detected in remission BM before BMT constituted a poor prognostic indicator, providing the first evidence for the biologic significance and clinical value of in vitro T-lineage ALL LPC assays. The median value for the residual leukemia burden before BMT, was approximately 8.6 x 10(3) LPC/10(8) mononuclear cells (MNC) (approximately 0.0086% LPC). Patients with a residual leukemia burden less than this median value appeared to have a better outlook for remaining free of relapse after autologous BMT than patients with a greater leukemia burden (53 +/- 25% v 14 +/- 13%, P = .006, Mantel-Cox). By comparison, the log kill efficacy of purging, the remaining numbers of LPC in purged autografts, or the estimated numbers of reinfused LPC, did not correlate with the probability of disease-free survival (DFS). These results indicate that the primary reason for the recurrence of leukemia was inefficient pretransplant radiochemotherapy rather than inefficient purging of autografts.