Universität zu Köln

Wittekind, Paula (2023). The effect of JAK inhibitors baricitinib and tofacitinib on Th17 cells, regulatory T cells and the Th17/Treg balance in patients with rheumatoid arthritis. PhD thesis, Universität zu Köln.

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Abstract

Background: Rheumatoid arthritis (RA) is a systemic inflammatory autoimmune disease that leads to irreversible structural joint damage, progressive disability and severe systemic complications including pulmonary, cardiac, hematological and vascular disorders. It affects around 1% of the full age population and is the most common form of inflammatory arthritis worldwide. The first-line therapy with the immunosuppressant methotrexate is generally efficient and effective, but provokes a big range of adverse effects and, at times, shows inadequate treatment response. In order to improve the therapeutic management of Rheumatoid Arthritis, the JAK inhibitors baricitinib and tofacitinib - a new class of so called targeted Disease Modifying Anti-Rheumatic Drugs (DMARDs) - recently achieved RA approval, passing clinical phase III trials with high effectiveness and tolerable adverse effects. Due to their oral application, they have an additional advantage in clinical use. JAK inhibitors are immunomodulators that target cytokine receptor associated janus kinases and thereby affect the JAK/STAT-mediated differentiation and activation of different immune cells. Objective: Considering the pathogenesis of RA, especially the immunologic imbalances of pro-inflammatory Th17 cells and immunosuppressive regulatory T (Treg) cells seem to play a pivotal role in the development of disease. In RA patients, increased frequencies of Th17 cells are found while Treg cells are impaired, leading to a defective Th17/Treg ratio in favor of Th17 cells. With that in mind, this study was constituted in order to investigate the molecular effect of baricitinib and tofacitinib on JAK/STAT-mediated activation and differentiation of Th17 cells and Treg cells in patients with RA. We particularly focused the question, if JAK inhibitors are able to restore the impaired Th17/Treg balance. Moreover, we aimed to detect differences in the molecular effects of baricitinib and tofacitinib taking into account that they target different janus kinases. Baricitinib is known as a JAK1/JAK2 inhibitor, while tofacitinib was designed to inhibit primarily JAK1 and JAK3. Last, this study analyzed the effect of JAK inhibitor treatment on janus kinase expression levels, investigating if reduced JAK activation also affects its expression. Methods: The study was performed with patients of the rheumatologic ambulance of the University Hospital of Cologne, including patients with diagnosed rheumatoid arthritis and a ‘healthy’ control group without rheumatic diseases but similar epidemiological features. The RA patients were subclassified in 4 groups according to their treatment: baricitinib-treated, tofacitinib-treated, methotrexate-treated and untreated. Peripheral blood samples were taken, CD4+ T cells were isolated and then analyzed via flowcytometry and PCR in order to display the frequency of the different T cell subsets % in relation to CD4+ T cells on the one hand, and to evaluate the expression of janus kinases and T cell associated key-transcriptions factors on the other hand. Results: Patients with untreated rheumatoid arthritis showed significantly increased expression levels of the Th17 cell associated key cytokine IL-17 compared to healthy controls, indicating chronic inflammation in RA. We showed that both treatments, MTX and the JAK inhibitors baricitinib and tofacitinib, were highly effective in the reduction of IL-17 expression achieving similar expression levels as healthy controls. The study also revealed that the frequency of regulatory T cells is significantly reduced in untreated RA patients compared to healthy controls, confirming the pathologic imbalance of Th17 cells and Treg cells in disease. Interestingly, neither baricitinib nor tofacitinib was able to restore the impaired regulatory T cells, in contrast to MTX, which at least partially enhanced them. With regard to the Th17/Treg ratio, this study concludes that JAK inhibitors only partially restore the immunologic balance via their effect on Th17 cells. This finding might form a background for prospective improvements of JAK inhibitor treatment in the therapeutic management of rheumatoid arthritis and other inflammatory diseases. The analyzed gene expression levels of the janus kinases JAK1, JAK2, JAK3 and TYK2 slightly indicate that JAK expression is not increased in patients with rheumatoid arthritis compared to healthy controls. Further, it can be assumed, that the mechanism by which JAK inhibitors affect JAK-induced downstream signaling is probably rather related to reduced JAK activation than to reduced JAK expression. But further studies with larger data sets need to be addressed to make a precise statement on the molecular effect of JAK inhibitors on JAK gene expressions. Although baricitinib and tofacitinib target different janus kinases, this study did not show any differences in the effect of baricitinib and tofacitinib on JAK/STAT-mediated differentiation and activation of the analyzed T cells. Conclusion: The most intriguing finding of this study was the demonstration of low Treg cell numbers in peripheral blood after treatment with baricitinib and tofacitinib, which leads to a partially restored Th17/Treg balance that is mainly achieved by a strong therapeutic effect on pathogenic Th17 cells. Based on these results, we presume that novel therapeutic strategies with the ability to fully restore the Th17/Treg balance might bring additional benefit in the treatment of rheumatoid arthritis. Outlook: One approach could be the combination of JAK inhibitors with low-dose IL-2, considering that IL-2 receptor downstream signaling induces Treg cell differentiation. This might recover Treg cell frequencies and could thereby contribute to immune homeostasis. Another promising approach is the development of more selective JAK inhibitors such as a JAK 1 inhibitor upadacitinib and TYK2 inhibitor BMS-986165. These selective JAK inhibitors and its research will further improve our understanding of janus kinase mediated downstream signaling and bring new insights into immunologic processes of the pathogenesis of rheumatoid arthritis and other autoimmune diseases.

Item Type:	Thesis (PhD thesis)
Creators:	Creators Email ORCID ORCID Put Code Wittekind, Paula UNSPECIFIED UNSPECIFIED UNSPECIFIED
URN:	urn:nbn:de:hbz:38-709842
Date:	2023
Language:	English
Faculty:	Faculty of Medicine
Divisions:	Faculty of Medicine > Innere Medizin > Klinik I für Innere Medizin - Hämatologie und Onkologie
Subjects:	Medical sciences Medicine
Uncontrolled Keywords:	Keywords Language rheumatoid arthritis English JAK inhibitors English Th17/Treg balance English
Date of oral exam:	14 February 2023
Referee:	Name Academic Title Kofler, David M.G. Privatdozent Dr. med. Stripecke, Renata Universitätsprofessorin Dr. rer. nat.
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/70984