Immunohistochemical Evaluation of Cathepsin B, L, and S Expression in Breast Cancer Patients

Cysteine cathepsins are proteases that play a role in normal cellular physiology and neoplastic transformation. Elevated expression and enzymatic activity of cathepsins in breast cancer (BCa) indicates their potential as a target for tumor imaging. In particular cathepsin B (CTSB), L (CTSL), and S (CTSS) are used as targets for near-infrared (NIR) fluorescence imaging (FI), a technique that allows real-time intraoperative tumor visualization and resection margin assessment. Therefore, this immunohistochemical study explores CTSB, CTSL, and CTSS expression levels in a large breast cancer patient cohort, to investigate in which BCa patients the use of cathepsin-targeted NIR FI may have added value.

Protein expression was analyzed in tumor tissue microarrays (TMA) of BCa patients using immunohistochemistry and quantified as a total immunostaining score (TIS), ranging from 0-12. In total, the tissues of 557 BCa patients were included in the TMA.

CTSB, CTSL, and CTSS were successfully scored in respectively 340, 373 and 252 tumors. All tumors showed CTSB, CTSL, and/or CTSS expression to some extent (TIS > 0). CTSB, CTSL, and CTSS expression was scored as high (TIS > 6) in respectively 28%, 80%, and 18% of tumors. In 89% of the tumors scored for all three cathepsins, the expression level of one or more of these proteases was scored as high (TIS > 6). Tumors showed significantly higher cathepsin expression levels with advancing Bloom-Richardson grade (p < 0.05). Cathepsin expression was highest in estrogen receptor (ER)-negative, human epidermal growth factor receptor 2(HER2)-positive and triple-negative (TN) tumors. There was no significant difference in cathepsin expression between tumors that were treated with neoadjuvant systemic therapy and tumors that were not.

The expression of at least one of the cysteine cathepsins B, L and S in all breast tumor tissues tested suggests that cathepsin-activatable imaging agents with broad reactivity for these three proteases will likely be effective in the vast majority of breast cancer patients, regardless of molecular subtype and treatment status. Patients with high grade ER-negative, HER2-positive, or TN tumors might show higher imaging signals.