Mechanisms of proteasome inhibitor PS-341-induced G₂-M-phase arrest and apoptosis in human non-small cell lung cancer cell lines

Purpose: PS-341 is a novel dipeptide boronic acid proteasome inhibitor with in vitro and in vivo antitumor activity that induces mechanisms of apoptosis by unknown mechanisms. Experimental Design: Human non-small cell lung cancer cell lines were used to investigate effects PS-341 on cell proliferation, cell cycle progression, and the induction of apoptosis. Results: PS-341 was 38-360-fold more cytotoxic against H460 cells when compared with the proteasome inhibitors MG-132 and PSI. Differential PS-341 cytotoxic effects were found with respect to P53 function: H322 cells (p53 mutant) were 6-fold less sensitive as compared with H460 cells (p53 wild type); and H358 cells (p53 null) were 1.6-fold more sensitive as compared with H460 cells (p53 wild type). A concentration- and time-dependent cell cycle blockade at G₂-M phase was seen for H460 cells without any direct effects on microtubule polymerization or depolymerization. PS-341 exposure in H460 cells led to stabilization of p53, induction of p21_cip/waf-1 and MDM2 expression, an increase in cyclin B and cyclin A, and the activation of cyclin B and cyclin A kinases. MDM2 induction was found only in H460 cells, whereas in H322 and H358 cells, G₂-M-phase arrest, p21^cip/waf-1 induction, and an increase in cyclin B1 were found. The commitment of G₂-M-phase cells to apoptosis was verified by the activation of caspase-3 and cleavage of poly(ADP-ribose) polymerase in drug-free medium. Conclusions: Our data suggest that the PS-341-induced G₂-M-phase arrest may be associated with the inhibition of degradation of cell cycle regulators and that the up-regulation of p21^cip/waf-1 expression may be via p53-dependent and/or -independent pathways. The resulting disturbance of cell cycle progression leads either to growth inhibition or to the initiation of apoptotic pathways.