Abstract
Triple-negative breast cancer (TNBC) is a highly aggressive tumour that lacks marker for targeted diagnosis. Recently, it was reported that toll-like receptor 5 (TLR5) was associated with some kind of tumours, especially in TNBC, but whether it could be used as a non-invasive monitoring target is not fully understood. Here, we established TLR5(-) 4T1 cell line with lentivirus-shRNA-TLR5 knock-down transfection (with tag GFP, green fluorescent protein, TLR5(-) 4T1) and control TLR5(+) 4T1 cell line with negative control lentivirus transfection. The effect of TLR5 down-regulation was detected with qPCR and Western blot. (125) I-anti-TLR5 mAb and control isotype (125) I-IgG were prepared and injected to TLR5(+/-) 4T1-bearing mice models, respectively. Whole-body phosphor-autoradiography, fluorescence imaging and biodistribution were performed. Furthermore, ex vivo tumour TLR5 expression was proved through immunohistochemistry staining. We found that (125) I-anti-TLR5 mAb could bind to TLR5(+) 4T1 with high affinity and specificity. Whole-body phosphor-autoradiography after (125) I-anti-TLR5 mAb injection showed TLR5(+) 4T1 tumour images in 24 hours, more clearly in 48 hours. Radioactivities in tumour tissues were positively related with TLR5 expression. Biodistribution assay showed that (125) I-anti-TLR5 mAb was mainly metabolized through the liver and kidney, and (125) I-anti-TLR5 mAb was much more accumulated in TLR5(+) 4T1 tumour than TLR5(-) 4T1. In vivo fluorescence imaging successfully showed tumour tissues clearly both in TLR5(+) and TLR5(-) 4T1 mice compared with lentivirus untreated 4T1 tumour. Immunohistochemistry staining showed that TLR5 expression in tumours was indeed down-regulated in TLR5(-) 4T1 mice. Our results indicated that (125) I-antiTLR5 mAb was an ideal agent for non-invasive imaging of TLR5(+) tumours; TLR5 may be as a novel molecular target for TNBC non-invasive diagnosis.