Journal Articles

TAA/TSA-based approaches

Cancer vaccines based on tumor-associated (TAA) or tumor-specific (TSA) antigens derived from germline genes, oncogenic viruses or endogenous retroviruses

Phase II trial of ipilimumab in melanoma patients with preexisting humoural immune response to NY-ESO-1

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BACKGROUND: Immune checkpoint therapy has dramatically changed treatment options in patients with metastatic melanoma. However, a relevant part of patients still does not respond to treatment. Data regarding the prognostic or predictive significance of preexisting immune responses against tumour antigens are conflicting. Retrospective data suggested a higher clinical benefit of ipilimumab in melanoma patients with preexisting NY-ESO-1-specific immunity. PATIENTS AND METHODS: Twenty-five patients with previously untreated or treated metastatic melanoma and preexisting humoural immune response against NY-ESO-1 received ipilimumab at a dose of 10 mg/kg in week 1, 4, 7, 10 followed by 3-month maintenance treatment for a maximum of 48 weeks. Primary endpoint was the disease control rate (irCR, irPR or irSD) according to immune-related response criteria (irRC). Secondary endpoints included the disease control rate according to RECIST criteria, progression-free survival and overall survival (OS). Humoural and cellular immune responses against NY-ESO-1 were analysed from blood samples. RESULTS: Disease control rate according to irRC was 52%, irPR was observed in 36% of patients. Progression-free survival according to irRC was 7.8 months, according to RECIST criteria it was 2.9 months. Median OS was 22.7 months; the corresponding 1-year survival rate was 66.8%. Treatment-related grade 3 AEs occurred in 36% with no grade 4-5 AEs. No clear association was found between the presence of NY-ESO-1-specific cellular or humoural immune responses and clinical activity. CONCLUSION: Ipilimumab demonstrated clinically relevant activity within this biomarker-defined population. NY-ESO-1 positivity, as a surrogate for a preexisting immune response against tumour antigens, might help identifying patients with a superior outcome from immune checkpoint blockade. CLINICAL TRIAL INFORMATION: NCT01216696.

Author Info: (1) Department of Medical Oncology, National Center for Tumor Diseases, University Hospital Heidelberg, Germany. Electronic address: GeorgMartin.Haag@med.uni-heidelberg.de. (2) Department of Medical Oncology, National Center for

Author Info: (1) Department of Medical Oncology, National Center for Tumor Diseases, University Hospital Heidelberg, Germany. Electronic address: GeorgMartin.Haag@med.uni-heidelberg.de. (2) Department of Medical Oncology, National Center for Tumor Diseases, University Hospital Heidelberg, Germany. (3) Department of Dermatology and National Center for Tumor Diseases, University Hospital Heidelberg, Germany. (4) Department of Medical Oncology, National Center for Tumor Diseases, University Hospital Heidelberg, Germany. (5) Department of Dermatology and National Center for Tumor Diseases, University Hospital Heidelberg, Germany. (6) Department of Dermatology and National Center for Tumor Diseases, University Hospital Heidelberg, Germany. (7) Translational Immunology, National Center for Tumor Diseases, Heidelberg, Germany. (8) Department of Medical Oncology, National Center for Tumor Diseases, University Hospital Heidelberg, Germany. (9) Department of Medical Oncology, National Center for Tumor Diseases, University Hospital Heidelberg, Germany. (10) Translational Immunology, National Center for Tumor Diseases, Heidelberg, Germany. (11) Translational Immunology, National Center for Tumor Diseases, Heidelberg, Germany. (12) Translational Immunology, National Center for Tumor Diseases, Heidelberg, Germany. (13) Institute of Transplant Immunology, IFB-Tx, Hannover Medical School, Hannover, Germany. (14) NCT Trial Center, National Center for Tumor Diseases, Heidelberg, Germany. (15) NCT Trial Center, National Center for Tumor Diseases, Heidelberg, Germany. (16) Translational Immunology, National Center for Tumor Diseases, Heidelberg, Germany; Regensburg Center for Interventional Immunology, University Hospital Regensburg, Germany. (17) Department of Dermatology and National Center for Tumor Diseases, University Hospital Heidelberg, Germany. (18) Department of Medical Oncology, National Center for Tumor Diseases, University Hospital Heidelberg, Germany; Clinical Cooperation Unit "Applied Tumor-Immunity", German Cancer Research Center (DKFZ), Heidelberg, Germany.

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EpCAM peptide-primed dendritic cell vaccination confers significant anti-tumor immunity in hepatocellular carcinoma cells

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Cancer stem-like cells (CSCs) may play a key role in tumor initiation, self-renewal, differentiation, and resistance to current treatments. Dendritic cells (DCs) play a vital role in host immune reactions as well as antigen presentation. In this study, we explored the suitability of using CSC peptides as antigen sources for DC vaccination against human breast cancer and hepatocellular carcinoma (HCC) with the aim of achieving CSC targeting and enhancing anti-tumor immunity. CD44 is used as a CSC marker for breast cancer and EpCAM is used as a CSC marker for HCC. We selected CD44 and EpCAM peptides that bind to HLA-A2 molecules on the basis of their binding affinity, as determined by a peptide-T2 binding assay. Our data showed that CSCs express high levels of tumor-associated antigens (TAAs) as well as major histocompatibility complex (MHC) molecules. Pulsing DCs with CD44 and EpCAM peptides resulted in the efficient generation of mature DCs (mDCs), thus enhancing T cell stimulation and generating potent cytotoxic T lymphocytes (CTLs). The activation of CSC peptide-specific immune responses by the DC vaccine in combination with standard chemotherapy may provide better clinical outcomes in advanced carcinomas.

Author Info: (1) Research Center, Dongnam Institute of Radiological & Medical Sciences, Busan, Republic of Korea. (2) Research Center, Dongnam Institute of Radiological & Medical Sciences, Busan

Author Info: (1) Research Center, Dongnam Institute of Radiological & Medical Sciences, Busan, Republic of Korea. (2) Research Center, Dongnam Institute of Radiological & Medical Sciences, Busan, Republic of Korea. (3) Research Center, Dongnam Institute of Radiological & Medical Sciences, Busan, Republic of Korea. (4) Research Center, Dongnam Institute of Radiological & Medical Sciences, Busan, Republic of Korea. (5) Research Center, Dongnam Institute of Radiological & Medical Sciences, Busan, Republic of Korea. Department of Radiation Oncology, Dongnam Institute of Radiological & Medical Sciences, Busan, Republic of Korea. Department of Radiation Oncology, Korea Institute of Radiological & Medical Sciences, Seoul, Republic of Korea. (6) Research Center, Dongnam Institute of Radiological & Medical Sciences, Busan, Republic of Korea.

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Phase I study of glypican-3-derived peptide vaccine therapy for patients with refractory pediatric solid tumors

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The carcinoembryonic antigen glypican-3 (GPC3) is a good target of anticancer immunotherapy against pediatric solid tumors expressing GPC3. In this non-randomized, open-label, phase I clinical trial, we analyzed the safety and efficacy of GPC3-peptide vaccination in patients with pediatric solid tumors. Eighteen patients with pediatric solid tumors expressing GPC3 underwent GPC3-peptide vaccination (intradermal injections every 2 weeks), with the primary endpoint being the safety of GPC3-peptide vaccination and the secondary endpoints being immune response, as measured by interferon (IFN)-gamma enzyme-linked immunospot assay and Dextramer staining, and the clinical outcomes of tumor response, progression free survival (PFS), and overall survival (OS). Our findings indicated that GPC3 vaccination was well tolerated. We observed disease-control rates [complete response (CR)+partial response+stable disease] of 66.7% after 2 months, and although patients in the progression group unable to induce GPC3-peptide-specific cytotoxic T lymphocytes (CTLs) received poor prognoses, patients in the partial-remission and remission groups or those with hepatoblastoma received good prognoses. The GPC3-peptide vaccine induced a GPC3-specific CTL response in seven patients, with PFS and OS significantly longer in patients with high GPC3-specific CTL frequencies than in those with low frequencies. Furthermore, we established GPC3-peptide-specific CTL clones from a resected-recurrent tumor from one patient, with these cells exhibiting GPC3-peptide-specific cytokine secretion. The results of this trial demonstrated that the GPC3-peptide-specific CTLs induced by the GPC3-peptide vaccine infiltrated tumor tissue, and use of the GPC3-peptide vaccine might prevent the recurrence of pediatric solid tumors, especially hepatoblastomas, after a second CR.

Author Info: (1) Division of Cancer Immunotherapy, Exploratory Oncology Research and Clinical Trial Center, National Cancer Center, Kashiwa, Chiba, Japan. (2) Division of Pediatric Oncology, National Cancer

Author Info: (1) Division of Cancer Immunotherapy, Exploratory Oncology Research and Clinical Trial Center, National Cancer Center, Kashiwa, Chiba, Japan. (2) Division of Pediatric Oncology, National Cancer Center Hospital East, Kashiwa, Chiba, Japan. (3) Division of Cancer Immunotherapy, Exploratory Oncology Research and Clinical Trial Center, National Cancer Center, Kashiwa, Chiba, Japan. (4) Division of Cancer Immunotherapy, Exploratory Oncology Research and Clinical Trial Center, National Cancer Center, Kashiwa, Chiba, Japan. (5) Division of Cancer Immunotherapy, Exploratory Oncology Research and Clinical Trial Center, National Cancer Center, Kashiwa, Chiba, Japan. (6) Division of Cancer Immunotherapy, Exploratory Oncology Research and Clinical Trial Center, National Cancer Center, Kashiwa, Chiba, Japan. (7) Department of Pediatric Hematology and Oncology, Osaka City General Hospital, Miyakojima-hondori, Miyakojima-ku, Osaka, Japan. (8) Department of Pediatric Hematology and Oncology, Osaka City General Hospital, Miyakojima-hondori, Miyakojima-ku, Osaka, Japan. (9) Department of Pediatrics, St Luke's International Hospital, 9-1 Akashi-cho, Chuo-ku, Tokyo, Japan. (10) Department of Pediatrics, St Luke's International Hospital, 9-1 Akashi-cho, Chuo-ku, Tokyo, Japan. (11) Department of Pediatrics, St Luke's International Hospital, 9-1 Akashi-cho, Chuo-ku, Tokyo, Japan. (12) Division of Pediatric Oncology, National Cancer Center Hospital, 5-1-1 Tsukiji, Chuo-ku, Tokyo, Japan. (13) Department of Pediatric Surgery, Kyushu University Hospital, 3-1-1 Maidashi, Higashi-ku, Fukuoka, Japan. (14) Department of Anatomic Pathology, Graduate School of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka, Japan. (15) Department of Biomedical Statistics, Innovative Clinical Research Center, Kanazawa University, 13-1, Takara-machi, Kanazawa, Ishikawa, Japan. (16) Division of Cancer Immunotherapy, Exploratory Oncology Research and Clinical Trial Center, National Cancer Center, Kashiwa, Chiba, Japan. (17) Division of Cancer Immunotherapy, Exploratory Oncology Research and Clinical Trial Center, National Cancer Center, Kashiwa, Chiba, Japan. (18) Division of Cancer Immunotherapy, Exploratory Oncology Research and Clinical Trial Center, National Cancer Center, Kashiwa, Chiba, Japan. (19) Division of Cancer Immunotherapy, Exploratory Oncology Research and Clinical Trial Center, National Cancer Center, Kashiwa, Chiba, Japan.

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Cancer Immunotherapy Targeted Glypican-3 or Neoantigens

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Immune checkpoint inhibitors have ushered a new era in cancer therapy, although other therapies or combinations thereof are still needed for the many patients for whom these drugs are ineffective. In this light, we have identified in glypican-3 an HLA-24, HLA-A2 restriction peptide with extreme cancer specificity. In this paper, we summarize results from a number of related clinical trials demonstrating that glypican-3 peptide vaccines induce specific cytotoxic T lymphocytes in most patients (UMIN Clinical Trials Registry: UMIN000001395, UMIN000005093, UMIN000002614, UMN000003696, UMIN000006357,). We also describe the current state of personalized cancer immunotherapy based on neoantigens, and assess, based on our own research and experience, the potential of such therapy to elicit cancer regression. Finally, we discuss the future direction of cancer immunotherapy. This article is protected by copyright. All rights reserved.

Author Info: (1) Division of Cancer Immunotherapy, Exploratory Oncology Research and Clinical Trial Center National Cancer Center, Kashiwa. Department of Gastroenterological Surgery, Yokohama City University Graduate School

Author Info: (1) Division of Cancer Immunotherapy, Exploratory Oncology Research and Clinical Trial Center National Cancer Center, Kashiwa. Department of Gastroenterological Surgery, Yokohama City University Graduate School of Medicine, Yokohama, Japan. (2) Division of Cancer Immunotherapy, Exploratory Oncology Research and Clinical Trial Center National Cancer Center, Kashiwa. (3) Division of Cancer Immunotherapy, Exploratory Oncology Research and Clinical Trial Center National Cancer Center, Kashiwa. (4) Division of Cancer Immunotherapy, Exploratory Oncology Research and Clinical Trial Center National Cancer Center, Kashiwa. Department of Gastroenterological Surgery, Yokohama City University Graduate School of Medicine, Yokohama, Japan. (5) Division of Cancer Immunotherapy, Exploratory Oncology Research and Clinical Trial Center National Cancer Center, Kashiwa. Department of Gastroenterological Surgery, Yokohama City University Graduate School of Medicine, Yokohama, Japan. (6) Department of Gastroenterological Surgery, Yokohama City University Graduate School of Medicine, Yokohama, Japan. (7) Division of Cancer Immunotherapy, Exploratory Oncology Research and Clinical Trial Center National Cancer Center, Kashiwa.

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The potential of the CMB305 vaccine regimen to target NY-ESO-1 and improve outcomes for synovial sarcoma and myxoid/round cell liposarcoma patients

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INTRODUCTION: Synovial Sarcoma (SS) and Myxoid Round Cell Liposarcoma (MRCL) are devastating sarcoma subtypes with few treatment options and poor outcomes in the advanced setting. However, both these diseases may be ideal for novel immunotherapies targeting the cancer-testis antigen, NY-ESO-1. Areas covered: In this review, we discuss the novel NY-ESO-1 targeted vaccine regimen, CMB305. This regimen uses a unique integration-deficient, dendritic-cell targeting lentiviral vector from the ZVex(R) platform, LV305, in order to prime NY-ESO-1 specific T cells. LV305 has single agent activity, and, in one case, caused a durable partial response in a refractory SS patient. CMB305 also includes a boost from a NY-ESO-1 protein vaccine given along with a potent toll-like-4 receptor agonist, glycopyranosyl lipid A. CMB305 induces NY-ESO-1 specific T cell responses in both SS and MRC patients and these patients had excellent overall survival (OS) outcomes in the initial phase I study. Expert commentary: CMB305 is a therapeutic vaccine regimen targeting NY-ESO-1 based on the lentiviral vaccine vector, LV305. Phase I studies have proven this vaccine is active immunologically. Data suggesting this vaccine may improve OS for SS and MRCL patients is exciting but early, and on-going work is testing the impact of CMB305 on patient outcomes.

Author Info: (1) a Clinical Research Division , Fred Hutchinson Cancer Research Center , Seattle , WA , USA. b Department of Medicine , University of Washington , Seattle , WA , USA.

Author Info: (1) a Clinical Research Division , Fred Hutchinson Cancer Research Center , Seattle , WA , USA. b Department of Medicine , University of Washington , Seattle , WA , USA.

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Targeting tumor-associated carbohydrate antigens: a phase I study of a carbohydrate mimetic-peptide vaccine in stage IV breast cancer subjects

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Tumor-associated carbohydrate antigens (TACAs) support cell survival that could be interrupted by anti-TACA antibodies. Among TACAs that mediate cell survival signals are the neolactoseries antigen Lewis Y (LeY) and the ganglioside GD2. To induce sustained immunity against both LeY and GD2, we developed a carbohydrate mimicking peptide (CMP) as a surrogate pan-immunogen that mimics both. This CMP, referred to as P10s, is the N-terminal half of a peptide vaccine named P10s-PADRE, the C-terminal half of which (PADRE) is a Pan-T-cell epitope. A Phase I dose-escalation trial of P10s-PADRE plus adjuvant MONTANIDE ISA 51 VG was conducted in subjects with metastatic breast cancer to test 300 and 500 mug/injection in two cohorts of 3 subjects each. Doses of the P10s-PADRE vaccine were administered to research participants subcutaneously on weeks 1, 2, 3, 7 and 19. Antibody responses to P10s, GD2, and LeY were measured by ELISA. The P10s-PADRE vaccine induced antibodies specifically reactive with P10s, LeY and GD2 in all 6 subjects. Serum antibodies displayed Caspase-3-dependent apoptotic functionality against LeY or GD2 expressing breast cancer cell lines. Immunization with the P10s-PADRE vaccine was well-tolerated and induced functional antibodies, and the data suggest potential clinical benefit.

Author Info: (1) Departments of Medicine, University of Arkansas for Medical Sciences, Little Rock, Arkansas, USA. Winthrop P. Rockefeller Cancer Institute, Little Rock, Arkansas, USA. (2) Departments

Author Info: (1) Departments of Medicine, University of Arkansas for Medical Sciences, Little Rock, Arkansas, USA. Winthrop P. Rockefeller Cancer Institute, Little Rock, Arkansas, USA. (2) Departments of Medicine, University of Arkansas for Medical Sciences, Little Rock, Arkansas, USA. Winthrop P. Rockefeller Cancer Institute, Little Rock, Arkansas, USA. (3) Departments of Medicine, University of Arkansas for Medical Sciences, Little Rock, Arkansas, USA. Winthrop P. Rockefeller Cancer Institute, Little Rock, Arkansas, USA. (4) Departments of Medicine, University of Arkansas for Medical Sciences, Little Rock, Arkansas, USA. Winthrop P. Rockefeller Cancer Institute, Little Rock, Arkansas, USA. (5) Departments of Biostatistics, University of Arkansas for Medical Sciences, Little Rock, Arkansas, USA. (6) Departments of Pathology, University of Arkansas for Medical Sciences, Little Rock, Arkansas, USA. (7) Department of Gastroenterology, Third Affiliated Hospital of Xi'an Jiaotong University, Xi'an, Shaanxi, China. (8) Stephan Angelov Institute of Microbiology, Bulgarian Academy of Sciences, Sofia, Bulgaria. (9) Departments of Pathology, University of Arkansas for Medical Sciences, Little Rock, Arkansas, USA. Winthrop P. Rockefeller Cancer Institute, Little Rock, Arkansas, USA. (10) Departments of Pathology, University of Arkansas for Medical Sciences, Little Rock, Arkansas, USA. Winthrop P. Rockefeller Cancer Institute, Little Rock, Arkansas, USA.

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Combined Extract of Heated 4T1 and a Heat-Killed Preparation of Lactobacillus Casei in a Mouse Model of Breast Cancer

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Background: The adjuvanticity potential of Lactobacillus casei was first suggested in an old survey. The present study was designed to investigate the efficacy of a new immunotherapy against breast cancer made by mixing an extract of heated 4T1 mammary carcinoma cell line and a heat-killed preparation of Lactobacillus casei. Methods: Female BALB/c mice (6-8 weeks old, n=40) were challenged subcutaneously in the right flanks with 4T1 cells. When all the animals developed a palpable tumor, they were allocated to 4 equal groups and immunotherapy was initiated. The tumor-bearing mice in the experimental groups received the extract of heated 4T1 or heated Lactobacillus casei and/or a combination of both, twice at a 1-week interval. The mice in the control group received phosphate-buffered saline. One week after the last immunotherapy, one half of the mice were euthanized to determine the immune response profile. The remaining animals were kept until death occurred spontaneously. Results: The animals receiving the combined treatment significantly showed more favorable survival curves and slower rates of tumor development than the tumor-bearing mice receiving only the heated 4T1 and/or the negative control mice. The combined immunization significantly amplified the production of nitric oxide and the cytotoxicity of natural killer cells in the spleen cell culture of the tumor-bearing mice. Moreover, the combined immunotherapy significantly increased the secretion of IFN-gamma and conversely diminished the secretion of IL-4 and TGF-beta in the splenocyte population compared to the splenocytes from the other groups. Conclusion: The combined immunotherapy with heated 4T1 cells and heated Lactobacillus casei conferred beneficial outcomes in our mouse model of breast cancer.

Author Info: (1) Division of Immunology, Department of Microbiology, Faculty of Veterinary Medicine, Urmia University, Urmia, Iran. (2) Division of Immunology, Department of Microbiology, Faculty of Veterinary

Author Info: (1) Division of Immunology, Department of Microbiology, Faculty of Veterinary Medicine, Urmia University, Urmia, Iran. (2) Division of Immunology, Department of Microbiology, Faculty of Veterinary Medicine, Urmia University, Urmia, Iran. (3) Division of Immunology, Department of Microbiology, Faculty of Veterinary Medicine, Urmia University, Urmia, Iran.

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Autologous dendritic cells pulsed with allogeneic tumor cell lysate in mesothelioma: From mouse to human

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PURPOSE: Mesothelioma has been regarded as a non-immunogenic tumor, which is also shown by the low response rates to treatments targeting the PD-1/PD-L1 axis. Previously, we demonstrated that autologous tumor lysate-pulsed dendritic cell (DC) immunotherapy increased T-cell response towards malignant mesothelioma. However, the use of autologous tumor material hampers implementation in large clinical trials, which might be overcome by using allogeneic tumor cell lines as tumor antigen source. The purpose of this study was to investigate if allogeneic lysate pulsed DC immunotherapy is effective in mice and safe in humans. EXPERIMENTAL DESIGN: Firstly, in two murine mesothelioma models, mice were treated with autologous DCs pulsed with either autologous or allogeneic tumor lysate, or injected with PBS (negative control). Survival and tumor-directed T cell responses of these mice were monitored. Results were taken forward in a first-in-human clinical trial, in which 9 patients were treated with 10, 25 or 50 million DC per vaccination. DC vaccination consisted of autologous monocyte-derived DC pulsed with tumor lysate from 5 mesothelioma cell lines. RESULTS: In mice, allogeneic lysate-pulsed DC immunotherapy induced tumor-specific T cells and led to an increased survival, to a similar extent as DC immunotherapy with autologous tumor lysate. In the first-in-human clinical trial, no dose limiting toxicities were established and radiographic responses were observed. Median PFS was 8.8 months (95% CI 4.1-20.3) and median OS not reached (median follow up 22.8 months). CONCLUSIONS: DC immunotherapy with allogeneic tumor lysate is effective in mice and safe and feasible in humans.

Author Info: (1) Pulmonary Medicine, Erasmus MC Rotterdam j.aerts@erasmusmc.nl. (2) Pulmonary Medicine, Erasmus MC Cancer Institute. (3) Pulmonary Medicine, Erasmus MC Cancer Institute. (4) Erasmus MC Cancer

Author Info: (1) Pulmonary Medicine, Erasmus MC Rotterdam j.aerts@erasmusmc.nl. (2) Pulmonary Medicine, Erasmus MC Cancer Institute. (3) Pulmonary Medicine, Erasmus MC Cancer Institute. (4) Erasmus MC Cancer Institute, Erasmus Medical Center Rotterdam. (5) Erasmus MC Cancer Institute, Erasmus Medical Center Rotterdam. (6) Pulmonary Diseases, Amphia Ziekenhuis. (7) Pulmonary Medicine, Erasmus MC Cancer Institute. (8) Laboratory of Tumor Immunology, Department of Medical Oncology, Erasmus MC Cancer Institute. (9) Department of Medical Oncology, Erasmus MC Cancer Institute. (10) Center for Experimental and Molecular Medicine, AMC. (11) Hematology, Erasmus MC Cancer Institute. (12) Hematology, Erasmus MC Cancer Institute. (13) Hospital Pharmacy, Erasmus MC. (14) Pulmonary Medicine, Erasmus MC Rotterdam. (15) Pulmonary Medicine, Erasmus MC Cancer Institute. (16) Pulmonology, ErasmusMC.

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PSMA-targeted polyinosine/polycytosine vector induces prostate tumor regression and invokes an antitumor immune response in mice

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There is an urgent need for an effective treatment for metastatic prostate cancer (PC). Prostate tumors invariably overexpress prostate surface membrane antigen (PSMA). We designed a nonviral vector, PEI-PEG-DUPA (PPD), comprising polyethylenimine-polyethyleneglycol (PEI-PEG) tethered to the PSMA ligand, 2-[3-(1, 3-dicarboxy propyl)ureido] pentanedioic acid (DUPA), to treat PC. The purpose of PEI is to bind polyinosinic/polycytosinic acid (polyIC) and allow endosomal release, while DUPA targets PC cells. PolyIC activates multiple pathways that lead to tumor cell death and to the activation of bystander effects that harness the immune system against the tumor, attacking nontargeted neighboring tumor cells and reducing the probability of acquired resistance and disease recurrence. Targeting polyIC directly to tumor cells avoids the toxicity associated with systemic delivery. PPD selectively delivered polyIC into PSMA-overexpressing PC cells, inducing apoptosis, cytokine secretion, and the recruitment of human peripheral blood mononuclear cells (PBMCs). PSMA-overexpressing tumors in nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice with partially reconstituted immune systems were significantly shrunken following PPD/polyIC treatment, in all cases. Half of the tumors showed complete regression. PPD/polyIC invokes antitumor immunity, but unlike many immunotherapies does not need to be personalized for each patient. The potent antitumor effects of PPD/polyIC should spur its development for clinical use.

Author Info: (1) Unit of Cellular Signaling, Department of Biological Chemistry, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 9190411, Israel. (2)

Author Info: (1) Unit of Cellular Signaling, Department of Biological Chemistry, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 9190411, Israel. (2) Department of Chemistry, The Hebrew University of Jerusalem, Jerusalem 9190411, Israel. (3) Department of Chemistry, The Hebrew University of Jerusalem, Jerusalem 9190411, Israel. (4) Unit of Cellular Signaling, Department of Biological Chemistry, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 9190411, Israel. (5) Unit of Cellular Signaling, Department of Biological Chemistry, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 9190411, Israel. (6) Department of Chemistry, The Hebrew University of Jerusalem, Jerusalem 9190411, Israel. (7) Unit of Cellular Signaling, Department of Biological Chemistry, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 9190411, Israel. (8) Unit of Cellular Signaling, Department of Biological Chemistry, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 9190411, Israel. (9) Unit of Cellular Signaling, Department of Biological Chemistry, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 9190411, Israel. (10) Unit of Cellular Signaling, Department of Biological Chemistry, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 9190411, Israel. (11) Unit of Cellular Signaling, Department of Biological Chemistry, The Alexander Silberman Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 9190411, Israel; alex.levitzki@mail.huji.ac.il.

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In Vitro Evaluation of Vegf-Pseudomonas Exotoxin: A Conjugated on Tumor Cells

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Background: Angiogenesis which occurs mandatory in solid tumors, is a critical step in malignancy progression. Vascular endothelial growth factor (VEGF) is mainly responsible for angiogenesis process and facilitates the formation of new vessels. Distribution of monoclonal antibodies against VEGF or VEGF receptor (VEGFR) into the solid tumors is limited because of their huge dimensions. Moreover, many investigations have demonstrated the usefulness of immunotoxins to halt angiogenesis in solid tumors. Materials and Methods: We designed, expressed and evaluated the cytotoxicity of a novel nano-immunotoxin composed of VEGF splice variant containing 121 amino acids (VEGF121) and truncated the exotoxin A of Pseudomonas aeruginosa (PE38-KDEL). The fusion protein VEGF121-PE38 was successfully cloned and expressed in Escherichia coli, purified by Ni(+ 2) affinity chromatography. The fusion protein was subsequently subjected to refolding using the reduced and oxidized glutathione. Results: The expression level of the fusion protein reached to 1 mg/ml. The VEGF121-PE38 immunotoxin showed a 59 KDa MW which had cytotoxic effect on HUVEC and 293/KDR cells as low and high expressing VEGFR2 cells, respectively. But the cytotoxicity on 293/KDR was 100 folds more than that of VEGFR2 low expressing cell HUVEC. Conclusion: The designed immunotoxin showed more selectivity for higher VEGFR2 expressing cells in vitro.

Author Info: (1) Department of Molecular Medicine, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran. (2) Department of Molecular Medicine, Biotechnology Research Center, Pasteur Institute of

Author Info: (1) Department of Molecular Medicine, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran. (2) Department of Molecular Medicine, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran. (3) Department of Parasitology, Pasteur Institute of Iran, Tehran, Iran. (4) Department of Genetics and Molecular Biology, Faculty of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran. Pediatric Inherited Diseases Research Center, Research Institute for Primordial Prevention of Noncommunicable Disease, Isfahan University of Medical Sciences, Isfahan, Iran. (5) Department of Molecular Medicine, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran. (6) Department of Biochemistry, Pasteur Institute of Iran, Tehran, Iran. (7) Department of Pilot Nanobiotechnology, Pasteur Institute of Iran, Tehran, Iran. (8) Department of Molecular Medicine, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran. (9) Department of Parasitology, Pasteur Institute of Iran, Tehran, Iran. (10) Department of Molecular Medicine, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran. (11) Department of Molecular Medicine, Biotechnology Research Center, Pasteur Institute of Iran, Tehran, Iran.

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