Interplay between IGFBP-2 and GRP78 in MCF-7 breast cancer cells

: does it play a role in IGF signalling?

Abstract

As a member of the RTK family, the IGF-1R can activate multiple signalling pathways by binding with IGF-I and data shows that targeting the IGF pathway may be beneficial in preventing breast tumorigenesis. The PI3K-AKT signalling pathway is a conserved signalling pathway involved in mediating the effects of IGFs on cell growth and survival. Although IGF-I and the IGF-1R demonstrate a high degree of correlation with breast cancer development, therapies targeting this pathway have not been very successful. This work therefore aimed to understand the role of two molecules linked with cancer progression, GRP78 and IGFBP-2, in the proliferative effects of IGF-I, with a view to identifying new biomarkers or ways to improve targeting of the IGF axis.
My data showed that in MCF-7 breast cancer cells, IGF-I induced a proliferative effect on the cells, that was associated with an increase in the IGF-1R, IGFBP-2 and GRP78. Silencing IGFBP-2 blocked the proliferative effects of IGF-1, and the ability of IGF-I to increase levels of GRP78. Conversely silencing GRP78 enhanced the effect of IGF-I on cell growth, suggesting an inhibitory role for GRP78, that was associated with increased levels of IGFBP-2 in the cell supernatants, increased activation of the IGF-1R, inactivation of PTEN and enhanced phosphorylation of AKT.
Preliminary work was performed on the potential role of cell surface (cs) GRP78 in this model as increased expression of GRP78 translocation to the surface has been reported in tumour cells. Using an N-terminal directed csGRP78 blocking antibody, a dose-dependent decrease in metabolic activity was observed. And blocking antibodies against the N and C terminals of GRP78, compared with silencing GRP78 with siRNA, appeared to give different effects. Silencing GRP78 reduced total cell number, as did the N-terminal antibody but the C-terminal antibody had no effect. The antibody against the N-terminal of GRP78 also appeared to induce apoptosis.
In terms of signalling, whilst siRNA reduced levels of GRP78 as anticipated, blocking csGRP78 with the N-terminal antibody increased levels of GRP78 but were levels were decreased in the presence of the C-terminal antibody. Different impacts of these antibodies were also seen with levels of IGFBP-2 and the IGF-1R, levels of each reduced with the N- terminal antibody and increased with the C-terminal antibody.
In summary GRP78 and IGFBP-2 appear to play a role in the proliferative effects of IGF- I and the mechanism by which this occurs is still to be determined.

Date of Award	5 Dec 2023
Original language	English
Awarding Institution	University of Bristol
Supervisor	Claire M Perks (Supervisor), Kalina Biernacka (Supervisor), Richard M Martin (Supervisor) & Paul Verkade (Supervisor)