Recombinant Bispecific Antibodies to the Human ErbB2 Receptor and Interferon-Beta

A. A. Panina; V. S. Rybchenko; O. N. Solopova; D. S. Balabashin; S. A. Yakimov; T. K. Aliev; D. A. Dolgikh; P. G. Sveshnikov; M. P. Kirpichnikov

doi:10.32607/actanaturae.10903

Recombinant Bispecific Antibodies to the Human ErbB2 Receptor and Interferon-Beta

Acta Naturae ◽

10.32607/actanaturae.11156 ◽

2020 ◽

Vol 12 (2) ◽

pp. 95-104

Author(s):

A. A. Panina ◽

V. S. Rybchenko ◽

O. N. Solopova ◽

D. S. Balabashin ◽

S. A. Yakimov ◽

...

Keyword(s):

Tumor Cells ◽

Interferon Beta ◽

Bispecific Antibody ◽

Type I Interferons ◽

Bispecific Antibodies ◽

Type I ◽

Systemic Action ◽

Erbb2 Receptor ◽

The Moment ◽

The One

The development of and research into new therapies that can selectively and effectively destroy tumor cells that overexpress the ErbB2 receptor is apressing task. Recently, research into the use of type I interferons in the treatment of cancer has intensified. Cytokine therapy is aimed at activating the cells of the immune system to fight tumors, but it has drawbacks that limit its use because of a number of side effectsthe severity of which varies depending on the dosage and type of used cytokine. At the moment, a number of studies are being conducted regarding the use of IFNin oncology. The studies areaimed at mitigating the systemic action of this cytokine. The immunocytokine complex made of a bispecific antibody against the ErbB2 receptor and recombinant IFNdeveloped in this study underlies themechanism meant to avoid the systemic action of this cytokine. Part of this study focuses on the development of full-length antibodies that bind to the ErbB2 receptor on the one hand, and bind and neutralize IFN, on the other hand, which allows us to consider the antibodies as a means of cytokine delivery to tumor cells.

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Bispecific Antibodies for IFN-β Delivery to ErbB2+ Tumors

Biomolecules ◽

10.3390/biom11121915 ◽

2021 ◽

Vol 11 (12) ◽

pp. 1915

Author(s):

Vladislav S. Rybchenko ◽

Anna A. Panina ◽

Teimur K. Aliev ◽

Olga N. Solopova ◽

Dmitry S. Balabashin ◽

...

Keyword(s):

Tumor Cells ◽

Targeted Delivery ◽

Interferon Beta ◽

Bispecific Antibody ◽

Chimeric Antibody ◽

Tumor Cell Lines ◽

Erbb2 Receptor ◽

Human Igg1 ◽

Covalent Complex ◽

Ht29 Cell Line

The main aim of our work was to create a full-length bispecific antibody (BsAb) as a vehicle for the targeted delivery of interferon-beta (IFN-β) to ErbB2+ tumor cells in the form of non-covalent complex of BsAb and IFN-β. Such a construct is a CrossMab-type BsAb, consisting of an ErbB2-recognizing trastuzumab moiety, a part of chimeric antibody to IFN-β, and human IgG1 Fc domain carrying knob-into-hole amino acid substitutions necessary for the proper assembly of bispecific molecules. The IFN-β- recognizing arm of BsAb not only forms a complex with the cytokine but neutralizes its activity, thus providing a mechanism to avoid the side effects of the systemic action of IFN-β by blocking IFN-β Interaction with cell receptors in the process of cytokine delivery to tumor sites. Enzyme sandwich immunoassay confirmed the ability of BsAb to bind to human IFN-β comparable to that of the parental chimeric mAb. The BsAb binds to the recombinant ErbB2 receptor, as well as to lysates of ErbB2+ tumor cell lines. The inhibition of the antiproliferative effect of IFN-β by BsAb (IC50 = 49,3 µg/mL) was demonstrated on the HT29 cell line. It can be proposed that the BsAb obtained can serve as a component of the immunocytokine complex for the delivery of IFN-β to ErbB2-associated tumor cells.

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Facile Generation of Potent Bispecific Fab via Sortase A and Click Chemistry for Cancer Immunotherapy

Cancers ◽

10.3390/cancers13184540 ◽

2021 ◽

Vol 13 (18) ◽

pp. 4540

Author(s):

Xuefei Bai ◽

Wenhui Liu ◽

Shijie Jin ◽

Wenbin Zhao ◽

Yingchun Xu ◽

...

Keyword(s):

T Cells ◽

Click Chemistry ◽

Cancer Immunotherapy ◽

Tumor Cells ◽

Bispecific Antibody ◽

Bispecific Antibodies ◽

Great Promise ◽

Sortase A ◽

Superior Efficacy ◽

Mouse Xenograft Model

Bispecific antibodies (BsAbs) for T cell engagement have shown great promise in cancer immunotherapy, and their clinical applications have been proven in treating hematological malignance. Bispecific antibody binding fragment (BiFab) represents a promising platform for generating non-Fc bispecific antibodies. However, the generation of BiFab is still challenging, especially by means of chemical conjugation. More conjugation strategies, e.g., enzymatic conjugation and modular BiFab preparation, are needed to improve the robustness and flexibility of BiFab preparation. We successfully used chemo-enzymatic conjugation approach to generate bispecific antibody (i.e., BiFab) with Fabs from full-length antibodies. Paired click handles (e.g., N3 and DBCO) was introduced to the C-terminal LPETG tag of Fabs via sortase A mediated transpeptidation, followed by site-specific conjugation between two click handle-modified Fabs for BiFab generation. Both BiFabCD20/CD3 (EC50 = 0.26 ng/mL) and BiFabHer2/CD3 exhibited superior efficacy in mediating T cells, from either PBMC or ATC, to kill target tumor cell lines while spared antigen-negative tumor cells in vitro. The BiFabCD20/CD3 also efficiently inhibited CD20-positive tumor growth in mouse xenograft model. We have established a facile sortase A-mediated click handle installation to generate homogeneous and functional BiFabs. The exemplary BiFabs against different targets showed superior efficacy in redirecting and activating T cells to specifically kill target tumor cells, demonstrating the robustness of sortase A-mediated “bio-click” chemistry in generating various potent BiFabs. This approach also holds promise for further efficient construction of a Fab derivative library for personalized tumor immunotherapy in the future.

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Effect of modulation of CD3 binding in a PSMAxCD3 T-cell engaging bispecific antibody on maintenance of efficient tumor cell kill and cytokine release.

Journal of Clinical Oncology ◽

10.1200/jco.2020.38.15_suppl.e17583 ◽

2020 ◽

Vol 38 (15_suppl) ◽

pp. e17583-e17583

Author(s):

Ben Buelow ◽

Kevin Dang ◽

Pranjali Dalvi ◽

Yuping Li ◽

Alexander Cheung ◽

...

Keyword(s):

Prostate Cancer ◽

T Cell ◽

Tumor Cell ◽

Tumor Cells ◽

Bispecific Antibody ◽

Cytokine Production ◽

Bispecific Antibodies ◽

Cytokine Release ◽

Human T Cells

e17583 Background: Castration resistant prostate cancer (CRPC) is an incurable disease and represents a significant unmet need. Prostate specific membrane antigen (PSMA) is a protein highly expressed on the surface of prostate cancer cells; expression has been shown to increase with disease progression. Therapies targeting PSMA, such as anti-PSMA radioligand conjugates, have shown promise in clinical trials, validating this target for CRPC. T-cell recruiting bispecific antibodies (T-BsAbs) have demonstrated potent tumor killing activity against multiple tumor types, but immune-mediated toxicities have hampered T-cell redirecting therapies to date. Using Teneobio’s unique antibody discovery platform, we have developed a CD3xPSMA bispecific antibody (TNB-585) that retains the potent cytotoxicity of other T-BsAbs but with significantly reduced cytokine release. Methods: Antibodies targeting CD3 and PSMA were generated via immunization of our proprietary transgenic animals. Candidate antibodies were selected by repertoire deep sequencing of B-cells from draining lymph nodes, followed by high-throughput gene assembly and recombinant expression. Multiple bispecific antibodies targeting CD3 and PSMA were assembled and evaluated for their ability to selectively activate primary human T-cells and mediate killing of PSMA+ tumor cells in vitro, ex vivo, and in vivo. T-cell activation surface markers, cytokine production, and tumor cell cytotoxicity were measured. Results: In co-culture experiments, primary human T-cells were activated only in the presence of both the bispecifics and PSMA+ cells. These bispecifics mediated potent and selective cytotoxicity against PSMA-positive tumor cells, prostate tumor cell lines, or primary human prostate tumor cells isolated from patients. From among these we identified TNB-585, which showed attenuated binding to CD3. TNB-585 mediated comparable tumor cell cytotoxicity to CD3xPSMA T-BsAbs containing a high affinity anti-CD3 domain but with significantly reduced cytokine production. TNB-585 also showed tumor growth inhibition in xenograft models of prostate cancer in vivo. Conclusions: We have developed a novel CD3xPSMA T-BsAb that mediates T-cell killing of PSMA+ tumor cells with minimal production of cytokines. This molecule may improve safety, efficacy, and offer opportunities for combination therapy to treat CRPC. A Phase 1 clinical trial of this compound in CRPC is scheduled to begin in Q1 2021.

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Shared and Unique Features of Human Interferon-Beta and Interferon-Alpha Subtypes

Frontiers in Immunology ◽

10.3389/fimmu.2020.605673 ◽

2021 ◽

Vol 11 ◽

Author(s):

Megen C. Wittling ◽

Shannon R. Cahalan ◽

Eric A. Levenson ◽

Ronald L. Rabin

Keyword(s):

Interferon Beta ◽

Viral Infections ◽

Expression Patterns ◽

Type I Interferons ◽

Human Interferon ◽

Type I ◽

Gene Products ◽

Innate And Adaptive Immunity ◽

Chronic Viral Infections ◽

Alternative Approach

Type I interferons (IFN-I) were first discovered as an antiviral factor by Isaacs and Lindenmann in 1957, but they are now known to also modulate innate and adaptive immunity and suppress proliferation of cancer cells. While much has been revealed about IFN-I, it remains a mystery as to why there are 16 different IFN-I gene products, including IFNβ, IFNω, and 12 subtypes of IFNα. Here, we discuss shared and unique aspects of these IFN-I in the context of their evolution, expression patterns, and signaling through their shared heterodimeric receptor. We propose that rather than investigating responses to individual IFN-I, these contexts can serve as an alternative approach toward investigating roles for IFNα subtypes. Finally, we review uses of IFNα and IFNβ as therapeutic agents to suppress chronic viral infections or to treat multiple sclerosis.

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Development of Autoimmune Hypothyroidism after Interferon-beta 1a Treatment in Patient with Multiple Sclerosis: A Case Report

Bangladesh Journal of Nuclear Medicine ◽

10.3329/bjnm.v19i2.36014 ◽

2018 ◽

Vol 19 (2) ◽

pp. 149

Author(s):

Md Abdul Awal ◽

Md Sunny Anam Chowdhury ◽

Md Hafizur Rahman

Keyword(s):

Multiple Sclerosis ◽

Interferon Beta ◽

Demyelinating Disease ◽

Type I Interferons ◽

Type I ◽

Interferon Β ◽

Autoimmune Hypothyroidism ◽

The Central Nervous System ◽

Multiple Sclerosis Patients ◽

Established Treatment

<p>Interferon beta therapy is a well-established treatment of patients suffering from multiple sclerosis, a demyelinating disease of the central nervous system. Since type I interferons have immunomodulatory properties, these cytokines may trigger several autoimmune disorders. In this case, we report the development of autoimmune hypothyroidism in a multiple sclerosis patients receiving interferon-β 1a.</p><p>Bangladesh J. Nuclear Med. 19(2): 149-151, July 2016</p>

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Multi-Omics Analysis of Glioblastoma Cells’ Sensitivity to Oncolytic Viruses

Cancers ◽

10.3390/cancers13215268 ◽

2021 ◽

Vol 13 (21) ◽

pp. 5268

Author(s):

Anastasiya V. Lipatova ◽

Alesya V. Soboleva ◽

Vladimir A. Gorshkov ◽

Julia A. Bubis ◽

Elizaveta M. Solovyeva ◽

...

Keyword(s):

Tumor Cells ◽

Oncolytic Viruses ◽

Type I Interferons ◽

Interferon Response ◽

Type I ◽

Antiviral Protein ◽

Factors Affecting ◽

Protein Levels ◽

Promising Tool ◽

Omics Analysis

Oncolytic viruses have gained momentum in the last decades as a promising tool for cancer treatment. Despite the progress, only a fraction of patients show a positive response to viral therapy. One of the key variable factors contributing to therapy outcomes is interferon-dependent antiviral mechanisms in tumor cells. Here, we evaluated this factor using patient-derived glioblastoma multiforme (GBM) cultures. Cell response to the type I interferons’ (IFNs) stimulation was characterized at mRNA and protein levels. Omics analysis revealed that GBM cells overexpress interferon-stimulated genes (ISGs) and upregulate their proteins, similar to the normal cells. A conserved molecular pattern unambiguously differentiates between the preserved and defective responses. Comparing ISGs’ portraits with titration-based measurements of cell sensitivity to a panel of viruses, the “strength” of IFN-induced resistance acquired by GBM cells was ranked. The study demonstrates that suppressing a single ISG and encoding an essential antiviral protein, does not necessarily increase sensitivity to viruses. Conversely, silencing IFIT3 and PLSCR1 genes in tumor cells can negatively affect the internalization of vesicular stomatitis and Newcastle disease viruses. We present evidence of a complex relationship between the interferon response genes and other factors affecting the sensitivity of tumor cells to viruses.

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