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Cancer Immunotherapy: Evolving Oncolytic viruses and CAR T-cells
Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology. (Magnus Essand)ORCID iD: 0000-0003-2685-0575
2016 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

In the last decade cancer immunotherapy has taken huge strides forward from bench to bedside and being approved as drugs. Cancer immunotherapy harnesses the power of patient’s own immune system to fight cancer. Approaches are diverse and include antibodies, therapeutic vaccines, adoptively transferred T-cells, immune checkpoint inhibitors, oncolytic viruses and immune cell activators such as toll-like receptor (TLR) agonists. Excellent clinical responses have been observed for certain cancers with checkpoint antibodies and chimeric antigen receptor (CAR)-engineered T-cells. It is however becoming evident that strategies need to be combined for broader effective treatment responses because cancers evolve to escape immune recognition. A conditionally replication-competent oncolytic adenovirus (Ad5PTDf35-[Δ24]) was engineered to secrete Helicobacter pylori Neutrophil Activating Protein (HP-NAP, a TLR-2 agonist) to combine viral oncolysis and immune stimulation. Treatment with Ad5PTDf35-[Δ24-sNAP] improved survival of mice bearing human neuroendocrine tumors (BON). Expression of HP-NAP in the tumor microenvironment promoted neutrophil infiltration, proinflammatory cytokine secretion and increased necrosis. We further studied the ability of HP-NAP to activate dendritic cells (DCs) a key player in priming T-cell responses. HP-NAP phenotypically matured and activated DCs to secrete the T-helper type-1 (Th-1) polarizing cytokine IL-12. HP-NAP-matured DCs were functional; able to migrate to draining lymph nodes and prime antigen-specific T-cell proliferation. CAR T-cells were engineered to secrete HP-NAP upon T-cell activation. Secreted HP-NAP was able to mature DCs, leading to a reciprocal effect on the CAR T-cells with improved cytotoxicity in vitro. Semliki Forest virus (SFV), an oncolytic virus with natural neuro-tropism was tagged with central nervous system (CNS)-specific microRNA target sequences for miR124, miR125 and miR134 to selectively attenuate virus replication in healthy CNS cells. Systemic infection of mice with the SFV4miRT did not cause encephalitis, while it retained its ability to replicate in tumor cells and cure a big proportion of mice bearing syngeneic neuroblastoma and gliomas. Therapeutic efficacy of SFV4miRT inversely correlated with type-I antiviral interferon response (IFN-β) mounted by tumor cells. In summary, combining immunotherapeutic strategies with HP-NAP is a promising approach to combat cancers and SFV4miRT is an excellent candidate for treatment of neuroblastomas and gliomas.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2016. , 77 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1258
Keyword [en]
Oncolytic virus, Adenovirus, Semliki Forest virus, Cancer immunology, Chimeric antigen receptor T-cells
National Category
Immunology in the medical area
Research subject
Immunology; Oncology; Biology with specialization in Molecular Biotechnology
Identifiers
URN: urn:nbn:se:uu:diva-302891ISBN: 978-91-554-9705-7OAI: oai:DiVA.org:uu-302891DiVA: diva2:998737
Public defence
2016-11-21, Rudbecksalen, Dag Hammarskjöldsväg 20, Uppsala, 09:15 (English)
Opponent
Supervisors
Available from: 2016-10-31 Created: 2016-09-12 Last updated: 2016-11-02
List of papers
1. An infection-enhanced oncolytic adenovirus secreting H. pylori neutrophil-activating protein with therapeutic effects on neuroendocrine tumors
Open this publication in new window or tab >>An infection-enhanced oncolytic adenovirus secreting H. pylori neutrophil-activating protein with therapeutic effects on neuroendocrine tumors
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2013 (English)In: Molecular Therapy, ISSN 1525-0016, E-ISSN 1525-0024, Vol. 21, no 11, 2008-2018 p.Article in journal (Refereed) Published
Abstract [en]

Helicobacter pylori Neutrophil Activating Protein (HP-NAP) is a major virulence factor involved in H. pylori infection. HP-NAP can mediate anti-tumor effects by recruiting neutrophils and inducing Th1-type differentiation in the tumor microenvironment. It therefore holds strong potential as a therapeutic gene. Here, we armed a replication-selective, infection-enhanced adenovirus with secretory HP-NAP, Ad5PTDf35-[Δ24-sNAP], and evaluated its therapeutic efficacy against neuroendocrine tumors. We observed that it could specifically infect and eradicate a wide range of tumor cells lines from different origin in vitro. Insertion of secretory HP-NAP did not affect the stability or replicative capacity of the virus and infected tumor cells could efficiently secrete HP-NAP. Intratumoral administration of the virus in nude mice xenografted with neuroendocrine tumors improved median survival. Evidence of biological HP-NAP activity was observed 24 hours after treatment with neutrophil infiltration in tumors and an increase of proinflammatory cytokines such as TNF-α and MIP2-α in the systemic circulation. Furthermore, evidence of Th1-type immune polarization was observed as a result of increase in IL-12/23 p40 cytokine concentrations 72 hours post-virus administration. Our observations suggest that HP-NAP can serve as a potent immunomodulator in promoting anti-tumor immune response in the tumor microenvironment and enhance the therapeutic effect of oncolytic adenovirus.

Keyword
Helicobacter pylori, Neutrophil Activating Protein, adenovirus, cancer therapy
National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Research subject
Molecular Medicine; Clinical Virology; Medical Virology; Clinical Immunology
Identifiers
urn:nbn:se:uu:diva-203649 (URN)10.1038/mt.2013.153 (DOI)000326937000007 ()23817216 (PubMedID)
Funder
Swedish Cancer Society, 10‐0105Swedish Cancer Society, 10‐0552Swedish Research Council, K2013‐55X‐22191‐01‐3
Note

De två (2) sista författarna delar sistaförfattarskapet.

Open access under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs 3.0 Unported License (CC BY-NC-ND). 2013.

Supported funds:

The Swedish Children Cancer Foundation(PROJ10/027), Gunnar Nilsson’s Cancer Foundation, Marcus and Marianne Wallenberg’sFoundation

Available from: 2013-07-16 Created: 2013-07-16 Last updated: 2016-09-30Bibliographically approved
2. Vector-Encoded Helicobacter pylori Neutrophil-Activating Protein Promotes Maturation of Dendritic Cells with Th1 Polarization and Improved Migration
Open this publication in new window or tab >>Vector-Encoded Helicobacter pylori Neutrophil-Activating Protein Promotes Maturation of Dendritic Cells with Th1 Polarization and Improved Migration
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2014 (English)In: Journal of Immunology, ISSN 0022-1767, E-ISSN 1550-6606, Vol. 193, no 5, 2287-2296 p.Article in journal (Refereed) Published
Abstract [en]

Helicobacter pylori neutrophil-activating protein (HP-NAP) is a major virulence factor involved in H. pylori infection. Both HP-NAP protein and oncolytic viruses encoding HP-NAP have been suggested as immunotherapeutic anticancer agents and adjuvants for vaccination but with little known about its mode of action to activate adaptive immunity. Dendritic cells (DCs) are key players in bridging innate and adaptive immune responses, and in this study we aim to evaluate the effect of HP-NAP on DC maturation, migration, and induction of adaptive immune response. Maturation markers CD83, CD80, CD86, HLA-DR, CD40, and CCR7 were upregulated on human DCs after treatment with supernatants from HP-NAP adenovirus-infected cells. HP-NAP-activated DCs had a Th1 cytokine secretion profile, with high IL-12 and relatively low IL-10 secretion, and migrated toward CCL19. Ag-specific T cells were efficiently expanded by Ag-presenting HP-NAP-activated DCs, which is an important property of functionally mature DCs. Furthermore, intradermal injections of HP-NAP-encoding adenovirus in C57BL/6 mice enhanced resident DC migration to draining lymph nodes, which was verified by imaging lymph nodes by two-photon microscopy and by phenotyping migrating cells by flow cytometry. In conclusion, therapeutic effects of HP-NAP are mediated by maturation of DCs and subsequent activation of Ag-specific T cells in addition to provoking innate immunity.

National Category
Immunology in the medical area
Identifiers
urn:nbn:se:uu:diva-232998 (URN)10.4049/jimmunol.1400339 (DOI)000341140600031 ()
Funder
Swedish Cancer Society, 12-0569Swedish Research Council, K2013-55X-22191-01-3
Available from: 2014-10-10 Created: 2014-09-29 Last updated: 2016-09-30Bibliographically approved
3. CD19 CAR T-cells with induced secretion of Helicobacter Pylori Neutrophil-Activating Protein (HP-NAP) yields improved anti-tumor activity and reduced immunosuppression
Open this publication in new window or tab >>CD19 CAR T-cells with induced secretion of Helicobacter Pylori Neutrophil-Activating Protein (HP-NAP) yields improved anti-tumor activity and reduced immunosuppression
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(English)Manuscript (preprint) (Other academic)
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:uu:diva-300198 (URN)
Available from: 2016-08-12 Created: 2016-08-05 Last updated: 2016-09-30
4. Safe and effective treatment of experimental neuroblastoma and glioblastoma using systemically administered triple microRNA-detargeted oncolytic Semliki Forest virus.
Open this publication in new window or tab >>Safe and effective treatment of experimental neuroblastoma and glioblastoma using systemically administered triple microRNA-detargeted oncolytic Semliki Forest virus.
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2016 (English)In: Clinical Cancer Research, ISSN 1078-0432, E-ISSN 1557-3265Article in journal (Refereed) Epub ahead of print
Abstract [en]

PURPOSE:

Glioblastoma multiforme (GBM) and high-risk neuroblastoma are cancers with poor outcome. Immunotherapy in the form of neurotropic oncolytic viruses is a promising therapeutic strategy for these malignancies. Here we evaluate the oncolytic potential of the neurovirulent and partly interferon (IFN)-β-resistant Semliki Forest virus (SFV)-4 in GBMs and neuroblastomas. To reduce neurovirulence we constructed SFV4miRT, which is attenuated in normal CNS cells through insertion of microRNA target sequences for miR124, miR125, miR134 Experimental Design:Oncolytic activity of SFV4miRT was examined in mouse neuroblastoma and GBM cell lines and in patient-derived human glioblastoma cell cultures (HGCC). In vivo neurovirulence and therapeutic efficacy was evaluated in two syngeneic orthotopic glioma models (CT-2A, GL261) and syngeneic subcutaneous neuroblastoma model (NXS2). The role of IFN-β in inhibiting therapeutic efficacy was investigated.

RESULTS:

The introduction of microRNA target sequences reduced neurovirulence of SFV4 in terms of attenuated replication in mouse CNS cells and ability to cause encephalitis when administered intravenously. A single intravenous injection of SFV4miRT prolonged survival and cured 4 of 8 mice (50%) with NXS2 and 3 of 11 mice (27%) with CT-2A, but not for GL261 tumor bearing mice. In vivo therapeutic efficacy in different tumor models inversely correlated to secretion of IFN-β by respective cells upon SFV4 infection in vitro Similarly, killing efficacy of HGCC lines inversely correlated to IFN-β response and interferon-α⁄β receptor (IFNAR)-1 expression.

CONCLUSIONS:

SFV4miRT has reduced neurovirulence, while retaining its oncolytic potential. SFV4miRT is an excellent candidate for treatment of GBMs and neuroblastomas with low IFN-β secretion.

Place, publisher, year, edition, pages
American Association for Cancer Research, 2016
Keyword
Semliki Forest virus, Glioblastoma, Neuroblastoma, Oncolytic virus immunotherapy, Type-I antiviral response
National Category
Other Basic Medicine
Research subject
Oncology; Biology with specialization in Molecular Biotechnology
Identifiers
urn:nbn:se:uu:diva-303633 (URN)10.1158/1078-0432.CCR-16-0925 (DOI)
Funder
Swedish Research Council, K2013-22191-01-3Swedish Cancer Society, CAN2013/373Swedish Childhood Cancer Foundation, PROJ12/082
Available from: 2016-09-21 Created: 2016-09-21 Last updated: 2016-09-30

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Ramachandran, Mohanraj
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Science for Life Laboratory, SciLifeLabDepartment of Immunology, Genetics and Pathology
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