Discussion
The goal of this study was to characterise the molecular pathways impacted by the pharmacological effects of the JAK1 and JAK2 inhibitor baricitinib in SLE using RNA and serum samples from a phase II study of baricitinib that successfully met its primary endpoint. Baseline gene expression profiling of patients demonstrated that there was an elevation of STAT1, STAT2 and multiple IFN responsive genes, when compared with healthy controls. Baricitinib treatment reduced the mRNA expression of functionally interconnected genes involved in SLE including STAT1-target, STAT2-target and STAT4-target genes and multiple IFN responsive genes.2 Furthermore, the study demonstrated that treatment with baricitinib 4 mg significantly decreased serum IL-12p40 and IL-6 cytokine levels at week 12, which persisted through week 24. Thus, treatment with baricitinib 4 mg induced significant reduction in the RNA expression of a network of genes associated with the JAK/STAT pathway, cytokine signalling and SLE pathogenesis, and it consistently reduced serum levels of two key disease-associated cytokines, IL-12p40 and IL-6. In this regard, a JAK1-associated gene profile has been identified correlating with treatment-associated changes found for the serum cytokine IL-6 and IL-12p40 responses, which will require further validation/replication. Notably, other cytokine signalling can be influenced indirectly by complex interaction of signalling networks that interface with JAK/STAT.
The demographics of patients with SLE in the phase II study JAHH were similar to other recent phase II and III clinical trials suggesting that the baseline findings here may be broadly applicable.3 4 The baseline gene expression profiles of the patients studied yielded findings similar to the much larger 1760 patient ILLUMINATE studies, as well as other smaller studies, where large fold differences from healthy controls in expression of type I IFN responsive genes were observed in 50%–75% of patients.3 4
A key finding of the present study was the identification of baseline elevations of STAT1, STAT2 and STAT4, and/or downstream genes linked to the JAK/STAT pathway. The observations on STAT1, STAT 2 and STAT4 are distinctive to this study and again highlight the known importance of these genes and their associated signalling pathways in SLE.22 23
A second important finding of the present study was that pharmacologically induced changes in gene expression following baricitinib therapy resulted in significant reduction in STAT1-targets. Of note, patients with a relative elevation in the expression of JAK1 pathway genes at baseline displayed a better response to baricitinib 4 mg according to changes in SLEDAI-2K (figure 2B) or SRI-4 (data not shown) over 24 weeks. While it was anticipated that baricitinib could reduce JAK/STAT targets expression, this is the first publication to directly demonstrate this in a clinical trial linking such changes to clinical improvement.
Gene network analysis revealed further that there were expression changes in genes that are involved directly in classical, canonical JAK/STAT signal transduction, including STAT1 and STAT4, where genetic polymorphisms have been identified as a risk factor for both disease susceptibility and severity in SLE.23 24
This study is among the largest published surveys that includes both whole blood RNA expression and serum cytokine levels in SLE.3 4 25 26 While gene expression studies allow for a comprehensive survey of cellular and molecular immune processes, it is an indirect measure of circulating serum cytokines. Based on precedents in the medical literature, the state of our current understanding of the pathogenesis of SLE and technical feasibility, the study attempted to measure a select number of key cytokines using serum or plasma samples from JAHH. These included: IFN-α, IFN-γ, IL-10, IL-12p40, IL-17A, IL-5 and IL-6 levels. Overall, with few exceptions, the cytokines tested were elevated in patients with SLE at baseline as compared with healthy controls. These results strongly support the hypothesis that SLE is not a single cytokine-driven disease but is characterised by global dysregulation of the cytokine signalling network.2 Therefore, a pharmacological approach targeting multiple cytokines at the same time, such as the inhibition of the JAK/STAT axis, could be highly beneficial in the treatment of SLE. In further support of this, it was found that baricitinib treatment was associated with statistically significant decreases of serum IL-12p40 and IL-6, supporting the concept that reducing the levels of these cytokines may be effective in treating SLE, as was suggested by previous clinical trials targeting these molecules.11 12 Various transcription factors have been implicated in the regulation of IL-6 and IL-12p40 expression, including AP-1, C/EBP and nuclear factor-κB but not the JAK/STAT axis. Therefore, the molecular mechanisms responsible for the baricitinib-induced reduction of cytokine expression are still unclear and may entail cross-talk between STATs and other transcription factors, or suppressive effects on the proliferation and survival of cytokine-producing cells. Changes of STAT4 possibly linked to IL-12p40 under baricitinib 4 mg provided some confirmation between transcriptional changes and reduced cytokines, which was not seen for STAT3 connected to IL-6. Although differences between transcriptional changes and related protein expression can be found, the overall findings clearly indicate comprehensive inhibition of inflammatory pathways.
In conclusion, the mechanism of action of baricitinib in SLE may be mediated through the inhibition of multiple immune-related genes and cytokines. The study findings confirm that the JAK/STAT pathways appear to have a central role in the pharmacological effect of baricitinib in SLE. Baricitinib induced changes in the expression of STAT1/STAT2 target genes, and these were associated with treatment response. Serum cytokine measurement support the hypothesis that IL-12p40 and IL-6 may have a role in SLE pathogenesis and contribute to the PD effect of baricitinib. Finally, the two ongoing phase III studies of baricitinib in SLE will help to further define the clinical benefits of baricitinib in SLE and the molecular mechanisms of its action.27 28