Abstract 3670: Toll-like receptor (TLR) 7 expression in the human tumor microenvironment

Following the success of imiquimod, agents targeting TLR7 are being progressed as potential immunotherapeutics. Stimulation of TLR7 initiates a plasmacytoid dendritic cell driven immune response which drives efficacy. In addition to a role in regulating immune cell responses, TLR7 has also been reported to play a role in regulating tumor cell function (1,2). To investigate the expression of TLR7, by both tumor cells and by tumor infiltrating immune cells, we have used immunohistochemistry (IHC). Initially, a comprehensive validation of commercially available anti-TLR7 antibodies was conducted revealing one antibody (Epitomics, 3923-1) with specificity for TLR7 in western blots of cell lysates, and by immunocytochemistry of formalin fixed paraffin embedded (FFPE) cell pellets, of HEK293 cells stably transfected with TLR7, mock-transfected or non-transfected. 3923-1 was validated further for IHC across FFPE sections of human spleen, lymph node and tonsil demonstrating expected tissue and cellular localization. A comparison of staining for TLR7 in whole sections of FFPE tumors revealed non-specific tumor staining using antibodies that failed the validation process compared with the staining pattern observed for 3923-1. Subsequently, five tissue microarrays (TMAs) comprising 18 different human tumor types (6-25 patients/tumor type, triplicate cores) and 14 normal tissues (5 donors/tissue type, duplicate cores) were stained by IHC for TLR7 expression using 3923-1. Pathologist scores for tumor cells revealed 5/18 tumor types negative for TLR7 (ovarian, glioma, thyroid, liver, renal) and 9/18 tumor types with a proportion of patients (4%-36%) exhibiting weak (1+) staining (breast, lung, colorectal, pancreatic, gastric, head & neck, melanoma, esophageal, endometrial). Furthermore, 11-17% of sarcoma, prostate and bladder tumors displayed moderate (2+) staining for TLR7, whereas corresponding normal tissue epithelium was largely negative for TLR7. Importantly, an increased density of immune infiltrates was observed in tumor tissues compared with normal tissues, and a greater proportion of the immune infiltrate in tumors was TLR7 positive. The conclusion drawn is that TLR7 may be less frequently expressed by tumor cells than reported previously and that tumors exhibit a marked TLR7 positive immune cell infiltrate. These data identify tumour types which might benefit from TLR7 therapy and may guide patient selection. 1. Grimm M et al. Eur J Cancer (2010):2849-57. 2. C