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Novel N-butyrylated hyaluronic acid analog treats rheumatoid arthritis through both anti-inflammatory and antioxidant mechanisms

Background

Rheumatoid arthritis (RA) is a disease characterized as a persistent synovial inflammation. RA has been found to affect 3.6% of women and 1.7% of men in the United States¹. It costs approximately $13,500 USD each year to treat a single RA patient, resulting in a significant economic burden².

Despite extensive research on this topic, the exact causes and mechanism of RA are not well understood. Inflammatory mediators result in elevated reactive oxygen species (ROS) in inflamed tissue. Both ROS and the inflammatory mediators have been found to trigger nuclear factor κB (NF-κB) and mitogen-activated protein kinase (MAPK) pathways, leading to further production of pro-inflammatory cytokines. These cytokines induce the production of metalloproteinases, adhesion molecules, regulating immune cells, synoviocytes and chondrocytes, causing cartilage to degrade and ultimately resulting in joint space narrowing and joint destruction³.

Currently, non-steroidal anti-inflammatory drugs (NSAIDs) are commonly used to relieve joint pain and swelling associated with RA. However, the adverse side-effects of these drugs towards the cardiovascular and gastrointestinal system limit their suitability as a long-term treatment for RA-induced chronic inflammation. Disease-modifying anti-rheumatic drugs (DMARDs) are the current gold standard with respect to controlling disease activity, but are also associated with many side effects including serious hepatotoxicity, bone marrow suppression, pulmonary complications and increased risk of infection4. Therefore, there is an urgent need for alternative anti-inflammatory methods which can effectively and efficiently treat RA-induced inflammation without exhibiting serious adverse effects.

Hyaluronic acid (HA) is a high molecular weight (> 1 million daltons) glycosaminoglycan (GAG). Significant research now indicates that HA is more than an inert extracelullar GAG. A number of studies have found elevated levels of HA in serum and synovial fluid from patients with RA. However, further consideration of the molecular size of HA has now revealed the negative effects of endogenous low molecular weight HA in arthritic diseases (LMWHA; MW < 500 kDa) including being pro-inflammatory. Indeed, LWMHA is not only increased in RA serum and synovial fluids but further increased in those fluids in active RA compared to inactive RA. LMWHA is produced by synthases (HAS1 and 3), by enzymatic cleavage, and by oxidative degradation. These negative effects are mediated by LMWHA binding proteins including toll-like receptor 4 (TLR4), CD44, RHAMM and others. For example, RHAMM is overexpressed in RA patient fibroblast-like synoviocytes and is implicated in RA progression, likely via specific binding of LMWHA^4,5. Studies have also found increases in the HA-degrading enzyme KIAA1199 (aka CEMIP or HYBID) and LMWHA in both serum and synovial fluids of RA patients⁶. This increase in LMWHA is likely due to KIAA1199 as it has been shown to produce HA fragments of < 100 kDa⁷.

1. Crowson C.S. et al. The lifetime risk of adult-onset rheumatoid arthritis and other inflammatory autoimmune rheumatic diseases. Arthritis and Rheumatism. 2011;63(3): 633-9.
2. Rudan I. et al. Prevalence of rheumatoid arthritis in low- and middle-income countries: A systematic review and analysis. Journal of Global Health. 2015;5(1): 010409.
3. Mateen S. et al. Understanding the role of cytokines in the pathogenesis of rheumatoid arthritis. Clinica Chimica Acta; International Journal of Clinical Chemistry. 2016;455: 161-71.
4. Sulaiman W. et al. The trends of DMARDS prescribed in rheumatoid arthritis patients in Malaysia. Oman Medical Journal. 2009;24(4): 260-06.
5. Wu J et al. RHAMM induces progression of rheumatoid arthritis by enhancing the functions of fibroblast-like synoviocytes. BMC Musculoskeletal Disorders, 19:455 (2018).
6. Kouvidi K et al. Role of receptor for hyaluronic acid-mediated motility (RHAMM) in low molecular weight hyaluronan (LMWHA)-mediated fibrosarcoma cell adhesion. J. Biol.Chem. 286(44):38509-38520, 2011.
7. Kohi S et al. KIAA1199 is induced by inflammation and enhances malignant phenotype in pancreatic cancer. Oncotarget, 8(10): 17156-17163, 2017.

Technology Overview

Queen’s researchers have created novel N-acyl analogs of HA. Studies with N-butyrylated HA (BHA) have been found to prevent activation of TLR4 by either LMWHA or lipopolysaccharide on human macrophage cell lines. Furthermore, preclinical studies in gouty arthritis showed significant reduction of swelling, and speed of reduction of swelling, compared to prophylactic colchicine. Concurrent to those effects were significant reductions in pro-inflammatory cytokines and increases in anti-inflammatory cytokines. In follow-on they investigated effects of BHA on rheumatoid arthritis.

Low (0.05 mg/mL) concentrations of BHA significantly reduced the expression of pro-inflammatory cytokines (TNFα, IL-1β and IL-6) in an adjuvant‑induced arthritis (AIA) rat model (Figure 1), suggesting that BHA exhibits anti-inflammatory activity against cytokines produced by macrophages. This dosage of BHA showed a superior anti-inflammatory efficacy than positive control dexamethasone (DXMS), suggesting a promising role for BHA in the treatment of inflammation-related diseases.