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

Abstract

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.

Effect of stem cell factor (c‐ kit ligand), granulocyte‐macrophage colony stimulating factor and interleukin 3 on hematopoietic progenitors in human long‐term bone marrow cultures

“…In control cultures an average of 27.7 rt 10% SD of CFU-GM in S phase within the stromal layer was observed at time of feeding (i.e., seven days after the last medium addition). This result is consistent with the data presented in our previous study [18]. The higher percentage of cycling progenitors compared with the previous study could be explained by the use of a different culture technique and by the lack of distinction among various myeloid colonies with regard to their size.…”

“…The cultures were established as previously described [18]. Briefly, red blood cell (RBC) depleted BM aspirates at concentrations of 3 x lo6 cells/ml were inoculated either in 25 cm2 tissue culture flasks (Falcon, Becton-Dickinson, Rutherford, NJ) or in gas-permeable plastic bags with a 300 ml capacity ("Lifecell" tissue culture [TC] bags; Fenwall Lab., Deerfield, IL).…”

Murine and humanized constructs of monoclonal antibody m195 (anti-cd33) for the therapy of acute myelogenous leukemia

“…Because M195 kills target cells with guinea pig and rabbit complement,' M195 has been used for ex vivo bone marrow purging of AML. 4 Although M195 is not intrinsically cytotoxic in vitro, an initial pilot trial of 10 patients with myeloid leukemias showed rapid and specific bone marrow targeting and efficient internalization of 1311-M195 into blast^.^ Using an optimal biologic dose of antibody for radioimmunotherapy from this original trial, a dose escalation trial of 13'I-M195 for relapsed or refractory myeloid leukemias was conducted and the results are described in this report. Safe, leukemic cytoreduction was seen with 13'I-M195.…”