Effects of anti-CD33 blocked ricin immunotoxin on the capacity of CD34+ human marrow cells to establish in vitro hematopoiesis in long-term marrow cultures

V. F. La Russa, J. D. Griffin, S. W. Kessler, M. A. Cutting, R. D. Knight, W. A. Blattler, J. M. Lambert, D. G. Wright

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24 Scopus citations

Abstract

Human marrow cells that express the CD34 antigen but lack CD33 are able to initiate sustained, multilineage in vitro hematopoiesis in long-term Dexter cultures and are believed to include the primitive stem cells responsible for effecting long-term hematopoietic reconstitution in vivo following marrow transplantation. In studies described in this report we investigated the effects of a novel anti-CD33 immunotoxin on the clonogenic potential of normal human CD34+ marrow cells and on the ability of these cells to initiate hematopoiesis in two-stage Dexter cultures (long-term marrow cultures, LTMC). This immunotoxin (anti-CD33-bR), shown previously to kill both clonogenic myelogenous leukemia cells and normal mature myeloid progenitor cells (granulocyte-macrophage colony-forming units, CFU-GM), consists of an anti-CD33 monoclonal antibody conjugated to purified ricin that has been modified by blocking the carbohydrate binding domains of the ricin B-chain to eliminate nonspecific binding. For our studies, normal CD34+ human marrow cells were isolated from the light-density (<1.070 g/ml) cells of aspirated marrow by positive selection with immunomagnetic beads linked to the monoclonal antibody K6.1. These cell isolates were highly enriched with both multipotential and lineage-restricted clonogenic, hematopoietic progenitors (mixed lineage colony-forming units, CFU-Mix; CFU-GM; and erythroid burst-forming units, BFU-E) which constituted ≥20% of the cells. Recovery of clonogenic progenitors from these CD34+ cell preparations, following treatment with anti-CD33-bR (10 nM), was reduced by ≥85% for CFU-GM and 20%-40% for CFU-Mix and BFU-E. However, the capacity of these cells to initiate hematopoietic LTMC was preserved. Indeed, the production of high proliferative potential (HPP) CFU-GM, BFU-E, and CFU-Mix in cultures seeded with 105 anti-CD33-bR-treated CD34+ marrow cells was substantially greater than that observed in LTMC seeded with equivalent numbers of untreated CD34+ cells. Moreover, concentrations of long-term culture initiating cells in CD34+ cell isolates, quantified by a limiting dilution technique, were found to be increased following anti-CD33-bR treatment. These findings support the potential usefulness of anti-CD33-bR for in vitro marrow purging or in vivo treatment to eliminate CD33+ leukemic clones, while sparing normal CD34+/CD33- stem cells that support normal hematopoiesis and hematopoietic reconstitution in vivo.

Original languageEnglish (US)
Pages (from-to)442-448
Number of pages7
JournalExperimental Hematology
Volume20
Issue number4
StatePublished - Jan 1 1992

Keywords

  • anti-CD33 immunotoxin
  • CD34
  • hematopoiesis
  • long-term marrow cultures
  • marrow transplantation

ASJC Scopus subject areas

  • Cancer Research
  • Cell Biology
  • Genetics
  • Hematology
  • Oncology
  • Transplantation

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    La Russa, V. F., Griffin, J. D., Kessler, S. W., Cutting, M. A., Knight, R. D., Blattler, W. A., Lambert, J. M., & Wright, D. G. (1992). Effects of anti-CD33 blocked ricin immunotoxin on the capacity of CD34+ human marrow cells to establish in vitro hematopoiesis in long-term marrow cultures. Experimental Hematology, 20(4), 442-448.