Kingdom; 2School of Medicine, Kings College London, London,. United Kingdom. .... 1Oncology, University of Alberta, Edmonton, AB, Canada;. 2Medical ...
Cancer – Immunotherapy, Cancer Vaccines II lymphodepleting regimens. The absence of TCRαΒ at their cell surface along with their drug-resistance properties could prevent their alloreactivity and enable them to resist to lymphodepleting regimens that may be required to avoid their ablation via HvG reaction. By providing a basic frame work to develop a universal T cell compatible with allogeneic adoptive transfer, this work is laying the foundation stone of the large scale utilization of CAR T cell immunotherapies.
422. In a Suicide Gene Therapy Approach to Cancer, γ-Irradiation Can Completely Abolish the Bystander Effect of the PA-STK Ovarian Tumour Cell Line Due to Its Radiation Sensitivity, Resulting in Necrotic Cell Death Jehad Zweiri,1 Bernard Souberbielle,2 Farzin Farzaneh.2 1 School of Medicine, University of Liverpool, Liverpool, United Kingdom; 2School of Medicine, Kings College London, London, United Kingdom.
The effect of irradiation on HSV-TK-expressing tumour cells and their bystander killing has been examined. The data presented shows that the irradiation substantially reduced the ability of PA-STK ovarian cells to induce the bystander killing of unmodified tumour cells. PA-STK cells were found to be very sensitive to irradiation and appeared to die too rapidly (even after a low dose of 500-rad) to enable metabolic conversion of GCV to its phosphorylated products, and/or fail to form gap-junctions with their neighbouring cells. Characterisation of tumour cell death showed that PA-STK cells undergo picnosis (a form of necrosis) after irradiation. In contrast, the TK-modified human and mouse mesothelioma cells were found to retain their in vitro bystander killing effect after irradiation. These results suggest that PA-STK cells are not suitable for clinical trials if the cells need to be lethally irradiated before in vivo use. However, the human mesothelioma cell line CRL-5830-TK appears to retain its bystander killing potential after lethal irradiation. In clinical trials, it may therefore be a suitable vehicle for HSV-TK suicide gene therapy for mesothelioma. The low IC50 of GCV in these cells (about 1µM) makes them particularly attractive.
423. The Combination of p53 Dendritic Cell Vaccine and Ad-p53 Gene Therapy Against p53 Overexpressing and p53 Deleted Tumor Cell Lines
Hiroki Saito,1 Koichi Kitagawa,1 Risa Yamasaki,1 Nami Katai,2 Naoya Morishita,1 Masato Kawabata,1 Dody Bautista,3 Dante Dator,4 Toshiro Shirakawa.1,2 1 Division of Translational Research for Biologics, Kobe University Graduate School of Medicine, Kobe, Hyogo, Japan; 2Department of International Health, Kobe University Graduate School of Health Science, Kobe, Hyogo, Japan; 3MOdecs PI Inc, Quezon City, Philippines; 4National Kidney and Transplantation Center, Quezon City, Philippines. BACKGROUND DC are the most potent antigen-presenting cells and stimulate the immune system to protect the host from infectious and cancerous diseases. Adaptive immune system, especially CTL response, is important for cancer treatment. DC take up tumor antigens through phagocytosis, macropinocytosis or endocytosis utilizing Fc receptors. Upon uptake, DC process protein antigens into peptides that are loaded onto MHC molecules for presentation to T cells and are able to induce T cell dependent immune responses. In this study, we induced the p53 specific CTL by co-culturing the p53 DC vaccine, and examined the antitumor effect of p53 specific CTL against the cancer cell lines expressing p53 protein, and the p53-deleted cancer cell line with or without Ad-p53 infection. Molecular Therapy Volume 23, Supplement 1, May 2015 Copyright © The American Society of Gene & Cell Therapy
METHODS DC were generated from the peripheral blood mononuclear cells (PBMC) which were isolated from a healthy volunteer, incubated in medium for 2 hours at 37°. Following 2 hours incubation, only adherent cells were incubated in medium with IL-4 and GM-CSF for 6 days to generate immature DC. To generate p53 DC vaccine, the immature DC were infected with Ad-p53 to present p53 peptides on the MHC molecules. Presenting p53 peptides on the cell surface were examined by flow cytometry. To generate CTL, PBMCs were isolated from the healthy volunteer, and incubated in medium for 24 hours at 37°. Only suspended cells and the p53 DC vaccine were co-cultured to differentiate into CTL cells in medium with IL-2 and IL-7 for 9 days. Thereafter, the lymphocyte phenotype was analyzed by flow cytometry. The cytotoxicities of CTL incubated with the p53 DC vaccine, control of CTL incubated with the DC infected with recombinant adenovirus encoding LacZ gene (Ad-LacZ), and naive CTL, against human cervical carcinoma cell line, which expresses p53, human prostate cancer cell line, which deleted p53 gene and PC-3 infected with Ad-p53 were investigated by the in vitro cytotoxicity assay. RESULTS Ad-p53 infection significantly increased the rate of mature DC co-expressing p53 peptides and HLA-DR on the cell surface. The cytotoxicity of CTL incubated with the p53 DC vaccine against CaSki cells was significantly higher than that of naive CTL and CTL incubated with Ad-LacZ-DC. In addition, the cytotoxicity of the CTL incubated with the p53 DC vaccine against PC-3 cells was enhanced by Ad-p53 infection to PC-3 target cells. CONCLUSION These results indicated that the p53 vaccine could differentiate naive CTL to the p53-specific CTL, and the p53 specific CTL showed the higher cytotoxicity against p53-overexpressing cancer cells and Ad-p53 infected cancer cells. This finding suggested that the strategy of p53 DC vaccine immunotherapy with intra-tumoral injection of Ad-p53 gene therapy has a great potential to create a new paradigm for cancer therapies.
424. Successful Immunotherapy of Cancer Stem Cells Requires Memory and the Adaptive Immune System of the Host
Patrick Schmidt,1,2 Nicolas Desbaillets,2 Pierre Dessen,2 Nancy Thompson,2 Joerg Huelsken.2 1 Medical Oncology, Tumorimmunology, National Center for Tumor Diseases, Heidelberg, Germany; 2Cancer Stem Cell Laboratory, ISREC, EPFL, Lausanne, Switzerland.
The Cancer Stem Cell (CSC) hypothesis puts a, not necessarily rare, subpopulation of cells on the top of a tumor forming hierarchy, pointing to the importance of their destruction when aiming on a final cure. However, current studies targeting CSCs rather focus on the tumor initiating capacities of these cells than on their maintaining capacity in an established lesion. In this presentation we describe an immunotherapeutic approach of targeting the minor CSC subfraction by exploiting the stem cell related wnt-pathway. We found that a memory effect, provided by engineered T cells, is essential to keep the re-formation of CSCs from bulk tumor cells under control. Furthermore, our treatment setting is able to re-activate the host’s non engineered T cells against the majority of tumor cells, which leads to a long lasting tumor-free survival in our mouse model system. These findings implicate a leading role for immunotherapy when targeting the tumor stem cell fraction.
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Cancer – Oncolytic Viruses I The interim analysis of this trial supports the potential of the lentiviral vector platform developed by THERAVECTYS for the future development of therapeutic vaccines and immunotherapies in oncology and infectious diseases. As a second indication, THERAVECTYS is developing an anti-HTLV-1 vaccine candidate to treat patients who have developed a virally-induced adult T cell leukemia. This vaccine candidate should enter into the clinics in mid-2015. Other indications in the pipeline are urogenital cancers, triple-negative breast cancer, EBV-induced nasopharyngeal cancers, HPV-induced cancers (cervix, oropharyngeal and anal cancers) and multiple myeloma.
Cancer – Oncolytic Viruses I 425. Lentiviral Vector Platforrm To Develop AntiCancer Immunotherapies
Deborah Revaud,1 Sophie Agaugue,1 Emeline Sarry,1 Ana Bejanariu,1 Cecile Bauche.1 1 THERAVECTYS, Villejuif, France.
In immunotherapy of cancer, different vaccination strategies have been developed and evaluated in clinical trials, e.g. monoclonal antibodies, injection of DNA, peptides, proteins, autologous cells (tumoral cells, dendritic cells (DC) or T cells) processed in vitro or even direct injection of modified viral vectors. THERAVECTYS is a Paris-based, privately-owned, fully integrated discovery & clinical development biotech company, developping lentiviral vectors for human vaccination and immunotherapy applications. THERAVECTYS’ vaccine candidates based on a proprietary lentiviral vector technology enable a direct vaccination mechanism in patients by inducing a broad, intense and long-lasting T cell mediated response. Theravectys’ technology presents several advantages compared to these various vaccination approaches: - Our lentivectors encompasse up to 15kb DNA , the resulting polypeptide hence targeting a combination of different antigens which mobilizes different actors of the anti-tumor immune system - No selection of patients is needed according to HLA haplotype since epitopes are physiologically processed in vivo. - Unlike all other vaccination approaches, this technology allows the transduction of non-dividing cells such as DC, the most potent antigen-presenting cells (APC) of the immune system. - Break of immunotolerance: lentivectors not only provide sufficient amounts of antigens to DCs but also ensure an efficient processing and prolonged presentation of the antigens, an efficient maturation of DCs that possess all the stimulating abilities to induce an efficient activation of T cells - Non replicative and non persistent lentivectors-antigen expressing cells and transduced cells are eliminated by the immune response elicited after injection. - Direct injections in patients, without ex vivo process of autologous cells. - Different pseudotypes of lentivectors are developed to avoid specific immune reaction against the vector during boost injection(s). - Vaccination with lentiviral vectors are more potent and efficient than DNA or peptide or protein vaccination. The phase I/II clinical trial with our first vaccine candidate has been completed in HIV-infected patients. The vaccine is safe, as demonstrated by the clinical trial ongoing in 38 patients (no Serious Adverse Events have been reported during the trial). The interim analysis of the immunological data from the first two cohorts of patients performed by THERAVECTYS demonstrates the ability of the vaccine candidate to elicit multi-specific and poly-functional CD4+ and CD8+ cellular immune responses in vaccinated patients, even at the lowest dose. S168
426. Oncolytic Vaccinia Virus Has Significant Activity and Induces Anti-Tumor Immunity in Bladder Cancer
Kyle Potts,1 Nicole Favis,2 Chad Irwin,2 Ronald Moore,3 David Evans,2 Mary Hitt.1 1 Oncology, University of Alberta, Edmonton, AB, Canada; 2 Medical Microbiology and Immunology, University of Alberta, Edmonton, AB, Canada; 3Surgery, University of Alberta, Edmonton, AB, Canada. Introduction:Oncolytic viruses preferentially replicate in and lyse cancer cells while sparing normal cells. The majority of Bladder Cancers (BCa) are diagnosed as non-muscle invasive however, with a recurrence rate of up to 80%, many patients require multiple treatments that often fail, leading to disease progression. In particular, the current standard of care, Bacillus Calmette–Guérin (BCG) carries the risk of infection, including systemic, cystitis, and prostatitis and can be particularly dangerous for immunocompromised patients. Additionally, around 30% of patients fail BCG therapy and cystectomy remains the standard treatment in these cases. Here we examine the potential of an oncolytic vaccinia virus (VAC) in BCa. VAC replication requires an abundant supply of dNTPs, but because cellular enzymes that synthesize dNTPs are degraded at the end of Sphase, VAC expresses its own biosynthetic enzymes including both I4L (large, R1) and F4L (small, R2) subunits of the heterodimeric ribonucleotide reductase. We have shown that deleting the F4L gene inhibits virus replication in normal cells and reduces virulence in a mouse model while retaining replication proficiency in BCa cells. Methods: We developed VACs tagged with the gene encoding mCherry and deletions in F4L, J2R (viral thymidine kinase) or both. Activity of these VACs were evaluated in a panel of human and rodent BCa or normal bladder cell lines and primary fibroblasts in vitro. The immunocompetent orthotopic AY-27 rat and RT112-luc immunocompromised mouse models were used to assess the VACs in vivo. The AY-27 model is being used to determine the mechanism of antitumor immunity after VAC treatment. Finally, BCG resistant cells are being examined for their ability to support virus infection and replication. Results: Cytotoxicity assays show a high degree of cell killing in infections with VACs. Highly efficient VAC replication was seen in our BCa cells while limited replication was seen in normal bladder cells or primary fibroblasts. We show that our VACs selectively replicated in both the orthotopic AY-27 immunocompetent and RT112-luc immunocompromised models causing significant tumor regression or complete ablation with limited or no toxicity. Immunocompetent rats treated with the VACs developed a protective anti-tumor immunity that was evident by tumor rejection upon challenge. Finally, examination of VAC replication in BCG-refractory bladder cancer cell lines will be presented. Conclusions: Our data indicate that the VACs have retained their replication proficiency and cytotoxicity in BCa cells despite deletions Molecular Therapy Volume 23, Supplement 1, May 2015 Copyright © The American Society of Gene & Cell Therapy
