A role for caspase-mediated proteolysis of structural and adhesion proteins has been suggested in the morphological changes that characterise apoptotic cell death. After the initial phase of contraction and blebbing, caspase-mediated cleavage of actin monomers SAHA HDAC probably induces disassembly of actin filaments (Mashima et al, 1997; McCarthy et al, 1997). We found that after 48h of low concentration ZOL treatment, significant actin-cytoskeletal reorganization occurs in PC cells as demonstrated by actin staining. These modifications indicate that ZOL induces actin rearrangements into cortical rings and that these events may drive the cells to the apoptotic process. This specific effect indicates the intriguing possibility of positive interactions of BPs with drugs that interfere with actin polymerisation and depolymerisation such as taxoid or vinca alkaloid agents.

A synergistic effect of ZOL with paclitaxel has in fact been demonstrated in human breast cancer (Jagdev et al, 2001). Recent reports have also highlighted the role of signal transduction pathways controlled by the Rho family of small GTPases in regulating the architecture of the actin cytoskeleton and the morphological changes during apoptotic cell death (Coleman and Olson, 2002). An intriguing preliminary finding is that the small GTP-binding protein RAC, functionally related to cytoskeletal organisation, discloses enhanced (10-fold) expression in ZOL-exposed cancer cells as detected by Western blot analysis (data not shown). Based on these findings, further studies are presently in progress in order to understand the functional significance of these observations.

In conclusion, we have demonstrated that PC cells are highly sensitive to ZOL-induced growth perturbation and induction of apoptosis, which is caspase-9- caspase-6- and PARP-dependent. Moreover, our experimental findings suggest that inhibition of p21ras/Raf-1/MEK1/ERK signalling as well as PK-B/Akt inhibition might be relevant for the antitumour effects of ZOL. Although we have been capable of demonstrating that a short-term exposure is enough for activation of apoptosis, we think that a pharmacokinetic profile more relevant to the extra-bone antitumour effects of ZOL should be derived and might involve entrapping of BPs in lysosomes. An additional topic of current investigation is BP combination with cytotoxic drugs or selective signal transduction inhibitors.

Acknowledgments P Tassone was supported by a fellowship from FIRC (Fondazione Italiana per la Ricerca sul Cancro). This work AV-951 was supported in part by funds from Regione Calabria (POP 94/99) (Italy), MURST-CNR Biotechnology Program L. 95/95 (Italy), MURST (Cofin 98, 99), Consiglio Nazionale delle Ricerche (Special Project Biotechnology) and by a grant from the Italian Ministry of Health, FSN 2000.