ITK

Detailed Information

The ITK gene is located at position 5q31~5q32 on the human chromosome and consists of 5 distinct domains: pleckstrin homology (PH) domain, Tec homology (TH) domain, sarcoma homology 3 (SH3), SH2, and SH1 domains. The PH domain is located at the amino terminus of ITK, while the SH1 domain is at the carboxyl terminus. A flexible hinge connects the amino and carboxyl termini. The flexible hinge region connecting the two ends forms part of the adenosine triphosphate (ATP) binding site, and the activation loop is positioned between the two kinase domains. The PH domain of ITK is responsible for protein-lipid interactions and helps the kinase bind to the cell membrane. The TH domain comprises a conserved sequence of 27 amino acid residues, showing family recognition for ITK. The proline-rich region (PRR) of the TH domain binds to the SH3 domain, placing it in a self-inhibitory state. The SH3 domain contains a conserved amino acid residue Trp208, which is crucial for the binding of the SH3 domain to the PRR, resulting in a self-inhibited state of the kinase. The SH2 domain is responsible for protein-protein interactions, allowing the kinase to bind to phospholipase Cγ1 (PLCγ1). Any mutation or deletion of the SH2 and SH3 domains will inactivate the kinase. The SH1 domain (kinase or catalytic domain) is necessary for the catalytic activity of the kinase, particularly for activating PLCγ1 through phosphorylation of its Tyr775 and Tyr783 residues. The catalytic domain of the kinase includes an ATP-binding site that transfers phosphate groups from ATP molecules to substrates, resulting in substrate phosphorylation. This domain also contains an important amino acid residue, Tyr511, which is transphosphorylated by lymphocyte-specific protein tyrosine kinase (Lck) for initial activation of ITK.

Role of ITK in the TCR Signaling Pathway

ITK is a key component of TCR-mediated signal transduction. The TCR interacts with peptide-major histocompatibility complex (MHC) complexes on antigen-presenting cells (APCs), thereby activating Lck and leading to phosphorylation of the cluster of differentiation 3 (CD3) immunoreceptor tyrosine-based activation motifs (ITAMs). Subsequently, the 70 kDa zeta chain-associated protein kinase (Zap-70) binds to the phosphorylated ITAMs and is phosphorylated by Lck, activating Zap-70 and causing phosphorylation of the linker for activation of T cells (LAT) and SH2 domain-containing leukocyte protein of 76 kDa (SLP-76). Activation of Src kinases leads to activation of phosphoinositide 3-kinase (PI3K), followed by generation and accumulation of phosphatidylinositol-3,4,5-trisphosphate (PIP3) at the plasma membrane.

Figure 1. Figure 1. Targeting SLP76: ITK interaction. (Weeks S,et al., 2021)

Subsequently, ITK is recruited to the membrane through its PH domain, interacts with the phosphorylated SLP-76/LAT linker complex via its SH3 and SH2 domains, and is phosphorylated at Y511 and Y180. ITK also interacts with and directly phosphorylates its downstream target PLCγ1, leading to phospholipase activation. Activated PLCγ1 hydrolyzes phosphatidylinositol-4,5-bisphosphate (PIP2) to produce the second messengers inositol triphosphate (IP3) and diacylglycerol (DAG), both of which contribute to the transmission of downstream signals to regulate gene expression. Downstream signaling molecules include Ca²⁺ channels, extracellular regulated protein kinases (Erk) activation and transcription, cytokine release, and actin reorganization. The absence of ITK does not lead to the disappearance of these downstream signaling molecules, but significantly reduces them, resulting in altered development and differentiation of different T cell lineages. ITK is not essential for TCR signal transduction, but it catalyzes TCR signaling, accelerating the rate of cellular response to TCR stimulation.

ITK and Inflammation

ITK is associated with many inflammatory diseases. The balance between pro-inflammatory Th17 cells and anti-inflammatory regulatory T cells (Treg) is critical for generating protective immune responses while minimizing autoimmunity. The imbalance between them can lead to immune diseases. ITK regulates Th17 and Treg immune responses, with research showing that ITK positively regulates Th17 cells and negatively regulates Treg cells. Additionally, ITK negatively regulates Th1 cells, which also play an important role in inflammation.

Acute Kidney Injury

Innate or adaptive immune cells release various pro-inflammatory cytokines that migrate from systemic circulation to renal tissue, causing acute kidney injury (AKI) to renal endothelial cells and tubular epithelial cells. The CD4+ T cell subtypes Th1/Th17, when hyperactivated, lead to severe inflammatory responses associated with the pathogenesis of AKI. ITK in CD4+ T cells plays a key role in regulating inflammatory pathways during AKI. During AKI pathogenesis, ITK is activated in T cells, and inhibiting ITK may lead to downregulation of Th1/Th17 cytokines and upregulation of Treg cells, thereby improving AKI. The early injury phase is characterized by upregulation of interferon-γ (IFN-γ) expression, possibly produced by Th1 cells, while in the later phase, Th17 cells sustain injury and tissue fibrosis. Inhibition of ITK can alleviate AKI.

Multiple Sclerosis

Multiple sclerosis (MS) is a disease where Th1 and Th17 cells invade the central nervous system, causing myelin tissue damage, leading to paralysis as well as neuronal damage and loss. The histopathological pattern of MS lesions is heterogeneous and varies among patients and at different stages of the disease. Research indicates that ITK promotes CD4+ T cell migration to the central nervous system, leading to neuroinflammation. ITK may play a role in the migration of antigen-specific cells across the blood-brain barrier. Therefore, the absence of ITK reduces CD4+ T cell entry into the central nervous system, transport across the brain endothelial barrier, secretion of Th1 and Th17 effector cytokines, and promotes Treg cell expansion, inhibiting neuroinflammation. ITK can regulate inflammatory T cell entry into the central nervous system, becoming a therapeutic target for MS.

Psoriasis

Psoriasis is a chronic skin disease characterized by keratinocyte hyperproliferation and immune cell infiltration in the skin, resulting in distinct silvery scales on the hands, feet, back, and other skin areas. Research shows that applying imiquimod (IMQ) to human skin causes dendritic cells to release interleukin-23 (IL-23), promoting Th0 differentiation into Th17. Cytokines secreted by Th17 lead to keratinocyte dysfunction and recruitment of neutrophils/dendritic cells, resulting in psoriatic inflammation. By using ITK inhibitors to block the ITK signaling pathway, IMQ-initiated Th1/Th17 immune responses can be significantly weakened, and Treg cells expanded, ultimately alleviating psoriasis. Therefore, inhibiting ITK may be an effective approach to treating psoriatic inflammation.

Systemic Lupus Erythematosus

Systemic lupus erythematosus (SLE) is a chronic autoimmune disease that progressively weakens multiple organs. Complex interactions between genetic, environmental factors, and hormones lead to the breakdown of immune self-tolerance, damaging multiple organs including the skin, joints, kidneys, heart, and brain. CD4+ T cells and CD8+ T cells are key factors in SLE pathogenesis and inflammation. Th17 cells and the IL-17 family of cytokines play important roles in SLE. In SLE patients, there is an interaction between Th17/Treg cells, with Th17 cells possibly inhibiting Treg cell differentiation. T cells expressing ITK (including CD4+pITK+T, CD8+pITK+ T cells) are increased in SLE patients and positively correlate with SLE severity and Th17-related cytokines. Research found no significant association between ITK-expressing T cells and serum IL-10 levels, and this preliminary result cannot infer ITK's effect on Treg cells. The role of ITK in Treg cell differentiation in SLE patients requires further investigation. Currently, there are no studies on ITK inhibitors for SLE treatment.

Pneumonia

Research indicates that high expression of phosphorylated ITK (p-ITK) in Th17 cells during acute lung injury (ALI) is associated with upregulated neutrophil-related inflammatory responses and impaired lung epithelial permeability. ITK participates in lipopolysaccharide (LPS)-induced lung inflammation and epithelial barrier dysfunction by upregulating Th17 immune responses and oxidative stress. ITK inhibitors improve airway inflammation by reducing Th17 immune response/oxidative stress in ALI mouse lungs and upregulating Treg cells, thereby decreasing high epithelial barrier permeability. ITK blockade may be a potential therapeutic strategy to alleviate ALI-related airway inflammation. ITK can differentially regulate Th17 cytokine production depending on the type of lung inflammation. In Aspergillus polysaccharide-induced Th17-driven hypersensitivity pneumonitis (HP), the secretion of IL-17A by Th17 cells and cytokine production do not require ITK.

Ulcerative Colitis

Ulcerative colitis (UC) is a chronic inflammatory bowel disease characterized by mucosal inflammation starting from the rectum and extending proximally in the colon. Research results show increased ITK phosphorylation in mucosal T cells of UC patients. ITK regulates the spectrum of mucosal cytokines specific to UC, and the expansion of mucosal CD4+ T cells expressing p-ITK in UC correlates with disease severity. Selective targeting of ITK represents a new strategy for treating UC.