The good and the bad of chemokines/chemokine receptors in melanoma.

Chemokine ligand/receptor interactions affect melanoma cell growth, stimulate or inhibit angiogenesis, recruit leukocytes, promote metastasis, and alter the gene expression profile of the melanoma associated fibroblasts. Chemokine/chemokine receptor interactions can protect against tumor development/growth or can stimulate melanoma tumor progression, tumor growth and metastasis. Metastatic melanoma cells express chemokine receptors that play a major role in the specifying the organ site for metastasis, based upon receptor detection of the chemokine gradient elaborated by a specific organ/tissue.

The Duffy antigen receptor for chemokines transports chemokines and supports their promigratory activity.

The Duffy antigen receptor for chemokines (DARC) belongs to a family of 'silent' heptahelical chemokine receptors that do not couple to G proteins and fail to transmit measurable intracellular signals. DARC binds most inflammatory chemokines and is prominently expressed on venular endothelial cells, where its function has remained contentious. Here we show that DARC, like other silent receptors, internalized chemokines but did not effectively scavenge them.

Sunitinib reverses type-1 immune suppression and decreases T-regulatory cells in renal cell carcinoma patients.

Purpose: Immune dysfunction is well documented in renal cell carcinoma (RCC) patients and likely contributes to tumor evasion. This dysfunction includes a shift from a type-1 to a type-2 T-cell cytokine response and enhanced T-regulatory (Treg) cell expression. Given the antitumor activity of select tyrosine kinase inhibitors such as sunitinib in metastatic RCC (mRCC) patients, it is relevant to assess their effect on the immune system.

CXC chemokines and their receptors: a case for a significant biological role in cutaneous wound healing.

Wound healing requires a complex series of reactions and interactions among cells and their mediators, resulting in an overlapping series of events including coagulation, inflammation, epithelialization, formation of granulation tissue, matrix and scar formation. Cytokines and chemokines promote inflammation, angiogenesis, facilitate the passage of leukocytes from circulation into the tissue, and contribute to the regulation of epithelialization. They integrate inflammatory events and reparative processes that are important for modulating wound healing.

The carboxyl-terminal PDZ ligand motif of chemokine receptor CXCR2 modulates post-endocytic sorting and cellular chemotaxis.

Adaptor protein interaction with specific peptide motifs found within the intracellular, carboxyl terminus of chemokine receptor CXCR2 has been shown to modulate intracellular trafficking and receptor function. Efficient ligand-induced internalization of this receptor is dependent on the binding of adaptor protein 2 to the specific LLKIL motif found within the carboxyl terminus (1). In this study we show that the carboxyl-terminal type 1 PDZ ligand motif (-STTL) of CXCR2 plays an essential role in both proper intracellular receptor trafficking and efficient cellular chemotaxis.

Parallel phosphatidylinositol 3-kinase (PI3K)-dependent and Src-dependent pathways lead to CXCL8-mediated Rac2 activation and chemotaxis.

The requirement for phosphatidylinositol 3-kinase (PI3K) in the establishment of cell polarity and motility in a number of cell types has recently come into question. In this study, we demonstrate that inhibition of PI3K by wortmannin in neutrophil-like differentiated HL60 cells expressing CXCR2 resulted in reduced cell motility but normal chemotaxis in response to a gradient of CXCL8. However, wortmannin inhibition of PI3K did impair the ability of cells to re-orient their polarity and respond quickly to a change in the direction of the CXCL8 gradient.

Short tail with skin lesion phenotype occurs in transgenic mice with keratin-14 promoter-directed expression of mutant CXCR2.

CXCR2 plays an important role during cutaneous wound healing. Transgenic mice were generated using the keratin-14 promoter/enhancer to direct expression of wild-type human CXCR2 (K14hCXCR2 WT) or mutant CXCR2, in which the carboxyl-terminal domain (CTD) was truncated at Ser 331 and the dileucine AP-2 binding motif was mutated to alanine (K14hCXCR2 331T/LL/AA/IL/AA). Our results indicate that K14hCXCR2WT transgenic mice exhibited a normal phenotype, while K14hCXCR2 331T/LL/AA/IL/AA transgenic mice were born with tails of normal length, but three to eight days after birth their tails degenerated, leaving only a short tail stub.

Microfluidic switching system for analyzing chemotaxis responses of wortmannin-inhibited HL-60 cells.

The chemotaxis of phosphoinositide kinase-3 (PI3K)-inhibited differentiated HL-60 cells stably expressing CXCR2 was studied in a microfluidic switching gradient device that can generate stable and well-defined forward and reverse gradients. Wortmannin, a widely used PI3K inhibitor, was added during cell preparation and the experiment process. The studies quantify the chemotaxis gradient and the effects of a change in the direction of a CXCL-8 gradient on cell migration.

Abrogation of TGF beta signaling in mammary carcinomas recruits Gr-1+CD11b+ myeloid cells that promote metastasis.

Aberrant TGFbeta signaling is common in human cancers and contributes to tumor metastasis. Here, we demonstrate that Gr-1+CD11b+ myeloid cells are recruited into mammary carcinomas with type II TGF beta receptor gene (Tgfbr2) deletion and directly promote tumor metastasis. Gr-1+CD11b+ cells infiltrate into the invasive front of tumor tissues and facilitate tumor cell invasion and metastasis through a process involving metalloproteinase activity.

Opposing roles of murine duffy antigen receptor for chemokine and murine CXC chemokine receptor-2 receptors in murine melanoma tumor growth.

The Duffy antigen receptor for chemokines (DARC) has been classified as a "silent" receptor, as it can bind CXC and CC chemokines to undergo ligand-induced receptor internalization, but is not coupled to trimeric G proteins required for the classic G protein-coupled receptor-mediated signaling. CXC chemokine receptor-2 (CXCR2) has been shown to play a major role in tumor angiogenesis. To test the hypothesis that these two chemokine receptors might play opposing roles in the growth of melanoma tumors, we developed a transgenic mouse model, where the preproendothelin promoter/enhancer (PPEP) is used to drive expression of either murine DARC (mDARC) or murine CXCR2 (mCXCR2) in endothelial cells.

Expression of activated Akt in benign nevi, Spitz nevi and melanomas.

Background: Activated Akt expression (p-Akt) is reportedly increased in many melanomas as compared with benign nevi. The purpose of this study was to evaluate and compare p-Akt immunohistological staining in benign nevi, Spitz nevi and primary melanomas.

Methods: Immunostaining for phosphorylated Akt was performed in 41 melanocytic lesions previously classified as benign intradermal nevus (14 lesions), Spitz nevus (9 lesions) or melanoma (18 lesions).

Role of chemokines in tumor growth.

Chemokines play a paramount role in the tumor progression. Chronic inflammation promotes tumor formation. Both tumor cells and stromal cells elaborate chemokines and cytokines.

Hypoxia enhances CXCR4 expression in human microvascular endothelial cells and human melanoma cells.

The influence of environmental factors (cytokines, matrix components, serum factors and O(2) level) on expression of receptors for angiogenic versus angiostatic CXC chemokines in human microvascular endothelial cells has not been extensively investigated. Our semi-quantitative RT-PCR analysis demonstrated that TNF-alpha and IFN-gamma repressed CXCR4 mRNA levels in immortalized human microvascular endothelial HMEC-1 cells after 4 h, whereas only TNF-alpha displayed inhibitory activity in primary human microvascular endothelial cells (HMVEC). CXCR4 mRNA expression was not affected by VEGF, GM-CSF, IL-1beta or various basal membrane matrix components, but was significantly up-regulated after serum starvation and/or hypoxic treatment of the microvascular endothelial cells.

CCAAT displacement protein regulates nuclear factor-kappa beta-mediated chemokine transcription in melanoma cells.

Members of the nuclear factor-kappa beta (NF-kappaB) family maintain cellular homeostasis by enhancing the transcription of genes involved in inflammation, immune response, cell proliferation, and apoptosis. Melanoma tumor cells often express inflammatory mediators through enhanced activation of NF-kappaB. The NF-kappaB activation appears to result from the enhancer formation including NF-kappaB and lysine acetyl transferases such as p300, CREB (cyclic AMP-responsive element binding protein)-binding protein (CBP), and/or p300/CBP associating factor (PCAF).

Functional characterization of chimpanzee cytomegalovirus chemokine, vCXCL-1(CCMV).

Human cytomegaloviruses (HCMVs) are important pathogens in immunocompromised patients and newborns. The viral chemokine, vCXCL-1, of the Toledo (Tol) strain of HCMV has been implicated in HCMV virulence. Chimpanzee CMV (CCMV) has several genes with similarity to the vCXCL-1(Tol) gene, UL146.

Systemic targeting inhibitor of kappaB kinase inhibits melanoma tumor growth.

Constitutive activation of nuclear factor-kappaB (NF-kappaB) has been directly implicated in tumorigenesis of various cancer types, including melanoma. Inhibitor of kappaB kinase (IKK) functions as a major mediator of NF-kappaB activation. Thus, development of an IKK-specific inhibitor has been a high priority, although it remains unclear whether systemic inhibition of IKK will provide therapeutic benefit.

RhoB plays an essential role in CXCR2 sorting decisions.

The CXCR2 chemokine receptor is a G-protein-coupled receptor that undergoes clathrin-mediated endocytosis upon ligand binding. The trafficking of CXCR2 is crucial for cells to maintain a proper chemotactic response. The mechanisms that regulate the recycling/degradation sorting decision are unknown.

Antioxidants tiron and N-acetyl-L-cysteine differentially mediate apoptosis in melanoma cells via a reactive oxygen species-independent NF-kappaB pathway.

Tiron and N-acetyl-L-cysteine (NAC) have been recognized as potential antioxidants capable of inhibiting apoptosis induced by reactive oxygen species (ROS). Although the ROS-scavenging function of tiron and NAC is clear, the mechanism for their regulation of apoptosis is still elusive. Here we demonstrate that tiron increases nuclear factor-kappaB (NF-kappaB)/DNA binding and as a result enhances NF-kappaB transcriptional activity.

The Duffy antigen modifies systemic and local tissue chemokine responses following lipopolysaccharide stimulation.

The Duffy blood group Ag (dfy) binds selective CXC and CC chemokines at high affinity and is expressed on erythrocytes and endothelial cells. However, it does not transmit a signal via G proteins, as occurs with other seven-transmembrane receptors. We hypothesized that dfy functions as a chemokine reservoir and regulates inflammation by altering soluble chemokine concentrations in the blood and tissue compartments.

The IL sequence in the LLKIL motif in CXCR2 is required for full ligand-induced activation of Erk, Akt, and chemotaxis in HL60 cells.

The chemotaxis of differentiated HL60 cells stably expressing CXCR2 was examined in a microfluidic gradient device where the steepness of the CXCL8 chemokine gradient was varied from 2 pg/ml/mum (0-1 ng/ml over a width of 500 microm) to 50 pg/ml/microm (0-25 ng/ml over 500 microm). The differentiated HL60 cells stably expressing CXCR2 exhibited little chemotaxis in response to a 0-1 ng/ml gradient, but displayed an increasing chemotactic response as the gradient steepness increased from 0 to 5, 0 to 10, and 0 to 25 ng/ml, demonstrating that steepness of gradient is a major determinant of the relative ability of cells to persistently migrate up a chemotactic gradient. When HL60 cells expressed CXCR2 mutated in the C terminus LLKIL motif (IL to AA), ligand-induced internalization of receptors was reduced 50%, whereas cell migration along the gradient of CXCL8 was completely lost.

GM2 expression in renal cell carcinoma: potential role in tumor-induced T-cell dysfunction.

Multiple mechanisms have been proposed to account for immune escape by tumors. Although gangliosides have long been known to suppress T-cell immunity, few studies have examined the effect of human tumor-derived gangliosides on immune responses. Here, we show that gangliosides isolated from renal cell carcinoma (RCC) cell lines and clear cell tumor tissue can induce apoptosis in peripheral blood T cells.

Differential regulation of CXC ligand 1 transcription in melanoma cell lines by poly(ADP-ribose) polymerase-1.

The continuous production of the CXC ligand 1 (CXCL1) chemokine by melanoma cells is a major effector of tumor growth. We have previously shown that the constitutive expression of this chemokine is dependent upon transcription factors nuclear factor-kappa B (NF-kappaB), stimulating protein-1 (SP1), high-mobility group-I/Y (HMGI/Y), CAAT displacement protein (CDP) and poly(ADP-ribose) polymerase-1 (PARP-1). In this study, we demonstrate for the first time the mechanism of transcriptional regulation of CXCL1 through PARP-1 in melanoma cells.