Topoisomerase (
Topo) IIalpha has proven to be an adequate anticancer target for
tumors expressing this
enzyme. In this study, we elucidated the effect of 2-[4-(7-chloro-2-quinoxalinyloxyphenoxy]-
propionic acid (
XK469; a new
Topo IIbeta inhibitor) in the modulation of
Topo IIalpha levels and sensitivity to
Topo IIalpha
poisons. We demonstrate by Western blot analysis that indolent B-cell
tumors express undetectable levels of this
enzyme and are refractory to the effects of
Topo IIalpha
poisons such as
VP16. Using the
Waldenstrom's macroglobulinemia (WM) cell line WSU-WM, we show that
XK469 induced the expression of
Topo IIalpha
protein by 24 h compared with control. Immunofluorescence studies by confocal microscopy using a specific
monoclonal antibody against
Topo IIalpha supported the immunoblot findings with high intensity staining in
XK469-exposed cells. To determine the effect of up-regulating
Topo IIalpha on sensitivity of
Topo IIalpha-directed inhibitors, WSU-WM cells were exposed to simultaneous, sequential, and reverse order
XK469 and
VP16. We demonstrate that 24 h of exposure to
XK469 before
VP16 resulted in a maximum synergistic response. In contrast, simultaneous or reverse order exposure resulted in an antagonistic effect. A similar trend was observed with cells obtained from
chronic lymphocytic leukemia patients, but not in normal lymphocytes. This increase in
VP16 sensitivity after 24 h of
XK469 exposure was associated with
VP16-dependent DNA cleavage, as demonstrated by formation of a smeared
DNA band in a SDS-KCL DNA cleavage assay. From this study, we concluded that
XK469 up-regulates
Topo IIalpha levels and consequently sensitizes indolent malignant B cells to the cytotoxic effect of
VP16 in a schedule-dependent manner.