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Simultaneous targeting of TNF and Ang2 with a novel bispecific antibody enhances efficacy in an in vivo model of arthritis.

ABSTRACT: Despite the clinical success of anti-tumor necrosis factor (TNF) therapies in the treatment of inflammatory conditions such as rheumatoid arthritis, Crohn disease and psoriasis, full control of the diseases only occurs in a subset of patients and there is a need for new therapeutics with improved efficacy against broader patient populations. One possible approach is to combine biological therapeutics, but both the cost of the therapeutics and the potential for additional toxicities needs to be considered. In addition to the various mediators of immune and inflammatory pathways, angiogenesis is reported to contribute substantially to the overall pathogenesis of inflammatory diseases. The combination of an anti-angiogenic agent with anti-TNF into one molecule could be more efficacious without the risk of severe immunosuppression. To evaluate this approach with our Zybody technology, we generated bispecific antibodies that contain an Ang2 targeting peptide genetically fused to the anti-TNF antibody adalimumab (Humira®). The bispecific molecules retain the binding and functional characteristics of the anti-TNF antibody, but with additional activity that neutralizes Ang2. In a TNF transgenic mouse model of arthritis, the bispecific anti-TNF-Ang2 molecules showed a dose-dependent reduction in both clinical symptoms and histological scores that were significantly better than that achieved by adalimumab alone.

SUBMITTER: Kanakaraj P

PROVIDER: S-EPMC3499301 | biostudies-literature | 2012 Sep-Oct

REPOSITORIES: biostudies-literature

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Simultaneous targeting of TNF and Ang2 with a novel bispecific antibody enhances efficacy in an in vivo model of arthritis.

Kanakaraj Palanisamy P Puffer Bridget A BA Yao Xiao-Tao XT Kankanala Spandana S Boyd Ernest E Shah Rutul R RR Wang Geping G Patel Dimki D Krishnamurthy Rajesh R Kaithamana Shashi S Smith Rodger G RG LaFleur David W DW Barbas Carlos F CF Hilbert David M DM Kiener Peter A PA Roschke Viktor V VV

mAbs 20120806 5

Despite the clinical success of anti-tumor necrosis factor (TNF) therapies in the treatment of inflammatory conditions such as rheumatoid arthritis, Crohn disease and psoriasis, full control of the diseases only occurs in a subset of patients and there is a need for new therapeutics with improved efficacy against broader patient populations. One possible approach is to combine biological therapeutics, but both the cost of the therapeutics and the potential for additional toxicities needs to be c ...[more]

PMID: 22864384

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Project description:BackgroundRecently, therapeutic antibodies against programmed cell death 1 (PD-1) and its ligand (PD-L1) have exerted potent anticancer effect in a variety of tumors. However, blocking the PD-1/PD-L1 axis alone is not sufficient to restore normal immune response. Other negative regulators of antitumor immunity, like TGF-β and VEGFA, are also involved in immune escape of tumor cells and induce immunotherapy resistance.MethodsWe developed a novel anti-TGF-β/VEGF bispecific antibody Y332D based on the Nano-YBODY™ technology platform. The CCK-8, flow cytometry, SBE4 luciferase reporter assay, western blotting and transwell assays were used to measure the biological activities of the anti-TGF-β moiety. The NFAT luciferase reporter assay, luminescent cell viability assay and tube formation assay were used to measure the biological activities of the anti-VEGF moiety. The in vivo anticancer efficacy of Y332D alone or in combination with PD-1 blockade was evaluated in H22, EMT-6, 4T1, and AKT/Ras-driven murine hepatocellular carcinoma tumor models. Immunofluorescent staining, flow cytometry, RNA-seq and quantitative RT-PCR were adopted to analyze the alterations in the tumor microenvironment.ResultsY332D could maintain specific binding affinities for TGF-β and VEGFA. Y332D almost entirely counteracted the in vitro biological functions of TGF-β and VEGFA, including immunosuppression, activated TGF-β signaling, epithelial-mesenchymal transition (EMT), activated VEGF/VEGFR signaling, HUVEC proliferation and tube formation. The in vivo experiment data demonstrated that Y332D was more effective in inhibiting tumor growth and metastasis than anti-TGF-β and anti-VEGF monotherapies. In combination therapies, Y332D plus PD-1 blockade exhibited the most potent and durable anticancer effect. Mechanistically, Y332D plus PD-1 blockade upregulated the density and function of tumor-infiltrating lymphocytes and exerted reinvigorated antitumor immunity.ConclusionY332D could simultaneously block TGF-β and VEGF signalings. In comparison with the monotherapies, Y332D combined with PD-1 blockade exerts superior antitumor effect through improving immune microenvironment.

Project description:Natural killer group 2D (NKG2D) is an activating receptor of natural killer (NK) cells and other lymphocytes that mediates lysis of malignant cells through recognition of stress-induced ligands such as MICA and MICB. Such ligands are broadly expressed by cancer cells of various origins and serve as targets for adoptive immunotherapy with effector cells endogenously expressing NKG2D or carrying an NKG2D-based chimeric antigen receptor (CAR). However, shedding or downregulation of NKG2D ligands (NKG2DL) can prevent NKG2D activation, resulting in escape of cancer cells from NKG2D-dependent immune surveillance. To enable tumor-specific targeting of NKG2D-expressing effector cells independent of membrane-anchored NKG2DLs, we generated a homodimeric recombinant antibody which harbors an N-terminal single-chain fragment variable (scFv) antibody domain for binding to NKG2D, linked via a human IgG4 Fc region to a second C-terminal scFv antibody domain for recognition of the tumor-associated antigen ErbB2 (HER2). The ability of this molecule, termed NKAB-ErbB2, to redirect NKG2D-expressing effector cells to ErbB2-positive tumor cells of different origins was investigated using peripheral blood mononuclear cells, ex vivo expanded NK cells, and NK and T cells engineered with an NKG2D-based chimeric receptor. On its own, bispecific NKAB-ErbB2 increased lysis of ErbB2-positive breast carcinoma cells by peripheral blood-derived NK cells endogenously expressing NKG2D more effectively than an ErbB2-specific IgG1 mini-antibody able to induce antibody-dependent cell-mediated cytotoxicity via activation of CD16. Furthermore, NKAB-ErbB2 synergized with NK-92 cells or primary T cells engineered to express an NKG2D-CD3ζ chimeric antigen receptor (NKAR), leading to targeted cell killing and greatly enhanced antitumor activity, which remained unaffected by soluble MICA known as an inhibitor of NKG2D-mediated natural cytotoxicity. In an immunocompetent mouse glioblastoma model mimicking low or absent NKG2DL expression, the combination of NKAR-NK-92 cells and NKAB-ErbB2 effectively suppressed outgrowth of ErbB2-positive tumors, resulting in treatment-induced endogenous antitumor immunity and cures in the majority of animals. Our results demonstrate that combining an NKAB antibody with effector cells expressing an activating NKAR receptor represents a powerful and versatile approach to simultaneously enhance tumor antigen-specific as well as NKG2D-CAR and natural NKG2D-mediated cytotoxicity, which may be particularly useful to target tumors with heterogeneous target antigen expression.

Dataset Information

Simultaneous targeting of TNF and Ang2 with a novel bispecific antibody enhances efficacy in an in vivo model of arthritis.

Publications

Simultaneous targeting of TNF and Ang2 with a novel bispecific antibody enhances efficacy in an in vivo model of arthritis.

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