Involvement of cyclin-dependent kinase activities in CD437-induced apoptosis

A novel synthetic retinoid, 6-[3-(l-adamantyl) -4-hydroxyphenyl]-2-naphthal ene carboxylic acid (CD437), is a selective ligand of the RAR gamma nuclear receptor. We examined the in vitro effects of CD437 and found that CD437 i nduces S phase arrest within 24 to 48 h, followed by cell death, in the p53 -negative Hep3B and the p53-positive HepG2 human hepatoma cell Lines. Based on observations of cellular and nuclear fragmentation, chromatin condensat ion, and DNA fragmentation, the CD437-mediated cell-killing effect appears to be due to apoptosis. On morphological examination, a number of CD437-tre ated cells were found to have increased 5-to 10-fold in size and persisted as single giant cells without cell division, while the remainder underwent nuclear division (multiple nuclei) but were unable to complete cytokinesis, and finally all died by apoptosis, In HepG2 cells that possessed wild-type p53, CD437-induced S phase arrest and apoptosis were accompanied by the up -regulation of cyclin A, cyclin B, p53, p21(CIP1/Wuf1), Bad, and Bcl-Xs pro teins and by a decrease in Bcl-2 protein levels. In Hep3B cells, CD437-medi ated S phase arrest and apoptosis were also associated with a concomitant u p-regulation of cyclin A, cyclin B, Bad, and Bcl-Xs. However, Hep3B cells d id not express p53 or Bcl-2 messages. Olomoucine and roscovitine, the poten t p34(cdc2) and CDK2 inhibitors, effectively blocked CD437-mediated cyclin A- and B-dependent kinase activation and prevented CD437-induced cell death . Furthermore, antisense oligonucleotide complementary to cyclin A and B mR NA significantly rescued CD437-induced apoptosis. These findings suggest th at activation of cyclin A- and B-dependent kinases is a critical determinan t of apoptotic death mediated by CD437. (C) 1999 Academic Press.