Comparison of Cobimetinib versus Combination of Dasatinib and Quercetin in Relieving Inflammatory Chondrocyte Senescence

Abstract

BACKGROUND: Osteoarthritis is closely related to aging and characterized by degeneration of articular cartilage, subchondral bone sclerosis, and low-grade inflammatory responses. Aging and injury are significant triggers for inflammatory factors that mediate joint pathological changes. Cobimetinib, a MEK1 inhibitor, has an unclear effect on the inflammatory senescence of chondrocytes. OBJECTIVE: To compare the effects of cobimetinib and the classic anti-aging drug combination of dasatinib and quercetin (D+Q) in inhibiting inflammatory senescence of chondrocytes, and to explore its potential application in the treatment of osteoarthritis. METHODS: Primary chondrocytes were isolated and cultured, and cell viability was detected by CCK-8 assay. Cells were divided into control group, model group, D+Q group, and cobimetinib group. Except for the control group, cells in the other three groups were induced with interleukin-1β to establish an inflammatory senescence model, and then treated with cobimetinib or D+Q combination. Senescence phenotype was observed by β-galactosidase staining. The activation of MEK-ERK1/2 pathway was detected by real-time quantitative PCR, western blot, and immunofluorescence. The expression levels of senescence markers (P16, P21, P53) and senescence-associated secretory phenotype factors (inducible nitric oxide synthase, cyclooxygenase-2, chemokine ligand 3, interleukin-6) were measured, and the changes in extracellular matrix synthesis and degradation-related molecules (COL2A1, matrix metalloproteinase 13, matrix metalloproteinase 3) were evaluated. RESULTS AND CONCLUSION: Both cobimetinib and D+Q effectively inhibited the activation of MEK-ERK1/2 pathway, alleviated the senescence phenotype of chondrocytes, significantly downregulated the expression of P16, P21, and P53, reduced the levels of senescence-associated secretory phenotype factors, restored COL2A1 expression, and inhibited the expression of matrix metalloproteinase 13 and matrix metalloproteinase 3. These results indicate that cobimetinib effectively alleviates interleukin-1β-induced inflammatory senescence of chondrocytes by inhibiting the MEK-ERK1/2 pathway, and has a similar anti-aging effect to the D+Q combination, while showing greater potential in promoting matrix synthesis and inhibiting degradation.

1. Introduction

Osteoarthritis is a chronic joint disease closely related to aging, characterized by degeneration of articular cartilage, subchondral bone sclerosis, and low-grade persistent inflammation [1]. Aging and injury are important triggers for inflammatory factor-mediated joint pathological changes. Senescent chondrocytes exhibit growth arrest, enhanced resistance to apoptosis, increased β-galactosidase activity, and sustained high expression of cell cycle inhibitory proteins P21, P16INK4A (P16), and DNA damage response factor P53 [2]. Meanwhile, senescent chondrocytes maintain an inflammatory microenvironment by continuously expressing senescence-associated secretory phenotype (SASP) factors, promoting the senescence of neighboring chondrocytes and ultimately leading to irreversible joint destruction [3-4]. Pro-inflammatory cytokines such as interleukin-1β can activate nuclear factor κB and mitogen-activated protein kinase (MAPK) signaling pathways, inducing cell cycle arrest in chondrocytes and significantly upregulating inflammatory pathways, thereby driving the pathological progression of osteoarthritis [5].

In recent years, therapeutic strategies targeting senescent cells have made breakthrough progress in age-related diseases. Among them, senolytics have shown clinical application potential by specifically clearing P16-positive senescent cells or blocking the activity of anti-apoptotic proteins BCL-2/BCL-xL, effectively inducing programmed death of senescent 'zombie' chondrocytes and slowing the progression of age-related osteoarthritis [5-6]. The combination of dasatinib (a tyrosine kinase inhibitor) and quercetin (a natural flavonoid) was proposed by the Mayo Clinic in 2018 as a classic anti-aging treatment [7]. The D+Q combination is considered to have potential in treating chronic diseases such as atherosclerosis, fatty liver, and osteoporosis [8-11]. In human osteoarthritic synovial cell models and mouse knee osteoarthritis models, this drug combination has shown multiple effects of inhibiting the release of SASP factors from senescent cells and improving cartilage metabolic homeostasis [12]. D+Q targets senescent cells by regulating multiple pathways including transcription factors, cell cycle-related proteins, growth factors, and protein kinases [13], and inhibits SASP [14]. Although D+Q can reduce SASP to some extent, its clinical application is limited by side effects and bioavailability. Therefore, exploring more targeted and effective senolytic agents is of great significance.