Abstract 574: Allogenic HER2-CAR T cells overcome intrinsic trastuzumab resistance in vitro and in vivo in a preclinical model of breast cancer
Authors: Oswald Eva , Moiset Gemma , Lashuk Kanstantsin , Vermond Sophie , Hazenoot Monique , Schüler Julia 
Published: June 2022

Abstract
The advent of anti-HER2 therapy has prominently prolonged the time of disease progression and survival for metastatic breast cancer patients, where a decent proportion is suffering from a HER2+ tumor. Beside classical approaches via antibodies against HER2, the use of Chimeric Antigen Receptor T (CAR-T) cells also in a solid cancer context is getting more and more attention. In our study, we evaluated the efficacy of Trastuzumab versus HER2 targeting CAR-T cells in a panel of human cancer cell lines (SK-OV3, Hs578T and JIMT-1) with different HER2 expression levels in vitro in 2D as well as 3D and in vivo in immunocompromised mice. In vitro, the tumor growth and invasion of the CAR-T cells was measured via fluorescence-based live cell imaging. On the last experiment day, a metabolic read-out (CellTiter-Glo, CTG assay) was performed. In vivo, we measured tumor growth inhibition (TGI) via caliper measurement and tumor tissue and hematopoietic organs were analyzed by flow cytometry (FC), Immunohistochemistry (IHC) and multiplex cytokine analysis. The HER2 targeting CAR-T cells eradicated the 3D spheroids of the HER2+ SK-OV3 as well as JIMT-1 in a dose-dependent manner. The untransduced control T cells did not influence the tumor growth in vitro. Trastuzumab displayed efficacy in 2D and 3D in SKOV3. As expected, JIMT-1 cells were resistant to trastuzumab treatment despite their positive HER2 status. The HER2- cell line Hs578T served as a negative control and proved to be resistant to any treatment in this study. The HER2-targeting CAR-T cells were applied in vivo in two different doses to JIMT-1 tumor bearing NSG mice (n=8/group) and tumor volume was measured over time. Again, the CAR-T cells were able to induce a complete remission. In contrast, the tumors displayed progressive disease under therapy with the untransduced T cells or Trastuzumab. Interestingly, the CAR-T cells induced a tumor swelling between eight- and twelve-days post injection very similar to the situation in the patient. Four weeks post injection the human T cells could be detected in spleen, peripheral blood, liver and bone marrow of the JIMT- bearing mice. The implication of this finding for possible side effects of the HER2 CAR´s remains to be elucidated. A PK/PD study is currently ongoing for this purpose. Taken together the 3D live cell imaging platform proved to be a feasible tool for efficacy testing of biologics as well as cellular therapies. Our in house developed HER2 CAR-T cells proved to be specific and effective in eradicating the targeted cancer cells in vitro and in vivo. The mechanism behind the modulated sensitivity of the HER2+ JIMT-1 cells against HER2-targeted treatment will help to shed some light in possible resistance mechanism and hopefully have some translational value for patients suffering from this disease.
Citation Format: Eva Oswald, Gemma Moiset, Kanstantsin Lashuk, Sophie Vermond, Monique Hazenoot, Julia Schüler. Allogenic HER2-CAR T cells overcome intrinsic trastuzumab resistance in vitro and in vivo in a preclinical model of breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 574.