Discovery of New Benzothiazine Derivative as Modulator of Pro- and Anti-inflammatory Cytokines in Rheumatoid Arthritis

Abstract

The anti-inflammatory activities of benzothiazine and pyrazole derivatives are well documented. A series of novel N'-arylmethylidene-2-(3,4-dimethyl-5,5-dioxidopyrazolo(4,3 c)(1,2) benzothiazin-2(4H)yl) acetohydrazide compounds were previously synthesized by combining benzothiazine and pyrazole moieties into a single nucleus. The current study investigates the anti-arthritic potential of 3-ethoxy-4-hydroxyphenyl derivative (EHP) and its possible mechanism in arthritic rat model. Sprague-Dawley rats were induced rheumatoid arthritis with Freund's complete adjuvant and treated with EHP and piroxicam. At the end of the study, arthritic score was calculated, and ankle joint histopathology was performed using hematoxylin and eosin staining. Real-time reverse transcription polymerase chain reaction (qRT-PCR) and enzyme-linked immunosorbent assay (ELISA) were used to determine mRNA expression and protein levels of various inflammatory markers, respectively. In vitro concanavalin A (ConA)-stimulated splenocyte proliferation was measured. Serum levels of C-reactive protein (CRP), urea, creatinine, aspartate aminotransferase (AST), alanine transaminase (ALT), and alkaline phosphatase (ALP) were also determined. EHP significantly attenuated macroscopic arthritic score, joint histopathological lesions, and CRP levels. Treatment with EHP significantly reduced pro-inflammatory tissue necrosis factor-α (TNF-α), nuclear factor-kappa B (NF-кB), interleukin-17 (IL-17), and prostaglandin-E2 (PGE2) levels and increased the levels of anti-inflammatory interleukin-4 (IL-4) and interleukin-10 (IL-10). ConA-stimulated splenocyte proliferation was also significantly suppressed by treatment with EHP. Normalizing all hematological markers and ALP levels, EHP did not display any sign of nephrotoxicity and hepatotoxicity as determined by urea, creatinine, ALT, and AST levels. In conclusion, EHP possesses significant anti-arthritic property which may be attributed to its anti-inflammatory and immunomodulatory effects.

Keywords: TNF-α; ankle joint; immunomodulatory; inflammation; rheumatoid arthritis.