TLR expression profiles are a function of disease status in rheumatoid arthritis and experimental arthritis.

The role of the innate immune system has been established in the initiation and perpetuation of inflammatory disease, but less attention has been paid to its role in the resolution of inflammation and return to homeostasis. Toll-like receptor (TLR) expression profiles were analysed in tissues with differing disease status in rheumatoid arthritis (RA), ankylosing spondylitis (AS), and in experimental arthritis. TLR gene expression was measured in whole blood and monocytes, before and after TNF blockade. In RA and osteoarthritis synovia, the expression of TLRs was quantified by standard curve qPCR. In addition, four distinct stages of disease were defined and validated in collagen-induced arthritis (CIA), the gold standard animal model for RA - pre-onset, early disease, late disease and immunised mice that were resistant to the development of disease. TLR expression was measured in spleens, lymph nodes, blood cells, liver and the paws (inflamed and unaffected). In RA whole blood, the expression of TLR1, 4 and 6 was significantly reduced by TNF blockade but the differences in TLR expression profiles between responders and non-responders were less pronounced than the differences between RA and AS patients. In RA non-responders, monocytes had greater TLR2 expression prior to therapy compared to responders. The expression of TLR1, 2, 4 and 8 was higher in RA synovium compared to control OA synovium. Circulating cytokine levels in CIA resistant mice were similar to naïve mice, but anti-collagen antibodies were similar to arthritic mice. Distinct profiles of inflammatory gene expression were mapped in paws and organs with differing disease status. TLR expression in arthritic paws tended to be similar in early and late disease, with TLR1 and 2 moderately higher in late disease. TLR expression in unaffected paws varied according to gene and disease status but was generally lower in resistant paws. Disease status-specific profiles of TLR expression were observed in spleens, lymph nodes, blood cells and the liver. Notably, TLR2 expression rose then fell in the transition from naïve to pre-onset to early arthritis. TLR gene expression profiles are strongly associated with disease status. In particular, increased expression in the blood precedes clinical manifestation.

Drug targeting to monocytes and macrophages using esterase-sensitive chemical motifs.

The therapeutic and toxic effects of drugs are often generated through effects on distinct cell types in the body. Selective delivery of drugs to specific cells or cell lineages would, therefore, have major advantages, in particular, the potential to significantly improve the therapeutic window of an agent. Cells of the monocyte-macrophage lineage represent an important target for many therapeutic agents because of their central involvement in a wide range of diseases including inflammation, cancer, atherosclerosis, and diabetes. We have developed a versatile chemistry platform that is designed to enhance the potency and delivery of small-molecule drugs to intracellular molecular targets. One facet of the technology involves the selective delivery of drugs to cells of the monocyte-macrophage lineage, using the intracellular carboxylesterase, human carboxylesterase-1 (hCE-1), which is expressed predominantly in these cells. Here, we demonstrate selective delivery of many types of intracellularly targeted small molecules to monocytes and macrophages by attaching a small esterase-sensitive chemical motif (ESM) that is selectively hydrolyzed within these cells to a charged, pharmacologically active drug. ESM versions of histone deacetylase (HDAC) inhibitors, for example, are extremely potent anticytokine and antiarthritic agents with a wider therapeutic window than conventional HDAC inhibitors. In human blood, effects on monocytes (hCE-1-positive) are seen at concentrations 1000-fold lower than those that affect other cell types (hCE-1-negative). Chemical conjugates of this type, by limiting effects on other cells, could find widespread applicability in the treatment of human diseases where monocyte-macrophages play a key role in disease pathology.

Stromelysin-1 (matrix metalloproteinase-3) and tissue inhibitor of metalloproteinase-3 are overexpressed in the wall of abdominal aortic aneurysms.

BACKGROUND: Atherosclerosis is implicated in the pathogenesis of abdominal aortic aneurysm (AAA) but more often causes aortic occlusive disease (AOD). The matrix metalloproteinases (MMPs) degrade extracellular matrix and may play a central role in the pathogenesis of AAA. The aim of this study was to examine differences in the patterns of MMP and MMP inhibitor expression between AAA and AOD. METHODS AND RESULTS: The expression of mRNA for 14 MMPs and 4 tissue inhibitors of metalloproteinases (TIMPs) was estimated in samples of aortic wall from 8 patients with AAA and 8 with AOD using the reverse-transcriptase polymerase chain reaction with a synthetic multicompetitor standard. AAA wall expressed significantly more stromelysin-1 (MMP-3) (mean log(10) ratio [copy enzyme cDNA/copy GAPDH cDNA], -1.9; range, -3.3 to -0.7) than the AOD wall (mean, 4; range, -5.7 to -2.4), P<0.005. TIMP-3 expression was significantly higher in AAA (mean, -1.7; range, -2.9 to -1.0) than AOD (mean, -3.6; range, -5.7 to -1.8), P<0.01. Expression of 8 other MMPs (1, 2, 7, 9, 11, 12, 14, and 17) was detected and was similar in AAA and AOD. Expression of the remaining 5 MMPs (-8, -10, -13, -15, and -16) was not detected in any of the samples. CONCLUSIONS: Both AAA and AOD walls express similar levels of a wide range of MMPs, including cell membrane-bound MT-MMPs. Stromelysin-1 (MMP-3) and TIMP-3 were, however, over expressed in the AAA samples and may be involved aneurysm pathogenesis. Stromelysin-1 could provide a target for pharmacological inhibition.