Washington D.C.— Khaled Elsaid, PharmD, PhD, discussed synovial macrophages/monocytes and monosodium urate (MSU) crystal interactions at a Monday, Nov. 18, session of ACR Convergence, exploring potential new gout pharmacology targets via the lubricin/CD44/protein phosphatase 2A (PP2A) pathway.
Gout Treatment Approaches
Dr. Elsaid
Dr. Elsaid, an associate professor of pharmacology at Chapman University, Orange, Calif., first became interested in the lubricin/CD44/PP2A pathway during his earlier work on posttraumatic osteoarthritis, but more recently he has begun exploring possible applications to gout pathophysiology and targets for treatment development.1
Although the 21st century has witnessed considerable innovations in gout therapy, many patients still are not meeting their therapeutic goals. The burden of disease is increasing globally. Current treatment options for gout may be inadequate, especially as many patients have significant comorbid conditions such as osteoarthritis, heart disease and kidney disease, and some cannot safely benefit from standard treatments, e.g., nonsteroidal anti-inflammatory drugs for a gout flare. Thus, new pharmacological approaches are still needed.
“Gout patients have a lot of comorbidities that share a common nature in terms of monocyte-driven inflammation,” explained Dr. Elsaid.
Hyperuricemia is key in gout pathogenesis, as it can lead to the deposition of MSU crystals in the joints and soft tissues. There, macrophages/monocytes phagocytose the crystals, activating the NFκB inflammatory pathways and NOD-like receptor protein 3 (NLRP3) inflammasomes. These inflammasomes activate caspases, leading to an inflammatory cascade and release of cytokines, including activation and release of interleukin (IL)‐1β. Binding to endothelial IL-1β receptors leads to further inflammatory mediator release, neutrophil recruitment and consequent symptoms of gout flare.1,2
Thus, therapies which lower uric acid are critical for gout management, e.g., xanthine oxidase inhibitors to lower uric acid production, newer uricosuric drugs like lesinurad which inhibit urate absorption, and a uricolytic like pegloticase which converts uric acid to a more easily excreted form. The inflammatory response has also been an important target, from older drugs like corticosteroids or colchicine to newer drugs which neutralize IL-1β such as anakinra, as well as experimental drugs that inhibit the NLRP3 inflammasome such as dapansutrile.3
“None of these therapeutics tried to address the critical step of how the macrophages recognize the MSU crystals and engulf them in order to release IL-1,” said Dr. Elsaid. He and his team wanted to take a different approach, instead focusing on this macrophage level.
Lubricin/CD44/PP2A Axis and Synovial Homeostasis
Lubricin, also known as “proteoglycan-4” or “PRG4,” is a mucinous glycoprotein produced by synovial fibroblasts and superficial zone chondrocytes. Along with hyaluronic acid, it is one of the two major components of synovial fluid. Known for its lubricant proteins, it also plays a role in controlling the synovial response to injury.
The CD44 receptor is highly expressed on macrophages and other cells of the immune system. It is a phagocytic receptor which can also extracellularly interact with ligands such as lubricin and interact intracellularly to activate signaling pathways. PP2A is a highly-conserved member of a family of enzymes that regulate signal transduction of many cellular processes.
When lubricin binds CD44 receptors on synovial macrophages, this activates recruitment and binding of PP2A enzymes to intracellular CD44 glycoproteins. Ultimately, this leads to inhibition of the NFκB inflammatory pathway. At homeostasis, CD44 receptors are covered by lubricin, acting as a sort of shield to prevent macrophage activation, Dr. Elsaid explained.
However, an inflammatory environment can decrease lubricin: Stimulation with IL‐1β downregulates lubricin, and enzymes such as neutrophil elastases may degrade it. This loss of lubricin upregulates the density of CD44 on macrophages. This results in increased NFκB inflammatory pathway signaling and inflammasome activation, ultimately leading to higher levels of activated IL‐1β and other inflammatory signals.
Using mouse and human tissue models, Dr. Elsaid and colleagues have explored whether these components—CD44, lubricin and PP2A—contribute to gout pathogenesis.
CD44 in Gout Pathophysiology
Dr. Elsaid said, “We wanted to ask whether CD44 can mediate urate crystal phagocytosis and whether that is a new mechanism of urate crystal uptake.”
Dr. Elsaid shared data from bone-derived macrophages from CD44 knockout mice (mice not expressing the CD44 receptor) and wild type mice (expressing the CD44 receptor). The wild type mice showed a higher extent of uric crystal phagocytosis, suggesting that CD44 might be involved in the process of uptake.4
“We also wanted to think pharmacologically about whether targeting this receptor is going to give us a new mechanism in terms of addressing apical events in terms of urate crystal uptake,” said Dr. Elsaid.
Thus, they used a monoclonal CD44 antibody which induces the shedding of the CD44 extracellular region, preventing MSU crystal phagocytosis. The CD44 antibody reduced neutrophil and monocyte infiltration as well as local IL‐1β release in vivo, illustrating its potential as a therapeutic target.4
PP2A in Gout Pathophysiology
Dr. Elsaid also shared findings about human monocyte experiments in which PP2A expression was knocked down via siRNA. He and his team found that when exposed to MSU crystals, monocytes with lower expression of PP2A showed increased IL‐1β gene expression and IL‐1β release compared to controls.5
They also utilized a drug known to increase PP2A activity, fingolimod, in the same model of human monocyte exposure to MSU crystals. Fingolimod, a sphingosine-1-phosphate receptor modulator with immunoregulatory properties, is already approved by the Food and Drug Administration for multiple sclerosis. The PP2A activator decreased IL‐1β release.5
Moreover, in a different study using the same model, the PP2A activator decreased neutrophil infiltration and infiltration of pro-inflammatory type “classical monocytes,” while increasing infiltration of anti-inflammatory “non-classical monocytes.”6
“So, we were able to show that the intracellular modulator PP2A does play a role in the degree of response in urate crystal inflammation,” said Dr. Elsaid.
Lubricin in Gout Pathophysiology
Dr. Elsaid and colleagues also studied urate crystal inflammation using recombinant human lubricin (rh-lubricin) using in vitro and in vivo models. They demonstrated a reduction in MSU crystal phagocytosis as well as decreased IL‐1β and IL-8 release compared to controls.7
They also examined the inflammasome in human macrophages, demonstrating that rh-lubricin resulted in decreased expression of inflammasome proteins, of IL‐1β and of caspase-1, a response that was specific to uric acid crystals.7
“Being a major component in synovial fluid, [lubricin] may play a role in terms of uric crystal aggregation and formation,” said Dr. Elsaid. To follow up on this idea, they applied rh-lubricin to bone marrow-derived macrophages, which reduced the activity of xanthine oxidase and of cellular uric acid. Moreover, they were able to reduce MSU crystal formation with increasing amounts of rh-lubricin.8
“Uric acid crystal uptake by synovial macrophages is the upstream hallmark for the gout pathophysiology,” concluded Dr. Elsaid. “That full circuit [of lubricin/CD44/PP2A] is an emerging multi-targeted pathway in gout pharmacology.”
Ruth Jessen Hickman, MD, a graduate of the Indiana University School of Medicine, is a medical and science writer in Bloomington, Ind.
References
- Elsaid KA, Jay, Liu-Bryan GD, Terkeltaub R. Proteoglycan 4 (PRG4)/lubricin and
the extracellular matrix in gout. Gout Urate Crys Depos Dis. 2023;1:122–136. - Dalbeth N, Gosling AL, Gaffo A, Abhishek A. Gout. Lancet. 2021;397(10287):1843–1855.
- James R, Paul BJ. New and emerging therapies in gout. Rheumatology and Autoimmunity. 2023;3:70–77.
- Bousoik E, Qadri M, Elsaid KA. CD44 receptor mediates urate crystal phagocytosis by macrophages and regulates inflammation in a murine peritoneal model of acute gout. Sci Rep. 2020;10(1):5748.
- Qadri M, Almadani S, Jay GD, Elsaid KA. Role of CD44 in Regulating TLR2 activation of human macrophages and downstream expression of proinflammatory cytokines. J Immunol. 2018 Jan 15;200(2):758–767.
- Elsayed S, Elsaid KA. Protein phosphatase 2A regulates xanthine oxidase-derived ROS production in macrophages and influx of inflammatory monocytes in a murine gout model. Front Pharmacol. 2022;13:1033520.
- Qadri M, Jay GD, Zhang LX, et al. Recombinant human proteoglycan-4 reduces phagocytosis of urate crystals and downstream nuclear factor kappa B and inflammasome activation and production of cytokines and chemokines in human and murine macrophages. Arthritis Res Ther. 2018;20(1):192.
- Elsaid K, Merriman TR, Rossitto LA, et al. Amplification of inflammation by lubricin deficiency implicated in incident, erosive gout independent of hyperuricemia. Arthritis Rheumatol. 2023;75(5):794–805.
