An Fc-free EGFR-specific 4-1BB-agonistic Trimerbody Displays Broad Antitumor Activity in Humanized Murine Cancer Models without Toxicity

Abstract

Purpose: The induction of 4-1BB signaling by agonistic antibodies can drive the activation and proliferation of effector T cells and thereby enhance a T-cell-mediated antitumor response. Systemic administration of anti-4-1BB-agonistic IgGs, although effective preclinically, has not advanced in clinical development due to their severe hepatotoxicity.Experimental Design: Here, we generated a humanized EGFRspecific 4-1BB-agonistic trimerbody, which replaces the IgG Fc region with a human collagen homotrimerization domain. It was characterized by structural analysis and in vitro functional studies. We also assessed pharmacokinetics, antitumor efficacy, and toxicity in vivo.Results: In the presence of a T-cell receptor signal, the trimerbody provided potent T-cell costimulation that was strictly depen-dent on 4-1BB hyperclustering at the point of contact with a tumor antigen-displaying cell surface. It exhibits significant antitumor activity in vivo, without hepatotoxicity, in a wide range of human tumors including colorectal and breast cancer cell-derived xenografts, and non-small cell lung cancer patient-derived xenografts associated with increased tumor-infiltrating CD8 þ T cells. The combination of the trimerbody with a PD-L1 blocker led to increased IFNg secretion in vitro and resulted in tumor regression in humanized mice bearing aggressive triple-negative breast cancer.Conclusions: These results demonstrate the nontoxic broad antitumor activity of humanized Fc-free tumor-specific 4-1BBagonistic trimerbodies and their synergy with checkpoint blockers, which may provide a way to elicit responses in most patients with cancer while avoiding Fc-mediated adverse reactions.

Half-Life Extended Nanobody-Based CD38-Specific Bispecific Killercell Engagers Induce Killing of Multiple Myeloma Cells

“…Our HLE-nano-BiKEs are composed only of lama-derived variable immunoglobulin domains (VHH or nanobody). Similar to therapeutics that incorporate murine VH and VL domains such as blinatumumab, a CD19-CD3 bispecific T-cell engager composed of two murine scFv ( 58 , 59 ) or rituximab ( 60 ), a chimeric antibody composed of murine VH and VL and human constant IgG1 domains, therapeutics composed of VHH domains such as caplacizumab ( 61 ) (a dimer of two llama VHH domains) show little if any immunogenicity in patients ( 62 ). Anti-drug antibodies in patients can abrogate the efficacy of antibody treatment; therefore antibody-constructs with a low immunogenicity are favorable ( 63 – 65 ).…”

Large Animal Models of Breast Cancer

“…The humanized mouse model (in which an immunocompromised mouse is engrafted with components of the human immune system) has been utilized in preclinical studies on immunotherapies and BC, particularly TNBC and HER2+ cancers (97). Humanized BC models have shown clinically relevant reduction of tumor growth in response to therapy (98,99). However, major concerns of these models include (1) a lack GM-CSF, which is important for the differentiation and maturation of the myeloid lineage; and (2) xenograft-versus-host disease, in which mature human T cells attack their murine host secondary to HLA mismatching between the hNSG and PDX components (99,100).…”

Engineered mRNA and the Rise of Next-Generation Antibodies

“…AFM13, a tetravalent bispecific anti-CD30 x anti-CD16A TandAb (tandem diabody), with a molecular weight of around 100 kDa, is in phase 2 trials for the treatment of peripheral T cell lymphoma [ 17 ]. Bispecific anti-CD137 x anti-EGFR (epidermal growth factor receptor) trimerbodies, with a molecular weight of 160 kDa, have been able to eradicate established tumors in several animal models [ 18 , 19 ]. Therefore, it seems that half-life extension and multimerization strategies would provide Fc-free antibodies with potential for therapeutic opportunities.…”