Abstract Programmed Cell Death Protein 1 (PD-1) is a crucial immune checkpoint receptor in the tumor microenvironment. Monoclonal blocking antibodies for this checkpoint were developed to restore exhausted T cells' ability to eliminate tumors. However, pharmaceutical companies outside Latin America produce most of these commercial antibodies in mammalian cells or hybridomas. The microbial expression system in Escherichia coli proposes a faster and lower-cost alternative for producing these molecules. In this scenario and in the context of improving public health and offering national biopharmaceuticals, this project aimed to develop and characterize the Fab version of the monoclonal Nivolumab (anti-PD-1). The synthetic pFab Nivolumab construction was transformed into E. coli standard strain BL21(DE3) for expression, followed by the soluble fraction protein A affinity chromatography to anti-PD-1 purification. The purification yield was 560 ug/L and analysis by SDS-PAGE of purified protein exhibited a single band at ∼25 kDa, suggesting high purity. The antibody functionality was first analyzed by indirect ELISA, evaluating that the least amount of Fab needed to obtain 50% of the PD-1 antigen blockade response (EC50) is ∼1018 nM. Functional analysis to evaluate its ability to modulate immune responses is ongoing. Our partial results demonstrate the successful expression and purification of the Fab anti-PD-1, if confirmed its ability to modulate immune responses, this molecule raises as a potential antitumor therapeutic tool. Citation Format: Marielly Câmara Rocha, Gabriel Correia Lima, Tobias Schatton, Daniela Luz. Development of the biosimilar Fab version of the monoclonal Nivolumab (anti-PD1) as an immunobiological tool for prospecting new antitumor strategies abstract. In: Proceedings of the AACR Immuno-Oncology Conference (AACR IO): Discovery and Innovation in Cancer Immunology: Revolutionizing Treatment through Immunotherapy; 2026 Feb 18-21; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Immunol Res 2026;14(2 Suppl):Abstract nr C054.
Rocha et al. (2026) studied this question.