We assessed the ability of
cryptophycin 52 (
LY355703), a novel antimicrotubule, to induce growth arrest and apoptosis in
prostate cancer cell lines and investigated potential molecular mechanisms of death. LNCaP (
androgen-dependent) and DU-145 (
androgen-independent) cells accumulated in G(2)-M phase of the cell cycle and progressively acquired sub-G(0)-G(1)
DNA content after 48 h of exposure to
cryptophycin 52 (1-10 pM). Induction of apoptosis was confirmed by
DNA ladder formation and detection of cytoplasmic
nucleosomes. PC-3 (
androgen-independent) cells were less responsive to
cryptophycin 52-induced death. Apoptosis was associated with proteolytic processing and activation of the caspase-3-like subfamily
proteins caspase-3 and
caspase-7 and cleavage of the
caspase substrate
poly(ADP-ribose) polymerase. The pan-
caspase inhibitor BOC-Asp(OMe)-fluoromethylketone effectively reduced
cryptophycin 52-induced caspase-3-like
protease activity and apoptosis in DU-145 cells. In contrast, BOC-Asp(OMe)-fluoromethylketone did not inhibit apoptosis induction in LNCaP cells by
cryptophycin 52, even though both
cryptophycin 52-induced caspase-3-like activity and
staurosporine-induced death were blocked under identical conditions.
Cryptophycin 52 induced phosphorylation of c-raf1 and bcl-2 and/or bcl-x(L) to comparable levels in all cell lines studied, and LNCaP cells overexpressing bcl-2 were more resistant to
cryptophycin 52-induced apoptosis. Up-regulation of p53, bax, and p21 expression was induced in wild-type p53-expressing LNCaP cells only after
cryptophycin 52 exposure. A sustained increase in c-Jun NH(2)-terminal
kinase phosphorylation was also observed, the levels of which strongly correlated with apoptosis. We conclude that apoptosis induced by
cryptophycin 52 in
prostate cancer cells is
androgen status independent, cell type specific for
caspase requirement, modulated by the bcl-2 family, linked to but not dependent on p53, and strongly correlated with c-Jun NH(2)-terminal
kinase phosphorylation.
Cryptophycin 52-induced apoptosis in
prostate cancer cells is therefore associated with multiple cell line-specific alterations in apoptosis-associated
proteins and pathways.