MODULATION OF THE ACTIVITY OF A HUMAN GRANULOCYTE-MACROPHAGE COLONY-STIMULATING FACTOR INTERLEUKIN-3 FUSION PROTEIN (PIXY-321) BY THE MACROCYCLIC LACTONE PROTEIN-KINASE-C ACTIVATOR BRYOSTATIN-1

We have examined the effect of the macrocyclic lactone protein kinase C (PK-C) activator bryostatin 1 on the proliferative capacity and line age commitment of CD34(+) human bone marrow cells exposed to the granu locyte-macrophage colony-stimulating factor/interleukin-3 (GM-CSF/IL-3 ) fusion protein pIXY 321, pIXY 321 administered at a dose of 10 ng/mL was as effective as the combination of plateau concentrations of reco mbinant (r) IL-3 and rGM-CSF (e.g., 50 ng/mL) in stimulating the growt h of day-14 committed myeloid progenitors (colony-forming units granul ocyte/macrophage [CFU-GM]). In the large majority of samples tested, c oadministration of 0.5 to 100 nM bryostatin 1 with either pIXY 321 or the combination of rIL-3 plus rGM-CSF led to modest but significant in creases (e.g., 30 to 75%) in the number of CFU-GM, compared to adminis tration of growth factors alone. The degree of bryostatin 1-induced po tentiation, however, was considerably less than that previously observ ed in the case of cells exposed to either rIL-3 or rGM-CSF, where incr eases of 100 to 150% were regularly noted. While at least 50% of day-1 4 CFU-GM stimulated by either pIXY 321 or the combination of rIL-3 plu s rGM-CSF were of the pure or mixed eosinophilic variety, coadministra tion of bryostatin 1 resulted in a dramatic inhibition of eosinophilic colonies and a corresponding increase in pure and mixed neutrophil an d macrophage colonies. Although coadministration of recombinant granul ocyte colony-stimulating factor (rG-CSF) or recombinant colony-stimula ting factor-1 (rCSF-1) mimicked the capacity of bryostatin 1 to increa se the total number of pIXY 321-induced day-14 CFU-GM, these growth fa ctors, unlike bryostatin 1, were not capable of inhibiting eosinophili c colony formation. Furthermore, whereas addition of neutralizing anti bodies to G-CSF or CSF-1 blocked the capacity of these growth factors to potentiate colony formation in the presence of pIXY 321, it did not abrogate the effect of bryostatin 1 on progenitor cell growth or line age commitment. Finally, in contrast to its effects on committed myelo id progenitors, bryostatin 1 did not increase the growth of erythroid (burst-forming units-erythroid [BFU-E]) and multipotent (multipotent c olony-forming units [CFU-GEMM]) progenitors stimulated by pIXY 321, bu t instead inhibited colony formation at higher concentrations (e.g., 1 0 to 100 nM). Together, these findings indicate that bryostatin 1 exer ts pleiotropic effects on the proliferative capacity and lineage commi tment of human hematopoietic progenitors exposed to the novel hybrid c ytokine pIXY 321, and provide further evidence of a role for PK-C in t he regulation of hematopoiesis vitro.