Journal Articles

Immunotherapy design

Novel approaches and designs of biologicals used for cancer immunotherapy as well as changes in the timing, combination sequence, adjuvant choice or route of immunization in immunotherapy regimens; biomarkers of response to therapy

Gastric cancer vaccines synthesized using a TLR7 agonist and their synergistic antitumor effects with 5-fluorouracil

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BACKGROUND: Vaccines play increasingly important roles in cancer treatment due to their advantages of effective targeting and few side effects. Our laboratory has attempted to construct vaccines by conjugating TLR7 agonists with tumor-associated antigens. Furthermore, immunochemotherapy has recently become an appealing approach to cancer therapy. 5-fluorouracil (5-FU), a commonly used chemotherapeutic agent, can reportedly potently and selectively kill tumor-associated MDSCs in vivo. METHODS: Gastric cancer vaccines were synthesized by the covalent attachment of our TLR7 agonist with the gastric cancer antigen MG7-Ag tetra-epitope, leading to T7 - ML (linear tetra-epitope) and T7 - MB (branched tetra-epitope). Cytokines induced by the vaccines in vitro were assessed by ELISA. A tumor challenge model was created by treating BALB/c mice on either a prophylactic or therapeutic vaccination schedule. 5-FU was simultaneously applied to mice in the combination treatment group. CTL and ADCC activities were determined by the LDH method, while CD3(+)/CD8(+), CD3(+)/CD4(+) T cells and MDSCs were evaluated by flow cytometry. RESULTS: In vitro, rapid TNF-alpha and IL-12 inductions occurred in BMDCs treated with the vaccines. In vivo, among all the vaccines tested, T7 - MB most effectively reduced EAC tumor burdens and induced CTLs, antibodies and ADCC activity in BALB/c mice. Immunization with T7 - MB in combination with 5-FU chemotherapy reduced tumor sizes and extended long-term survival rates, mainly by improving T cell responses, including CTLs, CD3(+)/CD8(+) and CD3(+)/CD4(+) T cells. 5-FU also enhanced the T7 - MB efficiency by reversing immunosuppressive factors, i.e., MDSCs, which could not be validly inhibited by the vaccines alone. In addition, T7 - MB repressed tumor growth and immune tolerance when the therapeutic schedule was used, although the effects were weaker than those achieved with either T7 - MB alone or in combination with 5-FU on the prophylactic schedule. CONCLUSIONS: A novel effective gastric cancer vaccine was constructed, and the importance of branched multiple antigen peptides and chemical conjugation to vaccine design were confirmed. The synergistic effects and mechanisms of T7 - MB and 5-FU were also established, observing mainly T cell activation and MDSC inhibition.

Author Info: (1) School of Pharmaceutical Sciences, Shenzhen University Health Science Center, Shenzhen, 518060, Guangdong, China. (2) The 3rd Affiliated Hospital of Shenzhen University, Shenzhen, 518001, Guangdong

Author Info: (1) School of Pharmaceutical Sciences, Shenzhen University Health Science Center, Shenzhen, 518060, Guangdong, China. (2) The 3rd Affiliated Hospital of Shenzhen University, Shenzhen, 518001, Guangdong, China. (3) Department of Biology and School of Medicine, Southern University of Science and Technology, Shenzhen, 518055, Guangdong, China. (4) School of Pharmaceutical Sciences, Shenzhen University Health Science Center, Shenzhen, 518060, Guangdong, China. (5) The 3rd Affiliated Hospital of Shenzhen University, Shenzhen, 518001, Guangdong, China. (6) School of Pharmaceutical Sciences, Shenzhen University Health Science Center, Shenzhen, 518060, Guangdong, China. (7) School of Pharmaceutical Sciences, Shenzhen University Health Science Center, Shenzhen, 518060, Guangdong, China. (8) School of Pharmaceutical Sciences, Shenzhen University Health Science Center, Shenzhen, 518060, Guangdong, China. (9) School of Pharmaceutical Sciences, Shenzhen University Health Science Center, Shenzhen, 518060, Guangdong, China. gyjin@szu.edu.cn. Cancer Research Center, Shenzhen University Health Science Center, Shenzhen, 518060, Guangdong, China. gyjin@szu.edu.cn.

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Cytomegalovirus: an unlikely ally in the fight against blood cancers

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CMV infection is a potentially fatal complication in patients receiving HSCT, but recent evidence indicates that CMV has strong anti-leukemia effects due in part to shifts in the composition of NK-cell subsets. NK-cells are the primary mediators of the anti-leukemia effect of allogeneic HSCT and infusion of allogeneic NK-cells has shown promise as a means of inducing remission and preventing relapse of several different hematologic malignancies. The effectiveness of these treatments is limited, however, when tumors express HLA-E, a ligand for the inhibitory receptor NKG2A which is expressed by the vast majority of post-transplant reconstituted and ex vivo expanded NK-cells. It is possible to enhance NK-cell cytotoxicity against HLA-E(pos) malignancies by increasing the proportion of NK-cells expressing NKG2C (the activating receptor for HLA-E) and lacking the corresponding inhibitory receptor NKG2A. The proportion of NKG2C(pos) /NKG2A(neg) NK-cells is typically low in healthy adults, but it can be increased by CMV infection or ex vivo expansion of NK-cells using HLA-E transfected feeder cells and IL-15. In this review, we will discuss the role of CMV-driven NKG2C(pos) /NKG2A(neg) NK-cell expansion on anti-tumor cytotoxicity and disease progression in the context of hematologic malignancies, and explore the possibility of harnessing NKG2C(pos) /NKG2A(neg) NK-cells for cancer immunotherapy. This article is protected by copyright. All rights reserved.

Author Info: (1) Laboratory of Integrated Physiology, Department of Health and Human Performance, University of Houston, 3875 Holman Street, Houston, Texas, 77204, USA. Department of Nutritional Sciences

Author Info: (1) Laboratory of Integrated Physiology, Department of Health and Human Performance, University of Houston, 3875 Holman Street, Houston, Texas, 77204, USA. Department of Nutritional Sciences, The University of Arizona, 1177 E. Fourth Street, Tucson, Arizona, 85721, USA. (2) Laboratory of Integrated Physiology, Department of Health and Human Performance, University of Houston, 3875 Holman Street, Houston, Texas, 77204, USA. Department of Nutritional Sciences, The University of Arizona, 1177 E. Fourth Street, Tucson, Arizona, 85721, USA. (3) Laboratory of Integrated Physiology, Department of Health and Human Performance, University of Houston, 3875 Holman Street, Houston, Texas, 77204, USA. Department of Nutritional Sciences, The University of Arizona, 1177 E. Fourth Street, Tucson, Arizona, 85721, USA.

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Induction of necrotic cell death and activation of STING in the tumor microenvironment via cationic silica nanoparticles leading to enhanced antitumor immunity

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Nanotechnology has demonstrated tremendous clinical utility, with potential applications in cancer immunotherapy. Although nanoparticles with intrinsic cytotoxicity are often considered unsuitable for clinical applications, such toxicity may be harnessed in the fight against cancer. Nanoparticle-associated toxicity can induce acute necrotic cell death, releasing tumor-associated antigens which may be captured by antigen-presenting cells to initiate or amplify tumor immunity. To test this hypothesis, cytotoxic cationic silica nanoparticles (CSiNPs) were directly administered into B16F10 melanoma implanted in C57BL/6 mice. CSiNPs caused plasma membrane rupture and oxidative stress of tumor cells, inducing local inflammation, tumor cell death and the release of tumor-associated antigens. The CSiNPs were further complexed with bis-(3'-5')-cyclic dimeric guanosine monophosphate (c-di-GMP), a molecular adjuvant which activates the stimulator of interferon genes (STING) in antigen-presenting cells. Compared with unformulated c-di-GMP, the delivery of c-di-GMP with CSiNPs markedly prolonged its local retention within the tumor microenvironment and activated tumor-infiltrating antigen-presenting cells. The combination of CSiNPs and a STING agonist showed dramatically increased expansion of antigen-specific CD8+ T cells, and potent tumor growth inhibition in murine melanoma. These results demonstrate that cationic nanoparticles can be used as an effective in situ vaccine platform which simultaneously causes tumor destruction and immune activation.

Author Info: (1) Department of Chemical Engineering and Materials Science, Wayne State University, Detroit, Michigan 48202, USA. haipengl.liu@wayne.edu. (2) Department of Chemical Engineering and Materials Science, Wayne

Author Info: (1) Department of Chemical Engineering and Materials Science, Wayne State University, Detroit, Michigan 48202, USA. haipengl.liu@wayne.edu. (2) Department of Chemical Engineering and Materials Science, Wayne State University, Detroit, Michigan 48202, USA. haipengl.liu@wayne.edu. (3) Department of Chemical Engineering and Materials Science, Wayne State University, Detroit, Michigan 48202, USA. haipengl.liu@wayne.edu. (4) Department of Oncology, Wayne State University, Detroit, Michigan 48201, USA and Tumor Biology and Microenvironment Program, Barbara Ann Karmanos Cancer Institute, Detroit, Michigan 48201, USA. (5) Department of Chemical Engineering and Materials Science, Wayne State University, Detroit, Michigan 48202, USA. haipengl.liu@wayne.edu. (6) Department of Oncology, Wayne State University, Detroit, Michigan 48201, USA and Tumor Biology and Microenvironment Program, Barbara Ann Karmanos Cancer Institute, Detroit, Michigan 48201, USA. (7) Department of Chemical Engineering and Materials Science, Wayne State University, Detroit, Michigan 48202, USA. haipengl.liu@wayne.edu and Department of Oncology, Wayne State University, Detroit, Michigan 48201, USA and Tumor Biology and Microenvironment Program, Barbara Ann Karmanos Cancer Institute, Detroit, Michigan 48201, USA.

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Empty conformers of HLA-B preferentially bind CD8 and regulate CD8+ T cell function

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When complexed with antigenic peptides, human leukocyte antigen (HLA) class I (HLA-I) molecules initiate CD8(+) T cell responses via interaction with the T cell receptor (TCR) and co-receptor CD8. Peptides are generally critical for the stable cell surface expression of HLA-I molecules. However, for HLA-I alleles such as HLA-B*35:01, peptide-deficient (empty) heterodimers are thermostable and detectable on the cell surface. Additionally, peptide-deficient HLA-B*35:01 tetramers preferentially bind CD8 and to a majority of blood-derived CD8(+) T cells via a CD8-dependent binding mode. Further functional studies reveal that peptide-deficient conformers of HLA-B*35:01 do not directly activate CD8(+) T cells, but accumulate at the immunological synapse in antigen-induced responses, and enhance cognate peptide-induced cell adhesion and CD8(+) T cell activation. Together, these findings indicate that HLA-I peptide occupancy influences CD8 binding affinity, and reveal a new set of regulators of CD8(+) T cell activation, mediated by the binding of empty HLA-I to CD8.

Author Info: (1) Department of Microbiology and Immunology, University of Michigan, Ann Arbor, United States. (2) Department of Microbiology and Immunology, Yerkes National Primate Research Center, Emory

Author Info: (1) Department of Microbiology and Immunology, University of Michigan, Ann Arbor, United States. (2) Department of Microbiology and Immunology, Yerkes National Primate Research Center, Emory University School of Medicine, Atlanta, United States. (3) Research and Development, Sirona Genomics, Immucor, Inc, Mountain View, United States. (4) Department of Microbiology and Immunology, University of Michigan, Ann Arbor, United States.

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A Novel Three-Dimensional Immune Oncology Model for High-Throughput Testing of Tumoricidal Activity

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The latest advancements in oncology research are focused on autologous immune cell therapy. However, the effectiveness of this type of immunotherapy for cancer remediation is not equivalent for all patients or cancer types. This suggests the need for better preclinical screening models that more closely recapitulate in vivo tumor biology. The established method for investigating tumoricidal activity of immunotherapies has been study of two-dimensional (2D) monolayer cultures of immortalized cancer cell lines or primary tumor cells in standard tissue culture vessels. Indeed, a proven means to examine immune cell migration and invasion are 2D chemotaxis assays in permeabilized supports or Boyden chambers. Nevertheless, the more in vivo-like three-dimensional (3D) multicellular tumor spheroids are quickly becoming the favored model to examine immune cell invasion and tumor cell cytotoxicity. Accordingly, we have developed a 3D immune oncology model by combining 96-well permeable support systems and 96-well low-attachment microplates. The use of the permeable support system enables assessment of immune cell migration, which was tested in this study as chemotactic response of natural killer NK-92MI cells to human stromal-cell derived factor-1 (SDF-1alpha). Immune invasion was assessed by measuring NK-92MI infiltration into lung carcinoma A549 cell spheroids that were formed in low-attachment microplates. The novel pairing of the permeable support system with low-attachment microplates permitted simultaneous investigation of immune cell homing, immune invasion of tumor spheroids, and spheroid cytotoxicity. In effect, the system represents a more comprehensive and in vivo-like immune oncology model that can be utilized for high-throughput study of tumoricidal activity.

Author Info: (1) Life Sciences Division, Corning Incorporated, Kennebunk, ME, United States. (2) Life Sciences Division, Corning Incorporated, Kennebunk, ME, United States. (3) Life Sciences Division, Corning

Author Info: (1) Life Sciences Division, Corning Incorporated, Kennebunk, ME, United States. (2) Life Sciences Division, Corning Incorporated, Kennebunk, ME, United States. (3) Life Sciences Division, Corning Incorporated, Kennebunk, ME, United States.

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Mycobacterium tuberculosis PPE60 antigen drives Th1/Th17 responses via Toll-like receptor 2-dependent maturation of dendritic cells

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Targeting of Mycobacterium tuberculosis (MTB) PE/PPE antigens that induce type 1 helper T cell (Th1) and Th17 responses represents a crucial strategy for the development of tuberculosis (TB) vaccines. However, only few PE/PPE antigens induce these responses. Here, we sought to determine how the cell wall-associated antigen PPE60 (Rv3478) activates dendritic cell (DC) maturation and T-cell differentiation. We observed that PPE60 induces DC maturation by augmenting the protein expression of cluster of differentiation 80 (CD80) and CD86, and major histocompatibility complex (MHC) class I and MHC class II on the cell surface. PPE60 also stimulated the production of tumor necrosis factor-alpha (TNF-alpha), interleukin (IL)-1beta, IL-6, IL-12p70, and IL-23p19, but not IL-10. This induction was mediated by Toll-like receptor 2 (TLR2) and followed by activation of p38, c-Jun N-terminal kinase (JNK), and NF-kappaB signaling. PPE60 enhanced MHC-II expression and promoted antigen processing by DCs in a TLR2-dependent manner. Moreover, PPE60-stimulated DCs directed naive CD4(+) T cells to produce IFN-gamma, IL-2, and IL-17A, expanding the Th1 and Th17 responses, along with activation of T-Bet and RAR-related orphan receptor C (RORgammat), but not GATA-3. Moreover, PPE60 activated the NLRP3 inflammasome, followed by caspase-1-dependent IL-1beta and IL-18 synthesis in DCs. Of note, pharmacological inhibition of NLRP3 activation specifically attenuated IFN-gamma and IL-17A secretion into the supernatant from CD4(+) T cells co-cultured with PPE60-activated DCs. These findings indicate that PPE60 induces Th1 and Th17 immune responses by activating DCs in a TLR2-dependent manner, suggesting PPE60's potential for use in MTB vaccine development.

Author Info: (1) GMU-GIBH Joint School of Life Science, Guangzhou Medical University, China. (2) Guangdong Second Provincial General Hospital, China. (3) GMU-GIBH Joint School of Life Science

Author Info: (1) GMU-GIBH Joint School of Life Science, Guangzhou Medical University, China. (2) Guangdong Second Provincial General Hospital, China. (3) GMU-GIBH Joint School of Life Science, Guangzhou Medical University, China. (4) GMU-GIBH Joint School of Life Science, Guangzhou Medical University, China. (5) State Key Laboratory of Genetic Engineering, Institute of Genetics, School of Life Science, Fudan University, China. (6) State Key Laboratory of Genetic Engineering, Institute of Genetics, School of Life Science, Fudan University, China. (7) Department of Laboratory Medicine and Central Laboratories, Guangdong Second Provincial General Hospital, China. (8) State Key Laboratory of Genetic Engineering, Institute of Genetics, School of Life Science, Fudan University, China.

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The Bacterial Toxin CNF1 Induces Activation and Maturation of Human Monocyte-Derived Dendritic Cells

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Cytotoxic necrotizing factor 1 (CNF1) is a bacterial protein toxin primarily expressed by pathogenic Escherichia coli strains, causing extraintestinal infections. The toxin is believed to enhance the invasiveness of E. coli by modulating the activity of Rho GTPases in host cells, but it has interestingly also been shown to promote inflammation, stimulate host immunity and function as a potent immunoadjuvant. The mechanisms underlying the immunostimulatory properties of CNF1 are, however, poorly characterized, and little is known about the direct effects of the toxin on immune cells. Here, we show that CNF1 induces expression of maturation markers on human immature monocyte-derived dendritic cells (moDCs) without compromising cell viability. Consistent with the phenotypic maturation, CNF1 further triggered secretion of proinflammatory cytokines and increased the capacity of moDCs to stimulate proliferation of allogenic naive CD4+ T cells. A catalytically inactive form of the toxin did not induce moDC maturation, indicating that the enzymatic activity of CNF1 triggers immature moDCs to undergo phenotypic and functional maturation. As the maturation of dendritic cells plays a central role in initiating inflammation and activating the adaptive immune response, the present findings shed new light on the immunostimulatory properties of CNF1 and may explain why the toxin functions as an immunoadjuvant.

Author Info: (1) Department of Immunology and Microbiology, University of Copenhagen, Norre Alle 14, 2200 Copenhagen, Denmark. lgmas@sund.ku.dk. (2) Italian Center for Global Health, Istituto Superiore di

Author Info: (1) Department of Immunology and Microbiology, University of Copenhagen, Norre Alle 14, 2200 Copenhagen, Denmark. lgmas@sund.ku.dk. (2) Italian Center for Global Health, Istituto Superiore di Sanita; Viale Regina Elena 299, 00161 Rome, Italy. alessia.fabbri@iss.it. (3) Department of Immunology and Microbiology, University of Copenhagen, Norre Alle 14, 2200 Copenhagen, Denmark. mnamini@sund.ku.dk. (4) Costerton Biofilm Center, Department of Immunology and Microbiology, University of Copenhagen, Norre Alle 14, 2200 Copenhagen, Denmark. mgivskov@sund.ku.dk. (5) Italian Center for Global Health, Istituto Superiore di Sanita; Viale Regina Elena 299, 00161 Rome, Italy. carla.fiorentini@iss.it. (6) Department of Immunology and Microbiology, University of Copenhagen, Norre Alle 14, 2200 Copenhagen, Denmark. thorkr@sund.ku.dk.

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PRAME peptide-specific CD8(+) T cells represent the predominant response against leukemia-associated antigens (LAAs) in healthy individuals

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Antigen-specific T cells isolated from healthy individuals (HIs) have shown great therapeutic potential upon adoptive transfer for the treatment of viremia in immunosuppressed patients. The lack of comprehensive data on the prevalence and characteristics of leukemia associated antigen (LAA)-specific T cells in HIs still limits such an approach for tumor therapy. Therefore, we have investigated T cell responses against prominent candidates comprising WT1, PRAME, Survivin, NY-ESO and p53 by screening PBMCs from HIs using intracellular IFN-gamma staining following provocation with LAA peptide mixes. Here, we found predominantly poly-functional effector/effector memory CCR7(-) /CD45RA(+/-) /CD8(+) LAA peptide-specific T cells with varying CD95 expression in 34 of 100 tested HIs, whereas CD4(+) T cells responses were restricted to 5. Most frequent LAA peptide-specific T cell responses were directed against WT1 and PRAME peptides with a prevalence of 20% and 17%, respectively, showing the highest magnitude (0.16% +/- 0.22% (mean+/-SD)) for PRAME peptides. Cytotoxicity of PRAME peptide-specific T cells was demonstrated by specific killing of PRAME peptide-pulsed T2 cells. Furthermore, the proliferative capacity of PRAME peptide-specific T cells was confined to HIs responsive towards PRAME peptide challenge corroborating the accuracy of the screening results. In conclusion, we identified PRAME as a promising target antigen for adoptive leukemia therapy. This article is protected by copyright. All rights reserved.

Author Info: (1) Experimental Transfusion Medicine, Medical Faculty Carl Gustav Carus, TU Dresden, Germany. Institute for Transfusion Medicine Dresden, German Red Cross Blood Donation Service North-East, Dresden

Author Info: (1) Experimental Transfusion Medicine, Medical Faculty Carl Gustav Carus, TU Dresden, Germany. Institute for Transfusion Medicine Dresden, German Red Cross Blood Donation Service North-East, Dresden, Germany. (2) Experimental Transfusion Medicine, Medical Faculty Carl Gustav Carus, TU Dresden, Germany. Institute for Transfusion Medicine Dresden, German Red Cross Blood Donation Service North-East, Dresden, Germany. (3) Center for Regenerative Therapies Dresden (CRTD), Dresden, Germany. (4) German Cancer Research Center (DKFZ), Heidelberg, Germany. German Cancer Consortium (DKTK), Dresden, Germany. Center for Regenerative Therapies Dresden (CRTD), Dresden, Germany. Institute of Immunology, Medical Faculty, TU Dresden, Dresden, Germany. National Center for Tumor Diseases, University Hospital Carl Gustav Carus, TU Dresden, Dresden, Germany. (5) German Cancer Research Center (DKFZ), Heidelberg, Germany. German Cancer Consortium (DKTK), Dresden, Germany. Center for Regenerative Therapies Dresden (CRTD), Dresden, Germany. Department of Medicine 1, University Hospital Carl Gustav Carus, TU Dresden, Dresden, Germany. National Center for Tumor Diseases, University Hospital Carl Gustav Carus, TU Dresden, Dresden, Germany. (6) Experimental Transfusion Medicine, Medical Faculty Carl Gustav Carus, TU Dresden, Germany. Institute for Transfusion Medicine Dresden, German Red Cross Blood Donation Service North-East, Dresden, Germany. German Cancer Research Center (DKFZ), Heidelberg, Germany. German Cancer Consortium (DKTK), Dresden, Germany. Center for Regenerative Therapies Dresden (CRTD), Dresden, Germany. (7) Experimental Transfusion Medicine, Medical Faculty Carl Gustav Carus, TU Dresden, Germany. Institute for Transfusion Medicine Dresden, German Red Cross Blood Donation Service North-East, Dresden, Germany.

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Carbonate Apatite Nanoparticles Act as Potent Vaccine Adjuvant Delivery Vehicles by Enhancing Cytokine Production Induced by Encapsulated Cytosine-Phosphate-Guanine Oligodeoxynucleotides

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Vaccine adjuvants that can induce not only antigen-specific antibody responses but also Th1-type immune responses and CD8(+) cytotoxic T lymphocyte responses are needed for the development of vaccines against infectious diseases and cancer. Of many available adjuvants, oligodeoxynucleotides (ODNs) with unmethylated cytosine-phosphate-guanine (CpG) motifs are the most promising for inducing the necessary immune responses, and these adjuvants are currently under clinical trials in humans. However, the development of novel delivery vehicles that enhance the adjuvant effects of CpG ODNs, subsequently increasing the production of cytokines such as type-I interferons (IFNs), is highly desirable. In this study, we demonstrate the potential of pH-responsive biodegradable carbonate apatite (CA) nanoparticles as CpG ODN delivery vehicles that can enhance the production of type-I IFNs (such as IFN-alpha) relative to that induced by CpG ODNs and can augment the adjuvant effects of CpG ODNs in vivo. In contrast to CpG ODNs, CA nanoparticles containing CpG ODNs (designated CA-CpG) induced significant IFN-alpha production by mouse dendritic cells and human peripheral blood mononuclear cells in vitro; and production of interleukin-12, and IFN-gamma was higher in CA-CpG-treated groups than in CpG ODNs groups. In addition, treatment with CA-CpG resulted in higher cytokine production in draining lymph nodes than did treatment with CpG ODNs in vivo. Furthermore, vaccination with CA-CpG plus an antigen, such as ovalbumin or influenza virus hemagglutinin, resulted in higher antigen-specific antibody responses and CD8(+) cytotoxic T lymphocyte responses in vivo, in an interleukin-12- and type-I IFN-dependent manner, than did vaccination with the antigen plus CpG ODNs; in addition, the efficacy of the vaccine against influenza virus was higher with CA-CpG as the adjuvant than with CpG ODNs as the adjuvant. These data show the potential of CA nanoparticles to serve as CpG ODN delivery vehicles that increase the production of cytokines, especially IFN-alpha, induced by CpG ODNs and thus augment the efficacy of CpG ODNs as adjuvants. We expect that the strategy reported herein will facilitate the design and development of novel adjuvant delivery vehicles for vaccines.

Author Info: (1) Vaccine Creation Project, BIKEN Innovative Vaccine Research Alliance Laboratories, Research Institute for Microbial Diseases, Osaka University, Suita, Japan. (2) Vaccine Creation Project, BIKEN Innovative

Author Info: (1) Vaccine Creation Project, BIKEN Innovative Vaccine Research Alliance Laboratories, Research Institute for Microbial Diseases, Osaka University, Suita, Japan. (2) Vaccine Creation Project, BIKEN Innovative Vaccine Research Alliance Laboratories, Research Institute for Microbial Diseases, Osaka University, Suita, Japan. (3) Vaccine Dynamics Project, BIKEN Innovative Vaccine Research Alliance Laboratories, Research Institute for Microbial Diseases, Osaka University, Suita, Japan. Vaccine Dynamics Project, BIKEN Center for Innovative Vaccine Research and Development, The Research Foundation for Microbial Diseases of Osaka University, Suita, Japan. (4) Vaccine Creation Project, BIKEN Center for Innovative Vaccine Research and Development, The Research Foundation for Microbial Diseases of Osaka University, Suita, Japan. (5) Division of Health Sciences, Department of Molecular Pathology, Graduate School of Medicine, Osaka University, Suita, Japan. (6) Division of Health Sciences, Department of Molecular Pathology, Graduate School of Medicine, Osaka University, Suita, Japan. (7) Laboratory of Adjuvant Innovation, Center for Vaccine and Adjuvant Research, National Institutes of Biomedical Innovation, Health and Nutrition, Ibaraki, Japan. Laboratory of Vaccine Science, WPI Immunology Frontier Research Center, Osaka University, Suita, Japan. (8) Laboratory of Adjuvant Innovation, Center for Vaccine and Adjuvant Research, National Institutes of Biomedical Innovation, Health and Nutrition, Ibaraki, Japan. Laboratory of Vaccine Science, WPI Immunology Frontier Research Center, Osaka University, Suita, Japan. (9) Division of Health Sciences, Department of Molecular Pathology, Graduate School of Medicine, Osaka University, Suita, Japan. (10) Vaccine Creation Project, BIKEN Innovative Vaccine Research Alliance Laboratories, Research Institute for Microbial Diseases, Osaka University, Suita, Japan. Vaccine Creation Project, BIKEN Center for Innovative Vaccine Research and Development, The Research Foundation for Microbial Diseases of Osaka University, Suita, Japan. Laboratory of Nano-Design for Innovative Drug Development, Graduate School of Pharmaceutical Sciences, Osaka University, Suita, Japan. Global Center for Medical Engineering and Informatics, Osaka University, Suita, Japan.

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Induction of Neoantigen-specific Cytotoxic T Cells and Construction of T-cell Receptor-engineered T cells for Ovarian Cancer

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PURPOSE: Current evolution of cancer immunotherapies, such as immune checkpoint blockade, has implicated neoantigens as major targets of anti-cancer cytotoxic T cells. Adoptive T cell therapy with neoantigen-specific T cell receptor (TCR)-engineered T cells would be an attractive therapeutic option for advanced cancers where the host anti-tumor immune function is strongly inhibited. We previously developed a rapid and efficient pipeline for production of neoantigen-specific TCR-engineered T cells using peripheral blood from an HLA-matched healthy donor. Our protocol required only two weeks from stimulation of T cells with neoantigen-loaded dendritic cells to the identification of neoantigen-specific TCRs. We conducted the pilot study to validate our protocol. EXPERIMENTAL DESIGN: We used tumors from 7 ovarian cancer patients to validate our protocol. RESULTS: We chose 14 candidate neoantigens from 7 ovarian tumors (1-3 candidates for each patient), and then successfully induced 3 neoantigen-specific T cells from one healthy donor and identified their TCR sequences. Moreover, we validated functional activity of the three identified TCRs by generating TCR-engineered T cells which recognized the corresponding neoantigens and showed cytotoxic activity in an antigen-dose-dependent manner. However, one case of neoantigen-specific TCR-engineered T cells showed cross-reactivity against the corresponding wild-type peptide. Conclusion/Discussions: This pilot study demonstrated the feasibility of our efficient process from identification of neoantigen to production of the neoantigen-targeting cytotoxic TCR-engineered T cells for ovarian cancer and revealed the importance of careful validation of neoantigen-specific-TCR-engineered T cells to avoid severe immune-related adverse events.

Author Info: (1) Department of Medicine, University of Chicago. (2) Institute of Immunology, Charite. (3) Department of Medicine, University of Chicago. (4) Department of Medicine, University of

Author Info: (1) Department of Medicine, University of Chicago. (2) Institute of Immunology, Charite. (3) Department of Medicine, University of Chicago. (4) Department of Medicine, University of Chicago. (5) Medicine, University of Chicago. (6) Department of Obstetrics and Gynecology, Faculty of Medicine, Nihon University. (7) Department of Medicine, University of Chicago. (8) Cancer Precision Medicine Center, Japanese Foundation for Cancer Research. (9) Medicine, University of Chicago. (10) Department of Medicine, University of Chicago. (11) Department of Medicine, University of Chicago ynakamura@medicine.bsd.uchicago.edu.

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