Page 51 - Haematologica Vol. 109 - July 2024

REVIEW ARTICLE - CXCL8 in primary myelofibrosis
G. Vermeersch et al.
less frequently a 79 amino acid (CXCL8(-2-77)), protein and can be produced by almost every cell type. CXCL8 is part of the CXC-chemokine family, which contains low molecular mass proteins (~8-10 kDa) that guide leuko- cyte migration during homeostasis and inflammatory states. The chemokine subfamily classification in CXC or CC chemokines is based on conserved cysteines along the protein structure. While CXC chemokines generally bind CXC receptors (CXCR) and CC chemokines bind CC receptors (CCR), chemokine redundancy is observed (i.e., several chemokine ligands attract the same leukocyte subtype because they bind to the same receptor).45 CXCL8 interacts with its chemokine receptors CXCR1 and CXCR2, previously known as IL-8RA and IL-8RB respec- tively. The human IL8RA and IL8RB genes are located on chromosome 2.46 Both receptors are distinguished by their ligand selectivity. CXCR1 shows high affinity for CXCL6 (granulocyte chemotactic protein-2 [GCP-2]) and CXCL8. In addition to these ligands, CXCR2 also binds CXCL1 (growth-related oncogene-α [GRO-α]), CXCL2 (GRO-b), CXCL3 (GRO-γ), CXCL5 (epithelial cell-derived neutro- phil-activating peptide-78 [ENA-78]), and CXCL7 (neutro- phil-activating peptide-2 [NAP-2]).47 Both receptors are predominantly expressed on neutrophils but also appear on other myeloid or lymphoid immune cells, such as ba- sophils, monocytes, and CD8+ T-lymphocytes.48 Aberrant CXCL8 signaling is present in various hyperinflammatory and fibrosis-related diseases, amongst which idiopathic pulmonary fibrosis.45,49 The production of CXCL8 may be increased in response to pro-inflammatory cytokines, such as IL-1 and TNF-α, which stimulate CXCL8 produc- tion by binding on their cognate receptors and activating the NF-κB pathway.46,48,50,51 CXCL8 activity is also influ- enced by post-translational changes, such as truncation by proteases. By example, truncation of CXCL8 (1-77) to CXCL8 (7-77) by gelatinase B, a matrix metalloproteinase (MMP-9) mainly produced by neutrophils, results in a 10- to 27-fold higher potency in neutrophil activation.52,53 The variable quaternary structures of chemokines (existing potentially as monomers, (hetero)dimers, multimers, or in association with soluble or cell-bound glycosamino- glycans) adds an extra complexity to the research into their functionalities and receptor interactions. These variables further explain why divergent effects may be observed with the same chemokine. In vitro experiments suggest CXCL8, which may exist as a monomer or dimer, to be more potent as a monomer. Nonetheless, the ex- act effect of its quaternary structure on functionality is not completely understood.53,54 It was recently shown that CXCL8 mainly tends to bind with CXCR2 as a dimer, whereas CXCR1 strongly binds CXCL8 as a monomer. In the case of the CXCL8 dimers, one monomer interacts with the chemokine recognition site 1 (CRS1) of CXCR2. CRS1 is located at the NH2-terminus of the receptor and originates from a conserved Pro-Cys (PC) motif. While
CRS1 is responsible for the initial recruitment of CXCL8, another region called CRS2 appears to be essential for activation of CXCR2 and interacts with the conserved Glu- Leu-Arg motif (ELR) of CXCL8. The ELR motif is located at the NH2-terminus of CXCL8 and is highly conserved amongst all CXC chemokines with neutrophil-activating characteristics.55 Contrary to other ELR+ chemokines (CXCL1/2/3/5/7), CXCL8 also binds to CXCR1. This speci- ficity of CXCL8 for CXCR1 can be explained by the higher number of polar residues within the CRS1 region of CXCR1 and the charged residues in the NH2-terminal regions of CXCL8; for example, a salt bridge is formed between D26 in CXCR1 and K16 in CXCL8(1-77). For the reader inter- ested in structural biology, we refer to the articles from Ishimoto et al. and Liu et al. wherein the structural basis of, respectively, CXCR1 and CXCR2 activation is presented by using cryo-electron microscopy.55,56
Classical chemokine receptors, including CXCR1 and CX- CR2, are G protein-coupled receptors (GPCR). Activation of GPCR results in the dissociation of the Gα and Gb/γ subunit.47 The separated Gb/γ subunit activates phospho- lipase C b2 (PLCb2), which hydrolyzes phosphatidylinosi- tol 4,5-bisphosphate (PIP2) and subsequently forms the secondary messenger molecules diacylglycerol (DAG) and inositol 1,4,5-triphosphate (IP3). The formation of IP3 even- tually results in the release of Ca2+ from the endoplasmic reticulum and activates protein kinases, such as protein kinase C (PKC), which is crucial for cellular migration, degranulation, and adhesion.48 Besides PLCb2, activation of CXCR1/CXCR2 may also initiate other pathways such as activation of phosphoinositide 3-kinase γ (PI3Kγ) and phospholipase D (PLD). PI3Kγ phosphorylates PIP2 into phosphatidylinositol (3,4,5)-trisphosphate (PIP3) and ac- tivates kinases such as AKT (protein kinase B), resulting in increased cellular proliferation and survival. Activation of PLD is specifically linked to CXCR1 and is associated with the production of reactive oxygen/nitrogen species (ROS/RNS), as well as the release of neutrophil extracel- lular traps (NET) (Figure 3).48,57-59 CXCL8 is also able to bind the Duffy antigen/receptor for chemokines (DARC), also known as atypical chemokine receptor ACKR1, which functions as a scavenging ‘sink’ receptor on red blood cells (RBC) and influences the plasma concentration of various chemokines. It is believed that DARC may play a critical role in preventing oncogenesis by reducing the load of protumorigenic/proangiogenic chemokines. As such, DARC status may provide a potential explanation for the higher incidence rates and more aggressive characteris- tics of breast cancer in Black/African-American women, who generally carry the Duffy null allele on RBC with higher frequency, compared to White/European-American women.47,60,61 Although DARC status was not investigated, Peseski et al. previously reported significantly reduced OS of Non-white compared to White patients with MPN.62 Contrary to humans, mice do not express CXCL8 but ex-
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