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Ixovex, a novel oncolytic E1B-mutated adenovirus
abstracts
Annals of Oncology
Part-time employment: Thermo Fisher Scientific. D. Topacio-Hall: Full / Part-time employment: Thermo Fisher Scientific. G. Lowman: Full / Part-time employment: Thermo Fisher Scientific. All other authors have declared no conflicts of interest.
1216P
Ixovex, a novel oncolytic E1B-mutated adenovirus
M. Anwar1, G. Alusi2, D. Oberg3 Ear Nose and Throat, Chelsea and Westminster Hospital - NHS Trust, London, UK, 2 Head and Neck, UCH, London, UK, 3Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala Biomedical Centre, Uppsala, Sweden
1
Background: There is a great demand for improved oncolytic viruses that selectively replicate inside cancer cells while sparing normal tissues. Here we describe a novel oncolytic adenovirus, Ixovex, that obtains a cancer selective replication phenotype by modulating the level of expression of the different alternatively spliced E1B mRNA isoforms. Methods: In our work to probe the function of the E1B proteins we generated a virus mutant, the Ixovex virus, in which the SA1 3’ splice acceptor site sequence CAG:GA was mutated to CgG:GA. This single nucleotide point mutation (genomic location 3216) changed aa 399 in the E1B-496R protein from an arginine (AGG) to a glycine (gGG). IxoVex was validated for its viral replication (TCID-50 assay) and cytotoxicity (MTS assay) in a panel of normal and cancer cell lines. Animal studies for anti-tumour efficacy studies were performed in nude mice. Results: Ixovex is a recombinant adenovirus that carries a single point mutation in the E1B-93R 3’ splice acceptor site that results in an overexpression of the E1B-156R splice isoform. Further, the mutation resulted in a loss of E1B-496R (E1B-55K) protein expression and as a consequence a failure of Ixovex to efficiently degrade the p53 tumour suppressor protein. Ixovex significantly inhibited tumour growth and prolonged survival of mice in an immune-deficient lung carcinoma tumour model. In complementation experiments overexpression of E1B-156R was shown to increase the oncolytic index of both Ad5wt and ONYX-015. In contrast to prior art viruses of similar type, Ixovex includes a functional E3B region for better in vivo efficacy. Ixovex virus, throughout this study has been proven to be by far the superior virus in potency compared to ONYX-015. Conclusions: The decrease in toxicity and the inhibition of replication in normal cells indicate an astounding safety profile of Ixovex. The observation that the ONYX-015 virus replicated better in normal cells compared to Ixovex is intriguing considering that the ONYX-015 virus carries a large deletion that removes the coding sequences and splice signals needed to express the E1B-496R, E1B-93R and E1B-156R proteins. Ixovex significantly inhibit tumour growth in a human lung carcinoma animal model and that ONYX-015 does not. Legal entity responsible for the study: The authors. Funding: IxoGen. Disclosure: M. Anwar: Shareholder / Stockholder / Stock options: IxoGen. G. Alusi: Shareholder / Stockholder / Stock options: IxoGen. D. Oberg: Shareholder / Stockholder / Stock options: IxoGen.
Volume 30 | Supplement 5 | October 2019
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Anti-CSPG4 DNA vaccination as a promising strategy for the treatment of CSPG41 tumours: A comparative oncology trial
F. Riccardo1, L. Tarone1, G. Barutello1, M. Arigoni1, D. Giacobino2, S. Iussich2, S. Occhipinti3, S. Ferrone4, P. Buracco2, F. Cavallo1 1 Department of Molecular Biotechnology And Health Sciences, Universit a degli Studi di a degli Studi di Torino, Torino, Turin, Italy, 2Department of Veterinary Sciences, Universit 3 Grugliasco, Italy, Center for Experimental Research and Medical Studies (CERMS), Universit a degli Studi di Torino, Turin, Italy, 4Department of Surgery, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA Background: Among the most interesting targets for immunotherapeutic approaches, the Chondroitin Sulfate Proteoglycan (CSPG)4 stands out, with low expression in healthy tissues, high expression in several solid tumors and a key role in cancer progression. Because of the translational power of dogs as pre-clinical models for human malignancies and the CSPG4 over-expression by both human and canine malignant melanoma (MM), we demonstrated the safety and the clinical effectiveness of a xenogeneic human (Hu)-CSPG4 DNA vaccine in client-owned canine patients with stage IIIII surgically resected CSPG4þ MM. However, Hu-CSPG4 vaccine was barely effective in activating human T cells from healthy donors in vitro. Based on these results, we aimed to increase the translational power of our approach and to extend it for the treatment of CSPG4þ tumors other than MM. Methods: We generated a hybrid plasmid, derived in part from the Hu- and in part from the dog (Do)-CSPG4 sequences (HuDo-CSPG4). We tested the safety, immunogenicity and anti-tumor potential of HuDo-CSPG4 DNA vaccine in mice, in dogs with stage II-IV surgically resected CSPG4þ MM and in a human setting in vitro. We evaluated the potential of CSPG4-immune-targeting also for osteosarcoma (OSA). Results: Chimeric HuDo-CSPG4 vaccination is strongly immunogenic in mice. In canine patients, the procedure is safe and induces antibodies (Ab) against both Huand Do-CSPG4, with a higher affinity and anti-tumor potential as compared to HuCSPG4. Clinically, HuDo-CSPG4 is effective in increasing the overall survival of vaccinated canine MM patients as compared to controls. Data obtained in vitro with T cells from human healthy donors suggest HuDo-CSPG4 is more immunogenic than HuCSPG4. Moreover, CSPG4 over-expression was demonstrated in human and canine OSA. Interestingly, anti-CSPG4 specific monoclonal Ab and sera derived from CSPG4 vaccinated canine MM patients showed to significantly inhibit human and canine CSPG4þ OSA cell proliferation, migration and osteospheres generation. Conclusions: These results provide the rationale to propose HuDo-CSPG4 vaccination for the treatment of canine CSPG4þ tumors, to be successfully translated in a human setting. Legal entity responsible for the study: Federica Cavallo. Funding: Associazione Italiana per la Ricerca sul Cancro (AIRC), Italian Ministry of Health. Disclosure: All authors have declared no conflicts of interest.
1218P
Antitumor activity, immunogenicity and safety of a novel PD-1 vaccine in combination with two chimeric HER-2 peptide vaccine in syngeneic Balb/c, C57Bl/6 models and in beagle dogs
P.T.P. Kaumaya1, J. Overholser2, L. Guo2, Y. Yang3, M. Penichet4, B. Sayanjali2, N. Ede5, L. Chong6, T. Bekaii-Saab7 1 OB/Gyn, James Comprehensive Cancer Center, Ohio State University Medical Center, Columbus, OH, USA, 2OB/Gyn, The Ohio State University Medical Center, Columbus, OH, USA, 3Neurology, The Ohio State University Medical Center, Columbus, OH, USA, 4 Medicine, UCLA, Los Angeles, CA, USA, 5Technology, Imugene, Carlton, Australia, 6 Imugene, Imugene, Sydney, Australia, 7Medical Oncology, Mayo Clinic Cancer Center, Phoenix, AZ, USA Background: Anti-PD-1 blocking monoclonal antibodies (mAbs) such as nivolumab or pembrolizumab have shown potent anti-tumor effects in different metastatic cancers. Similarly, anti-HER-2 mAbs such as Herceptin and Perjeta are approved for breast cancer. These mAbs are only effective in 10-25% of cancer patients, and are limited by high costs, side effects and development of resistance. Contrary to treatment with monoclonal antibodies, chimeric B-cell cancer vaccines incorporating a “promiscuous” T cell epitope have the advantage of safer and cheaper alternatives, producing a specific immune response that induces memory B & T cell responses, while reducing immune evasion, suppression and resistance. Methods: We have translated two HER-2 combination peptide vaccines (B-Vaxx) to the clinic in a Phase 1/2b trial to safely deliver curative and transformative cancer immunotherapies to advanced cancer patients. We have created and established the development of a novel B-cell peptide vaccine (Key-Vaxx) with high immunogenicity that binds to human PD-1 and produces tumor inhibition in vivo in two animal models of colon cancer. We will describe the CT-26, CT26-HER2 and MC38 tumor models in Balb/c and C57/Bl6 used to test for synergistic effects of anti-PD1 immunization therapy in combination with anti-HER2 immunization therapy. The immunogenicity/ optimal immunologic dose and toxicity profile was investigated in beagle dogs. Results: We show robust HER-2 and PD-1 antibody responses in mice, beagle dogs and non-human primates. The individual (PD-1) or combined vaccination (B-Vaxx)) was effective in reducing tumor growth in a Balb/c and C57Bl/6 syngeneic model of colon
doi:10.1093/annonc/mdz253 | v497
Downloaded from https://academic.oup.com/annonc/article-abstract/30/Supplement_5/mdz253.042/5577113 by guest on 27 October 2019
Methods: Total RNA was extracted from pre- and post-treatment PBL from 13 therapy recipients (6 responders, 7 non-Responders) including extended longitudinal samples for four responders. TCRB sequencing was performed via the Oncomine TCRB-LR assay using 25ng total RNA as input. We evaluated T cell clonal expansion and TCR convergence as potential biomarkers of response. Results: TCR convergence values were elevated in pretreatment PBL of responders compared to non-responders (mean frequency .012 vs .006; p¼.01, Wilcoxon), and remained elevated in responders up to 25 weeks post treatment. TCR evenness (normalized Shannon entropy) decreased at week 5 compared to baseline (p¼.01, one-sided student’s t-test), indicating increased clonal expansion following treatment. Conclusions: These data suggest that peripheral blood TCRB convergence may serve as a predictive or prognostic biomarker for response to dendritic cell-based immunotherapy. Our finding of increased T cell clonal expansion at week 5 of treatment supports the notion that TCR sequencing may serve as a tool for the measurement of pharmacodynamic markers of therapeutic agent activity. Ongoing and future studies will further clarify the utility of TCR convergence and clonal expansion as immune repertoire biomarkers. Clinical trial identification: NCT01876212. Legal entity responsible for the study: University of Pittsburgh School of Medicine & Hillman Cancer Center. Funding: Thermo Fisher Scientific. Disclosure: L. Quagliata: Full / Part-time employment: Thermo Fisher Scientific. T. Looney: Full /
Annals of Oncology
Part-time employment: Thermo Fisher Scientific. D. Topacio-Hall: Full / Part-time employment: Thermo Fisher Scientific. G. Lowman: Full / Part-time employment: Thermo Fisher Scientific. All other authors have declared no conflicts of interest.
1216P
Ixovex, a novel oncolytic E1B-mutated adenovirus
M. Anwar1, G. Alusi2, D. Oberg3 Ear Nose and Throat, Chelsea and Westminster Hospital - NHS Trust, London, UK, 2 Head and Neck, UCH, London, UK, 3Department of Medical Biochemistry and Microbiology, Uppsala University, Uppsala Biomedical Centre, Uppsala, Sweden
1
Background: There is a great demand for improved oncolytic viruses that selectively replicate inside cancer cells while sparing normal tissues. Here we describe a novel oncolytic adenovirus, Ixovex, that obtains a cancer selective replication phenotype by modulating the level of expression of the different alternatively spliced E1B mRNA isoforms. Methods: In our work to probe the function of the E1B proteins we generated a virus mutant, the Ixovex virus, in which the SA1 3’ splice acceptor site sequence CAG:GA was mutated to CgG:GA. This single nucleotide point mutation (genomic location 3216) changed aa 399 in the E1B-496R protein from an arginine (AGG) to a glycine (gGG). IxoVex was validated for its viral replication (TCID-50 assay) and cytotoxicity (MTS assay) in a panel of normal and cancer cell lines. Animal studies for anti-tumour efficacy studies were performed in nude mice. Results: Ixovex is a recombinant adenovirus that carries a single point mutation in the E1B-93R 3’ splice acceptor site that results in an overexpression of the E1B-156R splice isoform. Further, the mutation resulted in a loss of E1B-496R (E1B-55K) protein expression and as a consequence a failure of Ixovex to efficiently degrade the p53 tumour suppressor protein. Ixovex significantly inhibited tumour growth and prolonged survival of mice in an immune-deficient lung carcinoma tumour model. In complementation experiments overexpression of E1B-156R was shown to increase the oncolytic index of both Ad5wt and ONYX-015. In contrast to prior art viruses of similar type, Ixovex includes a functional E3B region for better in vivo efficacy. Ixovex virus, throughout this study has been proven to be by far the superior virus in potency compared to ONYX-015. Conclusions: The decrease in toxicity and the inhibition of replication in normal cells indicate an astounding safety profile of Ixovex. The observation that the ONYX-015 virus replicated better in normal cells compared to Ixovex is intriguing considering that the ONYX-015 virus carries a large deletion that removes the coding sequences and splice signals needed to express the E1B-496R, E1B-93R and E1B-156R proteins. Ixovex significantly inhibit tumour growth in a human lung carcinoma animal model and that ONYX-015 does not. Legal entity responsible for the study: The authors. Funding: IxoGen. Disclosure: M. Anwar: Shareholder / Stockholder / Stock options: IxoGen. G. Alusi: Shareholder / Stockholder / Stock options: IxoGen. D. Oberg: Shareholder / Stockholder / Stock options: IxoGen.
Volume 30 | Supplement 5 | October 2019
1217P
Anti-CSPG4 DNA vaccination as a promising strategy for the treatment of CSPG41 tumours: A comparative oncology trial
F. Riccardo1, L. Tarone1, G. Barutello1, M. Arigoni1, D. Giacobino2, S. Iussich2, S. Occhipinti3, S. Ferrone4, P. Buracco2, F. Cavallo1 1 Department of Molecular Biotechnology And Health Sciences, Universit a degli Studi di a degli Studi di Torino, Torino, Turin, Italy, 2Department of Veterinary Sciences, Universit 3 Grugliasco, Italy, Center for Experimental Research and Medical Studies (CERMS), Universit a degli Studi di Torino, Turin, Italy, 4Department of Surgery, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA Background: Among the most interesting targets for immunotherapeutic approaches, the Chondroitin Sulfate Proteoglycan (CSPG)4 stands out, with low expression in healthy tissues, high expression in several solid tumors and a key role in cancer progression. Because of the translational power of dogs as pre-clinical models for human malignancies and the CSPG4 over-expression by both human and canine malignant melanoma (MM), we demonstrated the safety and the clinical effectiveness of a xenogeneic human (Hu)-CSPG4 DNA vaccine in client-owned canine patients with stage IIIII surgically resected CSPG4þ MM. However, Hu-CSPG4 vaccine was barely effective in activating human T cells from healthy donors in vitro. Based on these results, we aimed to increase the translational power of our approach and to extend it for the treatment of CSPG4þ tumors other than MM. Methods: We generated a hybrid plasmid, derived in part from the Hu- and in part from the dog (Do)-CSPG4 sequences (HuDo-CSPG4). We tested the safety, immunogenicity and anti-tumor potential of HuDo-CSPG4 DNA vaccine in mice, in dogs with stage II-IV surgically resected CSPG4þ MM and in a human setting in vitro. We evaluated the potential of CSPG4-immune-targeting also for osteosarcoma (OSA). Results: Chimeric HuDo-CSPG4 vaccination is strongly immunogenic in mice. In canine patients, the procedure is safe and induces antibodies (Ab) against both Huand Do-CSPG4, with a higher affinity and anti-tumor potential as compared to HuCSPG4. Clinically, HuDo-CSPG4 is effective in increasing the overall survival of vaccinated canine MM patients as compared to controls. Data obtained in vitro with T cells from human healthy donors suggest HuDo-CSPG4 is more immunogenic than HuCSPG4. Moreover, CSPG4 over-expression was demonstrated in human and canine OSA. Interestingly, anti-CSPG4 specific monoclonal Ab and sera derived from CSPG4 vaccinated canine MM patients showed to significantly inhibit human and canine CSPG4þ OSA cell proliferation, migration and osteospheres generation. Conclusions: These results provide the rationale to propose HuDo-CSPG4 vaccination for the treatment of canine CSPG4þ tumors, to be successfully translated in a human setting. Legal entity responsible for the study: Federica Cavallo. Funding: Associazione Italiana per la Ricerca sul Cancro (AIRC), Italian Ministry of Health. Disclosure: All authors have declared no conflicts of interest.
1218P
Antitumor activity, immunogenicity and safety of a novel PD-1 vaccine in combination with two chimeric HER-2 peptide vaccine in syngeneic Balb/c, C57Bl/6 models and in beagle dogs
P.T.P. Kaumaya1, J. Overholser2, L. Guo2, Y. Yang3, M. Penichet4, B. Sayanjali2, N. Ede5, L. Chong6, T. Bekaii-Saab7 1 OB/Gyn, James Comprehensive Cancer Center, Ohio State University Medical Center, Columbus, OH, USA, 2OB/Gyn, The Ohio State University Medical Center, Columbus, OH, USA, 3Neurology, The Ohio State University Medical Center, Columbus, OH, USA, 4 Medicine, UCLA, Los Angeles, CA, USA, 5Technology, Imugene, Carlton, Australia, 6 Imugene, Imugene, Sydney, Australia, 7Medical Oncology, Mayo Clinic Cancer Center, Phoenix, AZ, USA Background: Anti-PD-1 blocking monoclonal antibodies (mAbs) such as nivolumab or pembrolizumab have shown potent anti-tumor effects in different metastatic cancers. Similarly, anti-HER-2 mAbs such as Herceptin and Perjeta are approved for breast cancer. These mAbs are only effective in 10-25% of cancer patients, and are limited by high costs, side effects and development of resistance. Contrary to treatment with monoclonal antibodies, chimeric B-cell cancer vaccines incorporating a “promiscuous” T cell epitope have the advantage of safer and cheaper alternatives, producing a specific immune response that induces memory B & T cell responses, while reducing immune evasion, suppression and resistance. Methods: We have translated two HER-2 combination peptide vaccines (B-Vaxx) to the clinic in a Phase 1/2b trial to safely deliver curative and transformative cancer immunotherapies to advanced cancer patients. We have created and established the development of a novel B-cell peptide vaccine (Key-Vaxx) with high immunogenicity that binds to human PD-1 and produces tumor inhibition in vivo in two animal models of colon cancer. We will describe the CT-26, CT26-HER2 and MC38 tumor models in Balb/c and C57/Bl6 used to test for synergistic effects of anti-PD1 immunization therapy in combination with anti-HER2 immunization therapy. The immunogenicity/ optimal immunologic dose and toxicity profile was investigated in beagle dogs. Results: We show robust HER-2 and PD-1 antibody responses in mice, beagle dogs and non-human primates. The individual (PD-1) or combined vaccination (B-Vaxx)) was effective in reducing tumor growth in a Balb/c and C57Bl/6 syngeneic model of colon
doi:10.1093/annonc/mdz253 | v497
Downloaded from https://academic.oup.com/annonc/article-abstract/30/Supplement_5/mdz253.042/5577113 by guest on 27 October 2019
Methods: Total RNA was extracted from pre- and post-treatment PBL from 13 therapy recipients (6 responders, 7 non-Responders) including extended longitudinal samples for four responders. TCRB sequencing was performed via the Oncomine TCRB-LR assay using 25ng total RNA as input. We evaluated T cell clonal expansion and TCR convergence as potential biomarkers of response. Results: TCR convergence values were elevated in pretreatment PBL of responders compared to non-responders (mean frequency .012 vs .006; p¼.01, Wilcoxon), and remained elevated in responders up to 25 weeks post treatment. TCR evenness (normalized Shannon entropy) decreased at week 5 compared to baseline (p¼.01, one-sided student’s t-test), indicating increased clonal expansion following treatment. Conclusions: These data suggest that peripheral blood TCRB convergence may serve as a predictive or prognostic biomarker for response to dendritic cell-based immunotherapy. Our finding of increased T cell clonal expansion at week 5 of treatment supports the notion that TCR sequencing may serve as a tool for the measurement of pharmacodynamic markers of therapeutic agent activity. Ongoing and future studies will further clarify the utility of TCR convergence and clonal expansion as immune repertoire biomarkers. Clinical trial identification: NCT01876212. Legal entity responsible for the study: University of Pittsburgh School of Medicine & Hillman Cancer Center. Funding: Thermo Fisher Scientific. Disclosure: L. Quagliata: Full / Part-time employment: Thermo Fisher Scientific. T. Looney: Full /