Chemokine-chemokine Receptor Research Articles

Inhibiting Vasculogenesis After Radiation: A New Paradigm to Improve Local Control by Radiotherapy

Tumors are supported by blood vessels, and it has long been debated whether their response to irradiation is affected by radiation damage to the vasculature. We have shown in preclinical models that, indeed, radiation is damaging to the tumor vasculature and strongly inhibits tumor angiogenesis. However, the vasculature can recover by colonization from circulating cells, primarily proangiogenenic CD11b+ monocytes or macrophages from the bone marrow. This secondary pathway of blood vessel formation, known as vasculogenesis, thus acts to restore the tumor vasculature and allows the tumor to recur following radiation. The stimulus for the influx of these CD11b+ cells into tumors following irradiation is the increased levels of hypoxia-inducible factor-1 in the tumor due to induced tumor hypoxia secondary to blood vessel loss. This increases tumor levels of the chemokine stromal cell-derived factor-1, which has chemokine receptors CXCR4 and CXCR7 on monocytes and endothelial cells thereby capturing these cells in the tumors. The increase in CD11b+ monocytes in tumors following irradiation can be prevented using antibodies or small molecules that inhibit hypoxia-inducible factor-1 or the interaction of stromal cell-derived factor-1 with its receptors. We show that the effect of inhibiting these chemokine-chemokine receptor interactions is a marked increase in the radiation response of transplanted or chemically induced tumors in mice and rats. This strategy of inhibiting vasculogenesis following tumor irradiation is a new paradigm in radiotherapy and suggests that higher levels of local control of tumors in several sites would be achievable with this strategy.

The multiple chemokine-binding bovine herpesvirus 1 glycoprotein G (BHV1gG) inhibits polymorphonuclear cell but not monocyte migration into inflammatory sites.

Chemokines facilitate the recruitment of inflammatory cells into tissues, contributing to target organ injury in a wide range of inflammatory and autoimmune diseases. Targeting either single chemokines or chemokine receptors alters the progression of disease in animal models of rheumatoid arthritis and lupus with varying degrees of efficacy but clinical trials in humans have been less successful. Given the redundancy of chemokine-chemokine receptor interactions, targeting of more than one chemokine may be required to inhibit active inflammatory disease. To test the effects of multiple-chemokine blockade in inflammation, we generated an adenovirus expressing bovine herpesvirus 1 glycoprotein G (BHV1gG), a viral chemokine antagonist that binds to a wide spectrum of murine and human chemokines, fused to the Fc portion of murine IgG2a. Administration of the adenovirus significantly inhibited thioglycollate-induced migration of polymorphonuclear leukocytes into the peritoneal cavity of BALB/c mice and reduced both clinical severity and articular damage in K/BxN serum transfer-induced arthritis. However, treatment with BHV1gG-Ig fusion protein did not prevent monocyte infiltration into the peritoneum in the thioglycollate model and did not prevent renal monocyte infiltration or nephritis in lupus-prone NZB/W mice. These observations suggest that the simultaneous inhibition of multiple chemokines by BHV1gG has the potential to interfere with acute inflammatory responses mediated by polymorphonuclear leukocytes, but is less effective in chronic inflammatory disease mediated by macrophages.

Treatment of inflammatory bowel disease by chemokine receptor-targeted leukapheresis

Leukapheresis removes circulating leukocytes en route to the target organ. Hitherto unspecific matrixes have been used to remove leukocytes in inflammatory bowel disease (IBD). This report describes a novel selective leukapheresis column based on chemokine-chemokine receptor interaction. We found an increased expression of the gut homing chemokine receptor CCR9 on CD14(+) monocytes and on CD3(+) T lymphocytes from IBD patients. Biologically active CCL25 was coupled to a Sepharose matrix and demonstrated to selectively remove CCR9-expressing cells leaving other cell populations largely unaffected. A patient with active ulcerative colitis, was subjected to CCL25-column leukapheresis. Four days after treatment, he experienced clinical improvement and stable disease improvement ensued. The study illustrates that specific cells can be targeted using high affinity interactions, i.e., CCL25-CCR9 interactions to remove pathogenic gut-homing cells. Leukapheresis using the bCCL25 column should be investigated in a clinical phase I trial of patients with inflammatory bowel disease.

The Multiple Chemokine-Binding Bovine Herpesvirus 1 Glycoprotein G (BHV1gG) Inhibits Polymorphonuclear Cell but Not Monocyte Migration into Inflammatory Sites

AbstractChemokines facilitate the recruitment of inflammatory cells into tissues, contributing to target organ injury in a wide range of inflammatory and autoimmune diseases. Targeting either single chemokines or chemokine receptors alters the progression of disease in animal models of rheumatoid arthritis and lupus with varying degrees of efficacy, but clinical trials in humans have been less successful. Given the redundancy of chemokine-chemokine receptor interactions, targeting of more than one chemokine may be required to inhibit active inflammatory disease. To test the effects of multiple chemokine blockade in inflammation, we generated an adenovirus expressing bovine herpesvirus 1 glycoprotein G (BHV1gG), a viral chemokine antagonist that binds to a wide spectrum of murine and human chemokines, fused to the fragment crystallizable (Fc) portion of murine immunoglobulin (IgG)2a. Administration of the adenovirus significantly inhibited thioglycollate-induced migration of polymorphonuclear leukocytes into the peritoneal cavity of BALB/c mice and reduced both clinical severity and articular damage in K/BxN serum transfer-induced arthritis. However, treatment with BHV1gG-Ig fusion protein did not prevent monocyte infiltration into the peritoneum in the thioglycollate model and did not prevent renal monocyte infiltration or nephritis in lupus-prone NZB/W mice. These observations suggest that the simultaneous inhibition of multiple chemokines by BHV1gG has the potential to interfere with acute inflammatory responses mediated by polymorphonuclear leukocytes, but is less effective in chronic inflammatory disease mediated by macrophages.

Antifibrotic effects of CXCR4 antagonist in bleomycin-induced pulmonary fibrosis in mice

Circulating fibrocytes had been reported to migrate into the injured lungs, and contribute to fibrogenesis via chemokine-chemokine receptor systems including CXCL12-CXCR4 axis. Here we hypothesized that blockade of CXCR4 might inhibit the migration of fibrocytes to the injured lungs and the subsequent pulmonary fibrosis. To explore the antifibrotic effects of blockade of CXCR4, we used a specific antagonist for CXCR4, AMD3100, in bleomycin-induced pulmonary fibrosis model in mice. Administration of AMD3100 significantly improved the loss of body weight of mice treated with bleomycin, and inhibited the fibrotic lesion in subpleural areas of the lungs. The quantitative analysis demonstrated that treatment with AMD3100 reduced the collagen content and fibrotic score (Aschcroft score) in the lungs. Although AMD3100 did not affect cell classification in bronchoalveolar lavage fluid on day 7, the percentage of lymphocytes was reduced by AMD3100 on day 14. AMD3100 directly inhibited the migration of human fibrocytes in response to CXCL12 in vitro, and reduced the trafficking of fibrocytes into the lungs treated with bleocmycin in vivo. These results suggest that the blockade of CXCR4 might be useful strategy for therapy of patients with pulmonary fibrosis via inhibiting the migration of circulating fibrocytes.

Tissue-Specific Homing of Immune Cells in Malignant Skin Tumors

Tissue-specific migration of immune cells involved both in physiological and pathological immune responses is a current research subject for medical science. Several homing molecules have been identified orchestrating extravasation of immune cells to certain peripheral non-lymphoid tissues such as gut, lung and skin. Regarding lymphocyte homing to skin, the first-line defense of human body cutaneous lymphocyte associated antigen (CLA) and a group of chemokine-chemokine receptor pairs are considered to be of crucial importance. The aim of the present review is to summarize existing knowledge about skin- and tumor-specific migration of immune cells playing a major pathogenetic role in host immune responses induced by non-lymphoid malignant skin tumors as well as in the development of primary cutaneous T-cell lymphomas (CTCL). Melanoma malignum, squamous and basal cell carcinoma evoke host immune responses and consequently a subset of reactive immune cells is recruited to site of the tumor. Regarding migratory process and exact functional role of these cells a growing number of data is available in literature. On the other hand tissue-specific immune cell homing is regarded as a key process in the pathogenesis of CTCL where malignant T-lymphocytes can be found in circulation and symptomatic skin. Hereby homing mechanism of malignant T-cells in mycosis fungoides and Sézary-syndrome as separate clinical entities of CTCL is discussed. A precise insight into the molecular background of skin- and tumor-specific immune cell migration can contribute to developing efficient vaccine therapies in non-lymphoid malignant skin tumors and beneficial treatment modalities in CTCL.

Abstract 301: Ccl17 and Ccr4 co-localize in mouse lung adenomas in vivo and are associated with epithelial cell migration in vitro

Abstract Lung cancer is the leading cause of cancer deaths among men and women in the United States and improved treatments are urgently needed. Like many cancers, long-term inflammation is now recognized to be a contributing factor in the progression of lung cancer from a benign to a malignant state. Exactly how inflammation contributes to lung cancer is incompletely understood. Chemokines are a group of proteins that function by attracting immune cells to areas of higher chemokine concentrations. Specific chemokine-chemokine receptor pairs control the migratory response of many inflammatory cell types. It is known that chemokines and chemokine receptors can also have direct effects on cancer cells. Using in situ hybridization, we determined that the chemokine Ccl17 and its receptor Ccr4 are coordinately induced in tumor cells in transgenic and benzo(a)pyrene-induced mouse models of lung cancer. In addition, multiple subsets of T lymphocytes within mouse lung tissue express Ccr4. Immunohistochemistry localized CCR4 to cancer cells in 11 of 32 (34%) human non-small cell lung cancer biopsies while staining was not detected in 12 non-neoplastic lung biopsies. We identified human lung cancer cell lines expressing CCR4 using flow cytometry and immunofluorescence. Treatment of human NCI-H292 lung cancer cells with CCL17 peptide induced migration in a collagen-dependent manner in a transwell migration assay. The presence of Ccr4 on both lung cancer cells and lymphocytes suggests that this receptor has multiple roles in regulating lung tumor promotion and progression. Characterization of these roles may provide novel targets to inhibit lung tumor progression. Future studies will directly address the function of Ccr4 during lung tumorigenesis in vivo. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 301. doi:1538-7445.AM2012-301

Hepatic recruitment of macrophages promotes nonalcoholic steatohepatitis through CCR2

Inflammatory cell infiltration in the liver is a hallmark of nonalcoholic steatohepatitis (NASH). The chemokine-chemokine receptor interaction induces inflammatory cell recruitment. CC-chemokine receptor (CCR)2 is expressed on hepatic macrophages and hepatic stellate cells. This study aims to investigate the therapeutic potential of CCR2 to NASH. Twenty-two weeks on a choline-deficient amino acid-defined (CDAA) diet induced steatosis, inflammatory cell infiltration, and liver fibrosis with increased CCR2 and monocyte chemoattractant protein (MCP)-1 expression in the wild-type livers. The infiltrated macrophages expressed CD68, CCR2, and a marker of bone marrow-derived monocytes, Ly6C. CCR2(-/-) mice had less steatosis, inflammatory cell infiltration, and fibrosis, and hepatic macrophages expressing CD68 and Ly6C were decreased. Toll-like receptor (TLR)4(-/-), TLR9(-/-), and MyD88(-/-) mice had reduced hepatic macrophage infiltration with decreased MCP-1 and CCR2 expression because TLR signaling is a potent inducer of MCP-1. To assess the role of Kupffer cells at the onset of NASH, Kupffer cells were depleted by liposomal clodronate. The Kupffer cell depletion ameliorated steatohepatitis with a decrease in the MCP-1 expression and recruitment of Ly6C-expressing macrophages at the onset of NASH. Finally, to test the therapeutic potential of targeting CCR2, a CCR2 inhibitor was administered to mice on a CDAA diet. The pharmaceutical inhibition of CCR2 prevented infiltration of the Ly6C-positive macrophages, resulting in an inhibition of liver inflammation and fibrosis. We concluded that CCR2 and Kupffer cells contribute to the progression of NASH by recruiting bone marrow-derived monocytes.

Natural Products to Improve Quality of Life Targeting for Colon Drug Delivery

The colon is largely being investigated as a site for administration of protein and peptides, which are degraded by digestive enzymes in the upper GIT. Also for local diseases of the colon such as inflammatory bowel disease, colorectal cancer and ameobiasis, drug administration to the site of action can not only reduce the dose to be administered, but also decrease the side effects. Inflammatory Bowel Disease (IBD) such as Ulcerative colitis and Crohn's disease are characterized by chronic intestinal inflammation. Intestinal bacteria initiate the activation of intestinal inflammatory processes, which are mediated by pro-inflammatory cytokines and chemokine. Increased chemokine expression has also been observed in epithelial cells, endothelial cells, and smooth muscle cells. Future trials of specific agents capable of inhibiting chemokine synthesis and secretion or blocking chemokine-chemokine receptor interaction will be important to study in patients with ulcerative colitis and Crohn's disease. Many important bioactive compounds have been discovered from natural sources using bioactivity directed fractionation and isolation (BDFl) Continuing discovery has also been facilitated by the recent development of new bioassay methods. These bioactive compounds are mostly plant secondary metabolites, and many naturally occurring pure compounds have become medicines, dietary supplements, and other useful commercial products. The present review includes various approaches investigated for colon drug delivery and their site specificity. To achieve successful colonic delivery, a drug needs to be protected from absorption and the environment of the upper gastrointestinal tract and then be abruptly released into the proximal colon, which is considered the optimum site for colon targeted delivery of drugs.

Molecular mechanisms of liver metastasis

Colorectal cancer is the second most common cancer, and is the third leading cause of cancer-related death in Japan. The majority of these deaths is attributable to liver metastasis. Recent studies have provided increasing evidence that the chemokine-chemokine receptor system is a potential mechanism of tumor metastasis via multiple complementary actions: (a) by promoting cancer cell migration, invasion, survival and angiogenesis; and (b) by recruiting distal stromal cells (i.e., myeloid bone marrow-derived cells) to indirectly facilitate tumor invasion and metastasis. Here, we discuss recent preclinical and clinical data supporting the view that chemokine pathways are potential therapeutic targets for liver metastasis of colorectal cancer.

Influence of Variations in CCL3L1 and CCR5 on Tuberculosis in a Northwestern Colombian Population

We investigated the association of polymorphisms in CCR5, the major human immunodeficiency virus (HIV)-1 coreceptor, and copy number of its potent ligand CCL3L1 with tuberculosis in 298 individuals from Colombia. The CCR5-HHD haplotype, a known genetic determinant of increased susceptibility to HIV-AIDS, and a high copy number of CCL3L1, a known genetic determinant of enhanced CCL3/CCL3L1 chemokine expression, each associated with presence of tuberculosis. Furthermore, CCR5-HHD was associated with higher CCR5 gene and surface expression. These results substantiate the strong link between the pro-inflammatory effects of CCR5 and its ligands with active tuberculosis and suggest that chemokine-chemokine receptor genetic determinants may influence tuberculosis in addition to HIV/AIDS.

Abstract 2780: Chemokine and chemokine receptor genetic variants as predictors of aggressive prostate cancer among European-American men

Abstract Background: Chemokines are essential mediators in the pathogenesis of inflammatory, autoimmune, vascular and neoplastic diseases. Consequently, investigators have evaluated the relationship between sequence variants in chemokines in relation to autoimmune or inflammatory disorders. However, the impact of inheriting multiple chemokine-chemokine receptor genetic susceptibilities on prostate cancer (PCa) clinical outcomes remains uncharacterized. Objectives & Hypothesis: Consequently, we systematically evaluated the joint modifying effects of 78 single nucleotide polymorphisms (SNPs) detected in 43 chemokine and chemokine receptor-associated genes. We hypothesized that individuals inheriting multiple chemokine-associated alleles linked with increased tumor metastasis, cell survival, or inflammation would have an increased risk of developing PCa or aggressive disease relative to those with the referent genotypes. Methods: To test the hypothesis, we used SNP profile data collected from 2277 European-American male participants of the Cancer Genetic Markers of Susceptibility (CGEMS) project, involving 1176 PCa cases and 1101 controls. Using a case-control and case-only designs, we evaluated the independent and joint modifying effects of 78 SNPs detected in chemokines, chemokine receptors and downstream signaling targets in relation to PCa risk and aggressive disease (tumor stage >2 and tumor grade >2). Main effects and complex interactions were assessed using logistic regression analysis, multi-factor dimensionality reduction (MDR) with covariate adjustment (i.e., age-group and family history of prostate cancer), and hierarchical entropy graphs. To control for multiple comparisons, we used 1000-fold permutation testing. Results: A three-SNP interaction [CCL21 promoter SNP at position -191 (rs11574914), CCR6 promoter SNP at position -397 (rs2023305), CCL17 intron 1+2217 (rs223895)] was the best prediction model in relation to aggressive disease, following restriction of the data analysis to the top 25th percentile of markers based on ReliefF scores (TBA = 56%, CVC = 90%, p = 0.024). Verification of this interaction using other bioinformatic techniques, including symbolic modeling, classification and regression trees (CART) and computational evolutionary systems modeling is currently underway. Conclusions: Our preliminary findings support the role of chemokine-related markers as significant predictors of aggressive prostate cancer among European-American men. Future Directions: Additional studies are needed to determine whether CCL21-CCR6-CCL17 interactions are predictive of disease prognosis within ethnically diverse sub-populations. In addition, the biological implications of the interaction require further investigation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2780. doi:10.1158/1538-7445.AM2011-2780

The Cyclophilin-Binding Agent Sanglifehrin A Is a Dendritic Cell Chemokine and Migration Inhibitor

Sanglifehrin A (SFA) is a cyclophilin-binding immunosuppressant but the immunobiology of action is poorly understood. We and others have reported that SFA inhibits IL-12 production and antigen uptake in dendritic cells (DC) and exhibits lower activity against lymphocytes. Here we show that SFA suppresses DC chemokine production and migration. Gene expression analysis and subsequent protein level confirmation revealed that SFA suppressed CCL5, CCL17, CCL19, CXCL9 and CXCL10 expression in human monocyte-derived DC (moDC). A systems biology analysis, Onto Express, confirmed that SFA interferes with chemokine-chemokine receptor gene expression with the highest impact. Direct comparison with the related agent cyclosporine A (CsA) and dexamethasone indicated that SFA uniquely suppresses moDC chemokine expression. Competitive experiments with a 100-fold molar excess of CsA and with N-Methyl-Val-4-cyclosporin, representing a nonimmunosuppressive derivative of CsA indicated chemokine suppression through a cyclophilin-A independent pathway. Functional assays confirmed reduced migration of CD4+ Tcells and moDCs to supernatant of SFA-exposed moDCs. Vice versa, SFA-exposed moDC exhibited reduced migration against CCL19. Moreover, SFA suppressed expression of the ectoenzyme CD38 that was reported to regulate DC migration and cytokine production. These results identify SFA as a DC chemokine and migration inhibitor and provide novel insight into the immunobiology of SFA.

2246 CCR5/CXCR3 BLOCKADE PROLONGED KIDNEY ALLOGRAFT SURVIVAL WITH OR WITHOUT CYCLOSPORINE VIA THE INHIBITION OF T CELL AND MACROPHAGE RECRUITMENT

You have accessJournal of UrologyTransplantation & Vascular Surgery: Renal Transplantation, Vascular Surgery1 Apr 20112246 CCR5/CXCR3 BLOCKADE PROLONGED KIDNEY ALLOGRAFT SURVIVAL WITH OR WITHOUT CYCLOSPORINE VIA THE INHIBITION OF T CELL AND MACROPHAGE RECRUITMENT Yoichi Kakuta, Masayoshi Okumi, Koichi Tsutahara, Toyofumi Abe, Koji Yazawa, Katsuyoshi Matsunami, Shuji Miyagawa, Shiro Takahara, and Norio Nonomura Yoichi KakutaYoichi Kakuta Suita, Japan More articles by this author , Masayoshi OkumiMasayoshi Okumi Suita, Japan More articles by this author , Koichi TsutaharaKoichi Tsutahara Suita, Japan More articles by this author , Toyofumi AbeToyofumi Abe Suita, Japan More articles by this author , Koji YazawaKoji Yazawa Suita, Japan More articles by this author , Katsuyoshi MatsunamiKatsuyoshi Matsunami Hiroshima, Japan More articles by this author , Shuji MiyagawaShuji Miyagawa Suita, Japan More articles by this author , Shiro TakaharaShiro Takahara Suita, Japan More articles by this author , and Norio NonomuraNorio Nonomura Suita, Japan More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2011.02.2488AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES In renal transplantation, the acute allograft rejection episodes is a major determinant of graft survival. In the cell-mediated rejection, a chemokine-chemokine receptor interactions is a key functional events in T cell recruitment and has attracted deep interest to potential therapeutic targets. We investigated the effects of CCR5/CXCR3 blockade in a rat kidney allotransplantation model. METHODS Left kidneys from male Dark Agouti rats were orthotopically transplanted into bilaterally nephrectomized male Lewis rats. Rats were treated with 10 mg/kg CCR5/CXCR3 blockde from day 0 to 14 (or until rejection) and 2.0 mg/kg cyclosporine (CsA) from day 0 to 5. The treatment groups were as follow: control group (n=7); treated with PBS, chemokine receptor blockade (CRB) group (n=6); treated with CCR5/CXCR3 blockade alone, CsA group (n=6); treated with CsA alone, combined therapy (CT) group (n=5); treated with both CCR5/CXCR3 blockade and CsA. RESULTS While the median survival time (MST) of the control group was 6.5 days, the MST of CRB group was 8.0 days (p<0.05 vs. control group). Compared with CsA group (MST: 9.0 days), the survival time of CT group was extremely prolonged (MST: 42.0 days, p<0.05 vs. CsA group) as shown in Figure. Pathological analysis revealed less signs of acute rejection in CRB, with significantly less numbers of CD4+, CD8+, and CCR5+ T cells compared with the control. There was no sing of acute rejection in CT group in day 5. RT-PCR examinations showed significant decrease in IFN-gamma, and IL-2 in CRB group, with no change of IL-4, or IL-17. Additionaly, when we evaluated the infiltration of macrophage, which act as effector cells in acute rejection, by ED-1 staining, the number of ED-1+ cells were significantly less in CRB group than those in control group. The expression of IL-18 was significantly less in CRB group. CONCLUSIONS Treatmant with CCR5/CXCR3 blockade prolonged allograft survival in a rat kidney transplantation model with or without cyclosporine, by preventing intragraft infiltration of T cell and macrophage. Further exploration of these effects will provide the novel treatment of the kidney allograft transplantation. © 2011 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 185Issue 4SApril 2011Page: e901 Peer Review Report Advertisement Copyright & Permissions© 2011 by American Urological Association Education and Research, Inc.MetricsAuthor Information Yoichi Kakuta Suita, Japan More articles by this author Masayoshi Okumi Suita, Japan More articles by this author Koichi Tsutahara Suita, Japan More articles by this author Toyofumi Abe Suita, Japan More articles by this author Koji Yazawa Suita, Japan More articles by this author Katsuyoshi Matsunami Hiroshima, Japan More articles by this author Shuji Miyagawa Suita, Japan More articles by this author Shiro Takahara Suita, Japan More articles by this author Norio Nonomura Suita, Japan More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

Abstract 5462: Enhancing efficacy of drugs by targeting CXCR2 receptor signaling for the treatment of malignant breast cancer

Abstract Breast cancer is the second most common cause of cancer death in women in the United States after lung cancer and is responsible for more than 40,000 deaths each year. Many challenges exist in the current management of advanced stage breast cancer as there are fewer recognized therapeutic strategies, often due to therapy resistance. Recent reports suggest that malignant cells that survive initial chemo- and radiation therapy often express inflammatory cytokines, which provides survival benefit making tumors resistant to further chemotherapy. The specific objective of the study is to develop strategy to manipulate chemokine-chemokine receptor network for effective therapy with limited toxicity against drug-resistant breast cancer. We investigated whether modulation of CXCR2-dependent signaling in malignant mammary tumor enhances therapeutic efficacy of chemotherapy drugs. We used mammary tumor cells expressing different levels of CXCR2 (Cl66-wt and Cl66-shCXCR2) and examined their response to Paclitaxel and Doxorubicin. We observed significant enhancement of paclitaxel and doxorubicin-mediated toxicity at concentrations 5 and 10nM in Cl66-shCXCR2 cells as compared to Cl66-wt cells. Moreover, the expression of CXCL1, a CXCR2 ligand, was increased in paclitaxel and doxorubicin treated mammary tumor cells. Paclitaxel and doxorubicin induced CXCL-1 levels was significantly lower in Cl66-shCXCR2 cells as compared to Cl66-wt cells. In vivo studies using wild type and knockdown cells showed no significant difference in the tumor growth. However, we observed a difference in the tumor growth between wild type and knockdown tumors in mice treated with 10mg/kg concentration of paclitaxel for 3 weeks after starting the treatment. When these studies were further extended there wasn't any difference between the two groups. Furthermore, we extended these studies using human breast cancer cell lines (MDA-MB231, MDA-MET, and SKBR3). MDA-MB231 and MDA-MET are triple negative and considered to be more metastatic and drug resistant than SKBR3 which is HER2-over expressing cell line. We observed differential expression of CXCR2 and its ligands in these cells. Our observation also showed significant difference in the paclitaxel and doxorubicin-mediated inhibitory response in MDA-MB231 and MDA-MET cells as compared to SKBR3 cells. Together these studies demonstrate that manipulating CXCR2-dependent activity in malignant breast cancer cells significantly enhanced therapeutic efficacy of paclitaxel and doxorubicin. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5462.

CCR1 Is Required for Treg-Mediated Protection From Lethal Acute Graft-Versus-Host Disease.

Abstract 1337Poster Board I-359 INTRODUCTIONGraft-versus-host disease (GVHD) is a major complication following allogeneic bone marrow transplantation (BMT). Despite advances in understanding the etiology of GVHD it remains a formidable obstacle to the widespread application of BMT. A number of studies have demonstrated that T regulatory (Treg) cells represent a potential therapy for GVHD as Tregs have been shown to inhibit GVHD while preserving the beneficial graft-versus-leukemia (GVL) effect. Numerous groups, including our own, have demonstrated the importance of T cell migration in the pathology of GVHD. Following conditioning, donor T cells migrate to secondary lymphoid tissues. Once activated in the lymphatics, T cells migrate to GVHD target organs including; the skin, liver, lung and the gastrointestinal (GI) tract in response to the local production of chemokines. Disruption of chemokine-chemokine receptor interactions has been demonstrated to affect the pathology of GVHD. Previously, we have shown that Tregs lacking the chemokine receptor CCR5, which binds CCL3, CCL4, and CCL5, do not protect animals from lethal GVHD as well as WT Tregs, due to their impaired migration to the liver and lung. Thus, a greater understanding of the function of chemokine receptors on Tregs is important in deciphering how Tregs function and whether targeting these cells to lymphoid tissue or GVHD target organs would be preferable for treating patients in clinical trials. METHODSWe utilized a parent into F1 haploidentical model to assess the role of CCR1 in Treg-mediated protection from GVHD. Here we demonstrate Tregs lacking CCR1, another receptor for CCL3 and CCL5, were unable to protect animals against lethal acute GVHD. While 67% of B6D2 recipients given 1×106 WT Tregs supplemented with 5×106 WT T cells and 3×106 B6 T cell-depleted BM cells survived, only 15% of the recipients given CCR1−/− Tregs survived (p < 0.03; Fisher's exact test). B6D2 recipient mice given WT Tregs had significantly reduced clinical scores for GVHD compared to B6D2 recipients of CCR1−/− Tregs (p <0.05) with elevated GVHD scores starting on day 28 post-transplant. Histopathology revealed significantly worse pathology in the liver (p < 0.03) and colon (p < 0.05) of CCR1−/− Treg recipients vs. WT Treg recipients. In vitro analysis demonstrated that CCR1−/− Tregs were capable of suppressing T cell responses to allo-antigen equally as well as WT Tregs, and CCR1−/− Tregs attained a normal activation phenotype. Interestingly, preliminary experiments suggested that CCR1−/− Tregs migrated to and/or expanded in GVHD target organs to a similar extent as WT Tregs. CONCLUSIONSTreg expression of CCR1 is required for the inhibition of GVHD. Tregs lacking CCR1 led to significantly more tissue destruction in the liver and colon, two predominant sites of GVHD pathology. Of interest, the migration of CCR1−/− Tregs to GVHD target organs and secondary lymphoid tissues did not appear to be compromised suggesting that CCR1 may be required for the function of Tregsin vivo. DisclosuresNo relevant conflicts of interest to declare.

Type I IFN-induced chemokine production and the regulation of immune effector cell trafficking from the bone marrow during acute MCMV infection (96.4)

Abstract Monocytes/macrophages are key innate effector cells in the response against murine cytomegalovirus (MCMV) infection. Activation of the chemokine receptor CCR2 on the monocyte/macrophage surface directs cell trafficking. In CCR2-deficient C57BL/6 mice, fewer monocytes/macrophages enter MCMV-infected livers. Consequently, downstream antiviral responses, such as NK cell recruitment, are impaired. Experiments were performed to better understand the origin of this defect. We hypothesized that CCR2 activation on bone marrow monocytes is required for release into the circulation, thus regulating their availability to the liver. We have demonstrated that monocytes accumulate in the bone marrow and are diminished in the circulation of CCR2-deficient mice. NK cells also accumulate in the bone marrow of CCR2-deficient mice, suggesting that impaired monocyte responses inhibit NK cell release. Monocyte mobilization from the bone marrow is likely coordinated by expression of the CCR2 ligands CCL2, CCL7 and CCL12. These chemokines are produced by F4/80+ bone marrow cells and detected in the serum in a type I IFN-dependent manner. These results further support the central role for type I IFN and chemokine-chemokine receptor interactions in establishing effective antiviral immune responses during MCMV infection. Supported by NIH R01CA-102708 and the Nicole Rosenthal Hartnett '91 Graduate Fellowship.

Rejection and acceptance of corneal allografts

Corneal transplantation is successful in the short-term, but the long-term prognosis has not improved over the past 20 years. Here, we review recent findings that may contribute to improved corneal allograft survival. A better understanding of the molecular pathways affecting corneal graft survival has led to more targeted approaches to immune modulation. Costimulatory molecule blockade, inhibition of chemokine-chemokine receptor interactions, modulation of apoptotic pathways, and reduction of corneal neovascularization and lymphangiogenesis have been shown to prolong corneal graft survival in animal models. Conventional immunosuppressive drugs have been tested in new combinations and formulations with some success. Two randomized prospective clinical trials in clinical penetrating corneal transplantation have been reported, but there remains little evidence on the long-term outcomes of the newer lamellar corneal graft procedures. New approaches to reducing the impact of rejection on corneal graft survival have focussed on topical rather than systemic therapies, and on component corneal transplantation. The most successful experimental strategies have been those in which more than one pathway has been targeted; it now seems likely that to improve clinical allograft survival, simultaneous modulation of multiple axes of the rejection process will be necessary.

Chemokine–Chemokine Receptors in Cancer Immunotherapy

A surge in interest in the chemokine-chemokine receptor network is probably related to the expanding roles that chemokines have now been identified to play in human biology, particularly immunity. Specific tissue microenvironments express distinct chemokines and both hematopoietic and nonhematopoietic cells have receptor expression profiles that permit the coordinated trafficking and organization of cells within these specific tissues. Since the chemokine network plays critical roles in both the function of the immune system and the progression of cancer, it is an attractive target for therapeutic manipulation. This review will focus on chemokine and chemokine receptor network-related therapeutic interventions that utilize host-tumor interactions particularly involving the immune system.

Microenvironmental influences in chronic lymphocytic leukaemia: the role of antigen stimulation

Several studies suggest that immune-mediated pathways are important in the pathogenesis of chronic lymphocytic leukaemia (CLL). The in vivo accumulation of leukaemic lymphocytes is facilitated by interactions of CLL cells with other cells and soluble factors that probably occur more often within the microenvironment through classical receptor-ligand interactions. These include CD40L-CD40 and chemokine-chemokine receptor interactions as well as B cell receptor (BCR) engagement by (auto)antigens. Indeed, the categorizations of CLL patients based on immunoglobulin heavy variable (IGHV) gene mutations and structure of the clone's BCR suggest that CLL patient outcome could be a reflection of ongoing BCR signalling in the context of other co-signals.