Toll-Like Receptor 9 Agonists Promote Cellular Invasion by Increasing Matrix Metalloproteinase Activity

Abstract

Toll-like receptor 9 (TLR9) recognizes microbial DNA. We show here that TLR9 protein is expressed in human breast cancer cells and clinical breast cancer samples. Stimulation of TLR9-expressing breast cancer cells with the TLR9 agonistic CpG oligonucleotides (1-10 Mmol/L) dramatically increased their in vitro invasion in both Matrigel assays and three-dimensional collagen cultures. Similar effects on invasion were seen in TLR9-expressing astrocytoma and glioblastoma cells and in the immortalized human breast epithelial cell line MCF-10A. This effect was not, however, dependent on the CpG content of the TLR9 ligands because the non-CpG oligonucleotides induced invasion of TLR9-expressing cells. CpG or non-CpG oligonucleotide-induced invasion in MDA-MB-231 cells was blunted by chloroquine and they did not induce invasion of TLR9 À breast cancer cells.Treatment of MDA-MB-231 cells with CpG or non-CpG oligonucleotides induced the formation of f50-kDa gelatinolytic band in zymograms. This band and the increased invasion were abolished by a matrix metalloproteinase (MMP) inhibitor GM6001 but not by a serine proteinase inhibitor aprotinin. Furthermore, CpG oligonucleotide treatment decreased tissue inhibitor of metalloproteinase-3 expression and increased levels of active MMP-13 in TLR9-expressing but not TLR9 À breast cancer cells without affecting MMP-8. Neutralizing anti-MMP-13 antibodies inhibited the CpG oligonucleotide-induced invasion. These findings suggest that infections may promote cancer progression through a novel TLR9-mediated mechanism. They also propose a new molecular target for cancer therapy, because TLR9 has not been associated with cancer invasiveness previously.

Circulating Methylated DNA: A New Generation of Tumor Markers

“…Although there is no direct proof, it is quite likely that the massive increase in free-circulating hypomethylated DNA, via activation of TLR-9, boosts the immune system. TLR-9 has also been identified, however, in human breast cancer cells and clinical breast cancer samples (29). Stimulation of TLR-9-expressing breast cancer cells with TLR-9 agonistic CpG oligonucleotides dramatically increases their in vitro invasion in both Matrigel assays and three-dimensional collagen cultures.…”

Expression of microRNA-146 suppresses NF-κB activity with reduction of metastatic potential in breast cancer cells

“…In this regard, it is noteworthy that epidermal growth factor receptor (EGFR), as a predicted target of miR-146 (miRGen database), coupled with the use of 10 ng/ml EGF as chemoattractant in the cell-invasion assays, is such a candidate pathway potentially impacted by miR-146 expression in MDA-MB-231 cells. Given the multiplicity of signals the aberrant regulation of which could provide a cancer cell with constitutively active NF-kB, it was perhaps surprising that expression of miR-146a/b in the highly metastatic human breast cancer cell line MDA-MB-231 produced suppression of NF-kB activity, although many tumor cells express IL-1 and Toll-like receptors (Singer et al, 2003;Merrell et al, 2006) and are thus potentially vulnerable to IRAK1 and TRAF6 suppression. MDA-MB-231 cells, in particular, have been shown to express the IL-1 receptor as well as Toll-like receptors 4 and 9 (Singer et al, 2003;Merrell et al, 2006).…”

“…Given the multiplicity of signals the aberrant regulation of which could provide a cancer cell with constitutively active NF-kB, it was perhaps surprising that expression of miR-146a/b in the highly metastatic human breast cancer cell line MDA-MB-231 produced suppression of NF-kB activity, although many tumor cells express IL-1 and Toll-like receptors (Singer et al, 2003;Merrell et al, 2006) and are thus potentially vulnerable to IRAK1 and TRAF6 suppression. MDA-MB-231 cells, in particular, have been shown to express the IL-1 receptor as well as Toll-like receptors 4 and 9 (Singer et al, 2003;Merrell et al, 2006). Future studies evaluating the susceptibility of various other cancer cell lines and tumors with constitutively active NF-kB programs to modulation by miR-146a/b expression will likely provide deeper insights and therapeutic opportunities for treating metastatic disease, a problem that has remained largely intractable.…”

Toll like receptor 7 expressed by malignant cells promotes tumor progression and metastasis through the recruitment of myeloid derived suppressor cells

“…In prostate and breast cancer, the stimulation of TLR9 induced an over-expression of matrix metalloproteinase 13 as well as an increase of cell invasion 31,32 . Finally, TLR4 stimulation increased tumor cells invasion in breast cancer 19 and in lung adenocarcinoma 33 .…”