Biological Evidence for Dual Antiangiogenic – Antiaromatase Activity of the VEGF-R Inhibitor PTK787/ZK222584.

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Abstract Background: Targeting the Vascular Endothelial Growth Factor (VEGF) and Estrogen Receptor (ER) signaling pathways concomitantly may provide enhanced therapeutic benefit in ER positive breast cancer. We have previously shown the VEGFR tyrosine kinase inhibitor, PTK787/ZK222584 (PTK/ZK) is a competitive aromatase inhibitor in vitro (Banerjee et al Cancer Res 2009 69: 4716-4723). Here we investigated whether (1) PTK/ZK shows both anti-angiogenic and aromatase inhibitory properties in in vivo models of ER positive breast cancer and (2) the combination of PTK/ZK and letrozole (LET) is superior to letrozole alone.Methods: Human breast cancer cell lines were engineered to express aromatase. Both MCF7 AROM1 (ER+HER2-) and BT474 AROM3 (ER+, HER2+) cells were grown as subcutaneous xenografts with androstenedione support in ovariectomized, nude mice. Mice bearing MCF7 AROM1 or BT474 AROM3 tumors were randomised to receive daily, by oral gavage, concentrations of vehicle, PTK/ZK (25, 50, or 100mg/kg), letrozole (1 mg/kg) or PTK/ZK (50mg/kg) in combination with letrozole (1 mg/kg) for 28 days. Mice were injected with FITC-lectin prior to sacrifice at weekly intervals. Tumor sections were stained for Ki67, apoptosis (TUNEL), CD31 and PgR. Mice bearing BT474 AROM3 xenografts were also treated twice daily with either vehicle or 50 mg/kg PTK/ZK for 3 days and uteri were then weighed and TFF1 and mRNA expression determined.Results: In MCF7 AROM1 xenografts, the daily growth rate was significantly reduced in all treatment groups compared to vehicle (p<0.05). Whilst letrozole caused tumor regression and PTK/ZK at 50 mg/kg and 100 mg/kg stabilized tumor volumes, there was no significant differences between the tumor volumes observed in the 50 mg/kg PTK/ZK and letrozole groups (p=0.14) or letrozole and combination group (p=0.14). The cell turnover index (CTI), a composite measurement of both proliferation and apoptosis significantly decreased (>50%) after 1 week with all treatments (p<0.05) except for 25 mg/kg PTK/ZK compared to the vehicle group The addition of PTK/ZK to letrozole did not lower the CTI further compared to letrozole alone. The vessel density was significantly reduced in the PTK/ZK (50 mg/kg and 100mg/kg) and combination groups compared to the vehicle group from as early as day 7 (p<0.05). However, in the letrozole group, there was no significant reduction in the vessel number until day 28 (p<0.05). PTK/ZK also significantly decreased the growth rate and vessel number compared to the vehicle (p<0.05) in BT474 AROM3 xenografts. PTK/ZK decreased PgR protein expression (p<0.01), TFF1 expression (p=0.002) and uterine weight (p=0.02) suggesting that PTK/ZK decreases E2 signaling in vivo.Conclusions: The VEGF-R inhibitor PTK/ZK showed effects on E2-dependent gene expression consistent with aromatase inhibition as well as anti-angiogenesis in xenograft models of breast cancer. Letrozole alone showed highly effective antitumor activity and the combination of PTK/ZK with letrozole was not superior. Nonetheless these results provide further support for a potential therapeutic approach of dual inhibition of VEGF and E2 signaling using a single agent. Citation Information: Cancer Res 2009;69(24 Suppl):Abstract nr 5139.