Oncolytic viruses encoding bispecific T cell engagers: a blueprint for emerging immunovirotherapies

Abstract

Bispecific T cell engagers (BiTEs) are an innovative class of immunotherapeutics that redirect T cells to tumor surface antigens. While efficacious against certain hematological malignancies, limited bioavailability and severe toxicities have so far hampered broader clinical application, especially against solid tumors. Another emerging cancer immunotherapy are oncolytic viruses (OVs) which selectively infect and replicate in malignant cells, thereby mediating tumor vaccination effects. These oncotropic viruses can serve as vectors for tumor-targeted immunomodulation and synergize with other immunotherapies. In this article, we discuss the use of OVs to overcome challenges in BiTE therapy. We review the current state of the field, covering published preclinical studies as well as ongoing clinical investigations. We systematically introduce OV-BiTE vector design and characteristics as well as evidence for immune-stimulating and anti-tumor effects. Moreover, we address additional combination regimens, including CAR T cells and immune checkpoint inhibitors, and further strategies to modulate the tumor microenvironment using OV-BiTEs. The inherent complexity of these novel therapeutics highlights the importance of translational research including correlative studies in early-phase clinical trials. More broadly, OV-BiTEs can serve as a blueprint for diverse OV-based cancer immunotherapies.

Keywords: Adenovirus; Bispecific T cell engagers; CAR T cells; Cancer immunotherapy; Immune checkpoint blockade; Measles virus; Oncolytic viruses; Tumor microenvironment; Vaccinia virus; Viral vectors.

Fig. 1

BiTE-encoding oncolytic viruses for cancer immunotherapy. Oncolytic viruses (OVs) selectively infect tumor cells, followed by lytic replication (1). In addition to direct tumor debulking, viral oncolysis triggers the release of danger- and pathogen-associated molecular patterns, cytokines, chemokines, and tumor-associated antigens (2). Upon immunogenic tumor cell death (ICD), local inflammation as well as innate and adaptive anti-tumor immune responses can set the stage for effective immunotherapy. Bispecific T cell engagers (BiTEs) redirect T cells to tumor cell surface antigens. OVs can be engineered for tumor-directed BiTE expression to benefit from high BiTE concentrations at the inflamed tumor site, while avoiding systemic toxicities (3). Preclinical studies have shown efficacy of this approach, utilizing BiTE-encoding OVs to engage endogenous or adoptively transferred T cells, including genetically modified CAR T cells (3a). Aside from direct tumor cell targeting, OV-BiTEs can also be used effectively to target immunosuppressive cells of the tumor microenvironment such as cancer-associated fibroblasts (3b) and tumor-associated macrophages (3c). Created with BioRender.com

Fig. 2

Oncolytic virus transgene cassettes encoding bispecific T cell engagers. Generally, BiTE sequences comprise single-chain variable fragments (scFvs) targeting CD3 (blue) and either a tumor-associated antigen (TAA, purple) or cell surface antigens expressed on cancer-associated fibroblasts (yellow) or tumor-associated macrophages (red). Variable heavy (V_H) and light (V_L) chains of scFvs are connected by flexible, non-immunogenic glycine-serine (G/S) linkers. Most constructs harbor peptide tags for detection and/or purification purposes (green). Transgenes are preceded by regulatory domains including promoters (F17R, SA, CMV, EF1, GRP78, GRP94), a Kozak sequence for efficient translation, and leader sequences coding for secretory signaling peptides derived from immunoglobulins (all in grey). a BiTEs specific for human Ephrin type 2 receptor (EphA2) [70] and murine fibroblast activation protein (FAP) [101], respectively, are encoded by oncolytic Vaccinia viruses (VV). b ICOVIR-15-derived adenoviral vectors have been engineered to encode BiTEs targeting human epithelial growth factor receptor (EGFR) (cBiTE) [73, 74, 86] or FAP (FBiTE, not shown) [107]. c Enadenotucirev (EnAd)-derived adenoviral vectors encode BiTEs targeting human epithelial cell adhesion molecule (EpCAM) [77], FAP (not shown) [106], or B. pertussis filamentous hemagglutinin adhesin (FHA, not shown) as a control, under control of either the constitutive cytomegalovirus (CMV) promoter or the adenoviral major late promoter via a splice acceptor (SA) site. d EnAd has also been engineered to express BiTEs specific for human folate receptor β (FRβ) or FHA (control, not shown), arranged in different orders with the CD3-targeting moiety being either C- or N-terminally [108]. e Four different BiTE transgene cassettes for oncolytic measles viruses (MV) have been designed, specific for either human or murine CD3 and either human carcinoembryonic antigen (CEA) or CD20 [82]. f Employing a combinatorial adenoviral vector system (CAd) with a replication-competent oncolytic adenovirus (not shown) and a helper-dependent vector, three immunomodulators have been encoded in cis; a BiTE targeting human CD44v6, a single-peptide interleukin-12 (IL-12p70), and an inhibitor of programmed death-ligand 1 (aPD-L1) [94]. TEA, T cell engager; F17R, late Vaccinia promoter; Ig, immunoglobulin; H-c, heavy chain; L-c, light chain; h, human; m, mouse; SA, splice acceptor for adenoviral major late promoter; CMV, cytomegalovirus promoter; HA tag, peptide from influenza A hemagglutinin; EF1, constitutive EF-1 α promoter; GRP78/94, commercial hamster and human promoters, respectively. Created with BioRender.com