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Anti-tumour activity of tivozanib, a pan-inhibitor of VEGF receptors, in therapy-resistant ovarian carcinoma cells

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ABSTRACT

Epithelial ovarian cancer (EOC) is the most fatal gynaecological malignancy. Despite initial therapeutic response, the majority of advanced-stage patients relapse and succumb to chemoresistant disease. Overcoming drug resistance is the key to successful treatment of EOC. Members of vascular endothelial growth factor (VEGF) family are overexpressed in EOC and play key roles in its malignant progression though their contribution in development of the chemoresistant disease remains elusive. Here we show that expression of the VEGF family is higher in therapy-resistant EOC cells compared to sensitive ones. Overexpression of VEGFR2 correlated with resistance to cisplatin and combination with VEGFR2-inhibitor apatinib synergistically increased cisplatin sensitivity. Tivozanib, a pan-inhibitor of VEGF receptors, reduced proliferation of the chemoresistant EOC cells through induction of G2/M cell cycle arrest and apoptotic cell death. Tivozanib decreased invasive potential of these cells, concomitant with reduction of intercellular adhesion molecule-1 (ICAM-1) and diminishing the enzymatic activity of urokinase-type plasminogen activator (uPA) and matrix metalloproteinase-2 (MMP-2). Moreover, tivozanib synergistically enhanced anti-tumour effects of EGFR-directed therapies including erlotinib. These findings suggest that the VEGF pathway has potential as a therapeutic target in therapy-resistant EOC and VEGFR blockade by tivozanib may yield stronger anti-tumour efficacy and circumvent resistance to EGFR-directed therapies.

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Tivozanib inhibits proliferation, clonal growth and anoikis resistance.(A) The effect of tivozanib on cell viability was estimated by MTT assay. (B,C) Clonogenic assay was conducted to evaluate the effect of tivozanib on clonal proliferation. (D) Anoikis resistance assay was performed with cell culture on poly-HEMA–coated culture dishes for 48 h and the proportion of viable cells was measured by MTT assay. (E) qRT-PCR was carried out to determine whether tivozanib-mediated decrease in the surviving fraction is due to down-regulation of the anoikis resistance marker BCL2. Gene expression levels were normalized to HPRT1. Data are given as mean ± SD, normalized to the vehicle-treated control group. Statistically significant values of *p < 0.05, **p < 0.01, and ***p < 0.001 were determined compared with the control.
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f2: Tivozanib inhibits proliferation, clonal growth and anoikis resistance.(A) The effect of tivozanib on cell viability was estimated by MTT assay. (B,C) Clonogenic assay was conducted to evaluate the effect of tivozanib on clonal proliferation. (D) Anoikis resistance assay was performed with cell culture on poly-HEMA–coated culture dishes for 48 h and the proportion of viable cells was measured by MTT assay. (E) qRT-PCR was carried out to determine whether tivozanib-mediated decrease in the surviving fraction is due to down-regulation of the anoikis resistance marker BCL2. Gene expression levels were normalized to HPRT1. Data are given as mean ± SD, normalized to the vehicle-treated control group. Statistically significant values of *p < 0.05, **p < 0.01, and ***p < 0.001 were determined compared with the control.

Mentions: MTT assay was carried out to determine the effect of tivozanib on proliferation of the therapy-resistant EOC cells. Treatment of these cells with tivozanib reduced their proliferation (Fig. 2A). Moreover, tivozanib diminished their clonogenic growth (Fig. 2B,C). In immortalized cells, detachment from the extracellular matrix induces anoikis, a special type of apoptosis29. Acquisition of resistance to anoikis is a prerequisite for EOC cells to survive in the ascites before forming metastatic foci30. Using an anoikis resistance assay, we found that tivozanib reduced surviving fraction of the EOC cells (Fig. 2D). Tivozanib decreased the expression of the anoikis resistance marker BCL231, suggesting that the diminished surviving fraction by tivozanib is due to enhanced anoikis (Fig. 2E).


Anti-tumour activity of tivozanib, a pan-inhibitor of VEGF receptors, in therapy-resistant ovarian carcinoma cells
Tivozanib inhibits proliferation, clonal growth and anoikis resistance.(A) The effect of tivozanib on cell viability was estimated by MTT assay. (B,C) Clonogenic assay was conducted to evaluate the effect of tivozanib on clonal proliferation. (D) Anoikis resistance assay was performed with cell culture on poly-HEMA–coated culture dishes for 48 h and the proportion of viable cells was measured by MTT assay. (E) qRT-PCR was carried out to determine whether tivozanib-mediated decrease in the surviving fraction is due to down-regulation of the anoikis resistance marker BCL2. Gene expression levels were normalized to HPRT1. Data are given as mean ± SD, normalized to the vehicle-treated control group. Statistically significant values of *p < 0.05, **p < 0.01, and ***p < 0.001 were determined compared with the control.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5382685&req=5

f2: Tivozanib inhibits proliferation, clonal growth and anoikis resistance.(A) The effect of tivozanib on cell viability was estimated by MTT assay. (B,C) Clonogenic assay was conducted to evaluate the effect of tivozanib on clonal proliferation. (D) Anoikis resistance assay was performed with cell culture on poly-HEMA–coated culture dishes for 48 h and the proportion of viable cells was measured by MTT assay. (E) qRT-PCR was carried out to determine whether tivozanib-mediated decrease in the surviving fraction is due to down-regulation of the anoikis resistance marker BCL2. Gene expression levels were normalized to HPRT1. Data are given as mean ± SD, normalized to the vehicle-treated control group. Statistically significant values of *p < 0.05, **p < 0.01, and ***p < 0.001 were determined compared with the control.
Mentions: MTT assay was carried out to determine the effect of tivozanib on proliferation of the therapy-resistant EOC cells. Treatment of these cells with tivozanib reduced their proliferation (Fig. 2A). Moreover, tivozanib diminished their clonogenic growth (Fig. 2B,C). In immortalized cells, detachment from the extracellular matrix induces anoikis, a special type of apoptosis29. Acquisition of resistance to anoikis is a prerequisite for EOC cells to survive in the ascites before forming metastatic foci30. Using an anoikis resistance assay, we found that tivozanib reduced surviving fraction of the EOC cells (Fig. 2D). Tivozanib decreased the expression of the anoikis resistance marker BCL231, suggesting that the diminished surviving fraction by tivozanib is due to enhanced anoikis (Fig. 2E).

View Article: PubMed Central - PubMed

ABSTRACT

Epithelial ovarian cancer (EOC) is the most fatal gynaecological malignancy. Despite initial therapeutic response, the majority of advanced-stage patients relapse and succumb to chemoresistant disease. Overcoming drug resistance is the key to successful treatment of EOC. Members of vascular endothelial growth factor (VEGF) family are overexpressed in EOC and play key roles in its malignant progression though their contribution in development of the chemoresistant disease remains elusive. Here we show that expression of the VEGF family is higher in therapy-resistant EOC cells compared to sensitive ones. Overexpression of VEGFR2 correlated with resistance to cisplatin and combination with VEGFR2-inhibitor apatinib synergistically increased cisplatin sensitivity. Tivozanib, a pan-inhibitor of VEGF receptors, reduced proliferation of the chemoresistant EOC cells through induction of G2/M cell cycle arrest and apoptotic cell death. Tivozanib decreased invasive potential of these cells, concomitant with reduction of intercellular adhesion molecule-1 (ICAM-1) and diminishing the enzymatic activity of urokinase-type plasminogen activator (uPA) and matrix metalloproteinase-2 (MMP-2). Moreover, tivozanib synergistically enhanced anti-tumour effects of EGFR-directed therapies including erlotinib. These findings suggest that the VEGF pathway has potential as a therapeutic target in therapy-resistant EOC and VEGFR blockade by tivozanib may yield stronger anti-tumour efficacy and circumvent resistance to EGFR-directed therapies.

No MeSH data available.


Related in: MedlinePlus