3 research outputs found
A genistein derivative, ITB-301, induces microtubule depolymerization and mitotic arrest in multidrug-resistant ovarian cancer
PURPOSE: To investigate the mechanistic basis of the anti-tumor effect of the compound ITB-301. METHODS: Chemical modifications of genistein have been introduced to improve its solubility and efficacy. The anti-tumor effects were tested in ovarian cancer cells using proliferation assays, cell cycle analysis, immunofluorescence, and microscopy. RESULTS: In this work, we show that a unique glycoside of genistein, ITB-301, inhibits the proliferation of SKOv3 ovarian cancer cells. We found that the 50% growth inhibitory concentration of ITB-301 in SKOv3 cells was 0.5Ā Ī¼M. Similar results were obtained in breast cancer, ovarian cancer, and acute myelogenous leukemia cell lines. ITB-301 induced significant time- and dose-dependent microtubule depolymerization. This depolymerization resulted in mitotic arrest and inhibited proliferation in all ovarian cancer cell lines examined including SKOv3, ES2, HeyA8, and HeyA8-MDR cells. The cytotoxic effect of ITB-301 was dependent on its induction of mitotic arrest as siRNA-mediated depletion of BUBR1 significantly reduced the cytotoxic effects of ITB-301, even at a concentration of 10Ā Ī¼M. Importantly, efflux-mediated drug resistance did not alter the cytotoxic effect of ITB-301 in two independent cancer cell models of drug resistance. CONCLUSION: These results identify ITB-301 as a novel anti-tubulin agent that could be used in cancers that are multidrug resistant. We propose a structural model for the binding of ITB-301 to Ī±- and Ī²-tubulin dimers on the basis of molecular docking simulations. This model provides a rationale for future work aimed at designing of more potent analogs
Activities of Topoisomerase I in Its Complex with SRSF1
Human DNA topoisomerase I (topo I) catalyzes DNA relaxation
and
phosphorylates SRSF1. Whereas the structure of topo I complexed with
DNA has been resolved, the structure of topo I in the complex with
SRSF1 and structural determinants of topo I activities in this complex
are not known. The main obstacle to resolving the structure is a contribution
of unfolded domains of topo I and SRSF1 in formation of the complex.
To overcome this difficulty, we employed a three-step strategy: identifying
the interaction regions, modeling the complex, and validating the
model with biochemical methods. The binding sites in both topo I and
SRSF1 are localized in the structured regions as well as in the unfolded
domains. One observes cooperation between the binding sites in topo
I but not in SRSF1. Our results indicate two features of the unfolded
RS domain of SRSF1 containing phosphorylated residues that are critical
for the kinase activity of topo I: its spatial arrangement relative
to topo I and the organization of its sequence. The efficiency of
phosphorylation of SRSF1 depends on the length and flexibility of
the spacer between the two RRM domains that uniquely determine an
arrangement of the RS domain relative to topo I. The spacer also influences
inhibition of DNA nicking, a prerequisite for DNA relaxation. To be
phosphorylated, the RS domain has to include a short sequence recognized
by topo I. A lack of this sequence in the mutants of SRSF1 or its
spatial inaccessibility in SRSF9 makes them inadequate as topo I/kinase
substrates