Role of stromal-derived IGF1 signalling in invasive lobular carcinoma

Abstract

Invasive lobular breast carcinoma (ILC) is the second most frequently diagnosed invasive breast cancer subtype. Besides E-cadherin loss, the prominent genetic hallmark of ILC, this subtype is characterised by unique pathologic, molecular, and clinical features, which differ from the other breast cancer subtypes. Recent studies have shown that loss of E-Cadherin, the hallmark of the ILC, sensitises the tumour cells to insulin-like growth factor 1 (IGF-1) signalling. However, the involvement of such activation in ILC tumourigenesis and metastasis remains unclear.

Here we aim to study this pathway further in human and mouse models of ILC and in a large cohort of patients with primary operable breast cancer with mature follow-up.

We initially characterised the pathway using quantitative RT-PCR, where significant expression of Igf1 was found in mouse ILC-derived cancer-associated fibroblasts (CAF), with no measurable expression in the mouse epithelial tumour cells. Both enzyme-linked immunosorbent assays (ELISA) and Forward phase protein arrays (FPPA) confirmed stromal secretion of IGF1 from the tumour cell-derived CAFs. Moreover, IGF-1 receptor stimulation in ILC tumour cells in response to recombinant IGF-1 or CAF-derived conditioned medium was observed. Such activation was successfully blocked upon treatment with an IGF1 receptor inhibitor.

Additionally, the effect of IGF-1 on human and mouse ILC cell proliferation was assessed in vitro, where we showed that both recombinant IGF-1 and CAF-derived conditioned medium resulted in the increased proliferation rate of the ILC tumour cells in comparison with the control cells. In contrast, adding an IGF-1 receptor inhibitor strongly reduced their proliferation.

As the role of the tumour microenvironment in tumour initiation and progression is increasingly recognised, an in vivo study of the ILC mouse model was performed. We injected ILC epithelial cells alone or with CAFs into mice. The co-injection with CAFs significantly accelerated tumour growth compared to mice injected with epithelial cells alone. Furthermore, significantly increased expression of Ki67 (proliferative marker) and CD31 (endothelial cell marker) was observed in the tumours of the co-injected mice.

This project also aimed to clarify the clinical significance of IGF-1 signalling in human breast cancer. We carried out immunohistochemical analysis in a large cohort of 850 samples of various breast cancer subtypes (lobular, ductal and others) for E-Cadherin, IGF-1 receptor and other members of the IGF-1 signalling pathway. These studies revealed differential patterns of expression of these proteins and highlighted their significant association with cancer-specific survival as well as with the other clinicopathological features of the patients.

To conclude, this work could provide deep insights into the importance of tumour-stromal IGF-1 signalling in breast cancer, particularly ILC tumourigenesis and metastasis. It will also help improve subsequent investigations and select patients for future targeting of IGF-1 signalling.