Novel Repurposed Drugs Against Joint Inflammation Reveal Potential Use for Gout Treatment: An In Silico, In Vitro and Clinical Study

Eloi Franco-Trepat¹, Ana Alonso-Pérez¹, Maria Guillán-Fresco¹, Miriam López-Fagundez¹, Andrés Pazos-Pérez¹, Ana Lois Iglesias², Susana Belén Bravo³, Alberto Jorge-Mora¹, JJ Gómez-Reino⁴ and Rodolfo Gómez¹, ¹IDIS-CHUS - Musculoskeletal Pathology Group, Santiago de Compostela, Spain, ²IDIS-CHUS - Musculoskeletal Pathology Group, A Coruna, Spain, ³IDIS-CHUS - Proteomics Unit, Santiago de Compostela, Galicia, Spain, ⁴IDIS-CHUS - Rheumatology Group, Santiago de Compostela, Spain

Meeting: ACR Convergence 2020

Keywords: gout, Inflammasome, innate immunity, Osteoarthritis, Synovitis

Session Information

Date: Friday, November 6, 2020

Session Title: Innate Immunity Poster

Session Type: Poster Session A

Session Time: 9:00AM-11:00AM

Background/Purpose: Joint inflammation is a common feature across multiple rheumatic diseases. To deal with the induction of innate immune factors, targeting therapeutic targets such as TLR4 and IL1R is required. We have recently reported the use of amitriptyline (AT), thalidomide (Th), and naloxone (NLX) to block TLR4 & IL1R-mediated innate immune responses in OA cartilage. Now we explore their use in synovia and gout inflammation.

Methods: The ethics committee approved (CAEIG 2016/258) the use of human samples and aggregated and dissociated clinical data from 440.000 citizens.

The activity and expression of TLR4, IL1R, and NLRP3 signaling pathways were determined by docking analysis (Autodocks-Vina), MTT cell viability assay, RT-PCR, and Western Blot, ELISA or MALDI-TOFF in human OA chondrocytes and synoviocytes (SW982 cell line). Statistics were performed using GraphPad Prism 9.0.

Results: The docking of AT, Th, and NLX towards TLR4 was confirmed by in silico binding affinity analysis. In human synoviocytes, the use of therapeutic doses of AT [0.1-1µM], NLX [1.4-100µM], and Th [3-500µM]) prevented the induction of several factors including NOS2, IL6, IL1B and MMP9 at the mRNA and protein level (up to -97%). The inhibition of the NLRP3 inflammasome pathway, which is linked to both synovitis and gout-associated inflammation, was verified by RT-PCR & MALDI-TOFF in OA chondrocytes and synoviocytes. Considering this link, we requested access to clinical data and found that amitriptyline consumers are less prone to require colchicine drug for gout treatment.

Conclusion: Amitriptyline, naloxone, and thalidomide prevent the induction of TLR4 & IL1R-mediated innate immune factors in OA chondrocytes and synoviocytes. Moreover, amitriptyline consumption reduced the dose of colchicine needed for gout treatment.

Considering that these drugs are being used in other indications, their repurpose might be a novel tool to manage inflammation across diverse rheumatic diseases.

Figure 1 – Amitriptyline blocked the proteome of IL1R–mediated innate immune response factors Fig. 1A.- Inflammatory-driven Pathway Enrichment Analysis of proteins quantified (SWATH) on primary human OA chondrocytes co-treated 24h with AT [1µM] and IL1β [0.1ng/ml]. Fig. 1B.- Blocking effects of amitriptyline (AT), naloxone (NLX) and thalidomide (Th) were evaluated on TLR4 (LPS [100ng/ml]) & IL1R (IL1β [0.1ng/ml])– activated human SW982 synoviocytes. Innate immune factors (NOS2, IL6, IL1β and MMP3) and NLRP3 inflammasome signalling were analysed by RT-PCR. Fig. 1C.- Contingency test comparing the number of patients requiring colchicine in both amitriptyline and non-amitriptyline consumers against the whole population (>65 years). Cumulative mean consumption of colchicine drug boxes in colchicine gout-patients who suffered of flare-like episodes, namely those who consumed >18 colchicine boxes in at least a single year. Statistics: SWATH data were filtered by FDR 1% and at least a p-value < 0,05. Data of three independent experiments was used and “Control” was used to normalise RT-PCR. Clinical data was obtained from the healthcare area of EOXI Santiago de Compostela (Spain) that covers a 400.000 citizen area. Contingency test was analysed by Fisher’s exact test. Cumulative mean tests were analysed by 2-way ANOVA Row factor, and other data was by one-way ANOVA. Tukey post-test was used, and results were expressed as the mean ± standard error of the mean (SEM) with the significance NEJM system (*p < 0,05; **p < 0,01; ***p < 0,001). Abbreviations: Toll like receptor 4 (TLR4), interleukin 1 receptor (IL1R), amitriptyline (AT), naloxone (NLX) and thalidomide (Th), human OA chondrocytes (hOC), lipopolysaccharide (LPS), interleukin 1 beta (IL1β), interleukin 6 (IL6),

Disclosure: E. Franco-Trepat, None; A. Alonso-Pérez, None; M. Guillán-Fresco, None; M. López-Fagundez, None; A. Pazos-Pérez, None; A. Lois Iglesias, None; S. Bravo, None; A. Jorge-Mora, None; J. Gómez-Reino, None; R. Gómez, None.

To cite this abstract in AMA style:

Franco-Trepat E, Alonso-Pérez A, Guillán-Fresco M, López-Fagundez M, Pazos-Pérez A, Lois Iglesias A, Bravo S, Jorge-Mora A, Gómez-Reino J, Gómez R. Novel Repurposed Drugs Against Joint Inflammation Reveal Potential Use for Gout Treatment: An In Silico, In Vitro and Clinical Study [abstract]. Arthritis Rheumatol. 2020; 72 (suppl 10). https://acrabstracts.org/abstract/novel-repurposed-drugs-against-joint-inflammation-reveal-potential-use-for-gout-treatment-an-in-silico-in-vitro-and-clinical-study/. Accessed February 26, 2021.

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