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Objective: NAFLD is a complex disease marked by cellular abnormalities leading to NASH. NAFLD patients manifest low hepatic levels of CEACAM1, a promoter of insulin clearance. Consistently, Cc1 -/- null mice displayed spontaneous hyperinsulinemia/insulin resistance and steatohepatitis. Liver-specific reconstitution of Ceacam1 reversed these metabolic anomalies in 8-month-old Cc1 -/-xliver+ mice fed a regular chow diet. The current study examined whether it would also reverse progressive hepatic fibrosis in mice fed a high-fat (HF) diet., Methods: 3-Month-old mice were fed a high-fat diet for 3-5 months, and metabolic and histopathological analysis were conducted to evaluate their NASH phenotype., Results: Reconstituting CEACAM1 to Cc1 -/- livers curbed diet-induced liver dysfunction and NASH, including macrovesicular steatosis, lobular inflammation, apoptosis, oxidative stress, and chicken-wire bridging fibrosis. Persistence of hepatic fibrosis in HF-fed Cc1 -/- treated with nicotinic acid demonstrated a limited role for lipolysis and adipokine release in hepatic fibrosis caused by Ceacam1 deletion., Conclusions: Restored metabolic and histopathological phenotype of HF-fed Cc1 -/-xliver+xliver+ assigned a critical role for hepatic CEACAM1 in preventing NAFLD/NASH including progressive hepatic fibrosis., Competing Interests: Declaration of competing interest None declared., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Carcinoembryonic antigen (CEA) is a tumor-specific antigen overexpressed in multiple cancers. CEA is expressed as a membrane protein, a part of which is cleaved from the cell membrane and secreted into blood. The soluble form of CEA (sCEA) has been shown to accelerate the clearance of anti-CEA antibody, which limits the antibody distribution in the tumor. To overcome this issue, we developed an anti-CEA monoclonal antibody, 15-1-32, which shows a strong affinity for membrane-bound CEA (mCEA) and relatively weak affinity for sCEA. In this study, we compared the effect of sCEA on the pharmacokinetics of 15-1-32 in mice with that of another anti-CEA monoclonal antibody, labetuzumab, showing less selectivity to mCEA than 15-1-32. As expected, the effect of sCEA on the serum concentration of 15-1-32 was much smaller than that of labetuzumab. The decrease in the area under the curve (AUC) of serum concentration was 22.5% for 15-1-32 when it was coadministered with sCEA, while that of labetuzumab was 79.9%. We also compared the pharmacokinetics of these two antibodies in CEA-positive tumor-bearing mice. The AUCs of 15-1-32 and labetuzumab were decreased in tumor-bearing mice compared with non-tumor-bearing mice to a similar extent (approximately 40% decrease). These results suggested that mCEA also contributes to the clearance of anti-CEA antibodies in CEA-positive tumor-bearing mice. Although the increased selectivity to mCEA minimized the effect of sCEA on the pharmacokinetics of 15-1-32, it may be insufficient to improve the pharmacokinetics in CEA-positive cancer patients. SIGNIFICANCE STATEMENT: Because previous studies reported the rapid clearance of anti-CEA antibodies mediated by soluble CEA, we obtained a monoclonal antibody, 15-1-32, selective to membrane-bound CEA and evaluated the effects of CEA on its pharmacokinetics. Although the effect of soluble CEA on the serum concentration of 15-1-32 was very small, the clearance of 15-1-32 in CEA-positive tumor-bearing mice was still rapid, suggesting membrane-bound CEA also contributes to the clearance of anti-CEA antibodies. These results indicated that increasing selectivity to membrane-bound CEA is not enough to improve the pharmacokinetics of anti-CEA antibody., (Copyright © 2019 by The American Society for Pharmacology and Experimental Therapeutics.)

BACKGROUND Glioblastoma multiforme (GBM) evades immune surveillance by inducing immunosuppression via receptor-ligand interactions between immune checkpoint molecules. T cell immunoglobulin and mucin domain 3 (Tim-3) is a key checkpoint receptor responsible for exhaustion and dysfunction of T cells and plays a critical role in immunosuppression. Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) has been recently identified as a heterophilic ligand for Tim-3. MATERIAL AND METHODS We established an intracranial GBM model using C57BL/6 mice and GL261 cells, and treated the mice with single or combined monoclonal antibodies (mAbs) against Tim-3/CEACAM1. The CD4+, CD8+, and regulatory T cells in brain-infiltrating lymphocytes were analyzed using flow cytometry, and the effector function of T cells was assessed using ELISA. We performed a rechallenge by subcutaneous injection of GL261 cells in the "cured" (>90 days post-orthotopic tumor implantation) and naïve mice. RESULTS The mean survival time in the control, anti-Tim-3, anti-CEACAM1, and combined treatment groups was 29.8, 43.4, 42.3, and 86.0 days, respectively, with 80% of the mice in the combined group becoming long-term survivors showing immune memory against glioma cells. Infiltrating CD4+ and CD8+ T cells increased and immunosuppressive Tregs decreased with the combined therapy, which resulted in a markedly elevated ratio of CD4+ and CD8+ cells to Tregs. Additionally, plasma IFN-γ and TGF-β levels were upregulated and downregulated, respectively. CONCLUSIONS Our data indicate that combined blockade of Tim-3 and CEACAM1 generates robust therapeutic efficacy in mice with intracranial tumors, and provides a promising option for GBM immunotherapy.

Objectives: CT texture analysis has shown promise to differentiate colorectal cancer patients with/without hepatic metastases., Aim: To investigate whether whole-liver CT texture analysis can also predict the development of colorectal liver metastases., Material and Methods: Retrospective multicentre study (n=165). Three subgroups were assessed: patients [A] without metastases (n=57), [B] with synchronous metastases (n=54) and [C] who developed metastases within ≤24 months (n=54). Whole-liver texture analysis was performed on primary staging CT. Mean grey-level intensity, entropy and uniformity were derived with different filters (σ0.5-2.5). Univariable logistic regression (group A vs. B) identified potentially predictive parameters, which were tested in multivariable analyses to predict development of metastases (group A vs. C), including subgroup analyses for early (≤6 months), intermediate (7-12 months) and late (13-24 months) metastases., Results: Univariable analysis identified uniformity (σ0.5), sex, tumour site, nodal stage and carcinoembryonic antigen as potential predictors. Uniformity remained a significant predictor in multivariable analysis to predict early metastases (OR 0.56). None of the parameters could predict intermediate/late metastases., Conclusions: Whole-liver CT-texture analysis has potential to predict patients at risk of developing early liver metastases ≤6 months, but is not robust enough to identify patients at risk of developing metastases at later stage., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Objective: To explore the predictive value of combination detection of Pgp1 expression in cancer tissue and serum CEA level for the prognosis of colorectal cancer (CRC) patients., Methods: Clinicopathological data, complete 5-year follow-up data and CRC tissue samples of 153 CRC patients with stage I( to II( tumor undergoing radical operation in our department from January 2004 to August 2006 were retrospectively collected. Immunohistochemical staining was used to detect the expression level of Pgp1. The combined evaluation of staining intensity and positive cell percentage was performed to determine the expression level of Pgp1. Pgp1 staining (-) and (+) was defined as low expression; and staining (++) and (+++) as high expression. Electrochemiluminescence immunoassay was used to detect the level of serum CEA. CEA > 5 μg/L was defined as positive. χ 2 and Fisher's exact test were performed to analyze the association of Pgp1 expression with CEA level and clinicopathological variables. Moreover, Kaplan-Meier method was used to analyze the survival. Univariate and multivariate Cox proportional hazard regression models were used to evaluate the roles of Pgp1 expression combined with serum CEA level in prognosis prediction., Results: Of 153 patients, 105 were males and 48 females with mean age of 59 (27 to 90) years; 41 cases were rectal cancer, and 112 cases colon cancer; 23 patients were TNM stage I( tumor, and 130 patients stage II( tumor; median follow-up time was 64 months; 30 cases were dead. Positive rate of Pgp1 expression in colorectal cancer tissues was 66.0%(101/153). The expression of Pgp1 was associated with gender, tumor location, and survival during the follow-up (all P<0.05). The preoperative positive rate of serum CEA was 28.1% (43/153). The preoperative serum CEA level was associated with tumor recurrence and survival (all P<0.05). Kaplan-Meier analysis showed the overall 5-year survival rate was 81.7%. The 5-year survival rate of patients with high expression of Pgp1 was 88.1%, which was significantly higher than 69.2% of those with low expression of Pgp1(P=0.003). The 5-year survival rate of patients with preoperative positive serum CEA was 72.1%, which was significantly lower than 86.1% of those with preoperative negative serum CEA(P=0.023). Furthermore, the 5-year survival rate of patients with negative Pgp1 plus positive CEA was 66.7%, which was significantly lower than 91.0% of those with positive Pgp1 plus negative CEA(P=0.002). Univariate analysis showed that gender, Pgp1 expression level, preoperative serum CEA level, and Pgp1 combined with CEA were significantly associated with the prognosis of patients(all P<0.05). Multivariate analysis showed that Pgp1 expression was an independent prognostic factor of CRC [HR(95%CI:1.261 to 64.224), P=0.028]., Conclusions: Low expression of Pgp1 in cancer tissue indicates poor prognosis in patients with stage I( and II( tumor. Combination detection of Pgp1 expression and serum CEA can be applied to predict the prognosis of patients with stage I( and II( colorectal cancer.

Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) regulates insulin sensitivity by promoting hepatic insulin clearance and mediating suppression of fatty acid synthase activity. Feeding C57BL/6J male mice with a high-fat (HF) diet for 3-4 weeks triggered a >60% decrease in hepatic CEACAM1 levels to subsequently impair insulin clearance and cause systemic insulin resistance and hepatic steatosis. This study aimed at investigating whether lipolysis drives reduction in hepatic CEACAM1 and whether this constitutes a key mechanism leading to diet-induced metabolic abnormalities. Blocking lipolysis with a daily intraperitoneal injection of nicotinic acid in the last two days of a 30-day HF feeding regimen demonstrated that white adipose tissue (WAT)-derived fatty acids repressed hepatic CEACAM1-dependent regulation of insulin and lipid metabolism in 3-month-old male C57BL/6J mice. Adenoviral-mediated CEACAM1 redelivery countered the adverse metabolic effect of the HF diet on insulin resistance, hepatic steatosis, visceral obesity, and energy expenditure. It also reversed the effect of HF diet on inflammation and fibrosis in WAT and liver. This assigns a causative role for lipolysis-driven decrease in hepatic CEACAM1 level and its regulation of insulin and lipid metabolism in sustaining systemic insulin resistance, hepatic steatosis, and other abnormalities associated with excessive energy supply., (Copyright © 2016 by the American Society for Biochemistry and Molecular Biology, Inc.)

Objective: To investigate the effects of long chain non-coding RNA urothelial carcinoembryonic antigen 1(UCA1) on invasion, migration and proliferation abilities in oral squamous cell carcinoma cell lines SCC15 and CAL27., Methods: Small interfering RNA of UCA1(UCA1-siRNA) was transfected into SCC15 and CAL27 cell lines by Lipofectamine(TM) 3000 to silence UCA1 , and transfected negtive control si-RNA served as a control. The effect of UCA1-siRNA was detected by quantitative real time-polymerase chain reaction(qRT-PCR) to confirm the successful inhibition of UCA1 by siRNA. The matrix metalloproteinase 9(MMP-9) protein level was detected by Western blotting analysis. The effect of siRNA on cell proliferation and invasion was assessed by transwell migration assay and wound healing assay. Cell counting kit-8(CCK-8) assay was carried out to estimate the proliferation of two cell lines with different expression levels of UCA1., Results: Expressions of UCA1 of SCC15 and CAL27 were successfully suppressed after transfected with siRNA which verified by qRT-PCR, and the efficiency of downregulation of SCC15 and CAL27 was 86.45%(P<0.001)and 78.24%(P<0.001), respectively. The migration, invasion and proliferation of SCC15 and CAL27 cell lines after transfected with siRNA were obviously restrained. The number of migration and invasion of CAL27 cells were 719.20±92.36 versus 208.00±25.58 (P=0.000 7) and 363.40 ± 45.96 versus 164.80 ± 24.68(P= 0.005 2), respectively, the number of migration and invasion of SCC15 cells were 437.20±54.75 vs 145.80±23.31(P=0.001 1) and 249.80±38.41 vs 63.80±11.11 (P=0.001 6), respectively (UCA1-si compare to negtive control), the relative proliferation rates of SCC15 and CAL27 were SCC15: R24 h=0.870, R48 h=0.863, R72 h=0.64, R96 h=0.732; CAL27: R24 h=0.913, R48 h=0.829, R72 h=0.756, R96 h= 0.705(P<0.05), and MMP-9 expression level was decreased by UCA1-siRNA compared with negative control., Conclusions: UCA1 could enhance the ability of invasion and migration of SCC15 and CAL27 cell lines via regulating MMP-9 protein expression, which suggests that UCA1 might play a pivotal role in oral squamous cell carcinoma invasion and progression.

Glioblastoma-initiating cells (GIC) are a tumorigenic cell subpopulation resistant to radiotherapy and chemotherapy, and are a likely source of recurrence. However, the basis through which GICs are maintained has yet to be elucidated in detail. We herein demonstrated that the carcinoembryonic antigen-related cell adhesion molecule Ceacam1L acts as a crucial factor in GIC maintenance and tumorigenesis by activating c-Src/STAT3 signaling. Furthermore, we showed that monomers of the cytoplasmic domain of Ceacam1L bound to c-Src and STAT3 and induced their phosphorylation, whereas oligomerization of this domain ablated this function. Our results suggest that Ceacam1L-dependent adhesion between GIC and surrounding cells play an essential role in GIC maintenance and proliferation, as mediated by signals transmitted by monomeric forms of the Ceacam1L cytoplasmic domain., (©2015 American Association for Cancer Research.)

B cells are essential for antiviral immune defence because they produce neutralizing antibodies, present antigen and maintain the lymphoid architecture. Here we show that intrinsic signalling of CEACAM1 is essential for generating efficient B-cell responses. Although CEACAM1 exerts limited influence on the proliferation of B cells, expression of CEACAM1 induces survival of proliferating B cells via the BTK/Syk/NF-κB-axis. The absence of this signalling cascade in naive Ceacam1(-/-) mice limits the survival of B cells. During systemic infection with cytopathic vesicular stomatitis virus, Ceacam1(-/-) mice can barely induce neutralizing antibody responses and die early after infection. We find, therefore, that CEACAM1 is a crucial regulator of B-cell survival, influencing B-cell numbers and protective antiviral antibody responses.

Oncomarkers play important roles in the detection and management of human malignancies. Carcinoembryonic antigen (CEA, CEACAM5) and epithelial cadherin (E-cadherin) are considered as independent tumor markers in monitoring metastatic colorectal cancer. They are both expressed by cancer cells and can be detected in the blood serum. We investigated the effect of CEA production by MIP101 colorectal carcinoma cell lines on E-cadherin adherens junction (AJ) protein complexes. No direct interaction between E-cadherin and CEA was detected; however, the functional relationships between E-cadherin and its AJ partners: α-, β- and p120 catenins were impaired. We discovered a novel interaction between CEA and beta-catenin protein in the CEA producing cells. It is shown in the current study that CEA overexpression alters the splicing of p120 catenin and triggers the release of soluble E-cadherin. The influence of CEA production by colorectal cancer cells on the function of E-cadherin junction complexes may explain the link between the elevated levels of CEA and the increase in soluble E-cadherin during the progression of colorectal cancer., (Copyright © 2014 Elsevier Inc. All rights reserved.)

The carcinoembryonic antigen-related cell adhesion molecule 1 regulates insulin sensitivity by promoting hepatic insulin clearance. Mice bearing a null mutation of Ceacam1 gene (Cc1(-/-)) develop impaired insulin clearance followed by hyperinsulinemia and insulin resistance, in addition to visceral obesity and increased plasma fatty acids. Because insulin resistance is associated with increased blood pressure, we investigated whether they develop higher blood pressure with activated renal renin-angiotensin system and whether this is mediated, in part, by the upregulation of renal (pro)renin receptor (PRR) expression. Compared with age-matched wild-type littermates, Cc1(-/-) mice exhibited increased blood pressure with increased activation of renal renin-angiotensin systems and renal PRR expression. Cytoplasmic and nuclear immunostaining of phospho-PI3K p85α and phospho-Akt was enhanced in the kidney of Cc1(-/-) mice. In murine renal inner medullary collecting duct epithelial cells with lentiviral-mediated small hairpin RNA knockdown of carcinoembryonic antigen-related cell adhesion molecule 1, PRR expression was upregulated and phosphorylation of PI3K (Tyr508), Akt (Ser473), NF-κB p65 (Ser276), cAMP response element-binding protein/activated transcription factor (ATF)-1 (Ser133), and ATF-2 (Thr71) was enhanced. Inhibiting PI3K with LY294002 or Akt with Akt inhibitor VIII attenuated PRR expression. In conclusion, global null deletion of Ceacam1 caused an increase in blood pressure with increased renin-angiotensin system activation together with upregulation of PRR via PI3K-Akt activation of cAMP response element-binding protein 1, ATF-1, ATF-2, and NF-κB p65 transcription factors.

Liver metastasis is the predominant cause of colorectal cancer (CRC)-related mortality in developed countries. Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) is a cell adhesion molecule with reduced expression in early phases of CRC development and thus functions as a tumor growth inhibitor. However, CEACAM1 is upregulated in metastatic colon cancer, suggesting a bimodal role in CRC progression. To investigate the role of this protein in the host metastatic environment, Ceacam1(-/-) mice were injected intrasplenically with metastatic MC38 mouse CRC cells. A significant reduction in metastatic burden was observed in Ceacam1(-/-) compared with wild-type (WT) livers. Intravital microscopy showed decreased early survival of MC38 cells in Ceacam1(-/-) endothelial environment. Metastatic cell proliferation within the Ceacam1(-/-) livers was also diminished. Bone marrow-derived cell recruitment, attenuation of immune infiltrates and diminished CCL2, CCL3 and CCL5 chemokine production participated in the reduced Ceacam1(-/-) metastatic phenotype. Transplantations of WT bone marrow (BM) into Ceacam1(-/-) mice fully rescued metastatic development, whereas Ceacam1(-/-) BM transfer into WT mice showed reduced metastatic burden. Chimeric immune cell profiling revealed diminished recruitment of CD11b(+)Gr1(+) myeloid-derived suppressor cells (MDSCs) to Ceacam1(-/-) metastatic livers and adoptive transfer of MDSCs confirmed the involvement of these immune cells in reduction of liver metastasis. CEACAM1 may represent a novel metastatic CRC target for treatment.

Background: Natural killer (NK) cells are lymphocytes of the innate immune system that play a crucial role in tumor immune surveillance. Accumulated data indicated that NK cells in the tumor microenvironment often display a suppressed function. However, the mechanism is not clear., Objective: In this study, the effects and relative mechanisms of malignant pleural effusion (MPE) from patients with lung cancer on NK cells were researched., Methods: MPE and peripheral blood (PB) samples were collected from patients with lung cancer. The cytotoxic activity of CD56(dim) NK cells in PB and MPE mononuclear cells was analyzed by flow cytometry., Results: It was observed that the percentages of total NK cells and a CD56(dim) NK subset in MPE reduced accompanying impaired cytotoxic activity compared with that in paired PB. Cell-free MPE treatment reduced both the proportion and cytotoxic activity of CD56(dim) NK cells in PB from healthy donors. The suppression effects were not based on soluble carcinoembryonic antigen and the inhibitory cytokines interleukin-10 and transforming growth factor-β1, but were dependent on the factor with a molecular weight >100 kDa., Conclusions: These results demonstrated that native soluble carcinoembryonic antigen does not suppress the activity of NK cells, and an unknown factor with a molecular weight >100 kDa plays a critical role in the impairment of CD56(dim) NK cells in MPE, which might lead to tumor progression., (Copyright © 2012 S. Karger AG, Basel.)

After separating from a primary tumor, metastasizing cells enter the circulatory system and interact with host cells before lodging in secondary organs. Previous studies have implicated the surface glycoproteins CD44 and carcinoembryonic antigen (CEA) in adhesion, migration, and invasion, suggesting that they may influence metastatic progression. To elucidate the role of these multifunctional molecules while avoiding the potential drawbacks of ectopic expression or monoclonal antibody treatments, we silenced the expression of CD44 and/or CEA in LS174T colon carcinoma cells and analyzed their ability to metastasize in 2 independent mouse models. Quantitative PCR revealed that CD44 knockdown increased lung and liver metastasis >10-fold, while metastasis was decreased by >50% following CEA knockdown. These findings were corroborated by in vitro experiments assessing the metastatic potential of LS174T cells. Cell migration was decreased as a result of silencing CEA but was enhanced in CD44-knockdown cells. In addition, CD44 silencing promoted homotypic aggregation of LS147T cells, a phenotype that was reversed by additional CEA knockdown. Finally, CD44-knockdown cells exhibited greater mechanical compliance than control cells, a property that correlates with increased metastatic potential. Collectively, these data indicate that CEA, but not CD44, is a viable target for therapeutics aimed at curbing colon carcinoma metastasis.

Background: Early diagnosis represents the best opportunity for cure of colorectal cancer. Current screening programmes use faecal occult blood testing for screening, which has limited sensitivity and poor specificity., Methods: In this study we looked at a series of previously described diagnostic markers utilising circulating free DNA (cfDNA), with a preparation method allowing small DNA fragments to be isolated. The Circulating free DNA was isolated from samples obtained from 85 patients, including 35 patients without endoscopic abnormality, a group of 26 patients with benign colorectal adenomas, and 24 patients with colorectal carcinomas. In each case, polymerase chain reaction (PCR) was performed for Line1 79 bp, Line1 300 bp, Alu 115 bp, Alu 247 bp, and mitochondrial primers. In addition, carcinoembryonic antigen (CEA) was measured by ELISA. Each marker was analysed between normal, polyp, and cancer populations, and the best performing analysed in combination by logistic regression., Results: The best model was able to discriminate normal from populations with adenoma or carcinoma using three DNA markers and CEA, showing an area under the receiver operator characteristic (ROC) curve of 0.855 with a positive predictive value of 81.1% for polyps and cancer diagnosis., Conclusion: These circulating markers in combination with other markers offer the prospect of a simple blood test as a possible secondary screen for colorectal cancers and polyps in patients with positive faecal occult blood tests.

Although carcinoembryonic antigen-related cell adhesion molecule-1 (CEACAM1) is an activation marker for neutrophils and delays neutrophil apoptosis, the role of CEACAM1 in granulopoiesis and neutrophil-dependent host immune responses has not been investigated. CEACAM1 expression correlated with granulocytic differentiation, and Ceacam1(-/-) mice developed neutrophilia because of loss of the Src-homology-phosphatase-1 (SHP-1)-dependent inhibition of granulocyte colony-stimulating factor receptor (G-CSFR) signal transducer and activator of transcription (Stat3) pathway provided by CEACAM1. Moreover, Ceacam1(-/-) mice were hypersensitive to Listeria Monocytogenes (LM) infection with an accelerated mortality. Reintroduction of CEACAM1 into Ceacam1(-/-) bone marrow restored normal granulopoiesis and host sensitivity to LM infection, while mutation of its immunoreceptor tyrosine-based inhibitory motifs (ITIMs) abrogated this restoration. shRNA-mediated reduction of Stat3 amounts rescued normal granulopoiesis, attenuating host sensitivity to LM infection in Ceacam1(-/-) mice. Thus, CEACAM1 acted as a coinhibitory receptor for G-CSFR regulating granulopoiesis and host innate immune response to bacterial infections., (Copyright © 2010 Elsevier Inc. All rights reserved.)

Murine coronavirus (mouse hepatitis virus, MHV) is a collection of strains that induce disease in several organ systems of mice. Infection with neurotropic strains JHM and A59 causes acute encephalitis, and in survivors, chronic demyelination, the latter of which serves as an animal model for multiple sclerosis. The MHV receptor is a carcinoembryonic antigen-related cell adhesion molecule, CEACAM1a; paradoxically, CEACAM1a is poorly expressed in the central nervous system (CNS), leading to speculation of an additional receptor. Comparison of highly neurovirulent JHM isolates with less virulent variants and the weakly neurovirulent A59 strain, combined with the use of reverse genetics, has allowed mapping of pathogenic properties to individual viral genes. The spike protein, responsible for viral entry, is a major determinant of tropism and virulence. Other viral proteins, both structural and nonstructural, also contribute to pathogenesis in the CNS. Studies of host responses to MHV indicate that both innate and adaptive responses are crucial to antiviral defense. Type I interferon is essential to prevent very early mortality after infection. CD8 T cells, with the help of CD4 T cells, are crucial for viral clearance during acute disease and persist in the CNS during chronic disease. B cells are necessary to prevent reactivation of virus in the CNS following clearance of acute infection. Despite advances in understanding of coronavirus pathogenesis, questions remain regarding the mechanisms of viral entry and spread in cell types expressing low levels of receptor, as well as the unique interplay between virus and the host immune system during acute and chronic disease.

Aim: To evaluate the inhibitory effect of lymphocytes activated by the superantigen of toxic shock syndrome toxin-1 (TSST-1), which is regulated synergistically by 5 copies of hypoxia-responsive element (5HRE) and promoter of carcino-embryonic antigen (CEAp), against the carcino-embryonic antigen (CEA)-positive human colon carcinoma cell line LoVo under hypoxia condition in vitro., Methods: The eukaryotic expressive vector was constructed with the transmembrane superantigen gene of 5HRE-CEAp-regulated TSST-1-linker-CD80TM, and was transfected into CEA-positive human colon carcinoma cell line LoVo and CEA-negative human cervical carcinoma cell line HeLa by Lipofectamine 2000. Stably transfected cell lines were selected by G418. RT-PCR was employed to examine the expression of TC mRNA. Peripheral blood lymphocytes(PBL) of healthy people were extracted and then activated by the lysates of tumor cells stably transfected with TC. Concurrently, PBL were cultured together with stably transfected tumor cells in order to kill these cells. Then the proliferative effect of TSST-1 on PBL, and the killing effect of PBL against the stably transfected tumor cells were detected by MTT., Results: Monoclonal LoVo and HeLa cells stably transfected with TC were successfully obtained. Expression of TC mRNA in monoclonal LoVo cells under hypoxia condition was significantly higher than those under normoxia condition as confirmed by RT-PCR. Monoclonal HeLa cells did not express TC under either hypoxia condition or normoxia condition. As is shown by MTT assay, TSST-1 expressed by the monoclonal LoVo cells could effectively activate PBL to proliferate under hypoxia condition, resulting in dose-dependent inhibition on the proliferation of LoVo cells that expressed TSST (P<0.05). But HeLa cells and wild LoVo cells could not activate PBL to proliferate. PBL could not inhibit proliferation of HeLa cells and wild LoVo cells, either., Conclusion: The superantigen of TSST-1 regulated dually by 5HRE and CEAp could activate human PBL to kill CEA-positive tumor cells specifically under hypoxia condition.

The efficiency of secondary tumor establishment is controlled by the ability of tumor cells to withstand a barrage of mechanical and immunological stresses during their passage through the circulatory system. Accumulating evidence suggests that the selectin-dependent interactions of circulating tumor cells with host cells promote their survival and extravasation from the vasculature, therefore representing a critical checkpoint for colonization of distant organs. These observations have motivated the identification and biochemical characterization of functional selectin ligands such as CD44 variant isoforms, carcinoembryonic antigen and podocalyxin-like protein, present on the surface of metastatic colon carcinoma cells. Understanding the molecular underpinnings of selectin-ligand interactions involved in heterotypic tumor cell-host cell adhesion events may provide guidelines for developing novel cancer diagnostics. Recent advancements in diagnostic device fabrication and design integrated with novel biomarkers exploiting the tissue specific biochemistry of malignant versus normal tissue-expressed selectin ligands may hold promise in providing effective alternatives to current cancer diagnostic technologies.

The carcinoembryonic antigen cell adhesion molecule 1 (CEACAM1) is downregulated in colonic and intestinal hyperplastic lesions as well as in other cancers, where it functions as a tumor suppressor. To investigate the functions of CEACAM1 in the normal intestine and in intestinal tumors, we generated a compound knockout mouse model and examined both Ceacam1(-/-) and Apc(1638N/+):Ceacam1(-/-) mice. Ceacam1(-/-) intestinal cells exhibited a significant decrease in apoptosis, with no change in proliferation or migration, however. Compound Apc(1638N/+):Ceacam1(-/-) mice demonstrated an increase in intestinal tumor multiplicity and tumor progression. Increases in intussusceptions and desmoid lesions were also observed. We have shown that CEACAM1-L associates with beta-catenin by co-immunoprecipitation and colocalization in CEACAM1-L-transfected CT26 and CT51 mouse colon carcinoma cells. Ceacam1(-/-) enterocytes displayed decreased glycogen synthase kinase 3-beta activity with corresponding nuclear localization of beta-catenin. Increased T-cell factor/Lef transcriptional activity was observed in CEACAM1-null CT51 colonic cells and in Caco2 colon cancer cells in which CEACAM1 was downregulated. A significant increased expression in c-Myc and cyclin D1 targets of the Wnt signaling pathway was also revealed in the Ceacam1(-/-) intestine. CEACAM1 therefore actively participates in Wnt signaling in intestinal cells and its downregulation in intestinal tissue contributes to malignancy by augmenting tumor multiplicity and progression.

The aim of this study was to verify through relative survival (an estimate of cancer-specific survival) the true prognostic factors of colorectal cancer. The study involved 506 patients who underwent locally radical resection. All the clinical, histological and laboratory parameters were prognostically analysed for both overall and relative survival. This latter was calculated from the expected survival of the general population with identical age, sex and calendar years of observation. Univariate and multivariate analyses were applied to the proportional hazards model. Liver metastases, age, lymph node involvement and depth of bowel wall involvement were independent prognosticators of both overall and relative survival, whereas carcinoembryonic antigen (CEA) was predictive only of relative survival. Increasing age was unfavourably related to overall survival, but mildly protective with regard to relative survival. Three out of the five prognostic factors identified are the cornerstones of the current staging systems, and were confirmed as adequate by the analysis of relative survival. The results regarding age explain the conflicting findings so far obtained from studies considering overall survival only and advise against the adoption of absolute age limits in therapeutic protocols. Moreover, the prechemotherapy CEA level showed a high clinical value.

A highly neurovirulent mouse hepatitis virus (MHV) JHMV strain (wt) with receptor (MHVR)-independent infection activity and its low-virulent mutant srr7 without such activity were found to attach to MHVR-negative, non-permissive BHK cells. To identify the molecule that interacts with JHMV, we focused on heparan sulfate (HS) since it works as a receptor of a mutant MHV-rec1 that infects in an MHVR-independent fashion. The present study indicates that HS interacts with both wt JHMV and srr7 but it does not function as an entry receptor as it apparently does for MHV-rec1. Furthermore, HS failed to serve as an entry receptor in the MHVR-independent infection of wt JHMV, indicating that HS is not a host factor that wt JHMV utilizes in an MHVR-independent infection.

GPI membrane anchors of cell surface glycoproteins have been shown to confer functional properties that are different from their transmembrane (TM)-anchored counterparts. For the human carcinoembryonic antigen (CEA) family, a subfamily of the immunoglobulin superfamily, conversion of the mode of membrane linkage from TM to GPI confers radical changes in function: from tumor suppression or neutrality toward inhibition of differentiation and anoikis and distortion of tissue architecture, thereby contributing to tumorigenesis. We show here that GPI anchorage in the CEA family evolved twice independently in primates, very likely from more primitive TM anchors, by different packages of mutations. Both mutational packages, one package found in many primates, including humans, and a second, novel package found only in the Cebidae radiation of New World monkeys, give rise to efficiently processed GPI-linked proteins. Both types of GPI anchors mediate inhibition of cell differentiation. The estimated rate of nonsynonymous mutations (Ka) in the anchor-determining domain for conversion from TM to GPI anchorage in the CEA family that were fixed during evolution in these primates is 7 times higher than the average Ka in primates, indicating positive selection. These results suggest therefore that the functional changes mediated by CEA GPI anchors, including the inhibition of differentiation and anoikis, could be adaptive and advantageous.

Carcinoembryonic antigen (CEA) and CEA family member CEACAM6 are glycophosphatidyl inositol (GPI)-anchored, intercellular adhesion molecules that are up-regulated in a wide variety of human cancers, including colon, breast, and lung. When over-expressed in a number of cellular systems, these molecules are capable of inhibiting cellular differentiation and anoikis, as well as disrupting cell polarization and tissue architecture, thus increasing tumorigenicity. The present study shows that perturbation of the major fibronectin receptor, integrin alpha5beta1, underlies some of these biological effects. Using confocal microscopy and specific antibodies, CEA and CEACAM6 were demonstrated to co-cluster with integrin alpha5beta1 on the cell surface. The presence of CEA and CEACAM6 was shown to lead to an increase in the binding of the integrin alpha5beta1 receptor to its ligand fibronectin, without changing its cell surface levels, resulting in increased adhesion of CEA/CEACAM6-expressing cells to fibronectin. More tenacious binding of free fibronectin to cells led to enhanced fibronectin matrix assembly and the formation of a polymerized fibronectin "cocoon" around the cells. Disruption of this process with specific monoclonal antibodies against either fibronectin or integrin alpha5beta1 led to the restoration of cellular differentiation and anoikis in CEA/CEACAM6 producing cells., (Copyright 2006 Wiley-Liss, Inc.)

The liver is the most common site for metastasis by colorectal cancer, and numerous studies have shown a relationship between serum carcinoembryonic antigen (CEA) levels and metastasis to this site. CEA activates hepatic macrophages or Kupffer cells via binding to the CEA receptor (CEA-R), which results in the production of cytokines and the up-regulation of endothelial adhesion molecules, both of which are implicated in hepatic metastasis. Since tissue macrophages implicated in the metastatic process can often be difficult to isolate, the aim of this study was to develop an in vitro model system to study the complex mechanisms of CEA-induced macrophage activation and metastasis. Undifferentiated, human monocytic THP-1 (U-THP) cells were differentiated (D-THP) to macrophages by exposure to 200 ng/ml phorbol myristate acetate (PMA) for 18 h. Immunohistochemistry showed two CEA-R isoforms present in both U- and D-THP cells. The receptors were localized primarily to the nucleus in U-THP cells, while a significant cell-surface presence was observed following PMA-differentiation. Incubation of D-THP-1 cells with CEA resulted in a significant increase in tumor necrosis factor-alpha (TNF-alpha) release over 24 h compared to untreated D-THP-1 or U-THP controls confirming the functionality of these cell surface receptors. U-THP cells were unresponsive to CEA. Attachment of HT-29 cells to human umbilical vein endothelial cells significantly increased at 1 h after incubation with both recombinant TNF-alpha and conditioned media from CEA stimulated D-THP cells by six and eightfold, respectively. This study establishes an in vitro system utilizing a human macrophage cell line expressing functional CEA-Rs to study activation and signaling mechanisms of CEA that facilitate tumor cell attachment to activated endothelial cells. Utilization of this in vitro system may lead to a more complete understanding of the expression and function of CEA-R and facilitate the design of anti-CEA-R therapeutic modalities that may significantly diminish the metastatic potential of CEA overexpressing colorectal tumors.

The neisserial opacity (Opa) proteins are phase-variable, antigenically distinct outer membrane proteins that mediate adherence to and invasion of human cells. We previously reported that Neisseria gonorrhoeae Opa protein expression appeared to be selected for or induced during experimental murine genital tract infection. Here we further defined the kinetics of recovery of Opa variants from the lower genital tracts of female mice and investigated the basis for this initial observation. We found that the recovery of different Opa phenotypes from mice appears cyclical. Three phases of infection were defined. Following intravaginal inoculation with primarily Opa- gonococci, the majority of isolates recovered were Opa+ (early phase). A subsequent decline in the percentage of Opa+ isolates occurred in a majority of mice (middle phase) and was followed by a reemergence of Opa+ variants in mice that were infected for longer than 8 days (late phase). We showed the early phase was due to selection for preexisting Opa+ variants in the inoculum by constructing a chloramphenicol-resistant (Cm(r)) strain and following Cm(r) Opa+ populations mixed with a higher percentage of Opa- variants of the wild-type (Cm(s)) strain. Reciprocal experiments (Opa- Cm(r) gonococci spiked with Opa+ Cm(s) bacteria) were consistent with selection of Opa+ variants. Based on the absence in mice of human carcinoembryonic antigen cell adhesion molecules, the major class of Opa protein adherence receptors, we conclude the observed selection for Opa+ variants early in infection is not likely due to a specific adherence advantage and may be due to Opa-mediated evasion of innate defenses.

Murine hepatitis coronavirus (MHV)-A59 infection depends on the interaction of its spike (S) protein with the cellular receptor mCEACAM1a present on murine cells. Human cells lack this receptor and are therefore not susceptible to MHV. Specific alleviation of the tropism barrier by redirecting MHV to a tumor-specific receptor could lead to a virus with appealing properties for tumor therapy. To demonstrate that MHV can be retargeted to a nonnative receptor on human cells, we produced bispecific adapter proteins composed of the N-terminal D1 domain of mCEACAM1a linked to a short targeting peptide, the six-amino-acid His tag. Preincubation of MHV with the adapter proteins and subsequent inoculation of human cells expressing an artificial His receptor resulted in infection of these otherwise nonsusceptible cells and led to subsequent production of progeny virus. To generate a self-targeted virus able to establish multiround infection of the target cells, we subsequently incorporated the gene encoding the bispecific adapter protein as an additional expression cassette into the MHV genome through targeted RNA recombination. When inoculated onto murine LR7 cells, the resulting recombinant virus indeed expressed the adapter protein. Furthermore, inoculation of human target cells with the virus resulted in a His receptor-specific infection that was multiround. Extensive cell-cell fusion and rapid cell killing of infected target cells was observed. Our results show that MHV can be genetically redirected via adapters composed of the S protein binding part of mCEACAM1a and a targeting peptide recognizing a nonnative receptor expressed on human cells, consequently leading to rapid cell death. The results provide interesting leads for further investigations of the use of coronaviruses as antitumor agents.

The carcinoembryonic antigen (CEA) is glycoprotein localized in the apical surface of mature enterocytes. The members of the CEA gene family are clustered on chromosome 19q13.2. It is formed by 29 genes, of which 18 are expressed. Many functions of CEA have been known in healthy indiuiduals, however its role as cell adhesion molecule is the most studied. Besides the colon, CEA is expressed in the stomach, tongue, oesophagus, cervix, and prostate. The most important clinical function is in colorectal, gastric and ovary cancer. It is used as prognosis marker, staging system, recurrence, treatment response and liver metastases. There are many non-neoplasic diseases that enhance CEA value. Actually, CEA is being studying as target of immunotherapy.

The NK killing activity is regulated by activating and inhibitory NK receptors. All of the activating ligands identified so far are either viral or stress-induced proteins. The class I MHC proteins are the ligands for most of the inhibitory NK receptors. However, in the past few years, several receptors have been identified that are able to inhibit NK killing independently of class I MHC recognition. We have previously demonstrated the existence of a novel inhibitory mechanism of NK cell cytotoxicity mediated by the homophilic carcinoembryonic Ag (CEA)-related cell adhesion molecule 1 (CEACAM1) interactions. In this study, we demonstrate that CEACAM1 also interacts heterophilically with the CEA protein. Importantly, we show that these heterophilic interactions of CEA and CEACAM1 inhibit the killing by NK cells. Because CEA is expressed on a wide range of carcinomas and commonly used as tumor marker, these results represent a novel role for the CEA protein enabling the escape of tumor cells from NK-mediated killing. We further characterize, for the first time, the CEACAM1-CEA interactions. Using functional and binding assays, we demonstrate that the N domains of CEACAM1 and CEA are crucial but not sufficient for both the CEACAM1-CEACAM1 homophilic and CEACAM1-CEA heterophilic interactions. Finally, we suggest that the involvement of additional domains beside the N domain in the heterophilic and homophilic interactions is important for regulating the balance between cis and trans interactions.

Purpose: Cancer vaccines targeting tumor-associated antigens are being investigated for the therapy of tumors. Numerous strategies, including the direct intratumoral (i.t.) vaccination route, have been examined. For tumors expressing carcinoembryonic antigen (CEA) as a model tumor-associated antigen, we previously designed poxviral vectors that contain the transgenes for CEA and a triad of T-cell costimulatory molecules, B7-1, intercellular adhesion molecule-1, (ICAM-1), and leukocyte function associated antigen-3 (LFA-3) (CEA/TRICOM). Two types of poxvirus vectors were developed: replication-competent recombinant vaccinia and replication-defective recombinant fowlpox. We have shown previously that a vaccine regimen composed of priming mice s.c. with recombinant vaccinia-CEA/TRICOM and boosting i.t. with recombinant fowlpox-CEA/TRICOM was superior to priming and boosting vaccinations using the conventional s.c. route in inducing T-cell responses specific for CEA. These studies also showed that CEA was needed to be present both in the vaccine and in the tumor for therapeutic effects., Experimental Design: To determine specific immune responses associated with vaccination-mediated tumor regression, CEA-transgenic mice bearing CEA(+) tumors were vaccinated with the CEA/TRICOM s.c./i.t. regimen, and T-cell immune responses were assessed., Results: In CEA(+) tumor-bearing mice vaccinated with the CEA/TRICOM s.c./i.t. regimen, T-cell responses could be detected not only to CEA encoded in vaccine vectors but also to other antigens expressed on the tumor itself: wild-type p53 and an endogenous retroviral epitope of gp70. Moreover, the magnitude of CD8(+) T-cell immune responses to gp70 was far greater than that induced to CEA or p53. Finally, the predominant T-cell population infiltrating the regressing CEA(+) tumor after therapy was specific for gp70., Conclusion: These studies show that the breadth and magnitude of antitumor immune cascades to multiple antigens could be critical in the therapy of established tumors.

CEACAM1a glycoproteins are members of the immunoglobulin (Ig) superfamily and the carcinoembryonic antigen family. Isoforms expressing either two or four alternatively spliced Ig-like domains in mice have been found in a number of epithelial, endothelial, or hematopoietic tissues. CEACAM1a functions as an intercellular adhesion molecule, an angiogenic factor, and a tumor cell growth inhibitor. Moreover, the mouse and human CEACAM1a proteins are targets of viral or bacterial pathogens, respectively, including the murine coronavirus mouse hepatitis virus (MHV), Haemophilus influenzae, Neisseria gonorrhoeae, and Neisseria meningitidis, as well as Moraxella catarrhalis in humans. We have shown that targeted disruption of the Ceacam1a (MHVR) gene resulting in a partial ablation of the protein in mice (p/p mice) led to reduced susceptibility to MHV-A59 infection of the modified mice in the BALB/c background. We have now engineered and produced a Ceacam1a-/- mouse that exhibits complete ablation of the CEACAM1a protein in every tissue where it is normally expressed. We report that 3-week-old Ceacam1a-/- mice in the C57BL/6 genetic background are fully resistant to MHV-A59 infection by both intranasal and intracerebral routes. Whereas virus-inoculated wild-type +/+ C57BL/6 mice showed profound liver damage and spinal cord demyelination under these conditions, Ceacam1a-/- mice displayed normal livers and spinal cords. Virus was recovered from liver and spinal cord tissues of +/+ mice but not of -/- mice. These results indicate that CEACAM1a is the sole receptor for MHV-A59 in both liver and brain and that its deletion from the mouse renders the mouse completely resistant to infection by this virus., (Copyright 2004 American Society for Microbiology)

Individual Neisseria gonorrhoeae colony opacity-associated (Opa) protein variants can bind up to four different carcinoembryonic antigen-related cellular adhesion molecule (CEACAM) receptors. Most human cells encountered by gonococci express a combination of CEACAM receptors, thereby complicating the elucidation of intracellular signaling pathways triggered by individual receptors. Here, we compare the process of bacterial engulfment by a panel of stably transfected HeLa epithelial cell lines expressing each CEACAM receptor in isolation. CEACAM1 and CEACAM3 each contain proteinaceous transmembrane and cytoplasmic domains; however, the processes of neisserial uptake mediated by these receptors differ with respect to their susceptibilities to both tyrosine kinase inhibitors and the actin microfilament-disrupting agent cytochalasin D. Neisserial uptake mediated by glycosylphosphatidylinositol (GPI)-anchored CEACAM5 and CEACAM6 was not significantly affected by any of a broad spectrum of inhibitors tested. However, cleavage of the GPI anchor by phosphatidylinositol-specific phospholipase C reduced bacterial uptake by HeLa cells expressing CEACAM5, consistent with a single zipper-like mechanism of uptake mediated by this receptor. Regardless of the CEACAM receptor expressed, internalized gonococci were effectively killed by a microtubule-dependent process that required acidification of the bacterium-containing phagosome. Given the phase-variable nature of neisserial Opa proteins, these results indicate that the mechanism of bacterial engulfment and the cellular response to gonococcal infection depend on both the receptor specificities of the neisserial Opa protein variants expressed and the spectrum of CEACAM receptors present on target cells, each of which determines the combination of receptors ultimately engaged.

Carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) are used by several human pathogens to anchor themselves to or invade host cells. Interestingly, human granulocytes express a specific isoform, CEACAM3, that participates together with CEACAM1 and CEACAM6 in the recognition of CEACAM-binding microorganisms. Here we show that CEACAM3 can direct efficient, opsonin-independent phagocytosis of CEACAM-binding Neisseria, Moraxella, and Haemophilus species. CEACAM3- but not CEACAM6-mediated uptake is blocked by dominant-negative versions of the small GTPase Rac. Moreover, CEACAM3 engagement triggers membrane recruitment and increased GTP loading of Rac that are not observed upon bacterial binding to CEACAM6. Internalization and Rac stimulation are also inhibited by compromising the integrity of an immunoreceptor tyrosine-based activation motif (ITAM)-like sequence in the cytoplasmic tail of CEACAM3 or by interference with Src family protein tyrosine kinases that phosphorylate CEACAM3. In contrast to interfering with CEACAM6, blockage of CEACAM3-mediated events reduces the ability of primary human granulocytes to internalize and eliminate CEACAM-binding bacteria, indicating an important role of CEACAM3 in the control of human-specific pathogens by the innate immune system.

CD98 is a multifunctional protein involved in amino acid transport and regulation of integrin-mediated cell adhesion. Herein, we demonstrated that CD98 stimulation by anti-CD98 antibodies induced CEA-CAM-1-mediated cell adhesion in BaF3 cells expressing CEA-CAM-1, and suggest that this might be responsible for compact clumping of F9 embryonic carcinoma cells by CD98 stimulation. CEA-CAM-1 was co-immunoprecipitated by anti-CD98 antibody. CD98 stimulation induced the translocation of cytoplasmic protein kinase Cdelta (PKCdelta) to the cell adhesion sites, and rottlerin that inhibited the PKCdelta translocation abolished the cell aggregation without affecting integrin activation. The results suggested that CD98 stimulation could activate CEA-CAM-1-mediated cell adhesion independently of integrins.

The external domains of Ig superfamily members are involved in multiple binding interactions, both homophilic and heterophilic, that initiate molecular events leading to the execution of diverse cell functions. Human carcinoembryonic antigen (CEA), an Ig superfamily cell surface glycoprotein used widely as a clinical tumor marker, undergoes homophilic interactions that mediate intercellular adhesion. Recent evidence supports the view that deregulated overexpression of CEA has an instrumental role in tumorigenesis through the inhibition of cell differentiation and the disruption of tissue architecture. The CEA-mediated block of the myogenic differentiation of rat L6 myoblasts depends on homophilic binding of its external domains. We show here that L6 transfectant cells expressing CEA can "trans-block" the myogenesis of juxtaposed differentiation-competent L6 transfectant cells expressing a deletion mutant of CEA (DeltaNCEA). This result implies the efficacy of antiparallel CEA-CEA interactions between cells in the differentiation block. In addition, DeltaNCEA can acquire differentiation blocking activity by cross-linking with specific anti-CEA antibodies, thus implying the efficacy of parallel CEA-CEA interactions on the same cell surface. The myogenic differentiation blocking activity of CEA was demonstrated by site-directed mutations to involve three subdomains of the amino-terminal domain, shown previously to be critical for its intercellular adhesion function. Monovalent Fab fragments of monoclonal antibodies binding to the region bridging subdomains 1 and 2 could both inhibit intercellular adhesion and release the myogenic differentiation block. Amino acid substitutions Q80A, Q80R, and D82N in subdomain 3, QNDTG, however, were found to completely ablate the differentiation blocking activity of CEA but had no effect on intercellular adhesion activity. A cyclized peptide representing this subdomain was the most effective at releasing the differentiation block.

CEACAM1 (carcinoembryonic antigen-related cell adhesion molecule 1) is a transmembrane glycoprotein belonging to the carcinoembryonic antigen (CEA) family and immunoglobulin superfamily. It is localized mainly in the apical domains of polarized epithelia, leukocytes and endothelia. With respect to this wide tissue distribution the research is focused on the study of its biological functions. Structural and functional analyses show that the extracellular domain of CEACAM1 participates in homotypic and heterotypic adhesion, whereas the cytoplasmic domain takes part in cell growth inhibition and signal transduction. Whereas CEA is highly expressed in adenocarcinomas, CEACAM1 expression is down regulated in many tumors and its tumor-supressive function was confirmed. CEACAM1 also takes part in insulin metabolism, acts as a promotor of cholesterol crystallization and serves as a binding receptor for certain bacterial strains in humans as well as hepatitis virus in mice.

Carcinoembryonic antigen (CEA), the first tumor-associated antigen to be described, was cloned in 1987 and is expressed on nearly 50% of all human tumors. The identification of T-cell specific epitopes within the coding region of the CEA protein has led to the development of various vaccine strategies that target CEA and CEA-expressing tumors. These vaccines have shown evidence of therapeutic effectiveness in animal models and are being evaluated in early phase clinical trials. Although trials are not designed to elucidate clinical responses, they have provided important information about the ability of individual vaccines to induce CEA-specific immune responses through the use of newer in vitro monitoring assays. Continued clinical testing in patients with less advanced disease and the administration of vaccines in combination with other standard therapy will help define the role of CEA-based vaccines in the treatment and prevention of human cancer.

Carcinoembryonic Ag-related cell adhesion molecule 1 (CEACAM1), the primordial carcinoembryonic Ag gene family member, is a transmembrane cell adhesion molecule expressed in leukocytes, epithelia, and blood vessel endothelia in humans and rodents. As a result of differential splicing, CEACAM1 occurs as several isoforms, the two major ones being CEACAM1-L and CEACAM1-S, that have long (L) or short (S) cytoplasmic domains, respectively. The L:S expression ratios vary in different cells and tissues. In addition to CEACAM1, human but not rodent cells express GPI-linked CEACAM members (CEACAM5-CEACAM8). We compared the expression patterns of CEACAM1-L, CEACAM1-S, CEACAM6, and CEACAM8 in purified populations of neutrophilic granulocytes, B lymphocytes, and T lymphocytes from rats, mice, and humans. Human granulocytes expressed CEACAM1, CEACAM6, and CEACAM8, whereas human B lymphocytes and T lymphocytes expressed only CEACAM1 and CEACAM6. Whereas granulocytes, B cells, and T cells from mice and rats expressed both CEACAM1-L and CEACAM1-S in ratios of 2.2-2.9:1, CEACAM1-S expression was totally lacking in human granulocytes, B cells, and T cells. Human leukocytes only expressed the L isoforms of CEACAM1. This suggests that the GPI-linked CEACAM members have functionally replaced CEACAM1-S in human leukocytes. Support for the replacement hypothesis was obtained from experiments in which the extracellular signal-regulated kinases (Erk)1/2 were activated by anti-CEACAM Abs. Thus, Abs against CEACAM1 activated Erk1/2 in rat granulocytes, but not in human granulocytes. Erk1/2 in human granulocytes could, however, be activated by Abs against CEACAM8. We demonstrated that CEACAM1 and CEACAM8 are physically associated in human granulocytes. The CEACAM1/CEACAM8 complex in human cells might accordingly play a similar role as CEACAM1-L/CEACAM1-S dimers known to occur in rat cells.

Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1), an Ig-like transmembrane protein, functions in cell adhesion, angiogenesis and epithelial cell morphogenesis, and has been identified as a tumor suppressor. For all of these functions, CEACAM1 requires signaling capabilities. However, the mechanisms of CEACAM1-mediated signaling are only poorly understood. Here we characterized for the first time CEACAM1 expression and signaling in the neuroendocrine rat pheochromocytoma PC12 cell line. Stimulation of CEACAM1 by ligation on the cell surface with antibodies induced formation of large CEACAM1 clusters and a rapid and transient CEACAM1 tyrosine dephosphorylation. Functionally, this dephosphorylation correlated with a reduced association between CEACAM1 and the tyrosine phosphatase SHP2. Clustering also stimulated binding of CEACAM1 to the actin cytoskeleton, measured by a partial translocation of CEACAM1 into the insoluble fraction after detergent extraction. Both tyrosine dephosphorylation and interaction with the cytoskeleton were sensitive to neuronal differentiation of PC12 cells. The first detected downstream activation of the mitogen-activated protein kinases ERK1 and ERK2, but not of JNK or p38, describes a novel target of CEACAM1-mediated signaling and contributes to the understanding of how CEACAM1 regulates cellular function.

Carcinoembryonic Ag cell adhesion molecule 1 (CEACAM1) consists of highly related homologs in humans and rodents that are characterized by significant alternate splicing generating isoforms capable of negative intracellular signaling by virtue of two immunoreceptor tyrosine-based inhibition motifs in its cytoplasmic (cyt) tail. Although human T cells have been recently observed to express CEACAM1, the expression and function of CEACAM1 in mouse T cells have not been defined. Although resting mouse spleen T cells exhibited no evidence of CEACAM1 on the cell surface, CEACAM1 was rapidly up-regulated on CD4+ and CD8+ T cells after activation with either Con A or anti-CD3 without a requirement for either de novo transcription or translation due to the fact that CEACAM1 was present intracellularly before activation. Using a GST-CEACAM1-cytoplasmic tail fusion protein, it was shown that the cytoplasmic tail of CEACAM1 bound the src homology domain-containing phosphatase 1 and adaptor protein 1 complex in its phosphorylated and nonphosphorylated states, respectively. CEACAM1 ligation with an anti-CEACAM1 mAb resulted in inhibition of an allogeneic MLR and anti-CD3 plus anti-CD28 Ab-induced proliferation of spleen T cells in vitro and inhibition of a delayed-type hypersensitivity response to oxazolone in vivo. Inhibition of the delayed-type hypersensitivity response required that the anti-CEACAM1-specific mAb be present at the time of T cell sensitization. These studies support a role for CEACAM1 as a novel class of immunoreceptor tyrosine-based inhibition motif-bearing regulatory molecules on T cells that are active during early phases of the immune response in mice.

It has been suggested that carcinoembryonic antigen (CEA) enhances metastatic seeding of colon cancer cells due to its homo- and heterophilic binding properties. Our recent finding that endogenous CEA protects colon cancer cells against apoptosis suggests a more complex role of CEA in cancer progression. In this study we compared the in vitro effects of endogenous CEA on tumor cell aggregation and cell cycle regulation of human HT29 colon cancer cells with the corresponding in vivo effects, i.e. tumor cell seeding and formation of metastatic lesions. Stable expression of CEA targeted ribozymes (Rz) under control of a tet-off promoter system allowed regulation of CEA levels on the mRNA and protein level by 50%. Downregulation of CEA levels inhibited tumor cell aggregation by 70%. In accordance with previous studies, reduction of CEA levels increased in vitro the apoptotic rate and reduced colony formation by 30% to 50%. To determine the in vivo effect of CEA-dependent aggregate formation and its growth regulating role under apoptotic stress, HT29 cells with high and low CEA levels, respectively, were injected into nude mice. Immunostaining of lung microsections revealed similar numbers of tumor cells one hour after injection. 24 h later virtually all cells were removed from the lung in both groups. However, after 6 weeks all doxycycline treated mice (Rz off = CEA high) showed 14.5 +/- 4.6 metastatic lung lesions/mouse while 0.2 +/- 0.2 lesions/mouse appeared in the untreated group (Rz on = CEA low) (P < 0.001). Our study demonstrates a multifunctional role of CEA and indicates a prometastatic role of CEA independent of its adhesive function possibly due to its anti-apoptotic function.

Biliary glycoprotein (BGP, CD66a, CEACAM1) is a member of the carcinoembryonic antigen family (CEA, CD66), a group of transmembrane proteins belonging to the immunoglobulin superfamily. The structural features surrounding the tyrosine residues in the cytoplasmic domain of BGP share similarity with the consensus sequence of the immunoreceptor tyrosine-based inhibition motif (ITIM), the docking site for SHIP, SHP-1, and SHP-2 molecules. Using the well-characterized inhibitory receptor, FcgammaRIIB, we constructed a FcgammaRIIB-BGPa chimeric molecule that contained the extracellular and transmembrane domain of FcgammaRIIB and the cytoplasmic tail of BGPa and expressed it in DT40 B cells. Our results showed that FcgammaRIIB-BGPa, just like the unmodified FcgammaRIIB molecule, inhibited calcium influx in activated DT40 B cells. Substitution of tyrosine with phenylalanine (Y459F) in FcgammaRIIB-BGPa completely abrogated its ability to inhibit calcium influx, indicating that the motif surrounding Y459 is ITIM. The presence of ITIM was also supported by showing that the FcgammaRIIB-BGPa-mediated inhibitory effect was reduced in SHP-1and SHP-2 mutant DT40 B cells and further diminished in a SHP-1/-2 double-deficient mutant line. The results suggest that SHP-1 and SHP-2 are required for the FcgammaRIIB-BGPa-mediated inhibitory signals.

Dendritic cells (DC) are important APCs that play a key role in the induction of an immune response. The signaling molecules that govern early events in DC activation are not well understood. We therefore investigated whether DC express carcinoembryonic Ag-related cell adhesion molecule 1 (CEACAM1, also known as BGP or CD66a), a well-characterized signal-regulating cell-cell adhesion molecule that is expressed on granulocytes, monocytes, and activated T cells and B cells. We found that murine DC express in vitro as well as in vivo both major isoforms of CEACAM1, CEACAM1-L (having a long cytoplasmic domain with immunoreceptor tyrosine-based inhibitory motifs) and CEACAM1-S (having a short cytoplasmic domain lacking phosphorylatable tyrosine residues). Ligation of surface-expressed CEACAM1 on DC with the specific mAb AgB10 triggered release of the chemokines macrophage inflammatory protein 1alpha, macrophage inflammatory protein 2, and monocyte chemoattractant protein 1 and induced migration of granulocytes, monocytes, T cells, and immature DC. Furthermore, the surface expression of the costimulatory molecules CD40, CD54, CD80, and CD86 was increased, indicating that CEACAM1-induced signaling regulates early maturation and activation of dendritic cells. In addition, signaling via CEACAM1 induced release of the cytokines IL-6, IL-12 p40, and IL-12 p70 and facilitated priming of naive MHC II-restricted CD4(+) T cells with a Th1-like effector phenotype. Hence, our results show that CEACAM1 is a signal-transducing receptor that can regulate early maturation and activation of DC, thereby facilitating priming and polarization of T cell responses.

Phagocytosis of Opa+ Neisseria gonorrhoeae (gonococcus, GC) by neutrophils is in part dependent on the interaction of Opa proteins with CGM1a (CEACAM3/CD66d) antigens, a neutrophil-specific receptor. However, the signaling pathways leading to phagocytosis have not been characterized. Here we show that interaction of OpaI bacteria with neutrophils or CGM1a-transfected DT40 cells induces calcium flux, which correlates with phagocytosis of bacteria. We identified an immunoreceptor tyrosine-based activation motif (ITAM) in CGM1a, and showed that the ability of CGM1a to transduce signals and mediate phagocytosis was abolished by mutation of the ITAM tyrosines. We also demonstrated that CGM1a-ITAM-mediated bacterial phagocytosis is dependent on Syk and phospholipase C activity in DT40 cells. Unexpectedly, the activation of the CGM1a-ITAM phagocytic pathway by Opa+ GC results in induction of cell death.

Background: Carcinoembryonic antigen (CEA) is one of the most widely used tumor markers worldwide. Its main application is mostly in gastrointestinal cancers, especially in colorectal malignancy. Although in use for almost 30 years, the clinical value of CEA in colorectal cancer is still not clear., Methods: The literature relevant to the clinical value of CEA in colorectal cancer was reviewed. Particular attention was paid to studies involving metaanalyses and guidelines issued by Expert Panels., Results: Although of little use in detecting early colorectal cancer, high preoperative concentrations of CEA correlate with adverse prognosis. Serial CEA measurements can detect recurrent colorectal cancer with a sensitivity of approximately 80%, a specificity of approximately 70%, and can provide a lead time of approximately 5 months. CEA is the most frequent indicator of recurrence in asymptomatic patients and currently is the most cost-effective test for the preclinical detection of resectable disease. CEA is most useful for the early detection of liver metastasis in patients with diagnosed colorectal cancer. Overall, however, little evidence is available that monitoring of all patients with diagnosed colorectal cancer leads to enhanced patient outcome or quality of life., Conclusions: Currently, the most useful application of CEA is in the detection of liver metastasis from colorectal cancers. Because of the relative success of surgery in resecting hepatic metastases, serial determinations of the marker are recommended for detecting cancer spread to the liver. In the future, preoperative concentrations of CEA may be included with the standard staging procedures for assessing prognosis.

Carcinoembryonic antigen (CEA) has been reported to promote the metastatic potential in some experimental tumors. Adhesion molecules are known to play an important role in the process of metastasis. Cytokines, including interleukin 1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha), which are produced by Kupffer cells, induce endothelial cells to express adhesion molecules. As a result, the present study was designed to investigate whether the interaction between CEA and Kupffer cells accelerated the metastatic potential of tumors in the liver. Kupffer cells isolated from the liver of male BALB/c mice were cultured with CEA, either with or without the addition of a cytokine inhibitor. The levels of IL-1beta and TNF-alpha were examined in a culture medium. An adhesion assay of colon cancer cell lines to human umbilical vein endothelial cells was also performed. When CEA was added to the Kupffer cell culture medium, cytokines were produced. Elevated levels of cytokines appeared to lead to increased rates of adhesion of cancer cells to endothelial cells. However, these phenomena were blocked by the addition of cytokine inhibitors. CEA stimulated Kupffer cells to produce cytokines. An elevated number of cytokines have been proven to promote the expression of adhesion molecules in endothelial cells. These processes are therefore considered to contribute to the metastasis of malignant cells to the liver. These results suggest that cytokine inhibitors may therefore play an important role in the inhibition of hepatic metastasis.

The domain(s) responsible for the specific heterophilic adhesion between two members of the carcinoembryonic antigen (CEA) family, CEACAM6 and CEACAM8, both of which with three extracellular domains, were investigated using Chinese hamster ovary (CHO) transfectants expressing chimeric antigens. Using a chimeric antigen in which the N-domain, a sole extracellular domain, of CEACAM3 was substituted with that of CEACAM6, it was shown that the N-domain of CEACAM6 alone was able to mediate specific adhesion to CEACAM8. Furthermore, the chimeric antigen was shown to bind significantly to chimeric CEA whose N-domain was substituted with that of CEACAM8, but not to unsubstituted CEA. These results demonstrate that the N-domain alone is sufficient and other domains of CEACAM6 or CEACAM8 are not required for this specific binding. We therefore propose a model of heterophilic interaction between the N-domains, which is distinct from that of CEA-CEA homophilic binding., (Copyright 2000 Academic Press.)

The therapeutic index of cancer gene therapy approaches will, at least in part, be dictated by the spatial and temporal control of expression of the therapeutic transgenes. Strategies which allow precise control of gene transcription are likely to play a crucial role in the future pre-clinical and clinical development of gene therapy. In this review, we discuss these issues as they relate to tissue and tumor specific promoters. In addition, the exciting opportunities offered by the development of regulated gene expression systems using small molecules, radiation and heat are reviewed. It is realistic to expect that the future offers the prospect of amalgamating elements of a number of these different systems in a co-ordinated gene delivery approach with the potential to increase the efficacy and reduce the toxicity of treatment.

Context: The "Standards, Options and Recommendations" (SOR) project, started in 1993, is a collaboration between the Federation of the French Cancer Centres (FNCLCC), the 20 French Cancer Centres and specialists from French Public Universities, General Hospitals and Private Clinics. The main objective is the development of clinical practice guidelines to improve the quality of health care and outcome for cancer patients. The methodology is based on literature review and critical appraisal by a multidisciplinary group of experts, with feedback from specialists in cancer care delivery., Objectives: To define, according to the definitions of the Standards, Options and Recommendations project, the characteristics of various tumour markers in breast cancer and the potential role of these markers in the management of patients with this malignancy., Methods: Data were identified by searching Medline and the personal reference lists of members of the expert groups. Once the guidelines were defined, the document was submitted for review to 43 independent reviewers, and to the medical committees of the 20 French Cancer Centres., Results: The main recommendations are: 1) CA 15.3 and CEA are the serum tumour markers most often used in breast cancer (standard). 2) If the CA 15.3 is raised at presentation, there is no place for the measurement of other tumour markers (standard, expert agreement). 3) All analyses for each patient must be performed in the same laboratory, using the same technique (standard, expert agreement). 4) CA 15.3 should not be used for screening or diagnosis. 5) The level of CA 15.3 before treatment is a recognised prognostic factor, the independent value of which has not been proven (standard, level of evidence C). 6) If the initial value of CA 15.3 is greater than 50 kU.L(-1), disseminated disease should be actively sought before any treatment decisions are made (standard, expert agreement). 7) An initial elevation of CA 15.3 that does not return to normal, reflects a lack of response to treatment and is a strong adverse prognostic factor (standard, level of evidence C). 8) The accuracy of tumours markers (especially CA 15.3) as early indicators of metastatic disease is well recognised (standard) but the clinical benefit has not been established. 9) There is a correlation between tumour markers and clinical response in the treatment of metastatic disease (level of evidence C). The level of CA 15.3 in metastatic disease does not predict response to treatment.