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Bryostatin-1 belongs to the family of macrocyclic lactones isolated from the marine bryozoan Bugula neritina and is a potent activator of protein kinase C (PKC). Bryostatin has been demonstrated to possess both in vivo and in vitro anti-leukaemic potential. In samples derived from chronic myeloid leukaemia (CML) patients, it has been demonstrated that bryostatin-1 induces a macrophage differentiation, suppresses colony growth in vitro and promotes cytokine secretion from accessory cells. We investigated the effect of bryostatin-1 treatment on colony-forming unit-granulocyte macrophage (CFU-GM) capacity in the presence of accessory cells, using mononuclear cells, as well as in the absence of accessory cells using purified CD34-positive cells. Cells were obtained from 14 CML patients as well as from nine controls. Moreover, CD34-positive cells derived from CML samples and controls were analysed for stem cell frequency and ability using the long-term culture initiating cell (LTCIC) assay at limiting dilution. Individual colonies derived from both the CFU-GM and LTCIC assays were analysed for the presence of the bcr-abl gene with fluorescence in situ hybridization (FISH) to evaluate inhibition of malignant colony growth. The results show that at the CFU-GM level bryostatin-1 treatment resulted in only a 1.4-fold higher reduction of CML colony growth as compared to the control samples, both in the presence and in the absence of accessory cells. However, at the LTCIC level a sixfold higher reduction of CML growth was observed as compared to the control samples. Analysis of the LTCICs at limiting dilution indicates that this purging effect is caused by a decrease in output per malignant LTCIC combined with an increase in the normal stem cell frequency. It is concluded that bryostatin-1 selectively inhibits CML growth at the LTCIC level and should be explored as a purging modality in CML. [ABSTRACT FROM AUTHOR]

The first step in the homing of hematopoietic progenitor cells (HPCs) from the peripheral blood to the bone marrow involves an adhesion molecule-dependent contact with human bone marrow endothelial cells (HBMECs). In the present study we describe the constitutive expression of two of these molecules, E-selectin and vascular cell adhesion molecule-1 (VCAM-1), on endothelial cells of hematopoietic tissues. Immunophenotypic analysis of tissue sections of hematopoietically active (human adult and fetal bone marrow, fetal spleen, fetal liver, and adult spleen with extramedullary hematopoiesis) and inactive tissues (human adult spleen, lymph node, appendix, and liver; and fetal lung and fetal intestine) revealed that E-selectin and VCAM-1 are selectively expressed on endothelial cells of adult and fetal hematopoietic organs. These results were validated by means of reverse-transcriptase polymerase chain reaction. Adhesion studies revealed that binding of normal mobilized peripheral blood HPCs to HBMECs was completely inhibited by preincubation of HBMECs with anti-E-selectin (ENA2), whereas no effect of anti-VCAM-1 (1G11B1) was detected. These results suggest that E-selectin plays a role in the homing of HPCs and that its constitutive expression on endothelial cells of hematopoietic organs may be essential in the initial step of the homing process.