by ecancer reporter Clare Sansom

The prognosis for women diagnosed with breast cancer has increased significantly in recent years, as treatment options have improved and more tumours have been identified early.

However, about 40% of patients with primary breast cancer still eventually relapse with metastatic disease, which is much harder to treat.

Small populations of cells within breast tumours that have stem cell like characteristics have been implicated in the development of resistance to therapy and thus in breast cancer recurrence.

Breast cancer stem cells (BCSCs) are more resistant to radiotherapy than other breast cancer cells; the proportion of these cells within a tumour increases after chemotherapy, as the tumour itself decreases in size.

The herceptin receptor (HER2) is over-expressed in about a quarter of all breast cancers.

These herceptin receptor positive (HER2⁺) cancers are usually aggressive but are amenable to treatment with the anti-HER2 antibody trastuzumab.

Some oestrogen receptor positive (ER⁺) tumours become amenable to trastuzumab therapy when they become resistant to hormone therapy, indicating that the HER2 receptor can become over-expressed in response to therapy in tumours that were originally HER2 negative (HER2^-/low).

The cells that express HER2 at the highest levels in resistant HER2^-/low tumours are those with stem-like characteristics.

Jian Jian Li of the University of California Davis, Sacramento, California, USA and his colleagues have investigated the mechanism through which HER2-associated signalling in BCSCs is associated with the development of resistance to therapy in HER2^-/low breast cancers.

Li and his co-workers first exposed both previously irradiated HER2^-/low breast tumour cells and a radio-resistant breast cancer cell line to ionizing radiation and confirmed that HER2 proteins were over-expressed in the cells that survived this radiation.

The researchers then identified HER2-expressing breast cancer cells with stem-like characteristics (HER2^-/low/CD44⁺/CD24^-/low cells) in these populations and isolated them for further analysis.

Before irradiation, about 1% of the cells in both populations were HER2^-/low/CD44⁺, but this proportion increased significantly afterwards.

These HER2-expressing stem cells were found using gap filling, Matrigel invasion, tumor-sphere formation and clonogenic survival assays to be more aggressive and invasive than HER2 negative cells with similar characteristics.

This aggressive phenotype could be reduced by inhibition of HER2 expression, either by treatment with trastuzumab or by reducing HER2 gene expression using siRNA.

Forty tissue samples were randomly selected from breast tumours diagnosed as of primary, metastatic or unknown status at the Emory University School of Medicine in Atlanta, Georgia for analysis of expression levels of HER2 and stem cell marker proteins. 

Samples taken from metastatic tumours were found to contain more cells expressing high levels of HER2 and CD44 proteins than those from primary breast tumours.

The researchers then compared the protein expression profiles of breast cancer stem cells that express HER2 (HER2⁺/CD44⁺/CD24^-/low cells) from those that do not (HER2^-/CD44⁺/CD24^-/low cells) using 2D gel electrophoresis and mass spectroscopy.

A total of 499 proteins were found to be differentially up-regulated in HER2⁺/CD44⁺ /CD24^-/low cells as compared to HER2^-/CD44⁺/CD24^-/low cells; 182 proteins were differentially up-regulated in the HER2 negative stem cells.

Proteins found to be differentially up-regulated in the HER2 positive stem cells included the signal transduction proteins STAT3 and Src.

HER2 is known to activate STAT3 in tumour cells through both JAK2- and Src- dependent pathways, leading to an aggressive tumour phenotype.

The researchers used Western blotting and immunofluorescence to confirm that STAT3 and HER2 were co-expressed in HER2 positive but not in HER2 negative stem cells.

Taken together, these results suggest that activation of the HER2-STAT3 signalling pathway in a subpopulation of stem cells is be responsible for the induction of an aggressive, radiation resistant phenotype in breast tumours that were originally oestrogen receptor positive and HER2 negative as they become resistant to therapy.

It is possible that this network will prove to be a useful drug target for recurrent oestrogen receptor positive breast tumours.

Reference

Duru, N., Fan, M., Candas, D. and 12 others (2012). HER2-Associated Radioresistance of Breast Cancer Stem Cells Isolated from HER2-Negative Breast Cancer Cells. Clin. Cancer Res. 18(24), 6634–47. doi: 10.1158/1078-0432.CCR-12-1436