Abstract P5-01-04: Activation of toll-like receptor 7 (TLR7) confers protection in human breast cancer
Authors: Obeid EI , Khramstova G , Zhang B , Fu Y-X , Mueller J , Nanda R , Olopade OI 
Published: August 2014

Background: Inflammation and angiogenesis in the tumor microenvironment play a role in human breast cancer. We previously showed that tumors with high M2 macrophages and high microvessel density (MVD) have a shorter overall survival. The antitumor effect of Toll-like receptor 7 (TLR7) has been previously described whereby its activation can alter the tumor microenvironment to adopt an antitumorigenic (Th-1) immune response and inhibit angiogenesis, with a potential immune mediated benefit in breast cancer. To our knowledge, there are no studies that evaluated TLR7 and its relationship to macrophages and angiogenesis in human breast cancer.

Methods: Breast tumors were obtained from the University of Chicago Breast Cancer SPORE tissue bank under IRB approved protocols. Tissue microarrays were constructed and molecular subtype was assigned based on immunohistochemical (IHC) staining into the following groups: luminal A (ER+, PR+, HER2-), luminal B (ER+, HER2+ or ER+, PR-), HER2-like (ER-, HER2+) and basal-like (ER-, HER2-, EGFR+ and/or CK5/6+). Macrophage density was determined using double staining with CD68/CD163. Microvessel density (MVD) was measured by IHC staining using anti-CD34. Staining quantification was performed by a trained pathologist and scoring performed independently by two pathologists. Based on median distribution, we assigned M2 macrophage content and MVD into high vs. low, and the TLR7 expression into faint vs. strong. To evaluate the association between M2 macrophages and TLR7, Spearman's rho correlation coefficients were calculated. Survival analysis was done using Tarone ware statistics.

Results: After removal of duplicate cores, we had 100 tumors available for scoring. There was an equal distribution of M2 macrophage content, MVD, and TLR7 expression (p = 0.074, p = 0.908, p = 0.145 respectively). The tumor subtypes were 53.2% luminal A, 34.0% basal-like, 7.4%, luminal B, and 5.3% Her2-like. We found a significant correlation between high number of M2 macrophages and high MVD (spearman's rho correlation coefficient = 0.343, p = 0.004). There was an association between M2 macrophages and TLR7 expression (Pearson χ2 = 4.425, p = 0.025). Compared to tumors with faint TLR7 expression, those with strong TLR7 expression exhibited a longer overall survival (Tarone ware statistic 5.053, p = 0.025) and this continued to be significant when adjusted for M2 macrophage content (Tarone ware statistic = 4.049, p = 0.044) and for MVD (Tarone ware statistic = 4.042, p = 0.044). There was no statistically significant association between TLR7 expression and molecular subtypes of breast cancer.

Conclusion: Tumor associated macrophages (M2 macrophages) and angiogenesis (high MVD) are known to contribute to worse outcomes in human breast cancer. Our results suggest that activation of TLR7 can have a protective effect in human breast cancer even in tumors harboring features of more aggressive disease.

Citation Information: Cancer Res 2013;73(24 Suppl): Abstract nr P5-01-04.