Abstract 5309: ATOR-4066, a Neo-X-Prime™ bispecific antibody targeting CD40 and CEACAM5, induces tumor localized immune cell activation in preclinical in vivo tumor model
Authors: Andersson Hampus , Uddbäck Ida , Celander Mona , Johansson Maria , Ljung Lill , Nilsson Anneli , Shetty Amulya Krishna , Jimenez David Gomez , Levin Mattias , Fritzell Sara , von Schantz Laura , Lindstedt Malin , Hägerbrand Karin , Sundstedt Anette , Ellmark Peter 
Published: March 2024

Abstract
ATOR-4066 is a preclinical stage bispecific antibody targeting CD40 and CEACAM5, based on Alligator’s novel Neo-X-Prime™ antibody technology platform. ATOR-4066 induces a neoantigen specific T-cell response by simultaneous binding to CD40 on myeloid cells such as antigen presenting cells (APCs) and to CEACAM5 on tumor cells. CEACAM5 is a tumor-associated antigen highly expressed on several cancer indications such as colorectal and gastric cancer, as well as on tumor-derived material. ATOR-4066 treatment results in (1) CEACAM5-conditional CD40-mediated activation of APCs and (2) enhanced uptake of tumor-derived material, leading to cross-presentation of neoantigen and priming of neoantigen-specific T cells, resulting in enhanced tumor cell killing. We have previously shown evidence of a potent anti-tumor effect of ATOR-4066 in vivo, superior to the effects observed with the corresponding CD40 mAb. Moreover, we have demonstrated CEACAM5-dependent activation of CD40-expressing cells in vitro, and an ability to mediate co-localization of CEACAM5-expressing tumor debris and CD40 expressing APCs. In addition, ATOR-4066 was shown to induce CEACAM5-conditional activation of tumor-infiltrating CD40-expressing immune cells in dissociated primary human tumor material from colorectal and gastric cancer patients. Here we present data further elucidating the mode of action of ATOR-4066 in vivo. Following ATOR-4066 treatment, tumors and lymph nodes were isolated from human CD40 transgenic mice bearing MC38 tumors expressing human CEACAM5. Flow cytometry analysis shows an increased number of tumor-infiltrating immune cells in mice treated with ATOR-4066 compared to treatments with CD40 mAb or vehicle. The study also reveals upregulation of co-stimulatory molecules such as CD86 on DCs and macrophages after treatment with ATOR-4066 compared to groups treated with CD40 mAb or vehicle. The effect was localized to the tumor microenvironment and not observed in tumor draining lymph nodes, confirming previous in vitro results demonstrating CEACAM5-conditional immune activation. Moreover, RNA-sequencing of tumor samples displayed distinct transcriptomic alterations following ATOR-4066 treatment vs. CD40 mAb or vehicle treatment in vivo. In contrast, RNA-sequencing of peripheral blood in ATOR-4066 treated mice showed only small transcriptomic differences compared to the vehicle treated group, but distinct from mice treated with a CD40 mAb, further demonstrating the CEACAM5 dependent response of ATOR-4066. In summary, these preclinical data demonstrate that ATOR-4066 activates infiltrating myeloid cells resulting in increases in activated tumor infiltrating T cells, thus creating a pro-inflammatory tumor microenvironment, which strongly supports further development and clinical testing of ATOR-4066.
Citation Format: Hampus Andersson, Ida Uddbäck, Mona Celander, Maria Johansson, Lill Ljung, Anneli Nilsson, Amulya Krishna Shetty, David Gomez Jimenez, Mattias Levin, Sara Fritzell, Laura von Schantz, Malin Lindstedt, Karin Hägerbrand, Anette Sundstedt, Peter Ellmark. ATOR-4066, a Neo-X-Prime™ bispecific antibody targeting CD40 and CEACAM5, induces tumor localized immune cell activation in preclinical in vivo tumor model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5309.