Chemokines (Greek -kinos, movement) are a family of small cytokines, or signaling proteins secreted by cells. Their name is derived from their ability to induce directed chemotaxis in nearby responsive cells; they are chemotactic cytokines. Cytokine proteins are classified as chemokines according to behavior and structural characteristics. In addition to being known for mediating chemotaxis, chemokines are all approximately 8-10 kilodaltons in mass and have four cysteine residues in conserved locations that are key to forming their 3-dimensional shape. All of these proteins exert their biological effects by interacting with G protein-linked transmembrane receptors called chemokine receptors that are selectively found on the surfaces of their target cells.
Figure 1. Typical structure of chemokines.
Chemokine receptors are cytokine receptors found on the surface of certain cells that interact with a type of cytokine called chemokine. There have been 20 distinct chemokine receptors discovered in humans. Each has a 7-transmembrane (7TM) structure and couples to G-protein for signal transduction within a cell, making them members of a large protein family of G protein-coupled receptors. Following interaction with their specific chemokine ligands, chemokine receptors trigger a flux in intracellular calcium (Ca2+) ions (calcium signaling). This causes cell responses, including the onset of a process known as chemotaxis that traffics the cell to a desired location within the organism.
Figure 2. Typical structure of a chemokine receptor.
Table 1. Chemokine related products
|CXCL4 / PF4||CXCL5||CXCL6|
|CXCL7 / PPBP||CXCL8 / IL8||CXCL9|
Chemokines have been classified into four main subfamilies: CXC, CC, CX3C and XC:
The CC chemokine (or β-chemokine) proteins have two adjacent cysteines (amino acids), near their amino terminus. There have been at least 27 distinct members of this subgroup reported for mammals, called CC chemokine ligands (CCL)-1 to -28; CCL10 is the same as CCL9. Chemokines of this subfamily usually contain four cysteines (C4-CC chemokines), but a small number of CC chemokines possess six cysteines (C6-CC chemokines). C6-CC chemokines include CCL1, CCL15, CCL21, CCL23 and CCL28. CC chemokines induce the migration of monocytes and other cell types such as NK cells and dendritic cells. Examples of CC chemokine include monocyte chemoattractant protein-1 (MCP-1 or CCL2) which induces monocytes to leave the bloodstream and enter the surrounding tissue to become tissue macrophages. CCL5 (or RANTES) attracts cells such as T cells, eosinophils and basophils that express the receptor CCR5. Increased CCL11 levels in blood plasma are associated with aging (and reduced neurogenesis) in mice and humans.
|C chemokines||The third group of chemokines is known as the C chemokines (or γ chemokines), and is unlike all other chemokines in that it only has two cysteines; one N-terminal cysteine and one cysteine downstream. Two chemokines have been described for this subgroup and are called XCL1 (lymphotactin-α) and XCL2 (lymphotactin-β).|
|CXC chemokines||The two N-terminal cysteines of CXC chemokines (or α-chemokines) are separated by one amino acid, represented in this name with an "X". There have been 17 different CXC chemokines described in mammals, that are subdivided into two categories, those with a specific amino acid sequence (or motif) of glutamic acid-leucine-arginine (or ELR for short) immediately before the first cysteine of the CXC motif (ELR-positive), and those without an ELR motif (ELR-negative). ELR-positive CXC chemokines specifically induce the migration of neutrophils, and interact with chemokine receptors CXCR1 and CXCR2. An example of an ELR-positive CXC chemokine is interleukin-8 (IL-8), which induces neutrophils to leave the bloodstream and enter into the surrounding tissue. Other CXC chemokines that lack the ELR motif, such as CXCL13, tend to be chemoattractant for lymphocytes. CXC chemokines bind to CXC chemokine receptors, of which seven have been discovered to date, designated CXCR1-7.|
|CX3C chemokines||The fourth group has also been discovered and members have three amino acids between the two cysteines and is termed CX3C chemokine (or d-chemokines). The only CX3C chemokine discovered to date is called fractalkine (or CX3CL1). It is both secreted and tethered to the surface of the cell that expresses it, thereby serving as both a chemoattractant and as an adhesion molecule.|
Chemokine receptors are divided into different families, CXC chemokine receptors, CC chemokine receptors, CX3C chemokine receptors and XC chemokine receptors that correspond to the 4 distinct subfamilies of chemokines they bind. Four families of chemokine receptors differ in spacing of cysteine residues near N-terminal of the receptor.
CXC chemokine receptors
|CXC chemokine receptors are integral membrane proteins that specifically bind and respond to cytokines of the CXC chemokine family. They represent one subfamily of chemokine receptors, a large family of G protein-linked receptors that are known as seven transmembrane (7-TM) proteins, since they span the cell membrane seven times. There are currently seven known CXC chemokine receptors in mammals, named CXCR1 through CXCR7.|
|CC chemokine receptors||CC chemokine receptors (or beta chemokine receptors) are integral membrane proteins that specifically bind and respond to cytokines of the CC chemokine family. They represent one subfamily of chemokine receptors, a large family of G protein-linked receptors that are known as seven transmembrane (7-TM) proteins since they span the cell membrane seven times. To date, ten true members of the CC chemokine receptor subfamily have been described. These are named CCR1 to CCR10 according to the IUIS/WHO subcommittee on chemokine nomenclature.|
|C chemokine receptors||The "C" sub-family of chemokine receptors contains only one member: XCR1, the receptor for XCL1 and XCL2 (or lymphotactin-1 and -2). XCR1 is also known as GPR5. XCL1 is expressed by medullary thymic epithelial T cells (mTECs) while XCR1 is expressed by thymic dendritic cells (tDCs). This communication helps with the destruction of cells that are not self-tolerant.|
|CX3C chemokine receptors||CX3C chemokine receptor 1 (CX3CR1) also known as the fractalkine receptor or G-protein coupled receptor 13 (GPR13) is a protein that in humans is encoded by the CX3CR1 gene. As the name suggests, this receptor binds the chemokine CX3CL1 (also called neurotactin or fractalkine).|
The major role of chemokines is to act as a chemoattractant to guide the migration of cells. Cells that are attracted by chemokines follow a signal of increasing chemokine concentration towards the source of the chemokine. Some chemokines control cells of the immune system during processes of immune surveillance, such as directing lymphocytes to the lymph nodes so they can screen for invasion of pathogens by interacting with antigen-presenting cells residing in these tissues. These are known as homeostatic chemokines and are produced and secreted without any need to stimulate their source cell(s).
Some chemokines have roles in development; they promote angiogenesis (the growth of new blood vessels), or guide cells to tissues that provide specific signals critical for cellular maturation. Other chemokines are inflammatory and are released from a wide variety of cells in response to bacterial infection, viruses and agents that cause physical damage such as silica or the urate crystals that occur in gout. Their release is often stimulated by pro-inflammatory cytokines such as interleukin 1. Inflammatory chemokines function mainly as chemoattractants for leukocytes, recruiting monocytes, neutrophils and other effector cells from the blood to sites of infection or tissue damage. Certain inflammatory chemokines activate cells to initiate an immune response or promote wound healing. They are released by many different cell types and serve to guide cells of both innate immune system and adaptive immune system.
Intracellular signaling by chemokine receptors is dependent on neighbouring G-proteins. G-proteins exist as a heterotrimer; they are composed of three distinct subunits. When the molecule GDP is bound to the G-protein subunit, the G-protein is in an inactive state. Following binding of the chemokine ligand, chemokine receptors associate with G-proteins, allowing the exchange of GDP for another molecule called GTP, and the dissociation of the different G protein subunits. The subunit called Gβ activates an enzyme known as Phospholipase C (PLC) that is associated with the cell membrane. PLC cleaves Phosphatidylinositol (4,5)-bisphosphate (PIP2) to form two second messenger molecules called inositol triphosphate (IP3) and diacylglycerol (DAG); DAG activates another enzyme called protein kinase C (PKC), and IP3 triggers the release of calcium from intracellular stores. These events promote many signaling cascades, effecting a cellular response. For example, when CXCL8 (IL-8) binds to its specific receptors, CXCR1 or CXCR2, a rise in intracellular calcium activates the enzyme phospholipase D (PLD) that goes on to initiate an intracellular signaling cascade called the MAP kinase pathway. At the same time the G-protein subunit Gα directly activates an enzyme called protein tyrosine kinase (PTK), which phosphorylates serine and threonine residues in the tail of the chemokine receptor, causing its desensitisation or inactivation. The initiated MAP kinase pathway activates specific cellular mechanisms involved in chemotaxis, degranulation, release of superoxide anions, and changes in the avidity of cell adhesion molecules called integrins. Chemokines and their receptors play a crucial role in cancer metastatis as they are involved in extravastation, migration, micrometastatis, and angiogenesis. This role of chemokine is strikingly similar to their normal function of localizing leukocytes to an inflammatory site.
Figure 3. Chemokine receptors in the metastic process.
Role in disease
It is clear from large clinical studies that selected chemokine receptors are often upregulated in a large number of common human cancers, including those of the breast, lung, prostate, colon, and melanoma. Chemokine receptors and their corresponding chemokine ligands have been demonstrated to play a number of nonredundant roles in cancer metastasis to vital organs as well as regional lymph nodes, the most frequent site of cancer metastasis. Chemokine receptors may potentially facilitate tumor dissemination at several key steps of metastasis, including adherence of tumor cells to endothelium, extravasation from blood vessels, metastatic colonization, angiogenesis, proliferation, and protection from the host response via activation of key survival pathways such as phosphatidylinositol-3 kinase and Akt.
Guleng et al. demonstrated that neutralization of CXCR4 with blocking antibodies resulted in a delay of tumor formation by CXCR4-expressing Colon38 tumor cells. It is surprising that these antibodies also effectively delayed tumor formation and prevented angiogenesis by Colon38 tumor cells in which CXCR4 expression was silenced by CXCR4-speciﬁc, inhibitory RNAs. Thus, CXCR4 inhibitors may be useful in preventing tumor formation by blocking CXCR4-dependent processes, including blood vessel formation, independent of CXCR4 expressed by the tumor cells.
Results from our in vitro studies suggested that inhibition of chemokine receptor signaling pathways (i.e., PI-3K) renders cancer cells susceptible to apoptosis induced by B16 melano-ma-specific CTL. Although it has been shown that chemokine receptor activation increases the resistance of cancer cells to death induced by specific CTL, it is unclear whether activation of chemokine receptors leads to resistance of cancer cells to apoptosis induced by other cancer treatment modalities(e.g., cytotoxic chemotherapeutic agents and radiation therapy). One can imagine that once these apoptotic triggers are identiﬁed, chemokine receptor antagonist pretreatment combined with the speciﬁc treatment may kill tumor cells better than the single treatment alone. It is interesting that chemokine receptor antagonists (even those that target receptors with broad distribution such as CXCR4) appear to have relatively little toxicity in clinical trials. Unlike traditional anticancer drugs, they do not have demonstrable cytotoxic effects in vitro.
Already, a large number of small molecule or peptide inhibitors of chemokine receptors (mostly targeted at CXCR4) have been shown to have effects on tumor growth in animal models. It should also be noted that the use of chemokine receptor antagonists in treating metastatic tumors may rely on the up-regulation of specific chemokine receptors (e.g.,CXCR4) by tumors in the presence of hypoxia or other cellular stress and on the dependence on these receptors for angiogenesis or survival. Thus, the chemokine receptors may be likened to an Achilles’ heel of cancer. When activated, they protect the tumor against a variety of apoptotic triggers, but inhibition of these chemokine receptors may leave cancer cells vulnerable to a number of existing anticancer treatments.
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