CXCR7 Is Highly Expressed in Acute Lymphoblastic Leukemia and Potentiates CXCR4 Response to CXCL12

Recently, a novel CXCL12-binding receptor, has been identified. This CXCL12-binding receptor commonly known as CXCR7 (CXC chemokine receptor 7), has lately, based on a novel nomenclature, has received the name ACKR3 (atypical chemokine receptor 3). In this study, we aimed to investigate the expression of CXCR7 in leukemic cells, as well as its participation in CXCL12 response. Interesting, we clearly demonstrated that CXCR7 is highly expressed in acute lymphoid leukemic cells compared with myeloid or normal hematopoietic cells and that CXCR7 contributed to T-acute lymphoid leukemic cell migration induced by CXCL12. Moreover, we showed that the cellular location of CXCR7 varied among T-lymphoid cells and this finding may be related to their migration capacity. Finally, we hypothesized that CXCR7 potentiates CXCR4 response and may contribute to the maintenance of leukemia by initiating cell recruitment to bone marrow niches that were once occupied by normal hematopoietic stem cells.     

CXCL14 is no direct modulator of CXCR4

C-X-C motif chemokine 12/C-X-C chemokine receptor type 4 (CXCL12/CXCR4) signaling is involved in ontogenesis, hematopoiesis, immune function and cancer. Recently, the orphan chemokine CXCL14 was reported to inhibit CXCL12-induced chemotaxis – probably by allosteric modulation of CXCR4. We thus examined the effects of CXCL14 on CXCR4 regulation and function using CXCR4-transfected human embryonic kidney (HEK293) cells and Jurkat T cells. CXCL14 did not affect dose–response profiles of CXCL12-induced CXCR4 phosphorylation, G protein-mediated calcium mobilization, dynamic mass redistribution, kinetics of extracellular signal-regulated kinase 1 (ERK1) and ERK2 phosphorylation or CXCR4 internalization. Hence, essential CXCL12-operated functions of CXCR4 are insensitive to CXCL14, suggesting that interactions of CXCL12 and CXCL14 pathways depend on a yet to be identified CXCL14 receptor.     

Fluorescent CXCL12AF647 as a novel probe for nonradioactive CXCL12/CXCR4 cellular interaction studies.

BACKGROUND: Chemokines drive the migration of leukocytes via interaction with specific G protein-coupled 7-transmembrane receptors. The chemokine ligand/receptor pair stromal cell-derived factor-1 (SDF-1, CXCL12)/CXCR4 is gaining increasing interest because of its involvement in the metastasis of several types of cancer and in certain inflammatory autoimmune disorders such as rheumatoid arthritis. In addition, CXCR4 serves as an important coreceptor for cellular entry of T-tropic strains of human immunodeficiency virus (HIV). Therefore, potent and specific CXCR4 antagonists may have therapeutic potential as anti-HIV, anti-cancer, and anti-inflammatory drugs. METHODS AND RESULTS: Chemokine receptor antagonists can be identified by their ability to inhibit ligand binding to the receptor protein. Until now, chemokine binding assays were mostly performed with radiolabeled chemokine ligands such as [(125)I]CXCL12. To overcome the practical problems associated with such radioactive chemokine binding assays, we have developed a flow cytometric technique using a new, commercially available Alexa Fluor 647 conjugate of CXCL12 (CXCL12(AF647)). Calcium flux, chemotaxis, and p44/42 mitogen-activated protein kinase phosphorylation assays showed that the agonistic activity of the fluorescent CXCL12 was unchanged as compared with that of unlabeled CXCL12. Human T-lymphoid (CXCR4(+)) SupT1 cells and CXCR4-transfected, but not CCR5- or CXCR3-transfected, human astroglioma U87.CD4 cells specifically bound CXCL12(AF647) in a concentration-dependent manner. Unlabeled CXCL12 and the well-known CXCR4 inhibitors, AMD3100 and T22, blocked the binding of CXCL12(AF647) to SupT1 cells with 50% inhibitory concentrations of 92, 13, and 8 ng/ml, respectively. We have also used this method to evaluate CXCL12 binding and CXCR4 expression level in different subsets of human peripheral blood mononuclear cells. CONCLUSION: CXCL12(AF647) is a valuable, more convenient alternative for [(125)I]CXCL12 in ligand/receptor interaction studies.     

The Chemokine Receptor CXCR4 and c-MET Cooperatively Promote Epithelial-Mesenchymal Transition in Gastric Cancer Cells

The C-X-C motif chemokine receptor 4 (CXCR4) pathway can promote tumor metastasis but is dependent on cross talk with other signaling pathways. The MET proto-oncogene (c-MET) participates in metastasis and is highly expressed in gastric cancer. However, the relationship between CXCR4 and c-MET signaling and their mechanisms of action in gastric cancer metastasis remain unclear. In this study, in vitro experiments demonstrated that C-X-C motif chemokine ligand 12 (CXCL12)/CXCR4 induces epithelial-mesenchymal transition (EMT) and promotes migration in gastric cancer cells, which is accompanied by c-MET activation. These phenomena were reversed by c-MET inhibition. Further investigation revealed that c-MET activation correlated with its interaction with caveolin 1 in lipid rafts, induced by CXCL12. In clinical samples, we observed a significant positive association between CXCR4 expression and c-MET phosphorylation (r = 0.259, P = .005). Moreover, samples expressing both receptors were found to indicate significantly poorer patient prognosis (P < .001). These results suggest that CXCL12 induces EMT at least partially through cross talk between CXCR4 and c-MET signaling. In addition, changes in these pathways could have clinical importance for the treatment of gastric cancer.     

Chemokine Transfer by Liver Sinusoidal Endothelial Cells Contributes to the Recruitment of CD4+ T Cells into the Murine Liver

Leukocyte adhesion and transmigration are central features governing immune surveillance and inflammatory reactions in body tissues. Within the liver sinusoids, chemokines initiate the first crucial step of T-cell migration into the hepatic tissue. We studied molecular mechanisms involved in endothelial chemokine supply during hepatic immune surveillance and liver inflammation and their impact on the recruitment of CD4⁺ T cells into the liver. In the murine model of Concanavalin A-induced T cell-mediated hepatitis, we showed that hepatic expression of the inflammatory CXC chemokine ligands (CXCL)9 and CXCL10 strongly increased whereas homeostatic CXCL12 significantly decreased. Consistently, CD4⁺ T cells expressing the CXC chemokine receptor (CXCR)3 accumulated within the inflamed liver tissue. In histology, CXCL9 was associated with liver sinusoidal endothelial cells (LSEC) which represent the first contact site for T-cell immigration into the liver. LSEC actively transferred basolaterally internalized CXCL12, CXCL9 and CXCL10 via clathrin-coated vesicles to CD4⁺ T cells leading to enhanced transmigration of CXCR4⁺ total CD4⁺ T cells and CXCR3⁺ effector/memory CD4⁺ T cells, respectively in vitro. LSEC-expressed CXCR4 mediated CXCL12 transport and blockage of endothelial CXCR4 inhibited CXCL12-dependent CD4⁺ T-cell transmigration. In contrast, CXCR3 was not involved in the endothelial transport of its ligands CXCL9 and CXCL10. The clathrin-specific inhibitor chlorpromazine blocked endothelial chemokine internalization and CD4⁺ T-cell transmigration in vitro as well as migration of CD4⁺ T cells into the inflamed liver in vivo. Moreover, hepatic accumulation of CXCR3⁺ CD4⁺ T cells during T cell-mediated hepatitis was strongly reduced after administration of chlorpromazine. These data demonstrate that LSEC actively provide perivascularly expressed homeostatic and inflammatory chemokines by CXCR4- and clathrin-dependent intracellular transport mechanisms thereby contributing to the hepatic recruitment of CD4⁺ T-cell populations during immune surveillance and liver inflammation.     

The peptidomimetic CXCR4 antagonist TC14012 recruits beta-arrestin to CXCR7: roles of receptor domains

CXCR7 is an atypical chemokine receptor that signals through β-arrestin in response to agonists without detectable activation of heterotrimeric G-proteins. Its cognate chemokine ligand CXCL12 also binds CXCR4, a chemokine receptor of considerable clinical interest. Here we report that TC14012, a peptidomimetic inverse agonist of CXCR4, is an agonist on CXCR7. The potency of β-arrestin recruitment to CXCR7 by TC14012 is much higher than that of the previously reported CXCR4 antagonist AMD3100 and differs only by one log from that of the natural ligand CXCL12 (EC(50) 350 nM for TC14012, as compared with 30 nM for CXCL12 and 140 μM for AMD3100). Moreover, like CXCL12, TC14012 leads to Erk 1/2 activation in U373 glioma cells that express only CXCR7, but not CXCR4. Given that with TC14012 and AMD3100 two structurally unrelated CXCR4 antagonists turn out to be agonists on CXCR7, this likely reflects differences in the activation mechanism of the arrestin pathway by both receptors. To identify the receptor domain responsible for these opposed effects, we investigated CXCR4 and CXCR7 C terminus-swapping chimeras. Using quantitative bioluminescence resonance energy transfer, we find that the CXCR7 receptor core formed by the seven-transmembrane domains and the connecting loops determines the agonistic activity of both TC14012 and AMD3100. Moreover, we find that the CXCR7 chimera bearing the CXCR4 C-terminal constitutively associates with arrestin in the absence of ligands. Our data suggest that the CXCR4 and CXCR7 cores share ligand-binding surfaces for the binding of the synthetic ligands, indicating that CXCR4 inhibitors should be tested also on CXCR7.     

Altered regulation of CXCR4 expression during aging contributes to increased CXCL12-dependent chemotactic migration of CD4(+) T cells

Chemokine‐dependent migration of T lymphocytes assures recirculation of naïve T cells to secondary lymphoid organs and tissue‐specific trafficking of memory‐effector T cells. Previous studies carried out in rodents have demonstrated age‐associated modulation of the expression of chemokine receptors such as CXCR4 and CCR5; however, little is known about the molecular mechanisms that regulate receptor expression and turnover in T cells, during advancing age in humans. Our recent results demonstrating increased chemotactic migration in response to CXCL12 in CD4⁺ T cells obtained from the elderly, as compared to those from young donors, led us to hypothesize that increase in surface expression, because of altered endocytic regulation of CXCR4 on T cells during aging, might be directly responsible for increased migration toward CXCL12. Studies presented here demonstrate a significant increase in the surface expression of CXCR4 in CD4⁺ T cells from elderly human donors, relative to those from the young. Additionally, CXCL12‐mediated endocytosis of CXCR4 was differentially regulated during aging, which could be attributed to alterations in the ubiquitination of CXCR4. Thus, altered ubiquitination of CXCR4 may contribute to the increased surface expression and enhanced T‐cell migration to chemotactic stimuli in the elderly.     

A point mutation (G574A) in the chemokine receptor CXCR4 detected in human cancer cells enhances migration

The chemokine receptor CXCR4 is widely expressed in human cancers and regulates cell invasion, proliferation and survival. Because mutations in the CXCR4 gene could regulate its function we sequenced the coding region of the CXCR4 gene in 18 human melanoma and 3 human colon carcinoma cell lines. The same somatic point mutation (G574A; V160I) in the fourth trans-membrane region of CXCR4 was detected in one colon cancer cell line (PD) and one melanoma cell line (LB). CXCR4 was expressed and functional in both PD and LB cells, PD and LB cells migrated specifically toward the receptor ligand, CXCL12 and P-Erk was specifically induced by CXCL12. To give insight into the function of the mutant CXCR4 receptor, human A431, epidermoid carcinoma cells, were stably transfected with both mutant and wild type CXCR4. In vitro, A431 cells harboring CXCR4^G574A migrated specifically toward CXCL12 and CXCL12 induced ERK phosphorylation. Interestingly, in vivo studies showed that the growth of A431 tumors harboring CXCR4G574A was delayed compared to those harboring WT CXCR4. As expected, treatment with AMD3100, a specific CXCR4 inhibitor, reduced the in vivo growth of CXCR4_^G574A tumor b^G574A rprisingly, increased the growth of CXCR4^G574A A431 cells. This is the first report of a spontaneously occurring, functionally active CXCR4 mutation in human cancer cells. While the mutation impairs cell growth in vivo, the CXCR4 inhibitor, AMD3100, stimulated the growth of cells harboring CXCR4^G574A.     

Chemokine receptor trio: CXCR3, CXCR4 and CXCR7 crosstalk via CXCL11 and CXCL12

Although chemokines are well established to function in immunity and endothelial cell activation and proliferation, a rapidly growing literature suggests that CXC Chemokine receptors CXCR3, CXCR4 and CXCR7 are critical in the development and progression of solid tumors. The effect of these chemokine receptors in tumorigenesis is mediated via interactions with shared ligands I-TAC (CXCL11) and SDF-1 (CXCL12). Over the last decade, CXCR4 has been extensively reported to be overexpressed in most human solid tumors and has earned considerable attention toward elucidating its role in cancer metastasis. To enrich the existing armamentarium of anti-cancerous agents, many inhibitors of CXCL12–CXCR4 axis have emerged as additional or alternative agents for neo-adjuvant treatments and even many of them are in preclinical and clinical stages of their development. However, the discovery of CXCR7 as another receptor for CXCL12 with rather high binding affinity and recent reports about its involvement in cancer progression, has questioned the potential of “selective blockade” of CXCR4 as cancer chemotherapeutics. Interestingly, CXCR7 can also bind another chemokine CXCL11, which is an established ligand for CXCR3. Recent reports have documented that CXCR3 and their ligands are overexpressed in different solid tumors and regulate tumor growth and metastasis. Therefore, it is important to consider the interactions and crosstalk between these three chemokine receptors and their ligand mediated signaling cascades for the development of effective anti-cancer therapies. Emerging evidence also indicates that these receptors are differentially expressed in tumor endothelial cells as well as in cancer stem cells, suggesting their direct role in regulating tumor angiogenesis and metastasis. In this review, we will focus on the signals mediated by this receptor trio via their shared ligands and their role in tumor growth and progression.     

Functional and structural consequences of chemokine (C-X-C motif) receptor 4 activation with cognate and non-cognate agonists

Chemokine (C-X-C motif) receptor 4 (CXCR4) regulates cell trafficking and plays important roles in the immune system. Ubiquitin has recently been identified as an endogenous non-cognate agonist of CXCR4, which activates CXCR4 via interaction sites that are distinct from those of the cognate agonist C-X-C motif chemokine ligand 12 (CXCL12). As compared with CXCL12, chemotactic activities of ubiquitin in primary human cells are poorly characterized. Furthermore, evidence for functional selectivity of CXCR4 agonists is lacking, and structural consequences of ubiquitin binding to CXCR4 are unknown. Here, we show that ubiquitin and CXCL12 have comparable chemotactic activities in normal human peripheral blood mononuclear cells, monocytes, vascular smooth muscle, and endothelial cells. Chemotactic activities of the CXCR4 ligands could be inhibited with the selective CXCR4 antagonist AMD3100 and with a peptide analogue of the second transmembrane domain of CXCR4. In human monocytes, ubiquitin- and CXCL12-induced chemotaxis could be inhibited with pertussis toxin and with inhibitors of phospholipase C, phosphatidylinositol 3 kinase, and extracellular signal-regulated kinase 1/2. Both agonists induced inositol trisphosphate production in vascular smooth muscle cells, which could be inhibited with AMD3100. In β-arrestin recruitment assays, ubiquitin did not sufficiently recruit β-arrestin2 to CXCR4 (EC₅₀ > 10 μM), whereas the EC₅₀ for CXCL12 was 4.6 nM (95% confidence interval 3.1–6.1 nM). Both agonists induced similar chemical shift changes in the ¹³C-¹H-heteronuclear single quantum correlation (HSQC) spectrum of CXCR4 in membranes, whereas CXCL11 did not significantly alter the ¹³C-¹H-HSQC spectrum of CXCR4. Our findings point towards ubiquitin as a biased agonist of CXCR4.     

Discovery and Computer Aided Potency Optimization of a Novel Class of Small Molecule CXCR4 Antagonists

Amongst the chemokine signalling axes involved in cancer, chemokine CXCL12 acting on chemokine receptor CXCR4 is particularly significant since it orchestrates migration of cancer cells in a tissue-specific metastatic process. High CXCR4 tumour expression is associated with poor prognosis of lung, brain, CNS, blood and breast cancers. We have identified a new class of small molecule CXCR4 antagonists based on the use of computational modelling studies in concert with experimental determination of in vitro activity against CXCL12-induced intracellular calcium mobilisation, proliferation and chemotaxis. Molecular modelling proved to be a useful tool in rationalising our observed potencies, as well as informing the direction of the synthetic efforts aimed at producing more potent compounds.     

SDF-1/CXCL12 regulates cAMP production and ion transport in intestinal epithelial cells via CXCR4

Human colonic epithelial cells express CXCR4, the sole cognate receptor for the chemokine stromal cell-derived factor (SDF)-1/CXC chemokine ligand (CXCL) 12. The aim of this study was to define the mechanism and functional consequences of signaling intestinal epithelial cells through the CXCR4 chemokine receptor. CXCR4, but not SDF-1/CXCL12, was constitutively expressed by T84, HT-29, HT-29/-18C1, and Caco-2 human colon epithelial cell lines. Studies using T84 cells showed that CXCR4 was G protein-coupled in intestinal epithelial cells. Moreover, stimulation of T84 cells with SDF-1/CXCL12 inhibited cAMP production in response to the adenylyl cyclase activator forskolin, and this inhibition was abrogated by either anti-CXCR4 antibody or receptor desensitization. Studies with pertussis toxin suggested that SDF-1/CXCL12 activated negative regulation of cAMP production through G(i)alpha subunits coupled to CXCR4. Consistent with the inhibition of forskolin-stimulated cAMP production, SDF-1/CXCL12 also inhibited forskolin-induced ion transport in voltage-clamped polarized T84 cells. Taken together, these data indicate that epithelial CXCR4 can transduce functional signals in human intestinal epithelial cells that modulate important cAMP-mediated cellular functions.     

趋化因子SD-1及其受体CXCR4在卵巢癌中的研究进展

基质细胞衍生因子-1(Stromal cell derived factor-1,SDF-1)是CXC趋化因子家族的重要成员,系统命名为CXCL12,能与它的唯一受体CXC趋化因子受体-4(CXC chemokine receptor-4,CXCR4)形成CXCL12-CXCR4生物学轴,CXCL12-CXCR4生物学轴在肿瘤生长、侵袭、转移过程中发生重要作用。到目前为止,已发现CXCL12-CXCR4在卵巢癌、胰腺癌、肝癌等多种肿瘤组织中表达。然而,国内目前还没有关于CXCL12-CXCR4与卵巢癌关系的相关综述,本文将从趋化因子CXCL12及其受体CXCR4,CXCL12/CXCR4轴与卵巢癌细胞系实验研究,CXCL12-CXCR4轴与卵巢癌的临床研究,CXCL12/CXCR4与卵巢癌预后,CXCL12/CXCR4与卵巢癌治疗展望等五个方面对CXCL12-CXCR4生物轴与卵巢癌的关系,及其在卵巢癌治疗中的应用展开综述。     

CXCR4 and CXCR7 transduce through mTOR in human renal cancer cells

Treatment of metastatic renal cell carcinoma (mRCC) has improved significantly with the advent of agents targeting the mTOR pathway, such as temsirolimus and everolimus. However, their efficacy is thought to be limited by feedback loops and crosstalk with other pathways leading to the development of drug resistance. As CXCR4–CXCL12–CXCR7 axis has been described to have a crucial role in renal cancer; the crosstalk between the mTOR pathway and the CXCR4–CXCL12–CXCR7 chemokine receptor axis has been investigated in human renal cancer cells. In SN12C and A498, the common CXCR4–CXCR7 ligand, CXCL12, and the exclusive CXCR7 ligand, CXCL11, activated mTOR through P70S6K and 4EBP1 targets. The mTOR activation was specifically inhibited by CXCR4 antagonists (AMD3100, anti-CXCR4-12G5 and Peptide R, a newly developed CXCR4 antagonist) and CXCR7 antagonists (anti-CXCR7-12G8 and CCX771, CXCR7 inhibitor). To investigate the functional role of CXCR4, CXCR7 and mTOR in human renal cancer cells, both migration and wound healing were evaluated. SN12C and A498 cells migrated toward CXCL12 and CXCL11; CXCR4 and CXCR7 inhibitors impaired migration and treatment with mTOR inhibitor, RAD001, further inhibited it. Moreover, CXCL12 and CXCL11 induced wound healing while was impaired by AMD3100, the anti CXCR7 and RAD001. In SN12C and A498 cells, CXCL12 and CXCL11 promoted actin reorganization characterized by thin spikes at the cell periphery, whereas AMD3100 and anti-CXCR7 impaired CXCL12/CXCL11-induced actin polymerization, and RAD001 treatment further reduced it. In addition, when cell growth was evaluated in the presence of CXCL12, CXCL11 and mTOR inhibitors, an additive effect was demonstrated with the CXCR4, CXCR7 antagonists and RAD001. RAD001-resistant SN12C and A498 cells recovered RAD001 sensitivity in the presence of CXCR4 and CXCR7 antagonists. In conclusion, the entire axis CXCR4–CXCL12–CXCR7 regulates mTOR signaling in renal cancer cells offering new therapeutic opportunities and targets to overcome resistance to mTOR inhibitors.Renal cell carcinoma (RCC) is the most lethal malignancy among urological cancers with a total of 64 770 new cases and 13 570 deaths estimated in the United States in 2012.¹ A growing understanding of the molecular biology of RCC changed the therapeutic approach toward target-based agents. Since 2005, the US Food and Drug Administration (FDA) has approved six new target agents for metastatic RCC that antagonize two principal signaling pathways: the vascular endothelial growth factor receptor (VEGF) and the mammalian target of rapamycin (mTOR).² The mTOR is an atypical intracellular serine/threonine protein kinase regulated by phosphatidylinositol 3-kinase (PI3K).³ mTOR exists in two distinct complexes termed mTOR complex 1 (mTORC1) comprising mTOR, mLST8 (also termed G-protein β-subunit-like protein, GβL, a yeast homolog of LST8), raptor (regulatory associated protein of mTOR) and PRAS40 (proline-rich Akt substrate, 40 kDa), and mTOR complex 2 (mTORC2) comprising mTOR, mLST8, rictor (rapamycin-insensitive companion of mTOR), mSin1 (mammalian stress-activated protein kinase (SAPK)-interacting protein 1), protor (protein observed with rictor) and PRR5 (proline-rich protein 5).⁴ mTORC1 responds to amino acids, stress, oxygen, energy and growth factors and is sensitive to rapamycin; when active, mTORC1 promotes cell growth and also drives cell-cycle progression. Alternatively, mTORC2 regulates cytoskeletal organization and cell survival/metabolism and is sensitive to rapamycin over longer incubation times or at higher doses.³ mTORC1 controls cell growth and translation through the phosphorylation of ribosomal protein S6 kinase (S6K) and of eukaryotic translation initiation factor 4EBP1, which regulate either the translation of ribosomal proteins or the cap-dependent translation by inhibition of eukaryotic translation initiation factor 4E, respectively.^{3, 4} The activated mTOR pathway has been identified in several human malignancies, thus being an attractive target for anticancer therapy. mTORC1 activity is inhibited by rapalogs such as rapamycin (sirolimus) and associated analogs (temsirolimus/CCI-779, RAD001, ridaforolimus/AP23573).⁵ These drugs suppress mTORC1 activity forming a complex with FK506-binding protein 12. Temsirolimus (rapamycin analog) was the first mTOR inhibitor approved as first-line treatment in patients with poor-prognosis metastatic RCC (mRCC) patients,³ ridaforolimus is currently tested in phase III clinical trials⁵ and RAD001 is indicated as second-line treatment in patients with RCC at failure of first-line treatment with sunitinib or sorafenib. Other indications are subependymal giant cell astrocytoma associated with tuberous sclerosis and progressive neuroendocrine tumors of pancreatic origin.⁵ Although mTOR inhibitors prolong progression-free survival in patients with advanced RCC, most patients develop resistance to mTOR-inhibiting agents, limiting their efficacy; the new frontier of inhibiting the mTOR pathway is to identify agents targeting the feedback loops and crosstalks with other pathways involved in the acquired resistance to mTOR inhibitors.⁶Chemokines and their receptors have been implicated in regulating RCC growth, angiogenesis and metastases.⁷ In RCC, VHL mutation resulted in HIF-dependent CXCR4 activation⁸ and CXCR4 expression predicted poor tumor-specific survival.^{8, 9, 10} Recently, CXCL12 was shown to bind with high affinity the orphan receptor CXCR7/RDC1, which also binds a second ligand in the form of interferon-inducible T-cell α chemoattractant (I-TAC/CXCL11).¹¹ Whereas the CXCR4 activity is primarily G-protein-mediated, CXCR7 is considered an atypical GPCR because ligand binding does not result in intracellular Ca2+ release.¹¹ Some studies provided evidence that CXCR7 represents a ‘decoy'' receptor, which is responsible for either sequestering extracellular CXCL12¹² or modulating CXCR4 signaling by forming CXCR7–CXCR4 heterodimers.¹³ In contrast, others demonstrated that CXCR7 relays intracellular signals^{14, 15, 16, 17} and promotes cell motility^{18, 13, 19} acting through β-arrestin.^{20, 21} CXCR7 is highly expressed in human cancers such as prostate, lung, glioma, ovarian, breast cancer cells and in tumor-associated blood vessels and seems to be essential for survival, adhesion and growth of tumor cells.^{11, 14, 15, 22, 23, 24} It was recently demonstrated that CXCR4 and CXCR7 predict prognosis in RCC.^{10, 25} CXCL12 activates CXCR4 and the derived signaling can transduce on the mTOR pathway in pancreatic cancer, gastric cancer and T-cell leukemia cells;^{26, 27, 28, 29} antagonists targeting PI3K and/or mTOR inhibited CXCL12-mediated cell migration and this effect was primarily attributed to the inhibition of mTORC1 and consequent decrease in RhoA, Cdc42 and Rac1 in human gastric carcinoma cells.²⁸Aim of the study was to evaluate interactions between the CXCL12–CXCR4–CXCR7 axis and the mTOR pathway in human renal cancer cells to identify new therapeutic opportunities and overcome resistance mechanisms.     

The role of polymorphisms of genes CXCL12/CXCR4 and MIF in the risk development IBD the Polish population

Inflammatory bowel disease (IBD) are characterized recurrent inflammation of gastrointestinal tract. The etiology and pathogenesis this disease is currently unclear, but it has become evident that immune and genetic factors are involved in this process. The aim of this study was to determine whether gene polymorphisms: MIF-173 G/C; CXCL12-801 G/A and CXCR4 C/T exon 2 position of rs2228014 is associated with susceptibility to IBD. A total of 286 patients were examined with IBD, including 152 patients with ulcerative colitis and 134 with Crohn’s disease (CD) and 220 healthy subjects were recruited from the Polish population. Genotyping for polymorphisms in CXCL12/CXCR4 and MIF was performed by RFLP-PCR. Statistical significance was found for polymorphisms CXCR4, a receptor gene for CXCL12 genotypes and alleles in CD and for genotype C/T and T allele in ulcerative colitis with respect to control. This confirms the effect of CXCL12 gene. The interplay between CXCL12 and its receptor CXCR4 affects homeostasis and inflammation in the intestinal mucosa. Three-gene analysis in CD confirmed the association of genotype GGGGCT. Statistical analysis of clinical data of patients with ulcerative colitis showed significant differences in the distribution of genotype C/T and T allele for CXCR4 in the left-side colitis. Having CXCR4/CXCL12 chemokine axis polymorphisms may predispose to the development of IBD. Activation can also be their defensive reaction to the long-lasting inflammation.     

The peculiarities of the SDF-1/CXCL12 system: in some cells,CXCR4 and CXCR7 sing solos,in others,they sing duets

The chemokine SDF-1/CXCL12 induces and modulates major steps of ontogenesis, regeneration and tumorigenesis. Depending on the organ or tissue, CXCL12 serves as a proliferation or cell survival factor, influences differentiation, induces adhesion and/or regulates cell migration. These functions are mediated by the two chemokine receptors, CXCR4 and CXCR7. Whereas CXCR4 is still viewed as the sole G-protein-activating and, hence, signaling receptor for CXCL12, CXCR7 is regarded as a non-classic scavenging or decoy receptor that modulates the function of CXCR4. However, this view might be too limited, since evidence has accumulated favoring a cell-type-specific mode of CXCL12 signaling. In addition to the “classic” CXCL12 signaling mode via CXCR4, CXCR4 and CXCR7 have to form a receptor unit for successful CXCL12 signaling in some cells. Moreover, examples exist whereby CXCL12 receptors split functions or switch roles, such that CXCR7 (instead of CXCR4) mediates signal transduction. The obvious lack of a universal mode of CXCL12 signaling urges a re-evaluation of the role of this chemokine in development, health and disease. This review depicts the exceptional characteristics of CXCL12-induced signal transduction in various cells and organs, points out remaining controversies and mentions consequences for therapeutic interventions.     

ZAP-70 Promotes the Infiltration of Malignant B-Lymphocytes into the Bone Marrow by Enhancing Signaling and Migration after CXCR4 Stimulation

ZAP-70 in chronic lymphocytic leukemia (CLL) is associated with enhanced response to microenvironmental stimuli. We analyzed the functional consequences of ZAP-70 ectopic expression in malignant B-cells in a xenograft mouse model of disseminated B-cell leukemia. Mice injected with B-cells expressing ZAP-70 showed a prominently higher infiltration of the bone marrow. In vitro analysis of the response of malignant B-cells to CXCL12, the main attracting chemokine regulating trafficking of lymphocytes to the bone marrow, or to bone marrow stromal cells, revealed that ZAP-70 induces an increased response in terms of signaling and migration. These effects are probably mediated by direct participation of ZAP-70 in CXCL12-CXCR4 signaling since CXCR4 stimulation led to activation of ZAP-70 and downstream signaling pathways, such as MAPK and Akt, whereas ZAP-70 did not alter the expression of the CXCR4 receptor. In addition, subclones of primary CLL cells with high expression of ZAP-70 also showed increased migrative capacity toward CXCL12. Neutralization of CXCR4 with a monoclonal antibody resulted in impaired in vitro responses to CXCL12 and bone marrow stromal cells. We conclude that ZAP-70 enhances the migration of malignant B-cells into the supportive microenvironment found in the bone marrow mainly by enhancing signaling and migration after CXCR4 stimulation.     

In vitro characterization and inhibition of the CXCR4/CXCL12 chemokine axis in human uveal melanoma cell lines

Purpose  

The CXCR4/CXCL12 chemokine axis may play a critical role in guiding CXCR4+ circulating malignant cells to organ specific locations that actively secrete its ligand CXCL12 (SDF-1) such as bone, brain, liver, and lungs. We sought to characterize the presence of the CXCR4/CXCL12 axis in five uveal melanoma (UM) cell lines in vitro. The ability of TN14003, a synthetic peptide inhibitor that targets the CXCR4 receptor complex, to inhibit this axis was also assessed.