**2. JAK-STAT Pathway**

JAKs are enzymes belonging to the tyrosine kinase family enzymes and their main function is to phosphorylate tyrosine residues to activate downstream signaling proteins and evoke physiological functions. When activated, they transfer extracellular signals provided by growth factors, cytokines, and chemokines that translate directly to the change of DNA transcription with the subsequent translation of several proteins. At the moment, four JAKs have been identified in mammalians (JAK1, JAK2, JAK3, and TYK2) which are specifically attached to receptors [11]. Activation of one specific JAK by the ligand-receptor can be recognized by the various receptor-ligand complexes as the one specific JAK and could be activated by several cytokines. *JAK1*, *JAK2*, and *TYK2* are expressed by many cells, contrary to this; hematopoietic, myeloid, and lymphoid cells express JAK3 [12]. The activation of JAK is a multi-step process. After a ligand is ligated to the receptor, the receptor's subunits dimerize and form an active receptor which is able to activate receptor-associated JAK [13]. Active phosphorylated JAK then phosphorylates tyrosine residues in the cytoplasmic part of the receptor enabling creation of docking sites for STAT. STATs are DNA-binding proteins which, when phosphorylated (activated), dimerize and translocate to the nucleus, followed by regulation of gene expression. It makes STATs the second key player in the transmission of signal. Currently, seven STAT proteins have been identified. The JAK/STAT system is responsible to transmit signals of more than 50 ligands and is recognized as one of the central communication systems of the immune response [14]. Active STATs then translocate to the nucleus where they interact with DNA regulatory elements, changing the expression of related genes [15,16].

JAK and more precisely JAK/STAT system is responsible for transmitting the signals provided by the wide spectrum of cytokines, which are ligands for class I and class II receptors. These receptors are protein complexes expressed on the surface of cells. They are built as one to four receptor chains. The typical structure of the receptor is formed from extracellular cytokine R homology domain (CHD) and a sequence acting as cytokine binding site. Slight structural differences in the CHD cytokine receptors enable to distinguish class I or class II family receptors [17]. Class I receptor family interacts with four cytokine families - gamma chains (γc), beta chains (βc), cytokines that utilize gp130 protein, and ILs that interact with a receptor's common subunit p40. Presence of γc in the receptor enables to interact with IL-2, IL-4, IL-7, IL-9, IL-15, and IL-21 [18], since βc is responsible for transducing the signals provided by granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-3, and IL-5. Several cytokines utilize gp130 protein as a component of their receptor. This cytokine subfamily consists of IL-6, IL-11, IL-27, and IL-31. This receptor complex is also used by ciliary neurotrophic growth factor, oncostatin M, cardiotrophin1 and cardiotrophin-like cytokine factor 1 [19–21]. Recently, two other cytokines namely IL-35 and IL-39 have been added to gp130 family due to the fact that they use gp130 as a signal transmitting unit in the receptor complex [22,23]. The last cytokine family that utilize class I receptor consists of IL-12 and IL-23 receptors for heterodimeric cytokines that

share the common subunit p40 [24,25]. Several hormone-like cytokines as growth hormone, leptin, erythropoietin, and thrombopoietin also transmit signals via the class I receptor. Plethora of cytokines, chemokines, and growth factors makes class I receptors the real crossroad of immune response, metabolism growth, tissue development, and indicates how important modulation of this pathway is. βc-the family of class II cytokines comprise a large group of signaling molecules. The most important members of this class are type I, II, and III IFNs (IFN-α, IFN-β, IFN- γ, IL-28, and IL-29) and cytokines belonging to IL-10-related family (IL-10, IL-19, IL-20, IL-22, IL-24, and IL-26) [17].

The transmission of signals from receptors requires two molecules of JAKs. JAK1 transmits signals provided by IL-6, IL-10, IL-11, IL-19, IL-20, and IL-22, and IFN-α, IFN-β, and IFN-γ, since JAK2 activation is responsible for the signaling of hormone-like cytokines-erythropoietin, thrombopoietin, growth hormone, GM-CSF, IL-3, and IL-5. [26]. JAK3 is exceptional member of JAK family which transmits signals as heterodimer of JAK1 and JAK3 molecules and is primarily expressed on hematopoietic cells. Attached to γ-chain transmits signals from IL-2, IL-4, IL-7, IL-9, IL-15, and IL-21 [26]. The last member of JAK family TYK2 facilitates signaling for IL-12, IL-23, and type I IFNs [27]. TYK2 creates heterodimers with either JAK1 or JAK2 [27]. Signaling the various cytokines by specific pairs of JAKs at least potentially creates a chance to target (inhibit) narrow branch of cytokines. However, one should remember that, contrary to biologic-targeted therapy, when one drug blocks only one cytokine, JAK inhibitor blocks many cytokines which utilize the same type of receptor.

#### **3. The Role of JAK**/**STAT Pathway in Immunity and Autoimmunity**

The results from many studies performed in the last two decades confirmed the involvement of the JAK/STAT pathway in several diseases associated with inflammation, cancer, immunity, and immune deficiency. This is not surprising as JAK mutations have been identified in immune deficiency syndromes including severe combined immune deficiency (SCID), in hematologic malignancies (leukemias and lymphomas) [28], and also in autoimmunity (hyper IgE-Job's syndrome) [29]. Some of the mutations in the JAK/STAT pathway directly increase the risk of developing well characterized autoimmune disorders like inflammatory bowel disease, psoriasis, ankylosing spondylitis, Behçet's disease [30–32], RA, Sjögren syndrome, or systemic lupus erythematosus (SLE) [33,34]. These findings underline the role of cytokine mediated regulation that affects such important pathophysiological processes as IFN-mediated immunity [35,36], T- and NK-cell-based immune response, regulation of function of lymphocytes, hematopoiesis, and nerve development. Recently, the role of IFNs in the development of several autoimmune disorders has been confirmed. In line with it, the term IFN signature has been coined indicating the special role of IFNs in the devolved of autoimmunity, specifically a prominent increase in the expression of type I IFN-regulated genes. This is especially a fact as far as SLE, inflammatory myopathies, and systemic sclerosis are concerned [37–41].

SLE is an autoimmune disease with a complex immunopathogenesis where B-cells have been implicated in humoral abnormalities and a prominent type I IFN signature is found in blood of majority of the SLE patients [42]. As the JAK/STAT cascade was identified to be responsible for the signal transduction from the activated IFN receptor to the nucleus, any disturbance in activity of this pathway may lead to the disease development. Indeed, the study on human lupus nephritis (LN) by Arakawa et al. [43] observed increased glomerular staining of STAT3 in renal biopsies of LN patients. Moreover, in patients with different types of glomerulonephritides, STAT3 activation highly correlated with glomerular and tubulointerstitial cell proliferation, interstitial fibrosis, and the level of renal injury. Obviously, the role of cytokines in the development of autoimmune diseases is not limited to IFNs. All known and perhaps not already known cytokines and chemokines create unique networks of self-interactions, activation, and regulatory loops. Any disturbance in this precise universe results directly to the development of autoimmunity, malignancy, allergy, or immunodeficiency. In addition, the second main player in autoimmunity, namely B-cells, are at least partially dependent upon cytokine stimulation utilizing the JAK/STAT system for signal transmission [44].

In the last four decades, the importance of cytokines in autoimmune diseases, as an executive arm of autoimmunity has been established. As the JAK/STAT pathway is one of the three most important signaling pathways in the cell, targeting of JAKs appears to be a rational strategy to stop the development of the diseases at a very early stage. In line with it, a great number of JAK inhibitors in various stages of preclinical development are being tested in clinical trials, and some of them have already been approved for the treatment.
