Journal Article

Effective cancer immunotherapy combining mRNA-encoded bispecific antibodies that induce polyclonal T cell engagement and PD-L1-dependent 4-1BB costimulation

Álvarez-Vallina, Luis, Frago, Susana, Rubio-Pérez, Laura, Smerdou, Cristian, Howard, Kenneth A., Hangiu, Oana, Gómez-Rosel, Marina, Díez-Alonso, Laura, Silva-Pilipich, Noelia, Compte, Marta, Navarro, Rocío, Tapia-Galisteo, Antonio, Vanrell, Lucía, Sanz, Laura

2025

Overview

Immune checkpoint inhibitors have revolutionized cancer therapy, but many patients fail to respond or develop resistance, often due to reduced T cell activity. Costimulation via 4-1BB has emerged as a promising approach to enhance the effector function of antigen-primed T cells. Bispecific T cell-engaging (TCE) antibodies are an effective way to provide tumor-specific T cell receptor-mediated signaling to tumor-infiltrating lymphocytes. mRNA-based delivery of bispecific antibodies, offer a novel approach to enhance tumor-specific immune responses while minimizing adverse effects. Two bispecific antibodies were generated: the EGFR x CD3 TCE antibody (LiTE) and the PD-L1 x 4-1BB costimulatory antibody (LiTCo), which was further fused to a high FcRn albumin variant (Albu-LiTCo). The mRNA encoding these bispecific antibodies contains an N1-methylpseudouridine modified nucleoside and regulatory sequences to ensure proper expression and stability. A series of assays and cell-based analyses were performed to characterize both antibodies. The efficacy of the mRNA-encoded bispecific antibodies was evaluated in xenograft tumor models expressing EGFR. We investigated the combined effect of two mRNA-encoded Fc-free bispecific antibodies with complementary mechanisms of action: an EGFR-targeting TCE and a half-life extended PD-L1 x 4-1BB costimulatory antibody. The mRNAs encoding both bispecific LiTE and Albu-LiTCo , showed similar binding specificity and function to their protein analogues. Pharmacokinetic studies demonstrated sustained expression of both bispecific antibodies following intravenous administration of the mRNAs formulated using a polymer/lipid-based nanoparticle (LNP) but different pharmacokinetic profiles, shorter for the TCE and longer for the PD-L1 x 4-1BB. When administered as a mRNA-LNP combination (Combo ), the growth of EGFR-positive tumors in immunocompetent mice was significantly inhibited, resulting in tumor regression in 20% of cases with no associated toxicity. Histological analysis confirmed increased T cell infiltration in the tumors treated with LITE and Combo . Repeated administration resulted in sustained production of bispecific antibodies with different exposure cycles and potent antitumor activity with a favorable safety profile. These results highlight the potential of combining two mRNA-encoded bispecific antibodies with different mechanisms of action and programmable half-life for cancer immunotherapy.

• Publisher: Frontiers Media SA,Frontiers Media S.A
• Subject: Animals / Antibiotics / Antibodies / Antibodies, Bispecific - genetics / Antibodies, Bispecific - immunology / Antibodies, Bispecific - pharmacology / Antigens / Antitumor activity / B7-H1 Antigen - immunology / B7-H1 Antigen - metabolism / Bispecific antibodies / Cancer / Cancer immunotherapy / Cancer therapies / CD3 antigen / Cell Line, Tumor / Chromatography / combined RNA / costimulatory antibody / Enzymes / Epidermal growth factor receptors / FDA approval / Female / Humans / Humidity / Immune checkpoint inhibitors / Immunology / Immunotherapy / Immunotherapy - methods / International organizations / Intravenous administration / Leukemia / Lymphocyte Activation - immunology / Lymphocytes / Lymphocytes T / Metastases / Mice / mRNA / mRNA encoded bispecific antibodies / Nanoparticles / Neoplasms - immunology / Neoplasms - therapy / PD-L1 protein / Penicillin / Pharmacokinetics / Regulatory sequences / RNA, Messenger - genetics / RNA, Messenger - immunology / T cell receptors / T-cell engager / T-Lymphocytes - immunology / T-Lymphocytes - metabolism / Toxicity / Trichloroethylene / Tumor Necrosis Factor Receptor Superfamily, Member 9 - immunology / Tumor Necrosis Factor Receptor Superfamily, Member 9 - metabolism / Tumor-infiltrating lymphocytes / Tumors / Xenograft Model Antitumor Assays