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To date, IgG in the tumor microenvironment (TME) has been considered a product of B cells and serves as an antitumor antibody. However, in this study, using a monoclonal antibody against cancer-derived IgG (Cancer-IgG), we found that cancer cells could secrete IgG into the TME. Furthermore, Cancer-IgG, which carries an abnormal sialic acid modification in the CH1 domain, directly inhibited effector T-cell proliferation and significantly promoted tumor growth by reducing CD4(+) and CD8(+) T-cell infiltration into tumor tissues. Mechanistic studies showed that the immunosuppressive effect of sialylated Cancer-IgG is dependent on its sialylation and binding to sialic acid-binding immunoglobulin-type lectins (Siglecs) on effector CD4(+) and CD8(+) T cells. Importantly, we show that several Siglecs are overexpressed on effector T cells from cancer patients, but not those from healthy donors. These findings suggest that sialylated Cancer-IgG may be a ligand for Siglecs, which may serve as potential checkpoint proteins and mediate tumor immune evasion.

Immunoglobulin (Ig&kappa, ) has been reported to be expressed in sorted liver epithelial cells of &mu, MT mice, and the sequence characteristics of hepatocyte-derived Ig&kappa, were different from those of classical B-cell-derived Ig&kappa, However, the physiological function of hepatocyte-derived Ig&kappa, is still unclear. The expression of Ig&kappa, was firstly identified in primary hepatocytes and normal liver cell line (NCTC1469), and hepatocyte-derived Ig&kappa, expression was elevated and displayed unique localization in hepatocytes of concanavalin A (ConA)-induced hepatitis model. Moreover, Ig&kappa, knockout mice were more sensitive to ConA-induced hepatitis and had higher serum aspartate aminotransferase (AST) levels, more severe histological injury and a greater number of terminal deoxynucleotide transferase-mediated deoxyuridine triphosphate nick end-labeling (TUNEL)-positive cells as compared with littermate controls. Furthermore, knockdown of Ig&kappa, in primary hepatocytes and NCTC1469 cells led to accelerated activation of the mitochondrial death pathway and caspase-3 cleavage in vitro, which might be related to inhibition of NF-&kappa, B signaling pathway and activation of JNK via the cytoskeleton dynamics. Taken together, these results indicate that hepatocyte-derived Ig&kappa, mediates cellular resistance to ConA-induced liver injury by inhibiting activation of caspase-3 and the mitochondrial death pathway, suggesting that Ig&kappa, plays an important role in hepatocyte survival and exerts a protective effect against ConA-induced liver injury in mice.

Data from clinical trials for hemophilia B using adeno-associated virus (AAV) vectors have demonstrated decreased transgenic coagulation factor IX (hFIX) expression 6-10 weeks after administration of a high vector dose. While it is likely that capsid-specific cytotoxic T lymphocytes eliminate vector-transduced hepatocytes, thereby resulting in decreased hFIX, this observation is not intuitively consistent with restored hFIX levels following prednisone application. Although the innate immune response is immediately activated following AAV vector infection via TLR pathways, no studies exist regarding the role of the innate immune response at later time points after AAV vector transduction. Herein, activation of the innate immune response in cell lines, primary human hepatocytes, and hepatocytes in a human chimeric mouse model was observed at later time points following AAV vector transduction. Mechanistic analysis demonstrated that the double-stranded RNA (dsRNA) sensor MDA5 was necessary for innate immune response activation and that transient knockdown of MDA5, or MAVS, decreased IFN-�� expression while increasing transgene production in AAV-transduced cells. These results both highlight the role of the dsRNA-triggered innate immune response in therapeutic transgene expression at later time points following AAV transduction and facilitate the execution of effective strategies to block the dsRNA innate immune response in future clinical trials.

// Qinyuan Liao 1, 2, * , Wei Liu 1, 2, * , Yang Liu 1, 2, * , Fulin Wang 4 , Chong Wang 1, 2 , Jingxuan Zhang 3 , Ming Chu 1, 2 , Dongyang Jiang 1, 2 , Lin Xiao 1, 2 , Wenwei Shao 1, 2 , Zhengzuo Sheng 1 , Xia Tao 5 , Lei Huo 6 , C. Cameron Yin 7 , Youhui Zhang 8 , Gregory Lee 9 , Jing Huang 1, 2 , Zihai Li 10 , Xiaoyan Qiu 1, 2, 3 1 Department of Immunology, School of Basic Medical Sciences, Peking University, Beijing, 100191, China 2 Peking University Center for Human Disease Genomics, Beijing, 100191, China 3 Key Laboratory of Medical Immunology, Ministry of Health, Beijing, 100191, China 4 Department of Pathology, Chinese PLA General Hospital, Beijing, 100853, China 5 Department of Gynecology, Peking University First Hospital, Beijing, 100034, China 6 Division of Pathology and Laboratory Medicine, The University of Texas MD Anderson Cancer Center, Houston, Texas, 77030, USA 7 Department of Hematopathology, The University of Texas MD Anderson Cancer Center, Houston, Texas, 77030, USA 8 Department of Immunology, Cancer Institute & Hospital, Chinese Academy of Medical Science, Beijing, 100021, China 9 Andrology Lab, University of British Columbia Centre for Reproductive Health, Vancouver, BC V5Z 4H4, Canada 10 Department of Microbiology and Immunology, Medical University of South Carolina, Charleston, SC 29425, USA * These authors have contributed equally to this work Correspondence to: Xiaoyan Qiu, e-mail: qiuxy@bjmu.edu.cn Keywords: IgG, RP215, epithelial stem/progenitor-like cells, tumor metastasis Received: July 05, 2015 Accepted: October 02, 2015 Published: October 12, 2015 ABSTRACT High expression of immunoglobulin G (IgG) in many non-B cell malignancies and its non-conventional roles in promoting proliferation and survival of cancer cells have been demonstrated. However, the precise function of non-B IgG remains incompletely understood. Here we define the antigen specificity of RP215, a monoclonal antibody that specifically recognizes the IgG in cancer cells. Using RP215, our study shows that IgG is overexpressed in cancer cells of epithelial lineage, especially cells with cancer stem/progenitor cell-like features. The RP215-recognized IgG is primarily localized on the cell surface, particularly lamellipodia-like structures. Cells with high IgG display higher migration, increased invasiveness and metastasis, and enhanced self-renewal and tumorgenecity ability in vitro and in vivo . Importantly, depletion of IgG in breast cancer leads to reduced adhesion, invasion and self-renewal and increased apoptosis of cancer cells. We conclude that high expression of IgG is a novel biomarker of tumor progression, metastasis and cancer stem cell maintenance and demonstrate the potential therapeutic benefits of RP215-recognized IgG targeted strategy.

CCDC134 is a poorly characterized secreted protein that may act as an immune cytokine. Here, we show that CCDC134 is differentially expressed on resting and activated immune cells and that it promotes CD8+ T-cell activation, proliferation, and cytotoxicity by augmenting expression of the T-cell effector molecules IFNγ, TNFα, granzyme B, and perforin. CCDC134 facilitated infiltration of CD8+ T cells with enhanced cytolytic activity into tumors, demonstrating strong antitumor effects in a CD8+ T-cell–dependent manner. Mechanistically, in CD8+ T cells, exposure to CCDC134 promoted cell proliferation through the JAK3–STAT5 pathway, a classic feature of many cytokines of the common γ-chain (γc) cytokine receptor family. Overall, our results provide evidence that CCDC134 may serve as a member of the γc cytokine family and illustrate its potent antitumor effects by augmenting CD8+ T-cell–mediated immunity. Cancer Res; 74(20); 5734–45. ©2014 AACR.

Background Immunoglobulins(Igs)play an important role in host defence and were initially thought to be expressed solely by B cells. However, recent data suggest that Igs are also expressed in other lineages. Recently, Ig transcripts were detected in cardiomyocytes, but whether the functional Ig protein is expressed by cardiomyocytes has not been thoroughly elucidated. Methods Gene Expression Omnibus (GEO) microarray database analysis was used to analyse IgM heavy chain expression in the myocardium of mice. Immunohistochemistry (IHC), ELISA and Western blot were used to identify IgM in cardiomyocytes of both Balb/c mice and μMT mice (B cell-deficient mice), as well as in cultured cardiomyocytes of neonatal mice and in the myocardial cell line HL-1. Moreover, RT-PCR and cDNA sequencing were used to determine the VDJ rearrangement of the IgM heavy chain. Results In this study, we first analysed transcription of the IgM heavy chain in heart tissue in mice by mining the GEO database, and we observed that IgM heavy chain transcripts were expressed in heart tissues. Subsequently, IgM was found to be expressed in cardiomyocytes in mice; the IgM was primarily localized on the cell membranes and intercalated discs of murine heart cells and in the cytoplasm and cell membranes of isolated cardiomyocytes and HL-1. Importantly, the functional IgM heavy chain transcripts exhibit a unique VDJ rearrangement pattern. Furthermore, IgM can be secreted and deposited in the extracellular space of the myocardium under ischaemic/hypoxic conditions. Conclusions Our data indicate for the first time that IgM is expressed by cardiomyocytes in mice and suggest that its physiological function warrants further investigation.

Numerous studies have demonstrated that free Ig light chain (FLC), a novel inflammation mediator, participates in many inflammatory diseases by activating mast cells and extending the survival of neutrophils. However, it remains unclear whether FLC is involved in colitis and colitis-associated colon carcinogenesis (CAC). In this study, we found a significant increase in FLC in murine models of DSS (Dextran Sulfate Sodium Salt)-induced colitis and CAC compared to controls. Peptide F991, a functional blocker of FLC, significantly attenuated colitis progression, which included abrogating the development of diarrhea and tumor burden, elevating survival rate, greatly reducing the infiltration of inflammatory cells (such as ROS+ active neutrophils), especially reducing tumorigenesis in CAC. Furthermore, we demonstrated that F991 inhibited the activation of the inflammasome by reducing the expression of cleaved caspase-1 and the maturation of IL-1β and IL-18. Altogether, our findings demonstrate that FLC can promote the pathogenesis of colitis and CAC and may be used as novel biomarker for the diagnosis of inflammatory bowel disease. Additionally, F991 may become a potential therapeutic option for colitis or colorectal cancer.

Growing evidence indicates that B cells are not the only source of immunoglobulin (Ig). To investigate this discovery further, we used μMT mice, which have a disruption of the first transmembrane exon of the μ heavy chain and do not express the membrane form of IgM. These mice lack mature B cells and thus serve as a good model to explore Ig expression by liver epithelial cells. We found that Ig heavy chains (μ, δ, γ and α) and light chains (κ and λ) were expressed in sorted liver epithelial cells of μMT mice. Surprisingly, each heavy chain class showed its respective variable region sequence characteristics in their variable region, instead of sharing the same VDJ usage, which suggests that class switching does not occur in liver epithelial cells. Moreover, the γ and α chains, but not the μ and δ chains, showed mutations in the variable region, thus indicating that different classes of Ig have different activities. Our findings support the concept that non-B cells, liver epithelial cells here, can produce different classes of Ig.

Currently, natural IgM antibodies are considered to be the constitutively secreted products of B-1 cells in mice and humans. In this study, we found that mouse epithelial cells, including liver epithelial cells and small intestinal epithelial cells (IECs), could express IgM that also showed natural antibody activity. Moreover, similar to the B-1 cell-derived natural IgM that can be upregulated by TLR9 agonists (mimicking bacterial infection), the expression of epithelial cell-derived natural IgM could also be significantly increased by TLR9 signaling. More importantly, the epithelial cell-derived IgM was polyreactive, and it could recognize single-stranded DNA (ssDNA), double-stranded DNA (dsDNA), lipopolysaccharide (LPS), and insulin with low affinity; additionally, TLR9 agonists could enhance it in a MyD88-dependent manner. Furthermore, epithelial cell-derived IgM could bind various bacteria; therefore, it could be involved in anti-infection responses. Together, these results highlight the fact that epithelial cells are an important source of natural IgM, in addition to that produced by B-1 cells, and IgM contributes to the innate immune responses in local tissues, further demonstrating that the epithelium is a first line of defense in the protection against invading microbes.

Mammalian IgG and IgE are thought to have evolved from IgY of nonmammalian tetrapods; however, no diversification of IgY subclasses has been reported in reptiles or birds, which are phylogenetically close to mammals. To our knowledge, we report the first evidence of the presence of multiple IgY-encoding (υ) genes in snakes. Two υ genes were identified in the snake Elaphe taeniura, and three υ genes were identified in the Burmese python (Python molurus bivittatus). Although four of the υ genes displayed a conventional four-H chain C region exon structure, one of the υ genes in the Burmese python lacked the H chain C region 2 exon, thus exhibiting a structure similar to that of the mammalian γ genes. We developed mouse mAbs specific for the IgY1 and IgY2 of E. taeniura and showed that both were expressed in serum; each had two isoforms: one full-length and one truncated at the C terminus. The truncation was not caused by alternative splicing or transcriptional termination. We also identified the μ and δ genes, but no α gene, in both snakes. This study provides valuable clues for our understanding of Ig gene evolution in tetrapods.