Exploring the role of αB-crystallin in resistance to anti-angiogenic therapies

Anti-VEGF therapies have not improved the overall survival in breast cancer and many patients show no response to these treatments. The causes of this innate resistance need to be investigated so that they can be targeted in order improve the efficacy of these treatments. Additionally, identification of biomarkers can be utilised to select patients who are more likely to respond to treatments. αB-crystallin is a small heat shock protein encoded by the CRYAB gene and known to function as a chaperone protein. Its function is to protect misfolded proteins from degradation and apoptosis under stress conditions. αB-crystallin is thought to protect VEGF from degradation and support its stability. αB-crystallin is significantly upregulated in tumour vasculature during angiogenesis and after anti-VEGF treatment. Protecting VEGF from degradation and increasing its stability may stimulate tumour growth and contribute to resistance to anti-VEGF therapies. Therefore, this project tests the hypothesis that αB-crystallin contributes to the resistance to anti-VEGF therapies in breast cancer. To test this hypothesis, I made transgenic triple negative, MDA-MB-231, breast cancer cells that produce different levels of αB-crystallin (MDA-MB-231/CRYAB) and control cells that do not produce αB-crystallin (MDA-MB-231/WT). These cells were compared with triple negative breast cancer cells that naturally produce high amounts of αB-crystallin (MDA-MB- 468) and MDA-MB-468 cells in which levels of αB-crystallin have been reduced by siRNA. In vitro, VEGF production from breast cancer cells expressing different levels of αB-crystallin were measured by ELISA after heat shock (42°C/24h), or hypoxia (0.1% O2/24h) and sensitivity to doxorubicin induced apoptosis was measured by flow cytometry. In vivo: MDA-MB-231/WT and MDA-MB-231/CRYAB cells were administered by intra-ductal injection into BALB/c nude mice 7-days prior to PBS (control), 4 mg/kg/week doxorubicin, 7.5 mg/kg/3X per week bevacizumab or a combination of both. Tumour growth was ...