Abstract
Breast cancer is the leading cause of cancer and cancer-related mortality in women globally and is the highest reported female cancer in New Zealand. Whilst a range of known risk factors are associated with the development of breast cancer, these do not explain all aspects of breast cancer epidemiology. It has been hypothesised that some breast cancers may be caused by late exposure to a common virus such as human cytomegalovirus (HCMV). This monocyte- attracted virus may use monocytes/macrophages as a vector for dissemination to tumour tissue, and can influence the differentiation of macrophages into a distinct macrophage phenotype, promoting a pro-inflammatory, pro-tumoural microenvironment within the breast tissue. The aim of this project was to investigate the presence of an HCMV gene product in human breast cancer cells and/or stromal cells within the breast tumour microenvironment of three clinically important breast cancer subtypes; ER/PR+, HER2-overexpressing and triple negative breast cancer. Utilising the RNAscope® in situ hybridisation assay, we aimed to detect the presence of the HCMV long non-coding RNA4.9 (lncRNA4.9) within formalin-fixed paraffin embedded tissue samples of breast cancer patients (n=75), representative of the three clinical subtypes, and normal breast tissue from mammary reduction surgery patients (n=5). Additionally, standard immunohistochemical staining techniques were used to identify the number and location of macrophages within the tumour microenvironment of breast tissue samples. This study showed that RNAscope® was able to detect HCMV lncRNA4.9 within normal and neoplastic breast tissue. HCMV was detected in different tissues and anatomical structures within ER+/PR+, HER2-overexpressing, and triple negative breast cancer. HCMV positive breast tissue structures included invasive tumour tissue and ductal carcinoma in situ as well as breast ducts, adipose tissue and stroma. Immunohistochemical staining identified macrophage populations in the majority of samples studied, with larger macrophage populations observed in RNAscope® positive samples, both in RNAscope® positive regions of tissue and/or the surrounding stroma. This is the first study to detect an HCMV gene product within breast tissue samples using the RNAscope® in situ hybridisation technique, and strengthens the evidence supporting the hypothesis that HCMV may be associated with initiation and progression of some breast cancers.