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Safe engineering of CAR T cells for adoptive cell therapy of cancer using long-term episomal gene transfer
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Uppsala University, Science for Life Laboratory, SciLifeLab. Uppsala University. (Magnus Essand group)
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Uppsala University, Science for Life Laboratory, SciLifeLab. (Magnus Essand group)
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Uppsala University, Science for Life Laboratory, SciLifeLab. (Magnus Essand group)
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Medicine, Department of Immunology, Genetics and Pathology, Clinical Immunology. Uppsala University, Science for Life Laboratory, SciLifeLab. (Magnus Essand group)
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2016 (English)In: EMBO Molecular Medicine, ISSN 1757-4676, E-ISSN 1757-4684, Vol. 8, no 7, 702-711 p.Article in journal (Refereed) Published
Abstract [en]

Chimeric antigen receptor (CAR) T-cell therapy is a new successful treatment for refractory B-cell leukemia. Successful therapeutic outcome depends on long-term expression of CAR transgene in T cells, which is achieved by delivering transgene using integrating gamma retrovirus (RV) or lentivirus (LV). However, uncontrolled RV/LV integration in host cell genomes has the potential risk of causing insertional mutagenesis. Herein, we describe a novel episomal long-term cell engineering method using non-integrating lentiviral (NILV) vector containing a scaffold/matrix attachment region (S/MAR) element, for either expression of transgenes or silencing of target genes. The insertional events of this vector into the genome of host cells are below detection level. CD19 CAR T cells engineered with a NILV-S/MAR vector have similar levels of CAR expression as T cells engineered with an integrating LV vector, even after numerous rounds of cell division. NILV-S/MAR-engineered CD19 CAR T cells exhibited similar cytotoxic capacity upon CD19(+) target cell recognition as LV-engineered T cells and are as effective in controlling tumor growth in vivo We propose that NILV-S/MAR vectors are superior to current options as they enable long-term transgene expression without the risk of insertional mutagenesis and genotoxicity.

Place, publisher, year, edition, pages
2016. Vol. 8, no 7, 702-711 p.
National Category
Immunology in the medical area
Identifiers
URN: urn:nbn:se:uu:diva-300197DOI: 10.15252/emmm.201505869PubMedID: 27189167OAI: oai:DiVA.org:uu-300197DiVA: diva2:951065
Note

GT och MR delar på andraförfattarskapet.

Available from: 2016-08-05 Created: 2016-08-05 Last updated: 2016-09-02Bibliographically approved
In thesis
1. Improvement of adoptive T-cell therapy for Cancer
Open this publication in new window or tab >>Improvement of adoptive T-cell therapy for Cancer
2016 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Cancer immunotherapy has recently made remarkable clinical progress. Adoptive transfer of T-cells engineered with a chimeric antigen receptor (CAR) against CD19 has been successful in treatment of B-cell leukemia. Patient’s T-cells are isolated, activated, transduced with a vector encoding the CAR molecule and then expanded before being transferred back to the patient. However some obstacles restrict its success in solid tumors. This thesis explores different aspects to improve CAR T-cells therapy of cancer.

Ex vivo expanded T-cells are usually sensitive to the harsh tumor microenvironment after reinfusion. We developed a novel expansion method for T-cells, named AEP, by using irradiated and preactivated allo-sensitized allogeneic lymphocytes (ASALs) and allogeneic mature dendritic cells (DCs). AEP-expanded T-cells exhibited better survival and cytotoxic efficacy under oxidative and immunosuppressive stress, compared to T-cells expanded with established procedures.

Integrating retro/lentivirus (RV/LV) used for CAR expressions randomly integrate in the T-cell genome and has the potential risk of causing insertional mutagenesis. We developed a non-integrating lentiviral (NILV) vector containing a scaffold matrix attachment region (S/MAR) element (NILV-S/MAR) for T-cells transduction. NILV-S/MAR-engineered CAR T-cells display similar cytotoxicity to LV-engineered CAR T-cells with undetectable level of insertional event, which makes them safer than CAR T-cells used in the clinic today.

CD19-CAR T-cells have so far been successful for B-cell leukemia but less successful for B-cell lymphomas, which present semi-solid structure with an immunosuppressive microenvironment. We have developed CAR T-cells armed with H. pylorineutrophil-activating protein (HP-NAP). HP-NAP is a major virulence factor and plays important role in T-helper type 1 (Th1) polarizing. NAP-CAR T-cells showed the ability to mature DCs, attract innate immune cells and increase secretion of Th1 cytokines and chemokines, which presumably leads to better CAR T-cell therapy for B-cell lymphoma.

Allogeneic-DCs (alloDCs) were used to further alter tumor microenvironment. The premise relies on initiation of an allo-reactive immune response for cytokine and chemokines secretion, as well as stimulation of T-cell response by bringing in tumor-associated antigen. We demonstrated that alloDCs promote migration and activation of immune cells and prolong the survival of tumor-bearing mice by attracting T-cells to tumors and reverse the immune suppressive tumor microenvironment.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis, 2016. 65 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, ISSN 1651-6206 ; 1247
Keyword
CAR T-cell therapy; AEP expansion protocol; scaffold matrix attachment region; non-integrating lentivirus; H. pylori Neutrophil-activating protein; allogeneic DCs
National Category
Immunology in the medical area
Research subject
Clinical Immunology
Identifiers
urn:nbn:se:uu:diva-300210 (URN)978-91-554-9661-6 (ISBN)
Public defence
2016-10-06, Rudbecksalen, Dag Hammarskjoldsv 20 Rudbeck laboratory, Uppsala, 09:30 (English)
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Available from: 2016-09-15 Created: 2016-08-05 Last updated: 2016-09-22

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