Abstract 4036: BRGSF-HIS and CD3Ε humanized mice: Translatable preclinical mouse models for assessment of T-cell engagers-induced CRS
Authors: Martin-Jeantet Perrine , Renart-Depontieu Florence , Bourre Ludovic , Martin Gaëlle , Pappalardo Angela , Campbell Dean O. , Doerner Astrid , Cherifi Yacine , Sônego Fabiane , Thiam Kader 
Published: June 2022

Abstract
T cell engagers show high efficacy in B cells malignances. High risk of immune-related adverse events, including cytokine release syndrome (CRS), is reported in patients due to on-target off-site effects of T cell engagers. Thus, reliable and translational mouse models are required to predict potential safety issues and investigate their rescue. Here we describe two preclinical models mimicking to some extent CRS induction upon activation with anti-CD3 antibodies.BRGSF (Balb/C Rag2-/-, IL2Rγ-/-, SIRPαNOD and Flt3-/-) is a highly immunodeficient mouse featuring reduced murine myeloid cells. Development of human lymphoid (B and T cells) and myeloid (NK, cDC, pDC and monocytes) compartments upon CD34+ HSC-engraftment are observed in blood, spleen and bone marrow and are stable for over a year, without side effects. Anti-hCD3 mAb, OKT3, administration in BRGSF-HIS mice induced a rapid release of human cytokines (i.e., IL-6, TNF-α, IFN-γ, IL-2) in serum. Pretreatment with Flt3L, enhancing the human myeloid compartment, boosted the production of cytokines. CRS clinical signs such as hypothermia and weight lost were reproduced in OKT3-injected BRGSF-HIS mice. Pretreatment with Tocilizumab (an anti-IL-6R) reduced cytokine production, suggesting a strong role of myeloid cells in the induced-CRS pathogenesis. The main limitation of this model is the lack of a fully functional crosstalk between human immune cells and mouse stroma, which can undermine endothelial cells contribution in CRS. Alternatively, the CD3ε N-terminal epitope knock-in mice expressing the humanized epitope of anti-CD3 clone SP34 (hCD3ε mice) enables the investigation of endothelial versus immune cells dialogue in CRS pathogenesis. hCD3ε mice display a functional CD3-TCR complex and a functional T and B cooperation, as shown by T cell proliferation upon SP34 activation and antigen-specific antibodies production, respectively. Cytotoxic responses were induced by various bispecific antibodies, ex vivo and in vivo. Splenocytes from hCD3ε mice activated with BsAb1 bispecific antibody targeting CD3ε and a tumor-associated antigen (TAA) induced mouse cytokines production (IFN-γ, TNF-α, IL-6, IL-1β, IL-10) in a concentration-dependent manner. A 2nd BSAb targeting CD3ε and an irrelevant TAA didn’t induce any cytokine release, confirming the specificity of the previous cytokine induction.These data suggest that both BRGSF-HIS and hCD3ε models could be used to assess T cell engager-induced CRS in a complementary setting: while BRGSF-HIS mice enable monitoring of any anti-hCD3 agent, the model might undermine the contribution of mouse endothelial cells due to specie specificity barrier of human cytokines and mouse cells. In contrast, hCD3ε mice enable monitoring of cytokine induction upon activation with SP34-derived anti-hCD3 taking into consideration a full crosstalk between immune and endothelial cells.
Citation Format: Perrine Martin-Jeantet, Florence Renart-Depontieu, Ludovic Bourre, Gaëlle Martin, Angela Pappalardo, Dean O. Campbell, Astrid Doerner, Yacine Cherifi, Fabiane Sônego, Kader Thiam. BRGSF-HIS and CD3Ε humanized mice: Translatable preclinical mouse models for assessment of T-cell engagers-induced CRS [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 4036.