The
retinoid 4-oxo-N-(4-hydroxyphenyl)retinamide (4-oxo-4-HPR), a metabolite of
fenretinide (4-HPR) present in plasma of 4-HPR-treated patients, is very effective in inducing growth inhibition and apoptosis in several
cancer cell lines.
4-Oxo-4-HPR and
4-HPR have different mechanisms of action because
4-oxo-4-HPR, unlike
4-HPR, causes marked cell accumulation in G2-M phase. Here, we investigated the molecular events involving 4-oxo-4-HPR-induced cell cycle perturbation in ovarian (A2780 and IGROV-1) and breast (T47D,
estrogen receptor+ and BT-20,
estrogen receptor-)
cancer cells.
4-Oxo-4-HPR induced a delay of mitosis (with mitotic index increasing 5- to 6-fold in all cell lines) without progression beyond the anaphase, as shown by
cyclin B1 expression.
4-Oxo-4-HPR induced multipolar spindle formation and phosphorylation of BUBR1, resulting in activation of the spindle checkpoint. Multipolar spindles were not due to impairment of pole-focusing process, loss of centrosome integrity, or modulation of the expression levels of molecules associated with spindle aberrations (Kif 1C, Kif 2A, Eg5, Tara,
tankyrase-1,
centractin, and TOGp). We show here that
4-oxo-4-HPR targets microtubules because, in treated cells, it interfered with the reassembly of cold-depolymerized spindle microtubules and decreased the polymerized
tubulin fraction. In cell-free assays,
4-oxo-4-HPR inhibited
tubulin polymerization (50% inhibition of microtubule assembly at 5.9 micromol/L), suggesting a direct molecular interaction with
tubulin. In conclusion, by showing that
4-oxo-4-HPR causes mitotic arrest through antimicrotubule activities, we delineate a new molecular mechanism for a
retinoid.