IRF7 Exacerbates Candida albicans Infection by Compromising CD209-Mediated Phagocytosis and Autophagy-Mediated Killing in Macrophages.

IFN regulatory factor 7 (IRF7) exerts anti-infective effects by promoting the production of IFNs in various bacterial and viral infections, but its role in highly morbid and fatal Candida albicans infections is unknown. We unexpectedly found that Irf7 gene expression levels were significantly upregulated in tissues or cells after C. albicans infection in humans and mice and that IRF7 actually exacerbates C. albicans infection in mice independent of its classical function in inducing IFNs production. Compared to controls, Irf7-/- mice showed stronger phagocytosis of fungus, upregulation of C-type lectin receptor CD209 expression, and enhanced P53-AMPK-mTOR-mediated autophagic signaling in macrophages after C. albicans infection. The administration of the CD209-neutralizing Ab significantly hindered the phagocytosis of Irf7-/- mouse macrophages, whereas the inhibition of p53 or autophagy impaired the killing function of these macrophages. Thus, IRF7 exacerbates C. albicans infection by compromising the phagocytosis and killing capacity of macrophages via regulating CD209 expression and p53-AMPK-mTOR-mediated autophagy, respectively. This finding reveals a novel function of IRF7 independent of its canonical IFNs production and its unexpected role in enhancing fungal infections, thus providing more specific and effective targets for antifungal therapy.

DHX9 contributes to the malignant phenotypes of colorectal cancer via activating NF-κB signaling pathway.

Colorectal cancer (CRC) is the leading cause of cancer-related mortality worldwide, which makes it urgent to identify novel therapeutic targets for CRC treatment. In this study, DHX9 was filtered out as the prominent proliferation promoters of CRC by siRNA screening. Moreover, DHX9 was overexpressed in CRC cell lines, clinical CRC tissues and colitis-associated colorectal cancer (CAC) mouse model. The upregulation of DHX9 was positively correlated with poor prognosis in patients with CRC. Through gain- and loss-of function experiments, we found that DHX9 promoted CRC cell proliferation, colony formation, apoptosis resistance, migration and invasion in vitro. Furthermore, a xenograft mouse model and a hepatic metastasis mouse model were utilized to confirm that forced overexpression of DHX9 enhanced CRC outgrowth and metastasis in vivo, while DHX9 ablation produced the opposite effect. Mechanistically, from one aspect, DHX9 enhances p65 phosphorylation, promotes p65 nuclear translocation to facilitate NF-κB-mediated transcriptional activity. From another aspect, DHX9 interacts with p65 and RNA polymerase II (RNA Pol II) to enhance the downstream targets of NF-κB (e.g., Survivin, Snail) expression to potentiate the malignant phenotypes of CRC. Together, our results suggest that DHX9 may be a potential therapeutic target for prevention and treatment of CRC patients.

In vivo characterization of emerging SARS-CoV-2 variant infectivity and human antibody escape potential.

As severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spreads, variants with enhanced virulence and transmissibility have emerged. Although in vitro systems allow rapid characterization, they do not fully recapitulate the dynamic interaction of virions and neutralizing antibodies in the airway. Here, we demonstrate that the N501Y variant permits respiratory infection in unmodified mice. We utilize N501Y to survey in vivo pseudovirus infection dynamics and susceptibility to reinfection with the L452R (Los Angeles), K417N + E484K (South Africa), and L452R + K417N + E484Q (India) variants. Human coronavirus disease 2019 (COVID-19)+ or vaccinated antibody isotypes, titers, variant receptor binding domain (RBD) binding, and neutralization potential are studied, revealing numerous significant correlations. Immune escape of the K417N + E484K variant is observed because infection can be appreciated in the nasopharynx, but not lungs, of mice transferred with low-antibody-tier plasma. Conversely, near-complete protection is observed in animals receiving high-antibody-tier plasma, a phenomenon that can only be appreciated in vivo.

Protein detection in blood with single-molecule imaging.

The ability to characterize individual biomarker protein molecules in patient blood samples could enable diagnosis of diseases at an earlier stage, when treatment is typically more effective. Single-molecule imaging offers a promising approach to accomplish this goal. However, thus far, single-molecule imaging methods have not been translated into the clinical setting. The detection limit of these methods has been confined to the picomolar (10-12 M) range, several orders of magnitude higher than the circulating concentrations of biomarker proteins present in many diseases. Here, we describe single-molecule augmented capture (SMAC), a single-molecule imaging technique to quantify and characterize individual protein molecules of interest down to the subfemtomolar (<10-15 M) range. We demonstrate SMAC in a variety of applications with human blood samples, including the analysis of disease-associated secreted proteins, membrane proteins, and rare intracellular proteins. SMAC opens the door to the application of single-molecule imaging in noninvasive disease profiling.

Albumin fusion with granulocyte-macrophage colony-stimulating factor acts as an immunotherapy against chronic tuberculosis.

A long duration of treatment and emerging drug resistance pose significant challenges for global tuberculosis (TB) eradication efforts. Therefore, there is an urgent need to develop novel strategies to shorten TB treatment regimens and to treat drug-resistant TB. Using an albumin-fusion strategy, we created a novel albumin-fused granulocyte-macrophage colony-stimulating factor (albGM-CSF) molecule that harnesses albumin's long half-life and targeting abilities to enhance the biostability of GM-CSF and direct it to the lymph nodes, where the effects of GM-CSF can increase dendritic cell populations crucial for eliciting a potent immune response. In this study, we demonstrate that albGM-CSF serves as a novel immunotherapy for chronic Mycobacterium tuberculosis (Mtb) infections by enhancing GM-CSF biostability in serum. Specifically, albumin is very safe, stable, and has a long half-life, thereby enhancing the biostability of GM-CSF. In the lungs and draining lymph nodes, albGM-CSF is able to increase the numbers of dendritic cells, which are crucial for the activation of naive T cells and for eliciting potent immune responses. Subcutaneous administration of albGM-CSF alone reduced the mean lung bacillary burden in mice with chronic tuberculosis infection. While GM-CSF administration was associated with IL-1ß release from Mtb-infected dendritic cells and macrophages, higher IL-1ß levels were observed in albGM-CSF-treated mice with chronic tuberculosis infection than in mice receiving GM-CSF. Albumin fusion with GM-CSF represents a promising strategy for the control of chronic lung tuberculosis infections and serves as a novel therapeutic vaccination platform for other infectious diseases and malignancies.

DNA sensors, crucial receptors to resist pathogens, are deregulated in colorectal cancer and associated with initiation and progression of the disease.

Background: DNA sensors are innate immune receptors that detect intracellular endogenous or exogenous DNA. They are critical to trigger immune response against DNA viral and intracellular bacterial infection, and are involved in inflammatory diseases and tumorigenesis. Recent accumulating evidences indicated that DNA sensors are also crucial for controlling the development of colorectal cancer (CRC). However, a systematic study on the expression profile of DNA sensors in CRC and their clinical significance are still lacking. Methods: We investigated the expression profile of DNA sensors in CRC and their clinical significance by taking advantage of clinical CRC samples, mouse AOM/DSS treatment model, and Oncomine ® bioinformatics platform. Results: Our study identified that the expression of DNA sensors, including AIM2, DAI, as well as inflammasome molecules ASC/IL-18, TLR9 and adaptor MyD88, and DDX60 decreased in human CRC, whereas the expression of DHX9, DHX36, and DDX41 significantly increased. Among them, the expression of AIM2/ASC/IL-18, MyD88, DAI, DHX36, and DDX60 were associated with cancer stages. In addition, we also performed correlation analysis between DNA sensors and their main signaling molecules to explore the possible mechanisms. The results showed that there were positive correlations between AIM2 and ASC/IL-18, DHX9 and MAVS, and TLR9 and MyD88 expression. In addition, the gene expression patterns of some DNA sensors were confirmed by Western-blot analysis. Conclusions: Our study revealed that the expression of multiple DNA sensors was deregulated in CRC and might be involved in tumor development. More importantly, the study identified that, among all these DNA sensors, AIM2, DAI, and DDX60 could be potentially critical for diagnosis, prognosis, and therapy of CRC and deserve further investigation.

The expression profile of RNA sensors in colorectal cancer and its correlation with cancer stages.

BACKGROUND: RNA sensors represent the most important pattern recognition receptors (PRRs) to defend against RNA pathogens, such as RNA viruses. Recent studies revealed their critical roles in inflammatory and autoimmune diseases. Furthermore, more recent evidences indicated that RNA sensors mediate the development of colitis or colorectal cancer (CRC). However, a systematic understanding of RNA sensors in CRC is still lacking, especially the expression patterns in CRC. METHODS: Here, we analyzed RNA sensor expression, clinical significance, and possible mechanisms in CRC by combining bioinformatic analysis and the analysis on pre-cancerous animal model and clinical tissue samples. RESULTS: We found that most of the members of RNA sensors, including RNA-sensing Toll-like receptors (TLR3, TLR7, and TLR8) and RIG-I-like receptors (MDA5 and RIG-I), were down-regulated in CRC, while the expression of DDX21 were up-regulated in human CRC. In addition, we also analyzed the correlation between gene expression and cancer stages. We found that the expression of RNA-sensing TLRs, RIG-I, and DDX21 in CRC were associated with cancer stages. Finally, in order to explore the possible mechanisms, the correlation between RNA sensors and the main downstream signaling molecules were analyzed. A positive correlation was observed in TLR7/8 and MyD88, RIG-I/MDA5 and LGP2, while a negative correlation was observed in RIG-I/MDA5 and MAVS. CONCLUSIONS: This study reveals the potential values of RNA-sensing genes including TLRs, RIG-I and DDX21 as biomarkers of CRC formation, progression and therapy.

Effects of the intestinal microbial metabolite butyrate on the development of colorectal cancer.

Colorectal cancer (CRC) is one of the major health threats in developed countries. Changes in dietary components, such as more protein and lipid intake, can increase the risk of CRC. Diet affects CRC in many ways. They regulate the composition and function of gut microbiota, which have an amazing metabolic capacity and can produce short chain fatty acids (SCFAs), such as propionate, acetate, and butyrate. Butyrate is a principal energy source for colonic epithelial cells and plays an important role in maintaining the stability of gut microbiota and the integrity of intestinal epithelium. However, there are few studies reviewing the anti-CRC potentials of butyrate. This review summarizes the recent research progresses in the effect of gut microbiota imbalance and the decrease in intestinal microbial metabolite butyrate caused by unbalanced diet on CRC development, and discusses the mechanisms of butyrate-induced anti-CRC activities, which may guide people to prevent CRC by improving diet structures.

Programmed self-assembly of peptide-major histocompatibility complex for antigen-specific immune modulation.

A technology to prime desired populations of T cells in the body-particularly those that possess low avidity against target antigen-would pave the way for the design of new types of vaccination for intractable infectious diseases or cancer. Here, we report such a technology based on positive feedback-driven, programmed self-assembly of peptide-major histocompatibility complex (pMHC) directly on the membrane of cognate T cells. Our design capitalizes on the unique features of the protein annexin V (ANXA5), which-in a concerted and synergistic manner-couples the early onset of TCR signaling by cognate pMHC with a surge in pMHC-TCR affinity, with repeated pMHC encounters, and with widespread TCR cross-linking. In our system, ANXA5 is linked to pMHC and firmly engages the plasma membrane of cognate T cells upon (and only upon) the early onset of TCR signaling. ANXA5, in turn, exerts a mechanical force that stabilizes interactions at the TCR-pMHC interface and facilitates repeated, serial pMHC encounters. Furthermore, ANXA5 quickly arranges into uniform 2D matrices, thereby prompting TCR cross-linking. Fusion of ANXA5 to pMHC augments lymphocyte activation by several orders of magnitude (>1,000-fold), bypasses the need for costimulation, and breaks tolerance against a model self-antigen in vivo. Our study opens the door to the application of synthetic, feedback-driven self-assembly platforms in immune modulation.

The research status of immune checkpoint blockade by anti-CTLA4 and anti-PD1/PD-l1 antibodies in tumor immunotherapy in China: A bibliometrics study.

PURPOSE: Using bibliometrics, we analyzed the research status of immune checkpoint blockade (ICB, a popular tumor immunotherapy method represented by antibodies targeted CTLA-4 and PD-1/PD-L1) in tumor immunotherapy in China during the past 2 decades. METHODS: Articles in Science Citation Index Expanded (SCI-EXPANDED), patents in Thomson Innovation, and drugs in Cortellis Competitive Intelligence in the field of ICB for tumor immunotherapy from 1996 to 2015 were the subjects of bibliometric analysis. Using database-attached software and Excel, quantitative analyses were performed including examination of the number of documents, citation frequency, h-index, key projects, quantity of publications, public patents, and status of new drug research. RESULTS: The number of publications from 1996 to 2015 in the field of ICB for tumor immunotherapy that came out of China was 380, which was 14.3% of the total publications worldwide and was second only to that of the USA. In the past decade, China has rapidly increased the number of publications and patents in this field. However, indicators of publication influence, such as citation frequency and h-index, were far behind other advanced countries. In addition, the total number of patents in China was much lower than that of the USA. China has introduced 5 drugs for ICB that are being developed for the healthcare market. CONCLUSION: Tumor immunotherapy research such as ICB in China has developed rapidly with increasing influence in the last 2 decades. However, there is still a relatively large gap compared with the USA. It is expected that China will have greater influence on tumor immunotherapy research in the near future.

Dock2 in the development of inflammation and cancer.

An atypical guanine exchange factor, Dock2 is specifically expressed in hematopoietic cells and regulates activation and migration of immune cells through activating Ras-related C3 botulinum toxin substrate (Rac). Dock2 was shown to be critical in the development of various inflammatory diseases, including allergic diseases, HIV infection, and graft rejection in organ transplantation. DOCK2 mutation in infants was recently identified to be associated with T and B cell combined immunodeficiency. Furthermore, Dock2 is involved in host protection during enteric bacterial infection and is also associated with the proliferation of cancer cells. It was also shown that patients with digestive tract cancer had high frequency mutation of DOCK2. This review summarizes the latest research progresses on the role of Dock2 for the development of various inflammatory diseases and cancers, and discusses the potential application of Dock2 modulators for patient treatment.

Anti-inflammatory and antioxidative effects of Camellia oleifera Abel components.

Camellia oleifera Abel is a member of Camellia, and its seeds are used to extract Camellia oil, which is generally used as cooking oil in the south of China. Camellia oil consists of unsaturated fatty acids, tea polyphenol, squalene, saponin, carrot element and vitamins, etc. The seed remains after oil extraction of C. oleifera Abel are by-products of oil production, named as Camellia oil cake. Its extracts contain bioactive compounds including sasanquasaponin, flavonoid and tannin. Major components from Camellia oil and its cake have been shown to have anti-inflammatory, antioxidative, antimicrobial and antitumor activities. In this review, we will summarize the latest advance in the studies on anti-inflammatory or antioxidative effects of C. oleifera products, thus providing valuable reference for the future research and development of C. oleifera Abel.

STING signaling in tumorigenesis and cancer therapy: A friend or foe?

Stimulator of interferon genes (STING) is a DNA sensor and an important cytoplasmic adaptor for other DNA sensors, such as Z-DNA binding protein 1 (DAI), DEAD-box helicase 41 (DDX41), and interferon-γ-inducible protein 16 (IFI16). The activation of STING signaling leads to the production of type I interferons and some other pro-inflammatory cytokines, which are critical for host defense against viral infection. Recent accumulating evidences suggest that STING is also involved in tumor development. However, the role of STING signaling in tumorigenesis is complicated, and a comprehensive review is still lacking. In this paper, we provided an overview of the dual role of STING signaling in tumor development from clinical significance to fundamental mechanisms, as well as its pre-clinical application in cancer therapy.

Nucleic acid sensing pattern recognition receptors in the development of colorectal cancer and colitis.

Colorectal cancer (CRC) is a leading cause of cancer-related deaths that is often associated with inflammation initiated by activation of pattern recognition receptors (PRRs). Nucleic acid sensing PRRs are one of the major subsets of PRRs that sense nucleic acid (DNA and RNA), mainly including some members of Toll-like receptors (TLR3, 7, 8, 9), AIM2-like receptors (AIM2, IFI16), STING, cGAS, RNA polymerase III, and DExD/H box nucleic acid helicases (such as RIG-I like receptors (RIG-I, MDA5, LPG2), DDX1, 3, 5, 7, 17, 21, 41, 60, and DHX9, 36). Activation of these receptors eventually leads to the release of cytokines and activation of immune cells, which are well known to play crucial roles in host defense against intracellular bacterial and virus infection. However, the functions of these nucleic acid sensing PRRs in the other diseases such as CRC and colitis remain largely unknown. Recent studies indicated that nucleic acid sensing PRRs contribute to CRC and/or colitis development, and therapeutic modulation of nucleic acid sensing PRRs may reduce the risk of CRC development. However, until now, a comprehensive review on the role of nucleic acid sensing PRRs in CRC and colitis is still lacking. This review provided an overview of the roles as well as the mechanisms of these nucleic acid sensing PRRs (AIM2, STING, cGAS, RIG-I and its downstream molecules, DDX3, 5, 6,17, and DHX9, 36) in CRC and colitis, which may aid the diagnosis, therapy, and prognostic prediction of CRC and colitis.

RPN13/ADRM1 inhibitor reverses immunosuppression by myeloid-derived suppressor cells.

Myeloid-derived-suppressor cells (MDSCs) are key mediators of immune suppression in the ovarian tumor microenvironment. Modulation of MDSC function to relieve immunosuppression may enhance the immunologic clearance of tumors. The bis-benzylidine piperidone RA190 binds to the ubiquitin receptor RPN13/ADRM1 on the 19S regulatory particle of the proteasome and directly kills ovarian cancer cells by triggering proteotoxic stress. Here we examine the effect of RA190 treatment on the immunosuppression induced by MDSCs in the tumor microenvironment, specifically on the immunosuppression induced by MDSCs. We show that RA190 reduces the expression of Stat3 and the levels of key immunosuppressive enzymes and cytokines arginase, iNOS, and IL-10 in MDSCs, while boosting expression of the immunostimulatory cytokine IL-12. Furthermore, we show that the RA190-treated MDSCs lost their capacity to suppress CD8+ T cell function. Finally, we show that RA190 treatment of mice bearing syngeneic ovarian tumor elicits potent CD8+ T cell antitumor immune responses and improves tumor control and survival. These data suggest the potential of RA190 for ovarian cancer treatment by both direct killing of tumor cells via proteasome inhibition and relief of MDSC-mediated suppression of CD8 T cell-dependent antitumor immunity elicited by the apoptotic tumor cells.

Buccal injection of synthetic HPV long peptide vaccine induces local and systemic antigen-specific CD8+ T-cell immune responses and antitumor effects without adjuvant.

BACKGROUND: Human Papillomavirus is responsible for over 99 % of cervical cancers and is associated with cancers of the head and neck. The currently available prophylactic vaccines against HPV do not generate therapeutic effects against established HPV infections and associated lesions and disease. Thus, the need for a therapeutic vaccine capable of treating HPV-induced malignancies persists. Synthetic long peptides vaccination is a popular antigen delivery method because of its safety, stability, production feasibility, and its need to be processed by professional antigen presenting cells before it can be presented to cytotoxic CD8+ T lymphocytes. Cancers in the buccal mucosa have been shown to elicit cancer-related inflammations that are capable of recruiting inflammatory and immune cells to generate antitumor effects. In the current study, we evaluated the therapeutic potential of synthetic HPV long peptide vaccination in the absence of adjuvant in the TC-1 buccal tumor model. RESULT: We show that intratumoral vaccination with E7 long peptide alone effectively controls buccal TC-1 tumors in mice. Furthermore, we observed an increase in systemic as well as local E7-specific CD8+ T cells in buccal tumor-bearing mice following the vaccination. Finally, we show that induction of immune responses against buccal tumors by intratumoral E7 long peptide vaccination is independent of CD4+ T cells, and that the phenomenon may be related to the unique environment associated with mucosal tissues. CONCLUSION: Our results suggest the possibility for clinical translation of the administration of adjuvant free therapeutic long peptide vaccine as a potentially effective and safe strategy for mucosal HPV-associated tumor treatment.

Toll-like receptor agonist imiquimod facilitates antigen-specific CD8+ T-cell accumulation in the genital tract leading to tumor control through IFNγ.

PURPOSE: Imiquimod is a Toll-like receptor 7 agonist used topically to treat external genital warts and basal cell carcinoma. We examined the combination of topical imiquimod with intramuscular administration of CRT/E7, a therapeutic human papillomavirus (HPV) vaccine comprised of a naked DNA vector expressing calreticulin fused to HPV16 E7. EXPERIMENTAL DESIGN: Using an orthotopic HPV16 E6/E7(+) syngeneic tumor, TC-1, as a model of high-grade cervical/vaginal/vulvar intraepithelial neoplasia, we assessed if combining CRT/E7 vaccination with cervicovaginal deposition of imiquimod could result in synergistic activities promoting immune-mediated tumor clearance. RESULTS: Imiquimod induced cervicovaginal accumulation of activated E7-specific CD8(+) T cells elicited by CRT/E7 vaccination. Recruitment was not dependent upon the specificity of the activated CD8(+) T cells, but was significantly reduced in mice lacking the IFNγ receptor. Intravaginal imiquimod deposition induced upregulation of CXCL9 and CXCL10 mRNA expression in the genital tract, which are produced in response to IFNγ receptor signaling and attract cells expressing their ligand, CXCR3. The T cells attracted by imiquimod to the cervicovaginal tract expressed CXCR3 as well as CD49a, an integrin involved in homing and retention of CD8(+) T cells at mucosal sites. Our results indicate that intramuscular CRT/E7 vaccination in conjunction with intravaginal imiquimod deposition recruits antigen-specific CXCR3(+) CD8(+) T cells to the genital tract. CONCLUSIONS: Several therapeutic HPV vaccination clinical trials using a spectrum of DNA vaccines, including vaccination in concert with cervical imiquimod, are ongoing. Our study identifies a mechanism by which these strategies could provide therapeutic benefit. Our findings support accumulating evidence that manipulation of the tumor microenvironment can enhance the therapeutic efficacy of strategies that induce tumor-specific T cells.

Direct T cell activation via CD40 ligand generates high avidity CD8+ T cells capable of breaking immunological tolerance for the control of tumors.

CD40 and CD40 ligand (CD40L) are costimulatory molecules that play a pivotal role in the proinflammatory immune response. Primarily expressed by activated CD4+ T cells, CD40L binds to CD40 on antigen presenting cells (APCs), thereby inducing APC activation. APCs, in turn, prime cytotoxic T lymphocytes (CTLs). Here, two tumor-associated antigen (TAA) animal models, p53-based and GP100-based, were utilized to examine the ability of CD40-CD40L to improve antigen-specific CTL-mediated antitumor immune responses. Although p53 and GP100 are self-antigens that generate low affinity antigen-specific CD8+ T cells, studies have shown that their functional avidity can be improved with CD40L-expressing APCs. Therefore, in the current study, we immunized mice with a DNA construct encoding a TAA in conjunction with another construct encoding CD40L via intramuscular injection followed by electroporation. We observed a significant increase in the antigen-specific CTL-mediated immune responses as well as the potent antitumor effects in both models. Antibody depletion experiments demonstrated that CD8+ T cells play a crucial role in eliciting antitumor effects in vaccinated mice. Furthermore, we showed that in vitro stimulation with irradiated tumor cells expressing both TAA and CD40L improved the functional avidity of antigen-specific CD8+ T cells. Thus, our data show that vaccination with TAA/CD40L DNA can induce potent antitumor effects against TAA-expressing tumors through the generation of better functioning antigen-specific CD8+ T cells. Our study serves as an important foundation for future clinical translation.

Control of spontaneous ovarian tumors by CD8+ T cells through NKG2D-targeted delivery of antigenic peptide.

BACKGROUND: There is an urgent need to develop targeted therapies for the control of advanced stage ovarian cancer because it is the most deadly gynecologic cancer. Antigen-specific immunotherapy is a promising approach because of the potential of the immune system to specifically target tumors without the toxicity associated with traditional chemoradiation. However, one of the major limitations for antigen-specific cancer immunotherapy is the pre-existing immune tolerance against endogenous targeted tumor antigens that frequently evolves during carcinogenesis. Here, we described the creation of a therapeutic agent comprised of a tumor-homing module fused to a functional domain capable of selectively rendering tumor cells sensitive to foreign antigen-specific CD8+ T cell-mediated immune attack, thereby circumventing many aspects of immune tolerance. The tumor-homing module, NKG2D, specifically binds to NKG2D ligand that is commonly overexpressed in ovarian tumors. The functional domain is comprised of the Fc portion of IgG2a protein and foreign immunogenic CD8+ T cell epitope flanked by furin cleavage sites (R), which can be recognized and cleaved by furin that is highly expressed in the tumor microenvironment. RESULTS: We show that this therapeutic chimeric protein specifically loaded antigenic epitope onto the surface of NKG2D ligand-expressing ovarian tumor cells, rendering ovarian tumors susceptible to antigen-specific CTL-mediated killing in vitro. Furthermore, we show that intraperitoneal administration of our therapeutic chimeric protein followed by adoptive transfer of antigen-specific CD8+ T cells generates potent antitumor effects and significant accumulation of antigen-specific CD8+ T cells in the tumor loci. CONCLUSIONS: Our findings have promise for bypassing immune tolerance to enhance cancer immunotherapy.

Creation of a Merkel cell polyomavirus small T antigen-expressing murine tumor model and a DNA vaccine targeting small T antigen.

BACKGROUND: Merkel cell polyomavirus (MCPyV) is a DNA virus expressing transcripts similar to the large T (LT) and small T (ST) transcripts of SV40, which has been implicated in the pathogenesis of Merkel cell carcinoma (MCC), a rare and highly aggressive neuroendocrine skin cancer. MCPyV LT antigen expression was found to be a requirement for MCC tumor maintenance and ST protein also likely contributes to the carcinogenesis of MCC. Previously, we have identified the probable immunodominant epitope of MCPyV LT and developed a DNA vaccine encoding this epitope linked to calreticulin. The LT-targeting DNA vaccine generated prolonged survival, decreased tumor size and increased LT-specific CD8+ T cells in tumor-bearing mice. RESULTS: In this study, we developed a MCPyV ST-expressing tumor cell line from B16 mouse melanoma cells. We then utilized this ST-expressing tumor cell line to test the efficacy of a DNA vaccine encoding ST. In ST-expressing tumor-bearing mice, this vaccine, pcDNA3-MCC/ST, generated a significant number of ST antigenic peptide-specific CD8+ T cells and experienced markedly enhanced survival compared to mice vaccinated with empty vector. CONCLUSIONS: The formation of an effective vaccine against MCPyV has the potential to advance the field of MCC therapy and may contribute to the control of this severe malignancy through immunotherapy. Both of the innovative technologies presented here provide opportunities to develop and test MCPyV-targeted therapies for the control of Merkel cell carcinoma.