CXCL16 Promotes Gastric Cancer Tumorigenesis via ADAM10-Dependent CXCL16/CXCR6 Axis and Activates Akt and MAPK Signaling Pathways

Abnormal expression of CXC motif chemokine ligand 16 (CXCL16) has been demonstrated to be associated with tumor progression and metastasis, served as a prognostic factor in many cancers, with higher relative expression behaving as a marker of tumor progression. However, its role and mechanisms underlying progression and metastasis of gastric cancer (GC) are yet to be elucidated. In our investigation, public datasets and human GC tissue samples were used to determine the CXCL16 expression levels. Our results revealed that CXCL16 was upregulated in GC. The high expression CXCL16 in GC was significantly associated with histologic poor differentiation and pTNM staging. And high CXCL16 was positively correlated with the poor survival of GC patients. Gain-and loss-of-function experiments were employed to investigate the biological role of CXCL16 in proliferation and migration both in vitro and in vivo. Mechanically, Gene set enrichment analysis (GSEA) revealed that the epithelial‑mesenchymal transition (EMT), Akt and MAPK signal pathway related genes were significantly enriched in the high CXCL16 group, which was confirmed by western blot. Moreover, overexpression CXCL16 promoted the disintegrin and metalloproteases (ADAM10) and the CXC motif chemokine receptor 6 (CXCR6) expression, which mediated the CXCL16/CXCR6 positive feedback loop in GC, with activating Akt and MAPK signaling pathways. Knocking down ADAM10 would interrupted the CXCL16/CXCR6 axis in the carcinogenesis and progression of GC. In conclusion, our findings offered insights into that CXCL16 promoted GC tumorigenesis by enhancing ADAM10-dependent CXCL16/CXCR6 axis activation.     

CXCR4/CXCL12 axis promotes VEGF-mediated tumor angiogenesis through Akt signaling pathway

CXC chemokine receptor 4 (CXCR4) has been shown to play a critical role in chemotaxis and homing, which are key steps in cancer metastasis. There is also increasing evidence that links this receptor to angiogenesis; however, its molecular basis remains elusive. Vascular endothelial growth factor (VEGF), one of the major angiogenic factors, promotes the formation of leaky tumor vasculatures that are the hallmarks of tumor progression. Here, we investigated whether CXCR4 induces the expression of VEGF through the PI3K/Akt pathway. Our results showed that CXCR4/CXCL12 induced Akt phosphorylation, which resulted in upregulation of VEGF at both the mRNA and protein levels. Conversely, blocking the activation of Akt signaling led to a decrease in VEGF protein levels; blocking CXCR4/CXCL12 interaction with a CXCR4 antagonist suppressed tumor angiogenesis and growth in vivo. Furthermore, VEGF mRNA levels correlated well with CXCR4 mRNA levels in patient tumor samples. In summary, our study demonstrates that the CXCR4/CXCL12 signaling axis can induce angiogenesis and progression of tumors by increasing expression of VEGF through the activation of PI3K/Akt pathway. Our findings suggest that targeting CXCR4 could provide a potential new anti-angiogenic therapy to suppress the formation of both primary and metastatic tumors.     

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。     

The CXCL8-CXCR1/2 pathways in cancer

Persistent infection or chronic inflammation contributes significantly to tumourigenesis and tumour progression. C-X-C motif ligand 8 (CXCL8) is a chemokine that acts as an important multifunctional cytokine to modulate tumour proliferation, invasion and migration in an autocrine or paracrine manner. Studies have suggested that CXCL8 and its cognate receptors, C-X-C chemokine receptor 1 (CXCR1) and C-X-C chemokine receptor 2 (CXCR2), mediate the initiation and development of various cancers including breast cancer, prostate cancer, lung cancer, colorectal carcinoma and melanoma. CXCL8 also integrates with multiple intracellular signalling pathways to produce coordinated effects. Neovascularisation, which provides a basis for fostering tumour growth and metastasis, is now recognised as a critical function of CXCL8 in the tumour microenvironment. In this review, we summarize the biological functions and clinical significance of the CXCL8 signalling axis in cancer. We also propose that CXCL8 may be a potential therapeutic target for cancer treatment.     

CXCR4 and CXCL12 are inversely expressed in colorectal cancer cells and modulate cancer cell migration, invasion and MMP-9 activation

Colorectal cancer (CRC) is characterized by a distinct metastatic pattern resembling chemokine-induced leukocyte trafficking. This prompted us to investigate expression, signal transduction and specific functions of the chemokine receptor CXCR4 in CRC cells and metastases. Using RT-PCR analysis and Western blotting, we demonstrated CXCR4 and CXCL12 expression in CRC and CRC metastases. Cell differentiation increases CXCL12 mRNA levels. Moreover, CXCR4 and its ligand are inversely expressed in CRC cell lines with high CXCR4 and low or not detectable CXCL12 expression. CXCL12 activates ERK-1/2, SAPK/JNK kinases, Akt and matrix metalloproteinase-9. These CXCL12-induced signals mediate reorganization of the actin cytoskeleton resulting in increased cancer cell migration and invasion. Moreover, CXCL12 increases vascular endothelial growth factor (VEGF) expression and cell proliferation but has no effect on CRC apoptosis. Therefore, the CXCL12/CXCR4 system is an important mediator of invasion and metastasis of CXCR4 expressing CRC cells.     

Inhibition of chemokine (CXC motif) ligand 12/chemokine (CXC motif) receptor 4 axis (CXCL12/CXCR4)-mediated cell migration by targeting mammalian target of rapamycin (mTOR) pathway in human gastric carcinoma cells

CXCL12/CXCR4 plays an important role in metastasis of gastric carcinoma. Rapamycin has been reported to inhibit migration of gastric cancer cells. However, the role of mTOR pathway in CXCL12/CXCR4-mediated cell migration and the potential of drugs targeting PI3K/mTOR pathway remains unelucidated. We found that CXCL12 activated PI3K/Akt/mTOR pathway in MKN-45 cells. Stimulating CHO-K1 cells expressing pEGFP-C1-Grp1-PH fusion protein with CXCL12 resulted in generation of phosphatidylinositol (3,4,5)-triphosphate, which provided direct evidence of activating PI3K by CXCL12. Down-regulation of p110β by siRNA but not p110α blocked phosphorylation of Akt and S6K1 induced by CXCL12. Consistently, p110β-specific inhibitor blocked the CXCL12-activated PI3K/Akt/mTOR pathway. Moreover, CXCR4 immunoprecipitated by anti-p110β antibody increased after CXCL12 stimulation and G(i) protein inhibitor pertussis toxin abrogated CXCL12-induced activation of PI3K. Further studies demonstrated that inhibitors targeting the PI3K/mTOR pathway significantly blocked the chemotactic responses of MKN-45 cells triggered by CXCL12, which might be attributed primarily to inhibition of mTORC1 and related to prevention of F-actin reorganization as well as down-regulation of active RhoA, Rac1, and Cdc42. Furthermore, rapamycin inhibited the secretion of CXCL12 and the expression of CXCR4, which might form a positive feedback loop to further abolish upstream signaling leading to cell migration. Finally, we found cells expressing high levels of cxcl12 were sensitive to rapamycin in its activity inhibiting migration as well as proliferation. In summary, we found that the mTOR pathway played an important role in CXCL12/CXCR4-mediated cell migration and proposed that drugs targeting the mTOR pathway may be used for the therapy of metastatic gastric cancer expressing high levels of cxcl12.     

SDF-1/CXCR4在肿瘤信号转导中作用的分子机制

CXC趋化因子受体4(CXCR4)是最主要的趋化因子受体之一,在多种类型细胞中均有表达,包括淋巴细胞、造血干细胞、内皮细胞和肿瘤细胞。CXCR4与其配体——基质细胞衍生因子1(SDF-1)(也称CXCL12)结合,能介导多种与细胞趋化、细胞存活或增殖相关信号传导通路。CXCR4与SDF-1轴涉及肿瘤的恶性演进、血管生成、转移和存活。因此,阻断CXCR4与SDF-1轴及下游信号通路成为相关治疗的分子靶标。     

The role of CXC chemokine ligand 4/CXC chemokine receptor 3-B in breast cancer progression

Chemokines and their receptors participate in the development of cancers by enhancing tumor cell proliferation, angiogenesis, invasion, metastasis and penetration of tumor immune cells. It remains unclear whether CXC chemokine ligand 4 (CXCL4)/CXC chemokine receptor 3-B (CXCR3-B) can be used as an independent molecular marker for establishing prognosis for breast cancer patients. We evaluated CXCL4 and CXCR3-B expression in 114 breast cancer tissues and 30 matched noncancerous tissues using immunohistochemistry and western blot, and determined the correlation between their expression and clinicopathologic findings. We observed that breast cancer tissues express CXCL4 strongly and CXCR3-B weakly compared to noncancerous tissues. Strong CXCL4 expression was detected in 94.7% and weak CXCR3-B expression was detected in 78.9% of the tissues. Therefore, CXCL4/CXCR3-B might play a crucial role in breast cancer progression. We found no significant correlation between CXCL4 and age, tumor stage, tumor grade or TNM stage. CXCR3-B was associated significantly with tumor grade. Moreover, the Chi-square test of association showed that the expression of CXCL4/CXCR3-B might be an independent prognostic marker for breast cancer. Therefore, we suggest that CXCR3-B is an indicator of poor prognosis and may also be a chemotherapeutic target.     

CXCR4/CXCL12 在肿瘤中的研究进展

研究表明趋化因子及其受体在胚胎发育、干细胞迁移以及各种免疫反应中发挥重要作用,是许多生理及病理过程中细胞运动的重要因素。趋化因子受体CXCR4是一个由352个氨基酸构成的、7次跨膜的G蛋白偶联受体。趋化因子CXCL12为其特异性受体。研究发现,CXCR4/CXCL12在多种肿瘤中都有表达,在肿瘤的生长、血管生成、转移等方面发挥着重要作用。与正常组织相比,肿瘤组织及转移灶CXCR4高表达。因此,对CXCR4/CXCL12轴在肿瘤病生理中的作用机制进行进一步研究,很可能为肿瘤的治疗及对肿瘤转移的预防提供一个新的思路。我们现在就对其在肿瘤病生理中的作用做一综述。     

The Chemokine CXCL16 and Its Receptor,CXCR6, as Markers and Promoters of Inflammation-Associated Cancers

Clinical observations and mouse models have suggested that inflammation can be pro-tumorigenic. Since chemokines are critical in leukocyte trafficking, we hypothesized that chemokines play essential roles in inflammation-associated cancers. Screening for 37 chemokines in prostate cancer cell lines and xenografts revealed CXCL16, the ligand for the receptor CXCR6, as the most consistently expressed chemokine. Immunohistochemistry and/or immunofluorescence and confocal imaging of 121 human prostate specimens showed that CXCL16 and CXCR6 were co-expressed, both on prostate cancer cells and adjacent T cells. Expression levels of CXCL16 and CXCR6 on cancer cells correlated with poor prognostic features including high-stage and high-grade, and expression also correlated with post-inflammatory changes in the cancer stroma as revealed by loss of alpha-smooth muscle actin. Moreover, CXCL16 enhanced the growth of CXCR6-expressing cancer and primary CD4 T cells. We studied expression of CXCL16 in an additional 461 specimens covering 12 tumor types, and found that CXCL16 was expressed in multiple human cancers associated with inflammation. Our study is the first to describe the expression of CXCL16/CXCR6 on both cancer cells and adjacent T cells in humans, and to demonstrate correlations between CXCL16 and CXCR6 vs. poor both prognostic features and reactive changes in cancer stoma. Taken together, our data suggest that CXCL16 and CXCR6 may mark cancers arising in an inflammatory milieu and mediate pro-tumorigenic effects of inflammation through direct effects on cancer cell growth and by inducing the migration and proliferation of tumor-associated leukocytes.     

Stromal cell-derived factor-1alpha (SDF-1alpha/CXCL12) stimulates ovarian cancer cell growth through the EGF receptor transactivation

Ovarian cancer (OC) is the leading cause of death in gynecologic diseases in which there is evidence for a complex chemokine network. Chemokines are a family of proteins that play an important role in tumor progression influencing cell proliferation, angiogenic/angiostatic processes, cell migration and metastasis, and, finally, regulating the immune cells recruitment into the tumor mass. We previously demonstrated that astrocytes and glioblastoma cells express both the chemokine receptor CXCR4 and its ligand stromal cell-derived factor-1 (SDF-1), and that SDF-1alpha treatment induced cell proliferation, supporting the hypothesis that chemokines may play an important role in tumor cells' growth in vitro. In the present study, we report that CXCR4 and SDF-1 are expressed in OC cell lines. We demonstrate that SDF-1alpha induces a dose-dependent proliferation in OC cells, by the specific interaction with CXCR4 and a biphasic activation of ERK1/2 and Akt kinases. Our results further indicate that CXCR4 activation induces EGF receptor (EGFR) phosphorylation that in turn was linked to the downstream intracellular kinases activation, ERK1/2 and Akt. In addition, we provide evidence for cytoplasmic tyrosine kinase (c-Src) involvement in the SDF-1/CXCR4-EGFR transactivation. These results suggest a possible important "cross-talk" between SDF-1/CXCR4 and EGFR intracellular pathways that may link signals of cell proliferation in ovarian cancer.     

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.     

趋化因子CXCL12及其受体CXCR4在胶质瘤中的研究进展

CXCL12是趋化因子家族成员之一,是能够特异性结合其受体CXCR4发挥趋化性作用的细胞因子。最初,CXCL12及CXCR4被发现于炎症细胞,参与机体炎症、免疫等病理反应。接下来的几年中发现,它在机体发育、成熟过程中也有重要作用。如今,大量研究表明它与肿瘤的生长、侵袭及转移密切相关。据报道,在乳腺癌、肺癌、卵巢癌等二十余种肿瘤组织中发现CXCL12及CXCR4的表达,其中也包括中枢系统肿瘤-胶质瘤。CXCL12/CXCR4参与胶质瘤生长过程的多个步骤,包括肿瘤增殖、侵袭、转移等。有实验指出,转移灶的CXCR4表达水平较原发灶高,CXCR4有可能成为抑制胶质瘤生长、转移的重要靶目标。     

趋化因子CXCL16与临床疾病

趋化因子在T细胞迁移过程中起重要作用,CXCL16是作为磷脂酰丝氨酸和ox-LDL的清道夫受体的多功能趋化因子。血管内皮细胞同时表达功能性地分泌型和膜结合型CXCL16分子,分泌型CXCL16参与激活T淋巴细胞趋化。膜结合型CXCL16作为粘附分子,通过它的趋化因子活性区参与活化T淋巴细胞与血管内皮细胞之间的识别和直接粘附;促进大量的特异性炎症细胞浸润。研究证明趋化因子CXCL16在多种临床疾病中扮演起重要作用。本文综述了CXCL16与临床疾病的关系及其研究进展。     

Stroma-derived factor (SDF-1/CXCL12) and human tumor pathogenesis

The chemokine stroma-derived factor (SDF-1/CXCL12) plays multiple roles in tumor pathogenesis. It has been demonstrated that CXCL12 promotes tumor growth and malignancy, enhances tumor angiogenesis, participates in tumor metastasis, and contributes to immunosuppressive networks within the tumor microenvironment. Therefore, it stands to reason that the CXCL12/CXCR4 pathway is an important target for the development of novel anti-cancer therapies. In this review, we consider the pathological nature and characteristics of the CXCL12/CXCR4 pathway in the tumor microenvironment. Strategies for therapeutically targeting the CXCL12/CXCR4 axis also are discussed. migration; immune suppression; tumor angiogenesis; tumor metastasis; stem cells     

CXCR4 chemokine receptor engagement modifies integrin dependent adhesion of renal carcinoma cells

The mechanisms leading to renal cell carcinoma (RCC) metastasis are incompletely understood. Although evidence shows that the chemokine receptor CXCR4 and its ligand CXCL12 may regulate tumor dissemination, their role in RCC is not clearly defined. We examined CXCR4 expression and functionality on RCC cell lines, and explored CXCL12-triggered tumor adhesion to human endothelium (HUVEC) or extracellular matrix proteins. Functional CXCR4 was expressed on A498 tumor cells, enabling them to migrate towards a CXCL12 gradient. CXCR4 engagement by CXCL12 induced elevated cell adhesion to HUVEC, to immobilized fibronectin, laminin or collagen. Anti-CXCR4 antibodies or CXCR4 knock down by siRNA applied prior to CXCL12 stimulation impaired CXCL12-triggered tumor adhesion. However, blocking CXCR4 subsequent to CXCL12 stimulation did not. This pointed to an indirect control of tumor cell adhesion by CXCR4. In fact, CXCR4 engagement by CXCL12 also induced alterations of receptors of the integrin family, notably alpha3, alpha5, beta1 and beta3 subunits, and blocking beta1 integrins with a function-blocking antibody prevented CXCL12-induced A498 adhesion. Focal adhesion kinase (total and activated) and integrin-linked kinase significantly increased in CXCL12-treated A498 cells, accompanied by a distinct up-regulation of ERK1/2, JNK and p38 phosphorylation. Therefore, CXCR4 may be crucial in controlling adhesion of A498 cells via cross talking with integrin receptors. These data show that CXCR4 receptors contribute to RCC dissemination and may provide a novel link between CXCR4 chemokine receptor expression and integrin triggered RCC adhesion to the vascular wall and subendothelial matrix components.     

Discovery of small molecule antagonists of chemokine receptor CXCR6 that arrest tumor growth in SK-HEP-1 mouse xenografts as a model of hepatocellular carcinoma

The chemokine system plays an important role in mediating a proinflammatory microenvironment for tumor growth in hepatocellular carcinoma (HCC). The CXCR6 receptor and its natural ligand CXCL16 are expressed at high levels in HCC cell lines and tumor tissues and receptor expression correlates with increased neutrophils in these tissues contributing to poor prognosis in patients. Availability of pharmacologcal tools targeting the CXCR6/CXCL16 axis are needed to elucidate the mechanism whereby neutrophils are affected in the tumor environment. We report the discovery of a series of small molecules with an exo-[3.3.1]azabicyclononane core. Our lead compound 81 is a potent (EC₅₀ = 40 nM) and selective orally bioavailable small molecule antagonist of human CXCR6 receptor signaling that significantly decreases tumor growth in a 30-day mouse xenograft model of HCC.