EFFECTS OF NORCANTHARIDIN, A PROTEIN PHOSPHATASE TYPE-2A INHIBITOR, ON THE GROWTH OF NORMAL AND MALIGNANT HEMATOPOIETIC-CELLS

Cantharidin is a natural toxin that inhibits protein phosphatase type 2A (PP2A) and has antitumour effects in man. We have studied the synth etic analogue, norcantharidin (NCTD), which has less nephrotoxic and p hlogogenic side-effects, investigating the effects on the normal haemo poietic system and leukaemia cell growth. Daily intraperitoneal (i.p.) injection of NCTD induced dose and circadian time-dependent transient leucocytosis in normal mice, but did not accelerate bone marrow (BM) regeneration, or have haemopoietic side-effects following chronic admi nistration. NCTD stimulated the cell cycle progression of granulocyte- macrophage colony-forming cells (GM-CFC), stimulated DNA synthesis and increased the frequency of mitotic cells in short-term human BM cultu res. NCTD also stimulated the production of interleukin (IL)-1 beta, c olony stimulating activity (CSA) and tumour necrosis factor (TNF)-alph a. Continuous in vitro NCTD treatment, however, inhibited both DNA syn thesis and GM-CFC growth. Fluorescence-activated cell sorting (FAGS) a nalysis of DNA profiles and cytological studies in HL-60, K-562 or MRC 5V2 (fibroblast) cells indicated that low doses of NCTD accelerated th e G(1)/S phase transition, while higher doses or prolonged incubations inhibited the cell cycle at the G(2)/M phases or during the formation of postmitotic daughter cells. Electron microscopy revealed that NCTD impaired the neogenesis of chromatin material and nuclear membrane du ring the M/G(1) phase transition in K-562 cells. The biphasic effect o f NCTD may be due to inhibition of PP2A activity, which regulates the cell cycle, both at the restriction point and at the G2 and M phases. Our data provide new insight into the cellular and molecular actions o f NCTD, and partly explain its therapeutical effects in cancer patient s.