title: NY-BR-1 specific CAR+ T cells engineered by S/MAR vectors prove effectiveness in pre-clinical models of breast cancer
creator: Berger, Aileen
subject: ddc-500
subject: 500 Natural sciences and mathematics
subject: ddc-570
subject: 570 Life sciences
description: Breast cancer is the most frequent cause of death among cancer diseases in women despite improved treatment options, which makes new and further development of additional therapeutic approaches urgently necessary. Immunotherapy, in particular CAR - based T cell therapy, became an increasingly important therapeutic tool in recent decades due to great success in the treatment of hematological diseases. However, this approach proved to be very challenging in the treatment of solid tumors due to the lack of tumor specific antigens and inefficient persistence and infiltration of CAR+ T cells into these tumors. The breast cancer antigen NY-BR-1, which is overexpressed in more than  70 % of breast cancer tumors and metastases, might serve as a target antigen for CAR - based immunotherapy. The feasibility of this hypothesis was investigated in this study using three different CAR candidates varying in their binding sites within the NY-BR-1 protein. During this process, the efficacy and safety of the individual CAR candidates was evaluated using various in vitro and in vivo  experiments. At the beginning of this work, human and murine NY-BR-1 - expressing cell lines were successfully generated by lentiviral transduction. However, surface expression of the NY-BR-1 protein decreased over time, mainly due to the dependency of NY-BR-1 expression on the G1 cell cycle phase  and degradation by proteasomes. For the generation of murine CAR+ T cells, a novel transfection strategy based on S/MAR DNA vectors (pS/MARter, NanoCMARter) was established. This innovative technology enabled a cost-efficient, fast and very effective generation of CAR - expressing murine T cells and was also successfully applied to human (CAR+) T cells. Indeed, all three CAR candidates, expressed in lentivirally transduced or (S/MAR vector) electroporated T cells, proved to be very effective in eliminating NY-BR-1 - expressing cell lines and primary pleural effusion cells isolated from  breast cancer patients. Additional cross-reactivity analyses with the NY-BR-1.1 protein, which exhibits a high homology to NY-BR-1 and is expressed in the breast tissue and brain, revealed a high safety risk for the application of one particular CAR candidate in clinical trials. The same CAR additionally caused strongly increased blood cytokine levels in NY-BR-1tg/- mice displaying low NY-BR-1 expression levels in the organs. In C57BL/6 wild-type mice, however, all CAR candidates proved to be safe. The CAR+ T cell treatment of tumor bearing mice resulted in significantly delayed tumor growth and prolonged overall survival, but the CAR+ T cells differed in their infiltration efficiency depending on the CAR type. This work demonstrated that NY-BR-1 is an attractive target antigen for CAR - based immunotherapy in  breast cancer patients. A direct comparison of the CAR candidates presented here revealed that one of these three candidates is particularly suitable for use in clinical trials due to its safety profile and high effectiveness.
date: 2024
type: Dissertation
type: info:eu-repo/semantics/doctoralThesis
type: NonPeerReviewed
format: application/pdf
identifier: https://archiv.ub.uni-heidelberg.de/volltextserverhttps://archiv.ub.uni-heidelberg.de/volltextserver/28760/7/Dissertation_AileenBerger.pdf
identifier: DOI:10.11588/heidok.00028760
identifier: urn:nbn:de:bsz:16-heidok-287609
identifier:   Berger, Aileen  (2024) NY-BR-1 specific CAR+ T cells engineered by S/MAR vectors prove effectiveness in pre-clinical models of breast cancer.  [Dissertation]     
relation: https://archiv.ub.uni-heidelberg.de/volltextserver/28760/
rights: info:eu-repo/semantics/openAccess
rights: http://archiv.ub.uni-heidelberg.de/volltextserver/help/license_urhg.html
language: eng