Caspase-3-dependent and caspase-3-independent pathways leading to chromatin DNA fragmentation in HL-60 cells

Apoptosis induced by etoposide (VP-16) in HL-60 cells was confirmed to be c aspase-dependent. It was fully inhibited by the broad-spectrum caspase inhi bitor Z-VAD-fmk. However, the caspase-3-specific inhibitor Z-DEVD-fmk only partially inhibited apoptosis. This indicated that a second caspase is requ ired in vivo for full activation of the apoptotic nucease CAD. Aurin tricar boxylic acid (ATA) did not inhibit VP-16-induced apoptosis. In contrast, ap optosis induced by hydroxychloroquine (HCQ) in HL-60 cells was caspase-3 in dependent and was fully inhibited by ATA. Thus, CAD does not appear to be i nvolved in chromatin DNA degradation in this case. A second apoptotic nucle ase is postulated to degrade the DNA, likely endo-exonuclease, an abundant nuclear enzyme that acts on both DNA and RNA and is present in latent form. HCQ, but not VP-16, stimulated DNA degradation ("laddering") in isolated n uclei. This indicates that the drug can act directly in the nuclei to trigg er activation of the second latent apoptotic nuclease.