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217 Multi-armored allogeneic MUC-1 CAR T-cells efficiently control triple negative breast cancer tumor growth
  1. Piril Erler,
  2. Tomasz Kurcon,
  3. Jordan Skinner,
  4. Chantel Dixon,
  5. Steven Grudman,
  6. Ben Mumford,
  7. Shipra Das,
  8. Alexander Boyne,
  9. Alexandre Juillerat,
  10. Roman Galetto,
  11. Julien Valton,
  12. Hana Cho,
  13. Laurent Poirot and
  14. Beatriz Aranda-Orgilles
  1. Cellectis, New York, NY, USA


Background CAR T-cell therapy success in solid tumors has been limited due to their complex biology. In solid tumors, lack of tumor-specific antigens, tumor heterogeneity, and immuno-inhibitory nature of tumor microenvironment (TME) demand an optimal therapeutic window where CAR T-cells can be highly efficient while ensuring safety. To address these challenges, we engineered CAR T-cells that i) use the tumor-specific MUC-1 antigen as a discriminatory target and ii) have enhanced therapeutic properties provided by multiple attributes. We focused on TNBC due to poor prognosis and overexpression of MUC1 (~67%) [1]. Here, we describe a universal CAR T-cell therapy for TNBC that can overcome both the host immune rejection and key inhibitory signals from the TME.

Methods We first screened several tumor-specific scFVs for MUC-1 CARs, assessing their binding and safety profiles. Then, we generated allogenic CAR T-cells by leveraging our TALEN® technology. TCR-alpha and B2M were knocked-out to prevent host-versus-graft disease, and to evade host T-cell attack. HLA-E was knocked-in at the B2M-KO site to provide resistance to host NK cell rejection. To increase activity and to overcome inhibitory signals from the TME, we introduced a PD-1 knock-out, a tumor-specific IL-12 release, and TGFBR2 knock-out. We tested these CAR T-cells in vitro using target specificity and cytotoxic assays, and in vivo by assessing tumor growth, survival, and tumor infiltration.

Results Three scFVs we prioritized showed efficient dose-dependent killing of breast cancer cells in vitro. Next, we used subcutaneous and orthotopic models to test CAR T-cells armored with IL-12 inducible release in vivo. Efficient tumor control and increased CAR-T cell infiltration extended mice survival, and notably antitumor response followed a dose-dependent pattern. Importantly, we introduced multiple edits in CAR-T cells with a high degree of efficacy, and precision using TALEN®. Functionally validating these edits, we demonstrated that TGFBR2-KO circumvents the inhibitory effects of TGFβ1, and IL-12 release follows a CAR T-cell activation pattern restricting it to the tumor site for increased safety.

Conclusions Overall, our data demonstrate that we can efficiently generate allogeneic CAR T-cells and equip them through complex engineering to overcome key challenges of solid tumors. We show that MUC-1 CAR T-cells control tumor growth, while infiltrating tumors more efficiently when enhanced with attributes catered towards the TME of TNBC tumors. Altogether, these pre-clinical data suggest that enhanced MUC-1 CAR T-cells could address some of the current challenges in development of CAR-Ts for TNBC patients with unmet medical needs.


  1. Siroy, A., Abdul-Karim, F. W., Miedler, J., Fong, N., Fu, P., Gilmore, H., & Baar, J. MUC1 is expressed at high frequency in early-stage basal-like triple-negative breast cancer. Human Pathology 2013; 44:54.

Ethics Approval All animals in this study were treated humanely and in agreement with IACUC guidelines. IACUC Protocol #:2019-05-10-CEL-02

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