BRYOSTATIN-1 ACTIVATES SPLENIC LYMPHOCYTES AND INDUCES SUSTAINED DEPLETION OF SPLENOCYTE PROTEIN-KINASE-C ACTIVITY IN-VIVO AFTER A SINGLE INTRAVENOUS ADMINISTRATION

Bryostatin 1 activates and subsequently down-regulates protein kinase C (PKC) in vitro and has potential use as an immunomodulator and as an anti-cancer agent. Despite extensive examination of its activities in vitro and anti-tumor effects in vivo, previous studies have failed to document that bryostatin 1 modulates total cellular PKC activity in t umor or normal tissues when administered in vivo, After a single bolus injection of bryostatin 1 (1.0 mu g) in normal C57BI/6 mice, blood wa s drawn at various intervals and assayed for bryostatin 1 levels. In a ddition, spleens from bryostatin-treated mice were harvested 10 min to 10 days after treatment, weighed and analyzed for cell numbers, PKC a ctivity and cell surface phenotypes, Bryostatin 1 levels in plasma ros e rapidly, reaching peak levels of 56.5 nM less than 1 min after injec tion, and then declined to undetectable levels by 1 h, A similar patte rn was observed when bryostatin 1 was incubated with leukemia cells in vitro, raising the possibility that the rapid fall in plasma levels r esults from intracellular uptake and binding. Bryostatin 1 induced mar ked depletion of total splenocyte PKC activity (as much as 69% relativ e to control values) at 24-96 h after drug administration, but not at earlier times (i.e. 1 h), A single injection of bryostatin 1 also indu ced expression of the T cell activation marker CD69, leading to positi vity in 53% of cells at 3-24 h versus 11% in control mice, and resulte d in marked splenomegaly, associated with increased numbers of nucleat ed cells at 48-96 h, Together, these studies demonstrate that despite rapid disappearance of the drug from plasma, a single i.v. dose of bry ostatin 1 exhibits significant and sustained effects on normal murine spleen cells, including early lymphocyte activation, prolonged depleti on of PKC activity, spenocyte proliferation and splenomegaly. These fi ndings may have implications for attempts to understand the in vivo ef fects of bryostatin 1 in normal host tissues.