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Peng JJ, Ding JY, Xu Y, Shih HP, Lin YN, Wu TY, Lo YF, Lo CC, Yeh CF, Kuo CY, Tu KH, Wang SY, Lei WT, Wu TS, Lin HS, Lee CH, Huang WC, Chen YC, Liu YM, Shi ZY, Chang YT, Syue LS, Chen PL, Teh SH, Chou CH, Ho MW, Chi CY, Ho PC, Ku CL. Chimeric autoantibody receptor T cells clonally eliminate B cells producing autoantibodies against IFN-γ. Sci Immunol 2025; 10:eadm8186. [PMID: 40344086 DOI: 10.1126/sciimmunol.adm8186] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2023] [Revised: 12/04/2024] [Accepted: 03/27/2025] [Indexed: 05/11/2025]
Abstract
Neutralizing anti-interferon-γ (IFN-γ) autoantibodies (nAIGAs) impair IFN-γ-mediated immunity, predisposing patients with nAIGAs to infection by nontuberculous mycobacteria, Talaromyces marneffei, and other intracellular pathogens. Current clinical management relies on continuous antimicrobial therapy, with no treatment offering sustained benefits. Here, we developed human chimeric autoantibody receptor (CAAR) T cells targeting autoreactive B cells expressing nAIGA B cell receptors (BCRs) using an IFN-γ receptor-irresponsive IFN-γ variant as bait. By exploiting a mouse model of nAIGA BCR-expressing B cell leukemia, we found that IFN-γ CAAR T cells lack off-target toxicity, including IFN-γ receptor cross-reactive toxicity and Fc-redirected toxicity. IFN-γ CAAR T cells substantially reduced circulating AIGAs secreted from target cells in vivo. Further, IFN-γ CAAR T cells effectively eliminated autoreactive B cells in ex vivo cultures of peripheral blood mononuclear cells from patients with nAIGAs. Together, these results demonstrate that IFN-γ CAAR T cells may be a promising strategy to ameliorate nAIGA-associated infections by eliminating autoreactive B cells.
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Affiliation(s)
- Jhan-Jie Peng
- Center for Molecular and Clinical Immunology, Chang Gung University, Taoyuan, Taiwan
- Laboratory of Human Immunology and Infectious Diseases, Graduate Institute of Clinical Medical Sciences, Chang Gung University, Taoyuan, Taiwan
- Ludwig Institute of Cancer Research, University of Lausanne, Lausanne, Switzerland
- Department of Fundamental Oncology, University of Lausanne, Lausanne, Switzerland
| | - Jing-Ya Ding
- Center for Molecular and Clinical Immunology, Chang Gung University, Taoyuan, Taiwan
- Laboratory of Human Immunology and Infectious Diseases, Graduate Institute of Clinical Medical Sciences, Chang Gung University, Taoyuan, Taiwan
| | - Yingxi Xu
- Ludwig Institute of Cancer Research, University of Lausanne, Lausanne, Switzerland
- Department of Fundamental Oncology, University of Lausanne, Lausanne, Switzerland
| | - Han-Po Shih
- Center for Molecular and Clinical Immunology, Chang Gung University, Taoyuan, Taiwan
- Laboratory of Human Immunology and Infectious Diseases, Graduate Institute of Clinical Medical Sciences, Chang Gung University, Taoyuan, Taiwan
| | - You-Ning Lin
- Center for Molecular and Clinical Immunology, Chang Gung University, Taoyuan, Taiwan
- Laboratory of Human Immunology and Infectious Diseases, Graduate Institute of Clinical Medical Sciences, Chang Gung University, Taoyuan, Taiwan
| | - Tsai-Yi Wu
- Center for Molecular and Clinical Immunology, Chang Gung University, Taoyuan, Taiwan
- Laboratory of Human Immunology and Infectious Diseases, Graduate Institute of Clinical Medical Sciences, Chang Gung University, Taoyuan, Taiwan
| | - Yu-Fang Lo
- Center for Molecular and Clinical Immunology, Chang Gung University, Taoyuan, Taiwan
- Laboratory of Human Immunology and Infectious Diseases, Graduate Institute of Clinical Medical Sciences, Chang Gung University, Taoyuan, Taiwan
| | - Chia-Chi Lo
- Center for Molecular and Clinical Immunology, Chang Gung University, Taoyuan, Taiwan
- Laboratory of Human Immunology and Infectious Diseases, Graduate Institute of Clinical Medical Sciences, Chang Gung University, Taoyuan, Taiwan
| | - Chu-Fu Yeh
- Laboratory of Human Immunology and Infectious Diseases, Graduate Institute of Clinical Medical Sciences, Chang Gung University, Taoyuan, Taiwan
- Division of Infectious Diseases, Department of Internal Medicine, Linkou Chang Gung Memorial Hospital, Taoyuan, Taiwan
| | - Chen-Yen Kuo
- Laboratory of Human Immunology and Infectious Diseases, Graduate Institute of Clinical Medical Sciences, Chang Gung University, Taoyuan, Taiwan
- Division of Infectious Diseases, Department of Pediatrics, Chang Gung Memorial Hospital, Taoyuan, Taiwan
- School of Traditional Chinese Medicine, College of Medicine, Chang Gung University, Taoyuan, Taiwan
| | - Kun-Hua Tu
- Center for Molecular and Clinical Immunology, Chang Gung University, Taoyuan, Taiwan
- Department of Nephrology, Chang Gung Memorial Hospital, Taoyuan, Taiwan
| | - Shang-Yu Wang
- Laboratory of Human Immunology and Infectious Diseases, Graduate Institute of Clinical Medical Sciences, Chang Gung University, Taoyuan, Taiwan
- Division of General Surgery, Department of Surgery, Chang Gung Memorial Hospital, Taoyuan, Taiwan
| | - Wei-Te Lei
- Laboratory of Human Immunology and Infectious Diseases, Graduate Institute of Clinical Medical Sciences, Chang Gung University, Taoyuan, Taiwan
- Department of Pediatrics, Division of Pediatric Allergy, Immunology, and Rheumatology, Hsinchu Municipal MacKay Children's Hospital, Hsinchu, Taiwan
| | - Ting-Shu Wu
- Division of Infectious Diseases, Department of Internal Medicine, Linkou Chang Gung Memorial Hospital, Taoyuan, Taiwan
- School of Medicine, College of Medicine, Chang Gung University, Taoyuan, Taiwan
| | - Huang-Shen Lin
- Division of Infectious Diseases, Department of Internal Medicine, Chang Gung Memorial Hospital, Chiayi, Taiwan
| | - Chen-Hsiang Lee
- Division of Infectious Diseases, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, Taiwan
| | - Wen-Chi Huang
- Division of Infectious Diseases, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, Taiwan
| | - Yi-Chun Chen
- Division of Infectious Diseases, Department of Internal Medicine, Kaohsiung Chang Gung Memorial Hospital, Kaohsiung, Taiwan
| | - Yuag-Meng Liu
- Division of Infectious Diseases, Department of Internal Medicine, Changhua Christian Hospital, Changhua, Taiwan
| | - Zhi-Yuan Shi
- Division of Infectious Diseases, Department of Internal Medicine, Taichung Veterans General Hospital, Taichung, Taiwan
| | - Ya-Ting Chang
- Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan
| | - Ling-Shan Syue
- Department of Internal Medicine, National Cheng Kung University Hospital, College of Medicine, National Cheng Kung University, Tainan, Taiwan
| | - Po-Lin Chen
- Department of Internal Medicine, National Cheng Kung University Hospital, College of Medicine, National Cheng Kung University, Tainan, Taiwan
| | - Soon-Hian Teh
- Division of Infectious Diseases, Department of Internal Medicine, Hualien Tzu Chi Hospital, Hualien, Taiwan
| | - Chia-Huei Chou
- Division of Infectious Diseases, Department of Internal Medicine, China Medical University Hospital, Taichung, Taiwan
- School of Medicine, College of Medicine, China Medical University, Taichung, Taiwan
| | - Mao-Wang Ho
- Division of Infectious Diseases, Department of Internal Medicine, China Medical University Hospital, Taichung, Taiwan
- School of Medicine, College of Medicine, China Medical University, Taichung, Taiwan
| | - Chih-Yu Chi
- Division of Infectious Diseases, Department of Internal Medicine, China Medical University Hospital, Taichung, Taiwan
- School of Medicine, College of Medicine, China Medical University, Taichung, Taiwan
| | - Ping-Chih Ho
- Ludwig Institute of Cancer Research, University of Lausanne, Lausanne, Switzerland
- Department of Fundamental Oncology, University of Lausanne, Lausanne, Switzerland
| | - Cheng-Lung Ku
- Center for Molecular and Clinical Immunology, Chang Gung University, Taoyuan, Taiwan
- Laboratory of Human Immunology and Infectious Diseases, Graduate Institute of Clinical Medical Sciences, Chang Gung University, Taoyuan, Taiwan
- Division of Infectious Diseases, Department of Pediatrics, Chang Gung Memorial Hospital, Taoyuan, Taiwan
- Department of Nephrology, Chang Gung Memorial Hospital, Taoyuan, Taiwan
- Elixiron Immunotherapeutics, Taipei, Taiwan
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Tomescu C, Ochoa-Ortiz A, Lu LD, Kong H, Riley JL, Montaner LJ. Gene-modified NK cells expressing CD64 and preloaded with HIV-specific BNAbs target autologous HIV-1-infected CD4+ T cells by ADCC. JOURNAL OF IMMUNOLOGY (BALTIMORE, MD. : 1950) 2025; 214:253-264. [PMID: 40073240 PMCID: PMC11878998 DOI: 10.1093/jimmun/vkae028] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 06/26/2024] [Accepted: 11/19/2024] [Indexed: 03/14/2025]
Abstract
Natural killer (NK) cells can efficiently mediate antibody-dependent cellular cytotoxicity (ADCC) of antibody coated target cells via the low-affinity Fc-receptor, CD16, but cannot retain antibodies over time. To increase antibody retention and facilitate targeted ADCC, we genetically modified human NK cells with the high-affinity Fc receptor, CD64, so that we could preload them with HIV-specific broadly neutralizing antibodies (BNAbs) and enhance their capacity to target HIV-infected cells via ADCC. Purified NK cells from the peripheral blood of control donors or persons living with HIV were activated with interleukin (IL)-2/IL-15/IL-21 cytokines and transduced with a lentivirus encoding CD64. High levels of CD64 surface expression were maintained for multiple weeks on NK cells and CD64-transduced NK cells were phenotypically similar to control NK cells with strong expression of CD56, CD16, NKG2A, NKp46, CD69, HLA-DR, CD38, and CD57. CD64-transduced NK cells exhibited significantly greater capacity to bind HIV-specific BNAbs in short-term antibody binding assay as well as retain the BNAbs over time (1-wk antibody retention assay) compared with control NK cells only expressing CD16. BNAb-preloaded CD64-transduced NK cells showed a significantly enhanced capacity to mediate ADCC against autologous HIV-1-infected CD4+ primary T cells in both a short-term 4 h degranulation assay as well as a 24 h HIV p24 HIV elimination assay when compared with control NK cells. A chimeric CD64 enhanced NK cell strategy (NuKEs [NK Enhancement Strategy]) retaining bound HIV-specific BNAbs represents a novel autologous primary NK cell immunotherapy strategy against HIV through targeted ADCC.
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Affiliation(s)
- Costin Tomescu
- HIV Immunopathogenesis Laboratory, BEAT-HIV Delaney Collaboratory, Wistar Institute, Philadelphia, PA, United States
| | - Adiana Ochoa-Ortiz
- HIV Immunopathogenesis Laboratory, BEAT-HIV Delaney Collaboratory, Wistar Institute, Philadelphia, PA, United States
| | - Lily D Lu
- Molecular Screening and Protein Expression Facility, Wistar Institute, Philadelphia, PA, United States
| | - Hong Kong
- Center for Cellular Immunotherapies, BEAT-HIV Delaney Collaboratory, Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, United States
| | - James L Riley
- Center for Cellular Immunotherapies, BEAT-HIV Delaney Collaboratory, Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, United States
| | - Luis J Montaner
- HIV Immunopathogenesis Laboratory, BEAT-HIV Delaney Collaboratory, Wistar Institute, Philadelphia, PA, United States
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Hrvat A, Benders S, Kimmig R, Brandau S, Mallmann-Gottschalk N. Immunoglobulins and serum proteins impair anti-tumor NK cell effector functions in malignant ascites. Front Immunol 2024; 15:1360615. [PMID: 38646521 PMCID: PMC11026578 DOI: 10.3389/fimmu.2024.1360615] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/23/2023] [Accepted: 03/20/2024] [Indexed: 04/23/2024] Open
Abstract
Introduction Malignant ascites indicates ovarian cancer progression and predicts poor clinical outcome. Various ascites components induce an immunosuppressive crosstalk between tumor and immune cells, which is poorly understood. In our previous study, imbalanced electrolytes, particularly high sodium content in malignant ascites, have been identified as a main immunosuppressive mechanism that impaired NK and T-cell activity. Methods In the present study, we explored the role of high concentrations of ascites proteins and immunoglobulins on antitumoral NK effector functions. To this end, a coculture system consisting of healthy donor NK cells and ovarian cancer cells was used. The anti-EGFR antibody Cetuximab was added to induce antibody-dependent cellular cytotoxicity (ADCC). NK activity was assessed in the presence of different patient ascites samples and immunoglobulins that were isolated from ascites. Results Overall high protein concentration in ascites impaired NK cell degranulation, conjugation to tumor cells, and intracellular calcium signaling. Immunoglobulins isolated from ascites samples competitively interfered with NK ADCC and inhibited the conjugation to target cells. Furthermore, downregulation of regulatory surface markers CD16 and DNAM-1 on NK cells was prevented by ascites-derived immunoglobulins during NK cell activation. Conclusion Our data show that high protein concentrations in biological fluids are able to suppress antitumoral activity of NK cells independent from the mechanism mediated by imbalanced electrolytes. The competitive interference between immunoglobulins of ascites and specific therapeutic antibodies could diminish the efficacy of antibody-based therapies and should be considered in antibody-based immunotherapies.
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Affiliation(s)
- Antonio Hrvat
- Experimental and Translational Research, Department of Otorhinolaryngology, University Hospital Essen, Essen, Germany
| | - Sonja Benders
- Experimental and Translational Research, Department of Otorhinolaryngology, University Hospital Essen, Essen, Germany
- Department for Trauma Surgery and Orthopedics, St. Joseph Hospital Kupferdreh, Essen, Germany
| | - Rainer Kimmig
- Department of Gynecology and Obstetrics, University Hospital Essen, Essen, Germany
| | - Sven Brandau
- Experimental and Translational Research, Department of Otorhinolaryngology, University Hospital Essen, Essen, Germany
- German Cancer Consortium, Partner Site Essen-Düsseldorf, Essen, Germany
| | - Nina Mallmann-Gottschalk
- Experimental and Translational Research, Department of Otorhinolaryngology, University Hospital Essen, Essen, Germany
- Department of Gynecology and Obstetrics, University Hospital Cologne, Cologne, Germany
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Boje AS, Pekar L, Koep K, Lipinski B, Rabinovich B, Evers A, Gehlert CL, Krohn S, Xiao Y, Krah S, Zaynagetdinov R, Toleikis L, Poetzsch S, Peipp M, Zielonka S, Klausz K. Impact of antibody architecture and paratope valency on effector functions of bispecific NKp30 x EGFR natural killer cell engagers. MAbs 2024; 16:2315640. [PMID: 38372053 PMCID: PMC10877975 DOI: 10.1080/19420862.2024.2315640] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/09/2023] [Accepted: 02/02/2024] [Indexed: 02/20/2024] Open
Abstract
Natural killer (NK) cells emerged as a promising effector population that can be harnessed for anti-tumor therapy. In this work, we constructed NK cell engagers (NKCEs) based on NKp30-targeting single domain antibodies (sdAbs) that redirect the cytotoxic potential of NK cells toward epidermal growth factor receptor (EGFR)-expressing tumor cells. We investigated the impact of crucial parameters such as sdAb location, binding valencies, the targeted epitope on NKp30, and the overall antibody architecture on the redirection capacity. Our study exploited two NKp30-specific sdAbs, one of which binds a similar epitope on NKp30 as its natural ligand B7-H6, while the other sdAb addresses a non-competing epitope. For EGFR-positive tumor targeting, humanized antigen-binding domains of therapeutic antibody cetuximab were used. We demonstrate that NKCEs bivalently targeting EGFR and bivalently engaging NKp30 are superior to monovalent NKCEs in promoting NK cell-mediated tumor cell lysis and that the architecture of the NKCE can substantially influence killing capacities depending on the NKp30-targeting sdAb utilized. While having a pronounced impact on NK cell killing efficacy, the capabilities of triggering antibody-dependent cellular phagocytosis or complement-dependent cytotoxicity were not significantly affected comparing the bivalent IgG-like NKCEs with cetuximab. However, the fusion of sdAbs can have a slight impact on the NK cell release of immunomodulatory cytokines, as well as on the pharmacokinetic profile of the NKCE due to unfavorable spatial orientation within the molecule architecture. Ultimately, our findings reveal novel insights for the engineering of potent NKCEs triggering the NKp30 axis.
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Affiliation(s)
- Ammelie Svea Boje
- Division of Antibody-Based Immunotherapy, Department of Internal Medicine II, University Medical Center Schleswig-Holstein and University of Kiel, Kiel, Germany
| | - Lukas Pekar
- Antibody Discovery & Protein Engineering, Merck Healthcare KGaA, Darmstadt, Germany
| | - Katharina Koep
- Drug Metabolism and Pharmacokinetics, Merck Healthcare KGaA, Darmstadt, Germany
| | - Britta Lipinski
- Antibody Discovery & Protein Engineering, Merck Healthcare KGaA, Darmstadt, Germany
| | - Brian Rabinovich
- Department of Oncology and Immuno-Oncology, EMD Serono Research & Development Institute Inc, 45A Middlesex Turnpike, Billerica, MA, USA
| | - Andreas Evers
- Antibody Discovery & Protein Engineering, Merck Healthcare KGaA, Darmstadt, Germany
| | - Carina Lynn Gehlert
- Division of Antibody-Based Immunotherapy, Department of Internal Medicine II, University Medical Center Schleswig-Holstein and University of Kiel, Kiel, Germany
| | - Steffen Krohn
- Division of Antibody-Based Immunotherapy, Department of Internal Medicine II, University Medical Center Schleswig-Holstein and University of Kiel, Kiel, Germany
| | - Yanping Xiao
- Department of Oncology and Immuno-Oncology, EMD Serono Research & Development Institute Inc, 45A Middlesex Turnpike, Billerica, MA, USA
| | - Simon Krah
- Antibody Discovery & Protein Engineering, Merck Healthcare KGaA, Darmstadt, Germany
| | - Rinat Zaynagetdinov
- Department of Oncology and Immuno-Oncology, EMD Serono Research & Development Institute Inc, 45A Middlesex Turnpike, Billerica, MA, USA
| | - Lars Toleikis
- Early Protein Supply & Characterization, Merck Healthcare KGaA, Darmstadt, Germany
| | - Sven Poetzsch
- Strategic Innovation, Merck Healthcare KGaA, Darmstadt, Germany
| | - Matthias Peipp
- Division of Antibody-Based Immunotherapy, Department of Internal Medicine II, University Medical Center Schleswig-Holstein and University of Kiel, Kiel, Germany
| | - Stefan Zielonka
- Antibody Discovery & Protein Engineering, Merck Healthcare KGaA, Darmstadt, Germany
- Institute for Organic Chemistry and Biochemistry, Technical University of Darmstadt, Darmstadt, Germany
| | - Katja Klausz
- Division of Antibody-Based Immunotherapy, Department of Internal Medicine II, University Medical Center Schleswig-Holstein and University of Kiel, Kiel, Germany
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Enell Smith K, Fritzell S, Nilsson A, Barchan K, Rosén A, Schultz L, Varas L, Säll A, Rose N, Håkansson M, von Schantz L, Ellmark P. ATOR-1017 (evunzekibart), an Fc-gamma receptor conditional 4-1BB agonist designed for optimal safety and efficacy, activates exhausted T cells in combination with anti-PD-1. Cancer Immunol Immunother 2023; 72:4145-4159. [PMID: 37796298 PMCID: PMC10700433 DOI: 10.1007/s00262-023-03548-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/06/2023] [Accepted: 09/11/2023] [Indexed: 10/06/2023]
Abstract
BACKGROUND 4-1BB (CD137) is a co-stimulatory receptor highly expressed on tumor reactive effector T cells and NK cells, which upon stimulation prolongs persistence of tumor reactive effector T and NK cells within the tumor and induces long-lived memory T cells. 4-1BB agonistic antibodies have been shown to induce strong anti-tumor effects that synergize with immune checkpoint inhibitors. The first generation of 4-1BB agonists was, however, hampered by dose-limiting toxicities resulting in suboptimal dose levels or poor agonistic activity. METHODS ATOR-1017 (evunzekibart), a second-generation Fc-gamma receptor conditional 4-1BB agonist in IgG4 format, was designed to overcome the limitations of the first generation of 4-1BB agonists, providing strong agonistic effect while minimizing systemic immune activation and risk of hepatoxicity. The epitope of ATOR-1017 was determined by X-ray crystallography, and the functional activity was assessed in vitro and in vivo as monotherapy or in combination with anti-PD1. RESULTS ATOR-1017 binds to a unique epitope on 4-1BB enabling ATOR-1017 to activate T cells, including cells with an exhausted phenotype, and NK cells, in a cross-linking dependent, FcγR-conditional, manner. This translated into a tumor-directed and potent anti-tumor therapeutic effect in vivo, which was further enhanced with anti-PD-1 treatment. CONCLUSIONS These preclinical data demonstrate a strong safety profile of ATOR-1017, together with its potent therapeutic effect as monotherapy and in combination with anti-PD1, supporting further clinical development of ATOR-1017.
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Affiliation(s)
| | | | | | | | | | | | | | - Anna Säll
- Alligator Bioscience AB, Lund, Sweden
| | | | | | | | - Peter Ellmark
- Alligator Bioscience AB, Lund, Sweden.
- Department of Immunotechnology, Lund University, Lund, Sweden.
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Bauer-Smith H, Sudol ASL, Beers SA, Crispin M. Serum immunoglobulin and the threshold of Fc receptor-mediated immune activation. Biochim Biophys Acta Gen Subj 2023; 1867:130448. [PMID: 37652365 PMCID: PMC11032748 DOI: 10.1016/j.bbagen.2023.130448] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/05/2023] [Revised: 08/23/2023] [Accepted: 08/23/2023] [Indexed: 09/02/2023]
Abstract
Antibodies can mediate immune recruitment or clearance of immune complexes through the interaction of their Fc domain with cellular Fc receptors. Clustering of antibodies is a key step in generating sufficient avidity for efficacious receptor recognition. However, Fc receptors may be saturated with prevailing, endogenous serum immunoglobulin and this raises the threshold by which cellular receptors can be productively engaged. Here, we review the factors controlling serum IgG levels in both healthy and disease states, and discuss how the presence of endogenous IgG is encoded into the functional activation thresholds for low- and high-affinity Fc receptors. We discuss the circumstances where antibody engineering can help overcome these physiological limitations of therapeutic antibodies. Finally, we discuss how the pharmacological control of Fc receptor saturation by endogenous IgG is emerging as a feasible mechanism for the enhancement of antibody therapeutics.
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Affiliation(s)
- Hannah Bauer-Smith
- School of Biological Sciences, University of Southampton, Southampton SO17 1BJ, UK; Centre for Cancer Immunology, School of Cancer Sciences, University of Southampton Faculty of Medicine, Southampton SO16 6YD, UK
| | - Abigail S L Sudol
- School of Biological Sciences, University of Southampton, Southampton SO17 1BJ, UK
| | - Stephen A Beers
- Centre for Cancer Immunology, School of Cancer Sciences, University of Southampton Faculty of Medicine, Southampton SO16 6YD, UK.
| | - Max Crispin
- School of Biological Sciences, University of Southampton, Southampton SO17 1BJ, UK.
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Hrvat A, Schmidt M, Wagner B, Zwanziger D, Kimmig R, Volbracht L, Brandau S, Mallmann-Gottschalk N. Electrolyte imbalance causes suppression of NK and T cell effector function in malignant ascites. J Exp Clin Cancer Res 2023; 42:235. [PMID: 37684704 PMCID: PMC10485936 DOI: 10.1186/s13046-023-02798-8] [Citation(s) in RCA: 10] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/18/2023] [Accepted: 08/13/2023] [Indexed: 09/10/2023] Open
Abstract
BACKGROUND Malignant ascites commonly occurs in advanced or recurrent stages of epithelial ovarian cancer during peritoneal carcinomatosis and is correlated with poor prognosis. Due to its complex composition of cellular and acellular components malignant ascites creates a unique tumor microenvironment, which mediates immunosuppression and promotes progression of disease. However, the immunosuppressive mechanisms remain poorly understood. METHODS In the present study, we explored the antitumor activity of healthy donor NK and T cells directed against ovarian cancer cells in presence of malignant ascites derived from patients with advanced or recurrent peritoneal carcinomatosis. A wide range of methods was used to study the effect of ascites on NK and T cells (FACS, ELISA, EliSpot, qPCR, Live-cell and confocal microscopy, Western blot and electrolyte flux assays). The ascites components were assessed using quantitative analysis (nephelometry, potentiometry and clinical chemistry) and separation methods (dialysis, ultracentrifugal filtration and lipid depletion). RESULTS Ascites rapidly inhibited NK cell degranulation, tumor lysis, cytokine secretion and calcium signaling. Similarly, target independent NK and T cell activation was impaired in ascites environment. We identified imbalanced electrolytes in ascites as crucial factors causing extensive immunosuppression of NK and T cells. Specifically, high sodium, low chloride and low potassium content significantly suppressed NK-mediated cytotoxicity. Electrolyte imbalance led to changes in transcription and protein expression of electrolyte channels and impaired NK and T cell activation. Selected inhibitors of sodium electrolyte channels restored intracellular calcium flux, conjugation, degranulation and transcript expression of signaling molecules. The levels of ascites-mediated immunosuppression and sodium/chloride/potassium imbalance correlated with poor patient outcome and selected molecular alterations were confirmed in immune cells from ovarian cancer patients. CONCLUSION Our data suggest a novel electrolyte-based mechanism of immunosuppression in malignant ascites of patients with peritoneal carcinomatosis. We show for the first time that the immunosuppression of NK cytotoxicity in coculture assays is correlated to patient poor survival. Therapeutic application of sodium channel inhibitors may provide new means for restoring immune cell activity in ascites or similar electrolyte imbalanced environments.
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Affiliation(s)
- Antonio Hrvat
- Experimental and Translational Research, Department of Otorhinolaryngology, University Hospital Essen, Hufelandstrasse 55, 45147, Essen, Germany
| | - Mathias Schmidt
- Experimental and Translational Research, Department of Otorhinolaryngology, University Hospital Essen, Hufelandstrasse 55, 45147, Essen, Germany
| | - Bernd Wagner
- Department of Clinical Chemistry, University Hospital Essen, 45147, Essen, Germany
| | - Denise Zwanziger
- Department of Clinical Chemistry, University Hospital Essen, 45147, Essen, Germany
- Department of Endocrinology, Diabetes and Metabolism, University Hospital Essen, 45147, Essen, Germany
| | - Rainer Kimmig
- Department of Gynecology and Obstetrics, University Hospital Essen, 45147, Essen, Germany
| | - Lothar Volbracht
- Department of Clinical Chemistry, University Hospital Essen, 45147, Essen, Germany
| | - Sven Brandau
- Experimental and Translational Research, Department of Otorhinolaryngology, University Hospital Essen, Hufelandstrasse 55, 45147, Essen, Germany.
- partner site Essen-Düsseldorf, German Cancer Consortium (DKTK), 45147, Essen, Germany.
| | - Nina Mallmann-Gottschalk
- Experimental and Translational Research, Department of Otorhinolaryngology, University Hospital Essen, Hufelandstrasse 55, 45147, Essen, Germany
- Department of Gynecology and Obstetrics, University Hospital of Cologne, 50931, Cologne, Germany
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8
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Zhang NZ, Zhao LF, Zhang Q, Fang H, Song WL, Li WZ, Ge YS, Gao P. Core fucosylation and its roles in gastrointestinal glycoimmunology. World J Gastrointest Oncol 2023; 15:1119-1134. [PMID: 37546555 PMCID: PMC10401475 DOI: 10.4251/wjgo.v15.i7.1119] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/18/2023] [Revised: 02/28/2023] [Accepted: 05/08/2023] [Indexed: 07/12/2023] Open
Abstract
Glycosylation is a common post-translational modification in eukaryotic cells. It is involved in the production of many biologically active glycoproteins and the regulation of protein structure and function. Core fucosylation plays a vital role in the immune response. Most immune system molecules are core fucosylated glycoproteins such as complements, cluster differentiation antigens, immunoglobulins, cytokines, major histocompatibility complex molecules, adhesion molecules, and immune molecule synthesis-related transcription factors. These core fucosylated glycoproteins play important roles in antigen recognition and clearance, cell adhesion, lymphocyte activation, apoptosis, signal transduction, and endocytosis. Core fucosylation is dominated by fucosyltransferase 8 (Fut8), which catalyzes the addition of α-1,6-fucose to the innermost GlcNAc residue of N-glycans. Fut8 is involved in humoral, cellular, and mucosal immunity. Tumor immunology is associated with aberrant core fucosylation. Here, we summarize the roles and potential modulatory mechanisms of Fut8 in various immune processes of the gastrointestinal system.
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Affiliation(s)
- Nian-Zhu Zhang
- Clinical Laboratory, The Second Hospital of Dalian Medical University, Dalian 116023, Liaoning Province, China
| | - Li-Fen Zhao
- Clinical Laboratory, The Second Hospital of Dalian Medical University, Dalian 116023, Liaoning Province, China
| | - Qian Zhang
- Department of Cell Therapy, Shanghai Tianze Yuntai Biomedical Co., Ltd., Shanghai 200100, China
| | - Hui Fang
- Graduate School of Comprehensive Human Sciences, University of Tsukuba, Tsukuba 305-0005, Ibaraki, Japan
| | - Wan-Li Song
- Guangdong Provincial Key Laboratory of Infectious Diseases and Molecular Immunopathology, Shantou University Medical College, Shantou 515041, Guangdong Province, China
| | - Wen-Zhe Li
- Guangdong Provincial Key Laboratory of Infectious Diseases and Molecular Immunopathology, Shantou University Medical College, Shantou 515041, Guangdong Province, China
| | - Yu-Song Ge
- Department of Neurology, The Second Hospital of Dalian Medical University, Dalian 116023, Liaoning Province, China
| | - Peng Gao
- Clinical Laboratory, The Second Hospital of Dalian Medical University, Dalian 116023, Liaoning Province, China
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9
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Lipinski B, Arras P, Pekar L, Klewinghaus D, Boje AS, Krah S, Zimmermann J, Klausz K, Peipp M, Siegmund V, Evers A, Zielonka S. NKp46-specific single domain antibodies enable facile engineering of various potent NK cell engager formats. Protein Sci 2023; 32:e4593. [PMID: 36775946 PMCID: PMC9951198 DOI: 10.1002/pro.4593] [Citation(s) in RCA: 10] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/25/2022] [Revised: 02/07/2023] [Accepted: 02/09/2023] [Indexed: 02/14/2023]
Abstract
Herein, we describe the generation of potent NK cell engagers (NKCEs) based on single domain antibodies (sdAbs) specific for NKp46 harboring the humanized Fab version of Cetuximab for tumor targeting. After immunization of camelids, a plethora of different VHH domains were retrieved by yeast surface display. Upon reformatting into Fc effector-silenced NKCEs targeting NKp46 and EGFR in a strictly monovalent fashion, the resulting bispecific antibodies elicited potent NK cell-mediated killing of EGFR-overexpressing tumor cells with potencies (EC50 killing) in the picomolar range. This was further augmented via co-engagement of Fcγ receptor IIIa (FcγRIIIa). Importantly, NKp46-specific sdAbs enabled the construction of various NKCE formats with different geometries and valencies which displayed favorable biophysical and biochemical properties without further optimization. By this means, killing capacities were further improved significantly. Hence, NKp46-specific sdAbs are versatile building blocks for the construction of different NKCE formats.
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Affiliation(s)
- Britta Lipinski
- Protein Engineering and Antibody TechnologiesMerck Healthcare KGaADarmstadtGermany
- Institute for Organic Chemistry and BiochemistryTechnical University of DarmstadtDarmstadtGermany
| | - Paul Arras
- Protein Engineering and Antibody TechnologiesMerck Healthcare KGaADarmstadtGermany
| | - Lukas Pekar
- Protein Engineering and Antibody TechnologiesMerck Healthcare KGaADarmstadtGermany
| | - Daniel Klewinghaus
- Protein Engineering and Antibody TechnologiesMerck Healthcare KGaADarmstadtGermany
| | - Ammelie Svea Boje
- Division of Antibody‐Based Immunotherapy, Department of Internal Medicine IIUniversity Hospital Schleswig‐Holstein and Christian‐Albrechts‐University KielKielGermany
| | - Simon Krah
- Protein Engineering and Antibody TechnologiesMerck Healthcare KGaADarmstadtGermany
| | - Jasmin Zimmermann
- Protein Engineering and Antibody TechnologiesMerck Healthcare KGaADarmstadtGermany
- Institute for Organic Chemistry and BiochemistryTechnical University of DarmstadtDarmstadtGermany
| | - Katja Klausz
- Division of Antibody‐Based Immunotherapy, Department of Internal Medicine IIUniversity Hospital Schleswig‐Holstein and Christian‐Albrechts‐University KielKielGermany
| | - Matthias Peipp
- Division of Antibody‐Based Immunotherapy, Department of Internal Medicine IIUniversity Hospital Schleswig‐Holstein and Christian‐Albrechts‐University KielKielGermany
| | | | - Andreas Evers
- Computational Chemistry and BiologyMerck Healthcare KGaADarmstadtGermany
| | - Stefan Zielonka
- Protein Engineering and Antibody TechnologiesMerck Healthcare KGaADarmstadtGermany
- Institute for Organic Chemistry and BiochemistryTechnical University of DarmstadtDarmstadtGermany
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10
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Whalen KA, Rakhra K, Mehta NK, Steinle A, Michaelson JS, Baeuerle PA. Engaging natural killer cells for cancer therapy via NKG2D, CD16A and other receptors. MAbs 2023; 15:2208697. [PMID: 37165468 PMCID: PMC10173799 DOI: 10.1080/19420862.2023.2208697] [Citation(s) in RCA: 19] [Impact Index Per Article: 9.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/04/2023] [Accepted: 04/26/2023] [Indexed: 05/12/2023] Open
Abstract
The field of immuno-oncology has revolutionized cancer patient care and improved survival and quality of life for patients. Much of the focus in the field has been on exploiting the power of the adaptive immune response through therapeutic targeting of T cells. While these approaches have markedly advanced the field, some challenges remain, and the clinical benefit of T cell therapies does not extend to all patients or tumor indications. Alternative strategies, such as engaging the innate immune system, have become an intense area of focus in the field. In particular, the engagement of natural killer (NK) cells as potent effectors of the innate immune response has emerged as a promising modality in immunotherapy. Here, we review therapeutic approaches for selective engagement of NK cells for cancer therapy, with a particular focus on targeting the key activating receptors NK Group 2D (NKG2D) and cluster of differentiation 16A (CD16A).
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Affiliation(s)
- Kerry A. Whalen
- Preclinical and Early Development, Cullinan Oncology, Inc, Cambridge, MA, USA
| | - Kavya Rakhra
- Preclinical and Early Development, Cullinan Oncology, Inc, Cambridge, MA, USA
| | - Naveen K. Mehta
- Preclinical and Early Development, Cullinan Oncology, Inc, Cambridge, MA, USA
| | - Alexander Steinle
- Institute for Molecular Medicine, Goethe-University Frankfurt, Frankfurt am Main, Germany
- Preclinical and Early Development, Frankfurt Cancer Institute, Frankfurt am Main, Germany
| | | | - Patrick A. Baeuerle
- Preclinical and Early Development, Cullinan Oncology, Inc, Cambridge, MA, USA
- Institute for Immunology, Ludwig Maximilians University, Munich, Germany
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11
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Yanaka S, Yogo R, Yagi H, Onitsuka M, Wakaizumi N, Yamaguchi Y, Uchiyama S, Kato K. Negative interference with antibody-dependent cellular cytotoxicity mediated by rituximab from its interactions with human serum proteins. Front Immunol 2023; 14:1090898. [PMID: 36761774 PMCID: PMC9905677 DOI: 10.3389/fimmu.2023.1090898] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/06/2022] [Accepted: 01/03/2023] [Indexed: 01/26/2023] Open
Abstract
Although interactions of small molecular drugs with serum proteins have been widely studied from pharmacokinetic and pharmacodynamic perspectives, there have been few reports on the effects of serum components on therapeutic antibody functions. This study reports the effect of abundant serum proteins on antibody-dependent cellular cytotoxicity (ADCC) mediated by rituximab and Fcγ receptor III (FcγRIII). Human serum albumin (HSA) and the Fab fragment from the pooled serum polyclonal IgG were found to compromise ADCC as non-competitive inhibitors. Our nuclear magnetic resonance data provided direct evidence for the interactions of HSA with both the Fab and Fc regions of rituximab and also with the extracellular region of FcγRIII (sFcγRIII). The degree of involvement in the interaction decreased in the order of rituximab-Fab > rituximab-Fc > sFcγRIII, suggesting preferential binding of HSA to net positively charged proteins. Although much less pronounced than the effect of HSA, polyclonal IgG-Fab specifically interacted with rituximab-Fc. The NMR data also showed that the serum protein interactions cover the Fc surface extensively, suggesting that they can act as pan-inhibitors against various Fc receptor-mediated functions and pharmacokinetics. Our findings highlight the importance of considering serum-protein interactions in the design and application of antibody-based drugs with increased efficacy and safety.
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Affiliation(s)
- Saeko Yanaka
- Exploratory Research Center on Life and Living Systems (ExCELLS), National Institutes of Natural Sciences, Okazaki, Japan.,Institute for Molecular Science (IMS), National Institutes of Natural Sciences, Okazaki, Japan.,Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya, Japan.,Graduate School of Pharmaceutical Sciences, Kyushu University, Fukuoka, Japan
| | - Rina Yogo
- Exploratory Research Center on Life and Living Systems (ExCELLS), National Institutes of Natural Sciences, Okazaki, Japan.,Institute for Molecular Science (IMS), National Institutes of Natural Sciences, Okazaki, Japan.,Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya, Japan
| | - Hirokazu Yagi
- Exploratory Research Center on Life and Living Systems (ExCELLS), National Institutes of Natural Sciences, Okazaki, Japan.,Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya, Japan
| | - Masayoshi Onitsuka
- Graduate School of Technology, Industrial and Social Sciences, Tokushima University, Tokushima, Japan
| | | | - Yuki Yamaguchi
- Graduate School of Engineering, University of Osaka, Osaka, Japan
| | - Susumu Uchiyama
- Graduate School of Engineering, University of Osaka, Osaka, Japan
| | - Koichi Kato
- Exploratory Research Center on Life and Living Systems (ExCELLS), National Institutes of Natural Sciences, Okazaki, Japan.,Institute for Molecular Science (IMS), National Institutes of Natural Sciences, Okazaki, Japan.,Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya, Japan
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12
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Ghaedrahmati F, Esmaeil N, Abbaspour M. Targeting immune checkpoints: how to use natural killer cells for fighting against solid tumors. Cancer Commun (Lond) 2022; 43:177-213. [PMID: 36585761 PMCID: PMC9926962 DOI: 10.1002/cac2.12394] [Citation(s) in RCA: 17] [Impact Index Per Article: 5.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/25/2022] [Revised: 10/08/2022] [Accepted: 11/15/2022] [Indexed: 01/01/2023] Open
Abstract
Natural killer (NK) cells are unique innate immune cells that mediate anti-viral and anti-tumor responses. Thus, they might hold great potential for cancer immunotherapy. NK cell adoptive immunotherapy in humans has shown modest efficacy. In particular, it has failed to demonstrate therapeutic efficiency in the treatment of solid tumors, possibly due in part to the immunosuppressive tumor microenvironment (TME), which reduces NK cell immunotherapy's efficiencies. It is known that immune checkpoints play a prominent role in creating an immunosuppressive TME, leading to NK cell exhaustion and tumor immune escape. Therefore, NK cells must be reversed from their dysfunctional status and increased in their effector roles in order to improve the efficiency of cancer immunotherapy. Blockade of immune checkpoints can not only rescue NK cells from exhaustion but also augment their robust anti-tumor activity. In this review, we discussed immune checkpoint blockade strategies with a focus on chimeric antigen receptor (CAR)-NK cells to redirect NK cells to cancer cells in the treatment of solid tumors.
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Affiliation(s)
- Farhoodeh Ghaedrahmati
- Department of ImmunologySchool of MedicineIsfahan University of Medical SciencesIsfahanIran
| | - Nafiseh Esmaeil
- Department of ImmunologySchool of MedicineIsfahan University of Medical SciencesIsfahanIran,Research Institute for Primordial Prevention of Non‐Communicable DiseaseIsfahan University of Medical SciencesIsfahanIran
| | - Maryam Abbaspour
- Department of Pharmaceutical BiotechnologyFaculty of PharmacyIsfahan University of Medical SciencesIsfahanIran
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13
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Hambach J, Mann AM, Bannas P, Koch-Nolte F. Targeting multiple myeloma with nanobody-based heavy chain antibodies, bispecific killer cell engagers, chimeric antigen receptors, and nanobody-displaying AAV vectors. Front Immunol 2022; 13:1005800. [PMID: 36405759 PMCID: PMC9668101 DOI: 10.3389/fimmu.2022.1005800] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/28/2022] [Accepted: 10/21/2022] [Indexed: 11/29/2022] Open
Abstract
Nanobodies are well suited for constructing biologics due to their high solubility. We generated nanobodies directed against CD38, a tumor marker that is overexpressed by multiple myeloma and other hematological malignancies. We then used these CD38-specific nanobodies to construct heavy chain antibodies, bispecific killer cell engagers (BiKEs), chimeric antigen receptor (CAR)-NK cells, and nanobody-displaying AAV vectors. Here we review the utility of these nanobody-based constructs to specifically and effectively target CD38-expressing myeloma cells. The promising results of our preclinical studies warrant further clinical studies to evaluate the potential of these CD38-specific nanobody-based constructs for treatment of multiple myeloma.
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Affiliation(s)
- Julia Hambach
- Institute of Immunology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany,Department of Diagnostic and Interventional Radiology and Nuclear Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Anna Marei Mann
- Institute of Immunology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Peter Bannas
- Department of Diagnostic and Interventional Radiology and Nuclear Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Friedrich Koch-Nolte
- Institute of Immunology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany,*Correspondence: Friedrich Koch-Nolte,
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14
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Klausz K, Pekar L, Boje AS, Gehlert CL, Krohn S, Gupta T, Xiao Y, Krah S, Zaynagetdinov R, Lipinski B, Toleikis L, Poetzsch S, Rabinovich B, Peipp M, Zielonka S. Multifunctional NK Cell–Engaging Antibodies Targeting EGFR and NKp30 Elicit Efficient Tumor Cell Killing and Proinflammatory Cytokine Release. THE JOURNAL OF IMMUNOLOGY 2022; 209:1724-1735. [DOI: 10.4049/jimmunol.2100970] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/08/2021] [Accepted: 08/23/2022] [Indexed: 01/04/2023]
Abstract
Abstract
In this work, we have generated novel Fc-comprising NK cell engagers (NKCEs) that bridge human NKp30 on NK cells to human epidermal growth factor receptor (EGFR) on tumor cells. Camelid-derived VHH single-domain Abs specific for human NKp30 and a humanized Fab derived from the EGFR-specific therapeutic Ab cetuximab were used as binding arms. By combining camelid immunization with yeast surface display, we were able to isolate a diverse panel of NKp30-specific VHHs against different epitopes on NKp30. Intriguingly, NKCEs built with VHHs that compete for binding to NKp30 with B7-H6, the natural ligand of NKp30, were significantly more potent in eliciting tumor cell lysis of EGFR-positive tumor cells than NKCEs harboring VHHs that target different epitopes on NKp30 from B7-H6. We demonstrate that the NKCEs can be further improved with respect to killing capabilities by concomitant engagement of FcγRIIIa and that soluble B7-H6 does not impede cytolytic capacities of all scrutinized NKCEs at significantly higher B7-H6 concentrations than observed in cancer patients. Moreover, we show that physiological processes requiring interactions between membrane-bound B7-H6 and NKp30 on NK cells are unaffected by noncompeting NKCEs still eliciting tumor cell killing at low picomolar concentrations. Ultimately, the NKCEs generated in this study were significantly more potent in eliciting NK cell–mediated tumor cell lysis than cetuximab and elicited a robust release of proinflammatory cytokines, both features which might be beneficial for antitumor therapy.
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Affiliation(s)
- Katja Klausz
- *Division of Antibody-Based Immunotherapy, Department of Internal Medicine II, University Hospital Schleswig-Holstein and Christian Albrechts University Kiel, Kiel, Germany
| | - Lukas Pekar
- †Protein Engineering and Antibody Technologies, Merck Healthcare KGaA, Darmstadt, Germany
| | - Ammelie Svea Boje
- *Division of Antibody-Based Immunotherapy, Department of Internal Medicine II, University Hospital Schleswig-Holstein and Christian Albrechts University Kiel, Kiel, Germany
| | - Carina Lynn Gehlert
- *Division of Antibody-Based Immunotherapy, Department of Internal Medicine II, University Hospital Schleswig-Holstein and Christian Albrechts University Kiel, Kiel, Germany
| | - Steffen Krohn
- *Division of Antibody-Based Immunotherapy, Department of Internal Medicine II, University Hospital Schleswig-Holstein and Christian Albrechts University Kiel, Kiel, Germany
| | - Tushar Gupta
- ‡Protein Engineering and Antibody Technologies, EMD Serono Research & Development Institute, Inc., Billerica, MA
| | - Yanping Xiao
- §Department of Oncology and Immuno-oncology, EMD Serono Research & Development Institute, Inc., Billerica, MA
| | - Simon Krah
- †Protein Engineering and Antibody Technologies, Merck Healthcare KGaA, Darmstadt, Germany
| | - Rinat Zaynagetdinov
- §Department of Oncology and Immuno-oncology, EMD Serono Research & Development Institute, Inc., Billerica, MA
| | - Britta Lipinski
- †Protein Engineering and Antibody Technologies, Merck Healthcare KGaA, Darmstadt, Germany
- ¶Institute for Organic Chemistry and Biochemistry, Technical University of Darmstadt, Darmstadt, Germany; and
| | - Lars Toleikis
- †Protein Engineering and Antibody Technologies, Merck Healthcare KGaA, Darmstadt, Germany
| | - Sven Poetzsch
- ‖Strategic Innovation, Merck Healthcare KGaA, Darmstadt, Germany
| | - Brian Rabinovich
- §Department of Oncology and Immuno-oncology, EMD Serono Research & Development Institute, Inc., Billerica, MA
| | - Matthias Peipp
- *Division of Antibody-Based Immunotherapy, Department of Internal Medicine II, University Hospital Schleswig-Holstein and Christian Albrechts University Kiel, Kiel, Germany
| | - Stefan Zielonka
- †Protein Engineering and Antibody Technologies, Merck Healthcare KGaA, Darmstadt, Germany
- ¶Institute for Organic Chemistry and Biochemistry, Technical University of Darmstadt, Darmstadt, Germany; and
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15
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Biological Therapies in the Treatment of Cancer-Update and New Directions. Int J Mol Sci 2021; 22:ijms222111694. [PMID: 34769123 PMCID: PMC8583892 DOI: 10.3390/ijms222111694] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/07/2021] [Revised: 10/23/2021] [Accepted: 10/25/2021] [Indexed: 12/22/2022] Open
Abstract
Biological therapies have changed the face of oncology by targeting cancerous cells while reducing the effect on normal tissue. This publication focuses mainly on new therapies that have contributed to the advances in treatment of certain malignancies. Immunotherapy, which has repeatedly proven to be a breakthrough therapy in melanoma, as well as B-ALL therapy with CAR T cells, are of great merit in this progress. These therapies are currently being developed by modifying bispecific antibodies and CAR T cells to improve their efficiency and bioavailability. Work on improving the therapy with oncolytic viruses is also progressing, and efforts are being made to improve the immunogenicity and stability of cancer vaccines. Combining various biological therapies, immunotherapy with oncolytic viruses or cancer vaccines is gaining importance in cancer therapy. New therapeutic targets are intensively sought among neoantigens, which are not immunocompromised, or antigens associated with tumor stroma cells. An example is fibroblast activation protein α (FAPα), the overexpression of which is observed in the case of tumor progression. Universal therapeutic targets are also sought, such as the neurotrophic receptor tyrosine kinase (NTRK) gene fusion, a key genetic driver present in many types of cancer. This review also raises the problem of the tumor microenvironment. Stromal cells can protect tumor cells from chemotherapy and contribute to relapse and progression. This publication also addresses the problem of cancer stem cells resistance to treatment and presents attempts to avoid this phenomenon. This review focuses on the most important strategies used to improve the selectivity of biological therapies.
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16
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Perera Molligoda Arachchige AS. NK cell-based therapies for HIV infection: Investigating current advances and future possibilities. J Leukoc Biol 2021; 111:921-931. [PMID: 34668588 DOI: 10.1002/jlb.5ru0821-412rr] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/08/2022] Open
Abstract
NK cells are well-known for their antiviral functions. Also, their role in HIV has been well established, with rapid responses elicited during early HIV infection. Most immune cells including CD4+ T cells, monocytes, Mϕs, and dendritic cells are readily infected by HIV. Recent evidence from multiple studies has suggested that similar to these cells, in chronic conditions like HIV, NK cells also undergo functional exhaustion with impaired cytotoxicity, altered cytokine production, and impaired ADCC. NK-based immunotherapy aims to successfully restore, boost, and modify their activity as has been already demonstrated in the field of cancer immunotherapy. The utilization of NK cell-based strategies for the eradication of HIV from the body provides many advantages over classical ART. The literature search consisted of manually selecting the most relevant studies from databases including PubMed, Embase, Google Scholar, and ClinicalTrial.gov. Some of the treatments currently under consideration are CAR-NK cell therapy, facilitating ADCC, TLR agonists, bNAbs, and BiKEs/TriKEs, blocking inhibitory NK receptors during infection, IL-15 and IL-15 superagonists (eg: ALT-803), and so on. This review aims to discuss the NK cell-based therapies currently under experimentation against HIV infection and finally highlight the challenges associated with NK cell-based immunotherapies.
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17
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Abstract
The aggregation of therapeutic antibodies is a major issue for the pharmaceutical industry leading to loss of drug quality, increased dosage, and unwanted immune responses such as the production of anti-drug antibodies (ADA). As aggregation can occur at various stages of development and storage, much work has been performed to reduce or eliminate it. In this report we analyzed four antibodies available in the PDB (1IGT, 1IGY, 1HZH, and 5DK3) using the online software UCSF Chimera to study the structural features of the proteins and the associated N-linked glycans in the CH2 domains of the Fc region. To study antibody aggregation in silico we used the online software TANGO and AGGRESCAN to identify aggregation prone regions (APR) in the antibodies and the influence of the Fc glycans on hydrophobic and aromatic residues present in the APRs. In the 3D structures of 1IGT and 1IGY the glycan chains are in close enough proximity to influence and protect these hydrophobic regions. However, in the 3D structures of 1HZH and 5DK3 the glycans do not appear to influence the likely APRs of the antibodies. Therefore, in these structures we modified the Fc glycan regions by adjusting the glycosylated asparagine side chains and glycosidic bonds. We successfully adjusted the glycan chains of 1HZH and 5DK3 and reduced the distance between them and the APRs to show potential influence on aggregation. However, similar to 5DK3, the influence of glycosylation on the APRs of the antibody was limited due to the size of the glycans present in the 3D structure. This report is based on in silico studies to show how antibody glycans can influence aggregation.
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18
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The Role of Fc Receptors on the Effectiveness of Therapeutic Monoclonal Antibodies. Int J Mol Sci 2021; 22:ijms22168947. [PMID: 34445651 PMCID: PMC8396266 DOI: 10.3390/ijms22168947] [Citation(s) in RCA: 74] [Impact Index Per Article: 18.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/29/2021] [Revised: 08/14/2021] [Accepted: 08/16/2021] [Indexed: 12/12/2022] Open
Abstract
Since the approval of the first monoclonal antibody (mAb) in 1986, a huge effort has been made to guarantee safety and efficacy of therapeutic mAbs. As of July 2021, 118 mAbs are approved for the European market for a broad range of clinical indications. In order to ensure clinical efficacy and safety aspects, (pre-)clinical experimental approaches evaluate the respective modes of action (MoA). In addition to antigen-specificity including binding affinity and -avidity, MoA comprise Fc-mediated effector functions such as antibody dependent cellular cytotoxicity (ADCC) and the closely related antibody dependent cellular phagocytosis (ADCP). For this reason, a variety of cell-based assays have been established investigating effector functions of therapeutic mAbs with different effector/target-cell combinations and several readouts including Fcγ receptor (FcγR)-mediated lysis, fluorescence, or luminescence. Optimized FcγR-mediated effector functions regarding clinical safety and efficacy are addressed with modification strategies such as point mutations, altered glycosylation patterns, combination of different Fc subclasses (cross isotypes), and Fc-truncation of the mAb. These strategies opened the field for a next generation of therapeutic mAbs. In conclusion, it is of major importance to consider FcγR-mediated effector functions for the efficacy of therapeutic mAbs.
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19
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Pekar L, Klausz K, Busch M, Valldorf B, Kolmar H, Wesch D, Oberg HH, Krohn S, Boje AS, Gehlert CL, Toleikis L, Krah S, Gupta T, Rabinovich B, Zielonka S, Peipp M. Affinity Maturation of B7-H6 Translates into Enhanced NK Cell-Mediated Tumor Cell Lysis and Improved Proinflammatory Cytokine Release of Bispecific Immunoligands via NKp30 Engagement. THE JOURNAL OF IMMUNOLOGY 2020; 206:225-236. [PMID: 33268483 DOI: 10.4049/jimmunol.2001004] [Citation(s) in RCA: 31] [Impact Index Per Article: 6.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/01/2020] [Accepted: 11/02/2020] [Indexed: 02/06/2023]
Abstract
Activating NK cell receptors represent promising target structures to elicit potent antitumor immune responses. In this study, novel immunoligands were generated that bridge the activating NK cell receptor NKp30 on NK cells with epidermal growth factor receptor (EGFR) on tumor cells in a bispecific IgG-like format based on affinity-optimized versions of B7-H6 and the Fab arm derived from cetuximab. To enhance NKp30 binding, the solitary N-terminal IgV domain of B7-H6 (ΔB7-H6) was affinity matured by an evolutionary library approach combined with yeast surface display. Biochemical and functional characterization of 36 of these novel ΔB7-H6-derived NK cell engagers revealed an up to 45-fold-enhanced affinity for NKp30 and significantly improved NK cell-mediated, EGFR-dependent killing of tumor cells compared with the NK cell engager based on the wild-type ΔB7-H6 domain. In this regard, potencies (EC50 killing) of the best immunoligands were substantially improved by up to 87-fold. Moreover, release of IFN-γ and TNF-α was significantly increased. Importantly, equipment of the ΔB7-H6-based NK cell engagers with a human IgG1 Fc part competent in Fc receptor binding resulted in an almost 10-fold superior killing of EGFR-overexpressing tumor cells compared with molecules either triggering FcγRIIIa or NKp30. Additionally, INF-γ and TNF-α release was increased compared with molecules solely triggering FcγRIIIa, including the clinically approved Ab cetuximab. Thus, incorporating affinity-matured ligands for NK cell-activating receptors might represent an effective strategy for the generation of potent novel therapeutic agents with unique effector functions in cancer immunotherapy.
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Affiliation(s)
- Lukas Pekar
- Protein Engineering and Antibody Technologies, Merck KGaA, D-64293 Darmstadt, Germany.,Discovery Pharmacology, Merck KGaA, D-64293 Darmstadt, Germany
| | - Katja Klausz
- Division of Stem Cell Transplantation and Immunotherapy, Department of Medicine II, University Hospital Schleswig-Holstein and Christian-Albrechts-University of Kiel, D-24105 Kiel, Germany
| | - Michael Busch
- Discovery Pharmacology, Merck KGaA, D-64293 Darmstadt, Germany
| | - Bernhard Valldorf
- Chemical and Pharmaceutical Development, Merck KGaA, D-64293 Darmstadt, Germany
| | - Harald Kolmar
- Institute for Organic Chemistry and Biochemistry, Technische Universität Darmstadt, D-64287 Darmstadt, Germany
| | - Daniela Wesch
- Institute of Immunology, University Hospital Schleswig-Holstein and Christian-Albrechts-University of Kiel, D-24105 Kiel, Germany; and
| | - Hans-Heinrich Oberg
- Institute of Immunology, University Hospital Schleswig-Holstein and Christian-Albrechts-University of Kiel, D-24105 Kiel, Germany; and
| | - Steffen Krohn
- Division of Stem Cell Transplantation and Immunotherapy, Department of Medicine II, University Hospital Schleswig-Holstein and Christian-Albrechts-University of Kiel, D-24105 Kiel, Germany
| | - Ammelie Svea Boje
- Division of Stem Cell Transplantation and Immunotherapy, Department of Medicine II, University Hospital Schleswig-Holstein and Christian-Albrechts-University of Kiel, D-24105 Kiel, Germany
| | - Carina Lynn Gehlert
- Division of Stem Cell Transplantation and Immunotherapy, Department of Medicine II, University Hospital Schleswig-Holstein and Christian-Albrechts-University of Kiel, D-24105 Kiel, Germany
| | - Lars Toleikis
- Protein Engineering and Antibody Technologies, Merck KGaA, D-64293 Darmstadt, Germany
| | - Simon Krah
- Protein Engineering and Antibody Technologies, Merck KGaA, D-64293 Darmstadt, Germany
| | - Tushar Gupta
- Department of Immuno-oncology, EMD Serono Research & Development Institute Inc., Billerica, MA 01821
| | - Brian Rabinovich
- Department of Immuno-oncology, EMD Serono Research & Development Institute Inc., Billerica, MA 01821
| | - Stefan Zielonka
- Protein Engineering and Antibody Technologies, Merck KGaA, D-64293 Darmstadt, Germany;
| | - Matthias Peipp
- Division of Stem Cell Transplantation and Immunotherapy, Department of Medicine II, University Hospital Schleswig-Holstein and Christian-Albrechts-University of Kiel, D-24105 Kiel, Germany;
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20
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Sustmann C, Dickopf S, Regula JT, Kettenberger H, Mølhøj M, Gassner C, Weininger D, Fenn S, Manigold T, Kling L, Künkele KP, Schwaiger M, Bossenmaier B, Griese JJ, Hopfner KP, Graff-Meyer A, Stahlberg H, Ringler P, Lauer ME, Brinkmann U, Schaefer W, Klein C. DuoMab: a novel CrossMab-based IgG-derived antibody format for enhanced antibody-dependent cell-mediated cytotoxicity. MAbs 2019; 11:1402-1414. [PMID: 31526159 PMCID: PMC6816436 DOI: 10.1080/19420862.2019.1661736] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/21/2019] [Revised: 08/08/2019] [Accepted: 08/26/2019] [Indexed: 01/16/2023] Open
Abstract
High specificity accompanied with the ability to recruit immune cells has made recombinant therapeutic antibodies an integral part of drug development. Here we present a generic approach to generate two novel IgG-derived antibody formats that are based on a modification of the CrossMab technology. MoAbs harbor two heavy chains (HCs) resulting in one binding entity and one fragment crystallizable region (Fc), whereas DuoMabs are composed of four HCs harboring two binding entities and two Fc regions linked at a disulfide-bridged hinge. The latter bivalent format is characterized by avidity-enhanced target cell binding while simultaneously increasing the 'Fc-load' on the surface. DuoMabs were shown to be producible in high yield and purity and bind to surface cells with affinities comparable to IgGs. The increased Fc load directed at the surface of target cells by DuoMabs modulates their antibody-dependent cell-mediated cytotoxicity competency toward target cells, making them attractive for applications that require or are modulated by FcR interactions.
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Affiliation(s)
- Claudio Sustmann
- Roche Pharma Research and Early Development (pRED), Large Molecule Research (LMR), Roche Innovation Center Munich, Penzberg, Germany
| | - Steffen Dickopf
- Roche Pharma Research and Early Development (pRED), Large Molecule Research (LMR), Roche Innovation Center Munich, Penzberg, Germany
| | - Jörg T. Regula
- Roche Pharma Research and Early Development (pRED), Large Molecule Research (LMR), Roche Innovation Center Munich, Penzberg, Germany
| | - Hubert Kettenberger
- Roche Pharma Research and Early Development (pRED), Large Molecule Research (LMR), Roche Innovation Center Munich, Penzberg, Germany
| | - Michael Mølhøj
- Roche Pharma Research and Early Development (pRED), Large Molecule Research (LMR), Roche Innovation Center Munich, Penzberg, Germany
| | - Christian Gassner
- Roche Pharma Research and Early Development (pRED), Large Molecule Research (LMR), Roche Innovation Center Munich, Penzberg, Germany
| | - Diana Weininger
- Roche Pharma Research and Early Development (pRED), Large Molecule Research (LMR), Roche Innovation Center Munich, Penzberg, Germany
| | - Sebastian Fenn
- Roche Pharma Research and Early Development (pRED), Large Molecule Research (LMR), Roche Innovation Center Munich, Penzberg, Germany
| | - Tobias Manigold
- Roche Pharma Research and Early Development (pRED), Discovery Oncology, Roche Innovation Center Basel, Basel, Switzerland
| | - Lothar Kling
- Roche Pharma Research and Early Development (pRED), Large Molecule Research (LMR), Roche Innovation Center Munich, Penzberg, Germany
| | - Klaus-Peter Künkele
- Roche Pharma Research and Early Development (pRED), Large Molecule Research (LMR), Roche Innovation Center Munich, Penzberg, Germany
| | - Manfred Schwaiger
- Roche Pharma Research and Early Development (pRED), Discovery Oncology, Roche Innovation Center Basel, Basel, Switzerland
| | - Birgit Bossenmaier
- Roche Pharma Research and Early Development (pRED), Large Molecule Research (LMR), Roche Innovation Center Munich, Penzberg, Germany
| | - Julia J. Griese
- Gene Center and Department of Biochemistry, Ludwig-Maximilians-University, Munich, Germany
| | - Karl-Peter Hopfner
- Gene Center and Department of Biochemistry, Ludwig-Maximilians-University, Munich, Germany
| | - Alexandra Graff-Meyer
- Center for Cellular Imaging and Nanoanalytics, Biozentrum, University of Basel, Basel, Switzerland
| | - Henning Stahlberg
- Center for Cellular Imaging and Nanoanalytics, Biozentrum, University of Basel, Basel, Switzerland
| | - Philippe Ringler
- Center for Cellular Imaging and Nanoanalytics, Biozentrum, University of Basel, Basel, Switzerland
| | - Matthias E. Lauer
- Roche Pharma Research and Early Development (pRED), Small Molecule Research, Roche Innovation Center Basel, Basel, Switzerland
| | - Ulrich Brinkmann
- Roche Pharma Research and Early Development (pRED), Large Molecule Research (LMR), Roche Innovation Center Munich, Penzberg, Germany
| | - Wolfgang Schaefer
- Roche Pharma Research and Early Development (pRED), Large Molecule Research (LMR), Roche Innovation Center Munich, Penzberg, Germany
| | - Christian Klein
- Roche Pharma Research and Early Development (pRED), Discovery Oncology, Roche Innovation Center Zurich, Schlieren, Switzerland
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21
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Gudelj I, Lauc G, Pezer M. Immunoglobulin G glycosylation in aging and diseases. Cell Immunol 2018; 333:65-79. [DOI: 10.1016/j.cellimm.2018.07.009] [Citation(s) in RCA: 206] [Impact Index Per Article: 29.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/12/2018] [Revised: 07/19/2018] [Accepted: 07/19/2018] [Indexed: 01/01/2023]
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22
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Li Y, Huang K, Liu L, Qu Y, Huang Y, Wu Y, Wei J. Effects of complement and serum IgG on rituximab-dependent natural killer cell-mediated cytotoxicity against Raji cells. Oncol Lett 2018; 17:339-347. [PMID: 30655772 PMCID: PMC6313095 DOI: 10.3892/ol.2018.9630] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/15/2017] [Accepted: 10/04/2018] [Indexed: 12/24/2022] Open
Abstract
Accumulating evidence indicates that the anti-CD20 monoclonal antibody rituximab significantly improves the clinical prognosis of patients with non-Hodgkin lymphoma and chronic lymphocytic leukemia. However, a number of patients relapse or fail to respond to rituximab. To further understand the cause of this, polymorphisms of FcγRIIIa were initially detected in healthy volunteers. Subsequently, the rituximab-dependent natural killer (NK) cell-mediated cytotoxicity of different FcγRIIIa genotypes was assessed by a cytotoxicity assay in vitro. Ultimately, the effect of human serum immunoglobulin (Ig) G and complement on rituximab-dependent NK cell-mediated cytotoxicity was evaluated in vitro. It was revealed that FcγRIIIa polymorphisms were associated with the antibody-dependent cell-mediated cytotoxicity (ADCC) of NK cells. In addition, the ADCC of NK cells with FcγRIIIa-158 V/V was increased compared with that of FcγRIIIa-158 V/F. The serum IgG and rituximab Fc segment was able to bind competitively with NK cell FcγRIIIa. It was observed that serum IgG inhibited, whereas complement enhanced rituximab-induced NK-cell mediated ADCC. Therefore, various agents administered synchronously with rituximab may modulate the efficacy of this agent and ultimately its toxicity against tumor cells.
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Affiliation(s)
- Yang Li
- Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, Guangdong 510120, P.R. China.,Department of Pediatric Hematology/Oncology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, Guangdong 510120, P.R. China
| | - Ke Huang
- Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, Guangdong 510120, P.R. China.,Department of Pediatric Hematology/Oncology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, Guangdong 510120, P.R. China
| | - Ling Liu
- Department of Pediatric, Affiliatied Hospital of Guangdong Medical University, Zhanjiang, Guangdong 524000, P.R. China
| | - Yuhua Qu
- Department of Pediatrics, Guang Zhou Women and Children's Medical Center, Guangzhou, Guangdong 510120, P.R. China
| | - Yan Huang
- Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, Guangdong 510120, P.R. China.,Department of Pediatric Hematology/Oncology, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, Guangdong 510120, P.R. China
| | - Yanfeng Wu
- Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, Guangdong 510120, P.R. China.,Biotherapy Center, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, Guangdong 510120, P.R. China
| | - Jing Wei
- Guangdong Provincial Key Laboratory of Malignant Tumor Epigenetics and Gene Regulation, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, Guangdong 510120, P.R. China.,Medical Research Center, Sun Yat-Sen Memorial Hospital, Sun Yat-Sen University, Guangzhou, Guangdong 510120, P.R. China
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23
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Dalziel M, Beers SA, Cragg MS, Crispin M. Through the barricades: overcoming the barriers to effective antibody-based cancer therapeutics. Glycobiology 2018; 28:697-712. [PMID: 29800150 DOI: 10.1093/glycob/cwy043] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/30/2018] [Accepted: 04/30/2018] [Indexed: 02/06/2023] Open
Abstract
Since the turn of the century, cancer therapy has undergone a transformation in terms of new treatment modalities and renewed optimism in achieving long-lived tumor control and even cure. This is, in large part, thanks to the widespread incorporation of monoclonal antibodies (mAbs) into standard treatment regimens. These new therapies have, across many settings, significantly contributed to improved clinical responses, patient quality of life and survival. Moreover, the flexibility of the antibody platform has led to the development of a wide range of innovative and combinatorial therapies that continue to augment the clinician's armory. Despite these successes, there is a growing awareness that in many cases mAb therapy remains suboptimal, primarily due to inherent limitations imposed by the immune system's own homeostatic controls and the immunosuppressive tumor microenvironment. Here, we discuss the principal barriers that act to constrain the tumor-killing activity of antibody-based therapeutics, particularly those involving antibody glycans, using illustrative examples from both pre-clinical and market approved mAbs. We also discuss strategies that have been, or are in development to overcome these obstacles. Finally, we outline how the growing understanding of the biological terrain in which mAbs function is shaping innovation and regulation in cancer therapeutics.
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Affiliation(s)
- Martin Dalziel
- Oxford Glycobiology Institute, Department of Biochemistry, South Parks Road, Oxford, UK
| | - Stephen A Beers
- Antibody & Vaccine Group, Cancer Sciences Unit, Faculty of Medicine, University of Southampton, Southampton General Hospital, Southampton, UK
| | - Mark S Cragg
- Antibody & Vaccine Group, Cancer Sciences Unit, Faculty of Medicine, University of Southampton, Southampton General Hospital, Southampton, UK
| | - Max Crispin
- Centre for Biological Sciences, University of Southampton, Southampton, UK
- Institute for Life Sciences, Highfield Campus, University of Southampton, Southampton, UK
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24
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The state-of-play and future of antibody therapeutics. Adv Drug Deliv Rev 2017; 122:2-19. [PMID: 27916504 DOI: 10.1016/j.addr.2016.11.004] [Citation(s) in RCA: 221] [Impact Index Per Article: 27.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/05/2016] [Revised: 11/26/2016] [Accepted: 11/28/2016] [Indexed: 12/22/2022]
Abstract
It has been over four decades since the development of monoclonal antibodies (mAbs) using a hybridoma cell line was first reported. Since then more than thirty therapeutic antibodies have been marketed, mostly as oncology, autoimmune and inflammatory therapeutics. While antibodies are very efficient, their cost-effectiveness has always been discussed owing to their high costs, accumulating to more than one billion dollars from preclinical development through to market approval. Because of this, therapeutic antibodies are inaccessible to some patients in both developed and developing countries. The growing interest in biosimilar antibodies as affordable versions of therapeutic antibodies may provide alternative treatment options as well potentially decreasing costs. As certain markets begin to capitalize on this opportunity, regulatory authorities continue to refine the requirements for demonstrating quality, efficacy and safety of biosimilar compared to originator products. In addition to biosimilars, innovations in antibody engineering are providing the opportunity to design biobetter antibodies with improved properties to maximize efficacy. Enhancing effector function, antibody drug conjugates (ADC) or targeting multiple disease pathways via multi-specific antibodies are being explored. The manufacturing process of antibodies is also moving forward with advancements relating to host cell production and purification processes. Studies into the physical and chemical degradation pathways of antibodies are contributing to the design of more stable proteins guided by computational tools. Moreover, the delivery and pharmacokinetics of antibody-based therapeutics are improving as optimized formulations are pursued through the implementation of recent innovations in the field.
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25
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Prior S, Hufton SE, Fox B, Dougall T, Rigsby P, Bristow A. International standards for monoclonal antibodies to support pre- and post-marketing product consistency: Evaluation of a candidate international standard for the bioactivities of rituximab. MAbs 2017; 10:129-142. [PMID: 28985159 PMCID: PMC5836816 DOI: 10.1080/19420862.2017.1386824] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/31/2023] Open
Abstract
The intrinsic complexity and heterogeneity of therapeutic monoclonal antibodies is built into the biosimilarity paradigm where critical quality attributes are controlled in exhaustive comparability studies with the reference medicinal product. The long-term success of biosimilars will depend on reassuring healthcare professionals and patients of consistent product quality, safety and efficacy. With this aim, the World Health Organization has endorsed the need for public bioactivity standards for therapeutic monoclonal antibodies in support of current controls. We have developed a candidate international potency standard for rituximab that was evaluated in a multi-center collaborative study using participants' own qualified Fc-effector function and cell-based binding bioassays. Dose-response curve model parameters were shown to reflect similar behavior amongst rituximab preparations, albeit with some differences in potency. In the absence of a common reference standard, potency estimates were in poor agreement amongst laboratories, but the use of the candidate preparation significantly reduced this variability. Our results suggest that the candidate rituximab standard can support bioassay performance and improve data harmonization, which when implemented will promote consistency of rituximab products over their life-cycles. This data provides the first scientific evidence that a classical standardization exercise allowing traceability of bioassay data to an international standard is also applicable to rituximab. However, we submit that this new type of international standard needs to be used appropriately and its role not to be mistaken with that of the reference medicinal product.
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Affiliation(s)
- Sandra Prior
- a Molecular Immunology Section, Biotherapeutics Division, National Institute for Biological Standards and Control, South Mimms , Potters Bar , Hertfordshire , United Kingdom
| | - Simon E Hufton
- a Molecular Immunology Section, Biotherapeutics Division, National Institute for Biological Standards and Control, South Mimms , Potters Bar , Hertfordshire , United Kingdom
| | - Bernard Fox
- a Molecular Immunology Section, Biotherapeutics Division, National Institute for Biological Standards and Control, South Mimms , Potters Bar , Hertfordshire , United Kingdom
| | - Thomas Dougall
- b Technology Development and Infrastructure Division, National Institute for Biological Standards and Control, South Mimms , Potters Bar , Hertfordshire , United Kingdom
| | - Peter Rigsby
- b Technology Development and Infrastructure Division, National Institute for Biological Standards and Control, South Mimms , Potters Bar , Hertfordshire , United Kingdom
| | - Adrian Bristow
- b Technology Development and Infrastructure Division, National Institute for Biological Standards and Control, South Mimms , Potters Bar , Hertfordshire , United Kingdom
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26
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Li W, Wu Y, Kong D, Yang H, Wang Y, Shao J, Feng Y, Chen W, Ma L, Ying T, Dimitrov DS. One-domain CD4 Fused to Human Anti-CD16 Antibody Domain Mediates Effective Killing of HIV-1-Infected Cells. Sci Rep 2017; 7:9130. [PMID: 28831040 PMCID: PMC5567353 DOI: 10.1038/s41598-017-07966-3] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/30/2017] [Accepted: 07/05/2017] [Indexed: 02/05/2023] Open
Abstract
Bispecific killer cells engagers (BiKEs) which can bind to natural killer (NK) cells through the activating receptor CD16A and guide them to cells expressing the HIV-1 envelope glycoprotein (Env) are a promising new weapon for elimination of infected cells and eradication of the virus. Here we report the design, generation and characterization of BiKEs which consist of CD16A binding human antibody domains fused through a flexible linker to an engineered one-domain soluble human CD4. In presence of cells expressing HIV-1 envelope glycoproteins (Envs), these BiKEs activated specifically CD16A-expressing Jurkat T cells, degranulated NK cells, induced cytokine production and killed Env-expressing cells. They also effectively mediated killing of chronically and acutely HIV-1 infected T cells by human peripheral blood mononuclear cells. The presumed ability of these CD4-based BiKEs to bind all HIV-1 isolates, their small size and fully human origin, combined with high efficacy suggest their potential for HIV-1 eradication.
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Affiliation(s)
- Wei Li
- Protein Interactions Section, Cancer and Inflammation Program, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, Maryland, 21702, USA.
| | - Yanling Wu
- Key Laboratory of Medical Molecular Virology of Ministries of Education and Health, School of Basic Medical Sciences, Fudan University, Shanghai, 200032, China
| | - Desheng Kong
- State Key Laboratory of Infectious Disease Prevention and Control, National Center for AIDS/STD Control and Prevention (NCAIDS), Chinese Center for Disease Control and Prevention, Beijing, China
| | - Hongjia Yang
- Palisades Charter High School, 15777 Bowdoin St, Pacific Palisades, CA, 90272, USA
| | - Yanping Wang
- Protein Interactions Section, Cancer and Inflammation Program, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, Maryland, 21702, USA
| | - Jiping Shao
- Protein Interactions Section, Cancer and Inflammation Program, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, Maryland, 21702, USA
- Hainan Medical University, Haikou City, Hainan Province, 571199, China
| | - Yang Feng
- Protein Interactions Section, Cancer and Inflammation Program, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, Maryland, 21702, USA
| | - Weizao Chen
- Protein Interactions Section, Cancer and Inflammation Program, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, Maryland, 21702, USA
| | - Liying Ma
- State Key Laboratory of Infectious Disease Prevention and Control, National Center for AIDS/STD Control and Prevention (NCAIDS), Chinese Center for Disease Control and Prevention, Beijing, China
| | - Tianlei Ying
- Key Laboratory of Medical Molecular Virology of Ministries of Education and Health, School of Basic Medical Sciences, Fudan University, Shanghai, 200032, China
| | - Dimiter S Dimitrov
- Protein Interactions Section, Cancer and Inflammation Program, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Frederick, Maryland, 21702, USA.
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27
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Järnum S, Runström A, Bockermann R, Winstedt L, Crispin M, Kjellman C. Enzymatic Inactivation of Endogenous IgG by IdeS Enhances Therapeutic Antibody Efficacy. Mol Cancer Ther 2017; 16:1887-1897. [DOI: 10.1158/1535-7163.mct-17-0108] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/01/2017] [Revised: 05/11/2017] [Accepted: 05/19/2017] [Indexed: 11/16/2022]
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28
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Yu X, Marshall MJE, Cragg MS, Crispin M. Improving Antibody-Based Cancer Therapeutics Through Glycan Engineering. BioDrugs 2017; 31:151-166. [DOI: 10.1007/s40259-017-0223-8] [Citation(s) in RCA: 45] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/19/2023]
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29
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Mittermayr S, Lê GN, Clarke C, Millán Martín S, Larkin AM, O’Gorman P, Bones J. Polyclonal Immunoglobulin G N-Glycosylation in the Pathogenesis of Plasma Cell Disorders. J Proteome Res 2016; 16:748-762. [DOI: 10.1021/acs.jproteome.6b00768] [Citation(s) in RCA: 24] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/25/2022]
Affiliation(s)
- Stefan Mittermayr
- NIBRT−The
National Institute for Bioprocessing Research and Training, Foster Avenue, Mount Merrion, Blackrock Co., Dublin A94 X099, Ireland
| | - Giao N. Lê
- NIBRT−The
National Institute for Bioprocessing Research and Training, Foster Avenue, Mount Merrion, Blackrock Co., Dublin A94 X099, Ireland
- Department
of Haematology, Mater Misericordiae University Hospital, Dublin D07 R2WY, Ireland
- National
Institute for Cellular Biotechnology, Dublin City University, Dublin D09 NR58, Ireland
| | - Colin Clarke
- NIBRT−The
National Institute for Bioprocessing Research and Training, Foster Avenue, Mount Merrion, Blackrock Co., Dublin A94 X099, Ireland
| | - Silvia Millán Martín
- NIBRT−The
National Institute for Bioprocessing Research and Training, Foster Avenue, Mount Merrion, Blackrock Co., Dublin A94 X099, Ireland
| | - Anne-Marie Larkin
- National
Institute for Cellular Biotechnology, Dublin City University, Dublin D09 NR58, Ireland
| | - Peter O’Gorman
- Department
of Haematology, Mater Misericordiae University Hospital, Dublin D07 R2WY, Ireland
| | - Jonathan Bones
- NIBRT−The
National Institute for Bioprocessing Research and Training, Foster Avenue, Mount Merrion, Blackrock Co., Dublin A94 X099, Ireland
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30
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Xu Y, Chen B, Zheng S, Wen Y, Xu A, Xu K, Li B, Liu C. IgG silencing induces apoptosis and suppresses proliferation, migration and invasion in LNCaP prostate cancer cells. Cell Mol Biol Lett 2016; 21:27. [PMID: 28536629 PMCID: PMC5415809 DOI: 10.1186/s11658-016-0029-6] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/26/2016] [Accepted: 11/24/2016] [Indexed: 01/10/2023] Open
Abstract
Immunoglobulin G (IgG) has been implicated in the progression of various cancers. This study explored the role of IgG in the proliferation, apoptosis, cell cycle and in vitro invasive properties of LNCaP prostate cancer cells. We used IGHG1 small interfering RNA to silence IgG1 expression in LNCaP cells. The efficacy of IgG1 gene knockdown was confirmed using qPCR and western blotting. The colony formation, proliferation, migration and invasion abilities of LNCaP cells after transfection were assessed using colony-forming, flow cytometry and transwell assays. The expressions of PCNA and caspase-3 proteins in LNCaP cells after transfection were detected with immunofluorescence staining and western blotting. IgG1 silencing significantly decreased the colony formation, survival, cell cycle progression, migration and invasion of LNCaP cells (p < 0.05). IgG1 silencing also reduced the amount of the proliferation marker PCNA and induced formation of the apoptotic marker caspase-3 (p < 0.05). Our results show that IgG1 produced by LNCaP cells confers advantages for tumor cell survival, proliferation, migration and invasion, suggesting that IgG1 is a potential target for prostate cancer treatment.
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Affiliation(s)
- Yawen Xu
- Department of Urology, Zhujiang Hospital, Southern Medical University, No. 253, Industrial Road, Guangzhou, Guangdong China
| | - Binshen Chen
- Department of Urology, Zhujiang Hospital, Southern Medical University, No. 253, Industrial Road, Guangzhou, Guangdong China
| | - Shaobo Zheng
- Department of Urology, Zhujiang Hospital, Southern Medical University, No. 253, Industrial Road, Guangzhou, Guangdong China
| | - Yong Wen
- Department of Urology, Zhujiang Hospital, Southern Medical University, No. 253, Industrial Road, Guangzhou, Guangdong China
| | - Abai Xu
- Department of Urology, Zhujiang Hospital, Southern Medical University, No. 253, Industrial Road, Guangzhou, Guangdong China
| | - Kai Xu
- Department of Urology, Zhujiang Hospital, Southern Medical University, No. 253, Industrial Road, Guangzhou, Guangdong China
| | - Bingkun Li
- Department of Urology, Zhujiang Hospital, Southern Medical University, No. 253, Industrial Road, Guangzhou, Guangdong China
| | - Chunxiao Liu
- Department of Urology, Zhujiang Hospital, Southern Medical University, No. 253, Industrial Road, Guangzhou, Guangdong China
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31
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Calow J, Behrens AJ, Mader S, Bockau U, Struwe WB, Harvey DJ, Cormann KU, Nowaczyk MM, Loser K, Schinor D, Hartmann MWW, Crispin M. Antibody production using a ciliate generates unusual antibody glycoforms displaying enhanced cell-killing activity. MAbs 2016; 8:1498-1511. [PMID: 27594301 PMCID: PMC5098438 DOI: 10.1080/19420862.2016.1228504] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022] Open
Abstract
Antibody glycosylation is a key parameter in the optimization of antibody therapeutics. Here, we describe the production of the anti-cancer monoclonal antibody rituximab in the unicellular ciliate, Tetrahymena thermophila. The resulting antibody demonstrated enhanced antibody-dependent cell-mediated cytotoxicity, which we attribute to unusual N-linked glycosylation. Detailed chromatographic and mass spectrometric analysis revealed afucosylated, oligomannose-type glycans, which, as a whole, displayed isomeric structures that deviate from the typical human counterparts, but whose branches were equivalent to fragments of metabolic intermediates observed in human glycoproteins. From the analysis of deposited crystal structures, we predict that the ciliate glycans adopt protein-carbohydrate interactions with the Fc domain that closely mimic those of native complex-type glycans. In addition, terminal glucose structures were identified that match biosynthetic precursors of human glycosylation. Our results suggest that ciliate-based expression systems offer a route to large-scale production of monoclonal antibodies exhibiting glycosylation that imparts enhanced cell killing activity.
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Affiliation(s)
| | - Anna-Janina Behrens
- b Oxford Glycobiology Institute, Department of Biochemistry, University of Oxford , Oxford , UK
| | | | | | - Weston B Struwe
- b Oxford Glycobiology Institute, Department of Biochemistry, University of Oxford , Oxford , UK
| | - David J Harvey
- b Oxford Glycobiology Institute, Department of Biochemistry, University of Oxford , Oxford , UK
| | - Kai U Cormann
- c Plant Biochemistry, Ruhr University Bochum , Bochum , Germany
| | - Marc M Nowaczyk
- c Plant Biochemistry, Ruhr University Bochum , Bochum , Germany
| | - Karin Loser
- d Department of Dermatology , University of Münster , Münster , Germany
| | - Daniel Schinor
- e Wessling GmbH, Pharmaanalytik Münster , Münster , Germany
| | | | - Max Crispin
- b Oxford Glycobiology Institute, Department of Biochemistry, University of Oxford , Oxford , UK
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32
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Dorokhov YL, Sheshukova EV, Kosobokova EN, Shindyapina AV, Kosorukov VS, Komarova TV. Functional role of carbohydrate residues in human immunoglobulin G and therapeutic monoclonal antibodies. BIOCHEMISTRY (MOSCOW) 2016; 81:835-57. [DOI: 10.1134/s0006297916080058] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/23/2022]
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33
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Mechanisms of disease: The human N-glycome. Biochim Biophys Acta Gen Subj 2016; 1860:1574-82. [DOI: 10.1016/j.bbagen.2015.10.016] [Citation(s) in RCA: 130] [Impact Index Per Article: 14.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/04/2015] [Revised: 10/03/2015] [Accepted: 10/15/2015] [Indexed: 12/20/2022]
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34
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Le NPL, Bowden TA, Struwe WB, Crispin M. Immune recruitment or suppression by glycan engineering of endogenous and therapeutic antibodies. BIOCHIMICA ET BIOPHYSICA ACTA 2016; 1860:1655-68. [PMID: 27105835 PMCID: PMC4922387 DOI: 10.1016/j.bbagen.2016.04.016] [Citation(s) in RCA: 38] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 02/25/2016] [Revised: 04/15/2016] [Accepted: 04/16/2016] [Indexed: 12/20/2022]
Abstract
Human serum IgG contains multiple glycoforms which exhibit a range of binding properties to effector molecules such as cellular Fc receptors. Emerging knowledge of how the Fc glycans contribute to the antibody structure and effector functions has opened new avenues for the exploitation of defined antibody glycoforms in the treatment of diseases. Here, we review the structure and activity of antibody glycoforms and highlight developments in antibody glycoengineering by both the manipulation of the cellular glycosylation machinery and by chemoenzymatic synthesis. We discuss wide ranging applications of antibody glycoengineering in the treatment of cancer, autoimmunity and inflammation. This article is part of a Special Issue entitled "Glycans in personalised medicine" Guest Editor: Professor Gordan Lauc.
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Affiliation(s)
- Ngoc Phuong Lan Le
- Oxford Glycobiology Institute, Department of Biochemistry, University of Oxford, South Parks Road, Oxford OX1 3QU, United Kingdom
| | - Thomas A Bowden
- Division of Structural Biology, University of Oxford, Wellcome Trust Centre for Human Genetics, Roosevelt Drive, Oxford OX3 7BN, United Kingdom
| | - Weston B Struwe
- Oxford Glycobiology Institute, Department of Biochemistry, University of Oxford, South Parks Road, Oxford OX1 3QU, United Kingdom
| | - Max Crispin
- Oxford Glycobiology Institute, Department of Biochemistry, University of Oxford, South Parks Road, Oxford OX1 3QU, United Kingdom.
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Vučković F, Theodoratou E, Thaçi K, Timofeeva M, Vojta A, Štambuk J, Pučić-Baković M, Rudd PM, Đerek L, Servis D, Wennerström A, Farrington SM, Perola M, Aulchenko Y, Dunlop MG, Campbell H, Lauc G. IgG Glycome in Colorectal Cancer. Clin Cancer Res 2016; 22:3078-86. [PMID: 26831718 PMCID: PMC5860729 DOI: 10.1158/1078-0432.ccr-15-1867] [Citation(s) in RCA: 105] [Impact Index Per Article: 11.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/08/2015] [Accepted: 12/31/2015] [Indexed: 11/16/2022]
Abstract
PURPOSE Alternative glycosylation has significant structural and functional consequences on IgG and consequently also on cancer immunosurveillance. Because of technological limitations, the effects of highly heritable individual variations and the differences in the dynamics of changes in IgG glycosylation on colorectal cancer were never investigated before. EXPERIMENTAL DESIGN Using recently developed high-throughput UPLC technology for IgG glycosylation analysis, we analyzed IgG glycome composition in 760 patients with colorectal cancer and 538 matching controls. Effects of surgery were evaluated in 28 patients sampled before and three times after surgery. A predictive model was built using regularized logistic regression and evaluated using a 10-cross validation procedure. Furthermore, IgG glycome composition was analyzed in 39 plasma samples collected before initial diagnosis of colorectal cancer. RESULTS We have found that colorectal cancer associates with decrease in IgG galactosylation, IgG sialylation and increase in core-fucosylation of neutral glycans with concurrent decrease of core-fucosylation of sialylated glycans. Although a model based on age and sex did not show discriminative power (AUC = 0.499), the addition of glycan variables into the model considerably increased the discriminative power of the model (AUC = 0.755). However, none of these differences were significant in the small set of samples collected before the initial diagnosis. CONCLUSIONS Considering the functional relevance of IgG glycosylation for both tumor immunosurveillance and clinical efficacy of therapy with mAbs, individual variation in IgG glycosylation may turn out to be important for prediction of disease course or the choice of therapy, thus warranting further, more detailed studies of IgG glycosylation in colorectal cancer. Clin Cancer Res; 22(12); 3078-86. ©2016 AACR.
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Affiliation(s)
| | - Evropi Theodoratou
- Centre for Population Health Sciences, University of Edinburgh, Edinburgh, United Kingdom. Colon Cancer Genetics Group, Institute of Genetics and Molecular Medicine, University of Edinburgh and Medical Research Council Human Genetics Unit, Edinburgh, United Kingdom
| | - Kujtim Thaçi
- Genos Glycoscience Research Laboratory, Zagreb, Croatia
| | - Maria Timofeeva
- Colon Cancer Genetics Group, Institute of Genetics and Molecular Medicine, University of Edinburgh and Medical Research Council Human Genetics Unit, Edinburgh, United Kingdom
| | | | - Jerko Štambuk
- Genos Glycoscience Research Laboratory, Zagreb, Croatia
| | | | - Pauline M Rudd
- National Institute for Bioprocessing Research and Training, Dublin, Ireland
| | - Lovorka Đerek
- Department of Medical Biochemistry and Laboratory Medicine, Clinical Hospital Merkur, Zagreb, Croatia
| | - Dražen Servis
- Allgemein- und Viszeralchirurgie, St. Anna Krankenhaus, Herne, Germany
| | - Annika Wennerström
- Department of Health, The National Institute for Health and Welfare, Helsinki, Finland
| | - Susan M Farrington
- Colon Cancer Genetics Group, Institute of Genetics and Molecular Medicine, University of Edinburgh and Medical Research Council Human Genetics Unit, Edinburgh, United Kingdom
| | - Markus Perola
- Department of Health, The National Institute for Health and Welfare, Helsinki, Finland
| | | | - Malcolm G Dunlop
- Colon Cancer Genetics Group, Institute of Genetics and Molecular Medicine, University of Edinburgh and Medical Research Council Human Genetics Unit, Edinburgh, United Kingdom
| | - Harry Campbell
- Centre for Population Health Sciences, University of Edinburgh, Edinburgh, United Kingdom. Colon Cancer Genetics Group, Institute of Genetics and Molecular Medicine, University of Edinburgh and Medical Research Council Human Genetics Unit, Edinburgh, United Kingdom
| | - Gordan Lauc
- Genos Glycoscience Research Laboratory, Zagreb, Croatia. Faculty of Pharmacy and Biochemistry, University of Zagreb, Zagreb, Croatia.
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Vučković F, Krištić J, Gudelj I, Teruel M, Keser T, Pezer M, Pučić-Baković M, Štambuk J, Trbojević-Akmačić I, Barrios C, Pavić T, Menni C, Wang Y, Zhou Y, Cui L, Song H, Zeng Q, Guo X, Pons-Estel BA, McKeigue P, Leslie Patrick A, Gornik O, Spector TD, Harjaček M, Alarcon-Riquelme M, Molokhia M, Wang W, Lauc G. Association of systemic lupus erythematosus with decreased immunosuppressive potential of the IgG glycome. Arthritis Rheumatol 2016. [PMID: 26200652 PMCID: PMC4626261 DOI: 10.1002/art.39273] [Citation(s) in RCA: 207] [Impact Index Per Article: 23.0] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
Objective Glycans attached to the Fc portion of IgG are important modulators of IgG effector functions. Interindividual differences in IgG glycome composition are large and they associate strongly with different inflammatory and autoimmune diseases. IKZF1, HLA–DQ2A/B, and BACH2 genetic loci that affect IgG glycome composition show pleiotropy with systemic lupus erythematosus (SLE), indicating a potentially causative role of aberrant IgG glycosylation in SLE. We undertook this large multicenter case–control study to determine whether SLE is associated with altered IgG glycosylation. Methods Using ultra‐performance liquid chromatography analysis of released glycans, we analyzed the composition of the IgG glycome in 261 SLE patients and 247 matched controls of Latin American Mestizo origin (the discovery cohort) and in 2 independent replication cohorts of different ethnicity (108 SLE patients and 193 controls from Trinidad, and 106 SLE patients and 105 controls from China). Results Multiple statistically significant differences in IgG glycome composition were observed between patients and controls. The most significant changes included decreased galactosylation and sialylation of IgG (which regulate proinflammatory and antiinflammatory actions of IgG) as well as decreased core fucose and increased bisecting N‐acetylglucosamine (which affect antibody‐dependent cell‐mediated cytotoxicity). Conclusion The IgG glycome in SLE patients is significantly altered in a way that decreases immunosuppressive action of circulating immunoglobulins. The magnitude of observed changes is associated with the intensity of the disease, indicating that aberrant IgG glycome composition or changes in IgG glycosylation may be an important molecular mechanism in SLE.
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Affiliation(s)
- Frano Vučković
- Genos Ltd., Glycoscience Research Laboratory, Zagreb, Croatia
| | | | - Ivan Gudelj
- Genos Ltd., Glycoscience Research Laboratory, Zagreb, Croatia
| | - Maria Teruel
- Pfizer-University of Granada-Junta de Andalucia Centre for Genomics and Oncological Research (GENYO), Granada, Spain
| | | | - Marija Pezer
- Genos Ltd., Glycoscience Research Laboratory, Zagreb, Croatia
| | | | - Jerko Štambuk
- Genos Ltd., Glycoscience Research Laboratory, Zagreb, Croatia
| | | | - Clara Barrios
- King's College London, London, UK, and Hospital del Mar and Institut Mar d'Investigacions Mediques, Barcelona, Spain
| | | | | | | | - Yong Zhou
- Beijing Tiantan Hospital and Capital Medical University, Beijing, China
| | - Liufu Cui
- Affiliated Kailuan General Hospital of Hebei United University, Tangshan, China
| | - Haicheng Song
- Affiliated Kailuan General Hospital of Hebei United University, Tangshan, China
| | - Qiang Zeng
- International Medical Center and Chinese People's Liberation Army General Hospital, Beijing, China
| | - Xiuhua Guo
- Capital Medical University, Beijing, China
| | | | | | | | | | | | | | - Marta Alarcon-Riquelme
- Pfizer-University of Granada-Junta de Andalucia Centre for Genomics and Oncological Research (GENYO), Granada, Spain, and Oklahoma Medical Research Foundation, Oklahoma City
| | | | - Wei Wang
- Capital Medical University, Beijing, China, and Edith Cowan University, Perth, Western Australia, Australia
| | - Gordan Lauc
- Genos Ltd., Glycoscience Research Laboratory, and University of Zagreb, Zagreb, Croatia
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Felices M, Lenvik TR, Davis ZB, Miller JS, Vallera DA. Generation of BiKEs and TriKEs to Improve NK Cell-Mediated Targeting of Tumor Cells. Methods Mol Biol 2016; 1441:333-46. [PMID: 27177679 PMCID: PMC5823010 DOI: 10.1007/978-1-4939-3684-7_28] [Citation(s) in RCA: 122] [Impact Index Per Article: 13.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/14/2023]
Abstract
Cancer immunotherapies have gained significant momentum over the past decade, particularly with the advent of checkpoint inhibitors and CAR T-cells. While the latter personalized targeted immunotherapy has revolutionized the field, a need for off-the-shelf therapies remains. The ability of NK cells to quickly lyse antibody-coated tumors and potently secrete cytokines without prior priming has made NK cells ideal candidates for antigen-specific immunotherapy. NK cells have been targeted to tumors through two main strategies: mono-specific antibodies and bi/tri-specific antibodies. Mono-specific antibodies drive NK cell antibody-dependent cell-mediated cytotoxicity (ADCC) of tumor cells. Bi/tri-specific antibodies drive re-directed lysis of tumor cells through binding of a tumor antigen and direct binding and crosslinking of the CD16 receptor on NK cells, thus bypassing the need for binding of the Fc portion of mono-specific antibodies. This chapter focuses on the generation of bi- and tri-specific killer engagers (BiKEs and TriKEs) meant to target NK cells to tumors. BiKEs and TriKEs are smaller molecules composed of 2-3 variable portions of antibodies with different specificities, and represent a novel and more versatile strategy compared to traditional bi- and tri-specific antibody platforms.
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Affiliation(s)
- Martin Felices
- Division of Hematology, Oncology, and Transplantation, Department of Medicine, University of Minnesota, Minneapolis, MN, USA
- Masonic Cancer Center, University of Minnesota, Minneapolis, MN, USA
| | - Todd R Lenvik
- Division of Hematology, Oncology, and Transplantation, Department of Medicine, University of Minnesota, Minneapolis, MN, USA
- Masonic Cancer Center, University of Minnesota, Minneapolis, MN, USA
| | - Zachary B Davis
- Division of Hematology, Oncology, and Transplantation, Department of Medicine, University of Minnesota, Minneapolis, MN, USA
- Masonic Cancer Center, University of Minnesota, Minneapolis, MN, USA
| | - Jeffrey S Miller
- Division of Hematology, Oncology, and Transplantation, Department of Medicine, University of Minnesota, Minneapolis, MN, USA.
- Masonic Cancer Center, University of Minnesota, Minneapolis, MN, USA.
| | - Daniel A Vallera
- Masonic Cancer Center, University of Minnesota, Minneapolis, MN, USA
- Department of Therapeutic Radiology-Radiation Oncology, University of Minnesota, Minneapolis, MN, USA
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Turini M, Chames P, Bruhns P, Baty D, Kerfelec B. A FcγRIII-engaging bispecific antibody expands the range of HER2-expressing breast tumors eligible to antibody therapy. Oncotarget 2015; 5:5304-19. [PMID: 24979648 PMCID: PMC4170649 DOI: 10.18632/oncotarget.2093] [Citation(s) in RCA: 34] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022] Open
Abstract
Trastuzumab is established as treatment of HER2high metastatic breast cancers but many limitations impair its efficacy. Here, we report the design of a Fab-like bispecific antibody (HER2bsFab) that displays a moderate affinity for HER2 and a unique, specific and high affinity for FcγRIII. In vitro characterization showed that ADCC was the major mechanism of action of HER2bsFab as no significant HER2-driven effect was observed. HER2bsFab mediated ADCC at picomolar concentration against HER2high, HER2low as well as trastuzumab-refractive cell lines. In vivo HER2bsFab potently inhibited HER2high tumor growth by recruitment of mouse FcγRIII and IV-positive resident effector cells and more importantly, exhibited a net superiority over trastuzumab at inhibiting HER2low tumor growth. Moreover, FcγRIIIA-engagement by HER2bsFab was independent of V/F158 polymorphism and induced a stronger NK cells activation in response to target cell recognition. Thus, taking advantage of its epitope specificity and affinity for HER2 and FcγRIIIA, HER2bsFab exhibits potent anti-tumor activity against HER2low tumors while evading most of trastuzumab Fc-linked limitations thereby potentially enlarging the number of patients eligible for breast cancer immunotherapy.
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Affiliation(s)
- Marc Turini
- INSERM, U1068, CRCM, Marseille, France. Institut Paoli-Calmettes, Marseille, France. Aix-Marseille Université, UM105, Marseille, France. CNRS, UMR7258, CRCM, Marseille, France
| | - Patrick Chames
- INSERM, U1068, CRCM, Marseille, France. Institut Paoli-Calmettes, Marseille, France. Aix-Marseille Université, UM105, Marseille, France. CNRS, UMR7258, CRCM, Marseille, France
| | - Pierre Bruhns
- Département d'Immunologie, Laboratoire Anticorps en Thérapie et Pathologie, Institut Pasteur, Paris, France. INSERM, U760, Paris, France
| | - Daniel Baty
- INSERM, U1068, CRCM, Marseille, France. Institut Paoli-Calmettes, Marseille, France. Aix-Marseille Université, UM105, Marseille, France. CNRS, UMR7258, CRCM, Marseille, France
| | - Brigitte Kerfelec
- INSERM, U1068, CRCM, Marseille, France. Institut Paoli-Calmettes, Marseille, France. Aix-Marseille Université, UM105, Marseille, France. CNRS, UMR7258, CRCM, Marseille, France
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Abstract
BACKGROUND Glycobiology is an underexplored research area in inflammatory bowel disease (IBD), and glycans are relevant to many etiological mechanisms described in IBD. Alterations in N-glycans attached to the immunoglobulin G (IgG) Fc fragment can affect molecular structure and immunological function. Recent genome-wide association studies reveal pleiotropy between IBD and IgG glycosylation. This study aims to explore IgG glycan changes in ulcerative colitis (UC) and Crohn's disease (CD). METHODS IgG glycome composition in patients with UC (n = 507), CD (n = 287), and controls (n = 320) was analyzed by ultra performance liquid chromatography. RESULTS Statistically significant differences in IgG glycome composition between patients with UC or CD, compared with controls, were observed. Both UC and CD were associated with significantly decreased IgG galactosylation (digalactosylation, UC: odds ratio [OR] = 0.71; 95% confidence interval [CI], 0.5-0.9; P = 0.01; CD: OR = 0.41; CI, 0.3-0.6; P = 1.4 × 10) and significant decrease in the proportion of sialylated structures in CD (OR = 0.46, CI, 0.3-0.6, P = 8.4 × 10). Logistic regression models incorporating measured IgG glycan traits were able to distinguish UC and CD from controls (UC: P = 2.13 × 10 and CD: P = 2.20 × 10), with receiver-operator characteristic curves demonstrating better performance of the CD model (area under curve [AUC] = 0.77) over the UC model (AUC = 0.72) (P = 0.026). The ratio of the presence to absence of bisecting GlcNAc in monogalactosylated structures was increased in patients with UC undergoing colectomy compared with no colectomy (FDR-adjusted, P = 0.05). CONCLUSIONS The observed differences indicate significantly increased inflammatory potential of IgG in IBD. Changes in IgG glycosylation may contribute to IBD pathogenesis and could alter monoclonal antibody therapeutic efficacy. IgG glycan profiles have translational potential as IBD biomarkers.
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Schirrmann T, Steinwand M, Wezler X, Ten Haaf A, Tur MK, Barth S. CD30 as a therapeutic target for lymphoma. BioDrugs 2015; 28:181-209. [PMID: 24043362 DOI: 10.1007/s40259-013-0068-8] [Citation(s) in RCA: 27] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022]
Abstract
Hodgkin's lymphoma (HL) and ALK(+) anaplastic large-cell lymphoma (ALCL) have become highly curable due to the success of modern regimens of chemotherapy and radiotherapy. However, up to one-third of the patients experience relapse or do not respond to first-line therapy, and half of them relapse again after secondary therapy with limited options for further treatment. In the last 15 years, monoclonal antibodies (mAbs) directed to surface receptors became a new and valuable therapeutic option in many hematologic malignancies. Due to its restricted expression on normal activated lymphocytes and its high expression on malignant cells, CD30 represents an attractive target molecule for HL and ALCL therapy. However, unconjugated CD30 mAbs have demonstrated a lack of objective clinical responses in patients with recurrent HL. CD30 exhibits complex signaling pathways, and binding of its natural ligand or anti-CD30 mAbs can induce apoptosis but may also promote proliferation and activation depending on the cellular context. Moreover, CD30 rapidly internalizes after crosslinking, which counteracts efficient recruitment of immunologic effectors but also provides the opportunity to transfer cytotoxic payloads coupled to CD30-specific mAbs into the tumor cells. Several tumor targeting approaches have been studied, including radio-immunoconjugates, immunotoxins, immunoRNases, immunokinases, and antibody drug conjugates (ADCs). In 2011, the ADC brentuximab-vedotin, consisting of the CD30-specific chimeric mAb cAC10 and the potent tubulin toxin monomethyl auristatin E, gained regulatory approval as a well tolerated and highly active drug in patients with refractory and relapsed HL and ALCL. SGN-35 is on the way to being incorporated in the standard management of CD30(+) lymphoma with significant therapeutic impact. This review gives a critical overview about anti-CD30 therapies with unconjugated, engineered, and conjugated mAbs and the therapeutic challenges of treatment of CD30(+) lymphoma.
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Affiliation(s)
- Thomas Schirrmann
- Department of Biotechnology, Institute of Biochemistry, Biotechnology and Bioinformatics, Technische Universität Braunschweig, Spielmannstr. 7, 38106, Braunschweig, Germany,
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Niwa R, Satoh M. The Current Status and Prospects of Antibody Engineering for Therapeutic Use: Focus on Glycoengineering Technology. J Pharm Sci 2015; 104:930-41. [DOI: 10.1002/jps.24316] [Citation(s) in RCA: 48] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/15/2014] [Revised: 11/09/2014] [Accepted: 12/02/2014] [Indexed: 12/31/2022]
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Deyev SM, Lebedenko EN, Petrovskaya LE, Dolgikh DA, Gabibov AG, Kirpichnikov MP. Man-made antibodies and immunoconjugates with desired properties: function optimization using structural engineering. RUSSIAN CHEMICAL REVIEWS 2015. [DOI: 10.1070/rcr4459] [Citation(s) in RCA: 49] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/08/2022]
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Abstract
The success of antibody therapy in cancer is consistent with the ability of these molecules to activate immune responses against tumors. Experience in clinical applications, antibody design, and advancement in technology have enabled antibodies to be engineered with enhanced efficacy against cancer cells. This allows re-evaluation of current antibody approaches dominated by antibodies of the IgG class with a new light. Antibodies of the IgE class play a central role in allergic reactions and have many properties that may be advantageous for cancer therapy. IgE-based active and passive immunotherapeutic approaches have been shown to be effective in both in vitro and in vivo models of cancer, suggesting the potential use of these approaches in humans. Further studies on the anticancer efficacy and safety profile of these IgE-based approaches are warranted in preparation for translation toward clinical application.
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Affiliation(s)
- Lai Sum Leoh
- Division of Surgical Oncology, Department of Surgery, David Geffen School of Medicine, University of California, Los Angeles, 10833 Le Conte Avenue, CHS 54-140, Box 951782, Los Angeles, CA 90095-1782, USA
| | - Tracy R. Daniels-Wells
- Division of Surgical Oncology, Department of Surgery, David Geffen School of Medicine, University of California, Los Angeles, 10833 Le Conte Avenue, CHS 54-140, Box 951782, Los Angeles, CA 90095-1782, USA
| | - Manuel L. Penichet
- Division of Surgical Oncology, Department of Surgery, David Geffen School of Medicine, University of California, Los Angeles, 10833 Le Conte Avenue, CHS 54-140, Box 951782, Los Angeles, CA 90095-1782, USA. Department of Microbiology, Immunology, and Molecular Genetics, University of California, 609 Charles E. Young Dr. East, 1602 Molecular Science Building, Los Angeles, CA 90095, USA. The Jonsson Comprehensive Cancer Center, University of California, 10833 Le Conte Ave, 8-684 Factor Building, Box 951781, Los Angeles, CA 90095, USA. The Molecular Biology Institute, University of California, 611 Charles E. Young Dr., Los Angeles, CA 90095, USA
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Theodoratou E, Campbell H, Ventham NT, Kolarich D, Pučić-Baković M, Zoldoš V, Fernandes D, Pemberton IK, Rudan I, Kennedy NA, Wuhrer M, Nimmo E, Annese V, McGovern DPB, Satsangi J, Lauc G. The role of glycosylation in IBD. Nat Rev Gastroenterol Hepatol 2014; 11:588-600. [PMID: 24912389 DOI: 10.1038/nrgastro.2014.78] [Citation(s) in RCA: 121] [Impact Index Per Article: 11.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/08/2023]
Abstract
A number of genetic and immunological studies give impetus for investigating the role of glycosylation in IBD. Experimental mouse models have helped to delineate the role of glycosylation in intestinal mucins and to explore the putative pathogenic role of glycosylation in colitis. These experiments have been extended to human studies investigating the glycosylation patterns of intestinal mucins as well as levels of glycans of serum glycoproteins and expression of glycan receptors. These early human studies have generated interesting hypotheses regarding the pathogenic role of glycans in IBD, but have generally been restricted to fairly small underpowered studies. Decreased glycosylation has been observed in the intestinal mucus of patients with IBD, suggesting that a defective inner mucus layer might lead to increased bacterial contact with the epithelium, potentially triggering inflammation. In sera, decreased galactosylation of IgG has been suggested as a diagnostic marker for IBD. Advances in glycoprofiling technology make it technically feasible and affordable to perform high-throughput glycan pattern analyses and to build on previous work investigating a much wider range of glycan parameters in large numbers of patients.
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Affiliation(s)
- Evropi Theodoratou
- Centre for Population Health Sciences, University of Edinburgh, Teviot Place, EH8 9AG, Edinburgh, UK
| | - Harry Campbell
- Centre for Population Health Sciences, University of Edinburgh, Teviot Place, EH8 9AG, Edinburgh, UK
| | - Nicholas T Ventham
- Centre for Molecular Medicine, University of Edinburgh, Western General Hospital, Crewe Road, Edinburgh EH4 2XU, Edinburgh, UK
| | - Daniel Kolarich
- Department of Biomolecular Systems, Max Planck Institute of Colloids and Interfaces, Am Mühlenberg 1 OT Golm, 14476, Potsdam, Germany
| | | | - Vlatka Zoldoš
- University of Zagreb, Faculty of Science, Horvatovac 102a, 10000 Zagreb, Croatia
| | | | - Iain K Pemberton
- IP Research Consulting SAS, 34 Rue Carnot, 93160 Noisy-le-Grand, Paris, France
| | - Igor Rudan
- Centre for Population Health Sciences, University of Edinburgh, Teviot Place, EH8 9AG, Edinburgh, UK
| | - Nicholas A Kennedy
- Centre for Molecular Medicine, University of Edinburgh, Western General Hospital, Crewe Road, Edinburgh EH4 2XU, Edinburgh, UK
| | - Manfred Wuhrer
- Center for Proteomics and Metabolomics, Leiden University Medical Center, Albinusdreef 2, 2333 ZA Leiden, Netherlands
| | - Elaine Nimmo
- Centre for Molecular Medicine, University of Edinburgh, Western General Hospital, Crewe Road, Edinburgh EH4 2XU, Edinburgh, UK
| | - Vito Annese
- Department of Medical and Surgical Specialities, Division of Gastroenterology, AOU Careggi University Hospital, Largo Brambilla 13, 50139 Florence, Italy
| | - Dermot P B McGovern
- F.Widjaja Family Foundation Inflammatory Bowel and Immunobiology Research Institute, Cedars-Sinai Medical Center, 8700 Beverly Blvd., Suite D4063, Los Angeles, CA 90048, USA
| | - Jack Satsangi
- Centre for Molecular Medicine, University of Edinburgh, Western General Hospital, Crewe Road, Edinburgh EH4 2XU, Edinburgh, UK
| | - Gordan Lauc
- Department of Biochemistry and Molecular Biology, University of Zagreb Faculty of Pharmacy and Biochemistry, Trg maršala Tita 14, 10000 Zagreb, Croatia
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Targeted therapy for HM1.24 (CD317) on multiple myeloma cells. BIOMED RESEARCH INTERNATIONAL 2014; 2014:965384. [PMID: 25143955 PMCID: PMC4124849 DOI: 10.1155/2014/965384] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 03/15/2014] [Accepted: 07/02/2014] [Indexed: 11/18/2022]
Abstract
Multiple myeloma (MM) still remains an incurable disease, at least because of the existence of cell-adhesion mediated drug-resistant MM cells and/or continuous recruitment of presumed MM cancer stem cell-like cells (CSCs). As a new alternative treatment modality, immunological approaches using monoclonal antibodies (mAbs) and/or cytotoxic T lymphocytes (CTLs) are now attracting much attention as a novel strategy attacking MM cells. We have identified that HM1.24 [also known as bone marrow stromal cell antigen 2 (BST2) or CD317] is overexpressed on not only mature MM cells but also MM CSCs. We then have developed a humanized mAb to HM1.24 and defucosylated version of the mAb to adapt to clinical practice. Moreover, we have successfully induced HM1.24-specific CTLs against MM cells. The combination of these innovative therapeutic modalities may likely exert an anti-MM activity by evading the drug resistance mechanism and eliminating presumed CSCs in MM.
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Anikeeva N, Steblyanko M, Fayngerts S, Kopylova N, Marshall DJ, Powers GD, Sato T, Campbell KS, Sykulev Y. Integrin receptors on tumor cells facilitate NK cell-mediated antibody-dependent cytotoxicity. Eur J Immunol 2014; 44:2331-9. [PMID: 24810893 DOI: 10.1002/eji.201344179] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/16/2013] [Revised: 04/03/2014] [Accepted: 05/06/2014] [Indexed: 12/21/2022]
Abstract
NK cells that mediate ADCC play an important role in tumor-specific immunity. We have examined factors limiting specific lysis of tumor cells by CD16.NK-92 cells induced by CNTO 95LF antibodies recognizing αV integrins that are overexpressed on many tumor cells. Although all tested tumor cells were killed by CD16.NK-92 effectors in the presence of the antibodies, the killing of target cells with a low level of ICAM-1 expression revealed a dramatic decrease in their specific lysis at high antibody concentration, revealing a dose limiting effect. A similar effect was also observed with primary human NK cells. The effect was erased after IFN-γ treatment of tumor cells resulting in upregulation of ICAM-1. Furthermore, killing of the same tumor cells induced by Herceptin antibody was significantly impaired in the presence of CNTO 95Ala-Ala antibody variant that blocks αV integrins but is incapable of binding to CD16. These data suggest that αV integrins on tumor cells could compensate for the loss of ICAM-1 molecules, thereby facilitating ADCC by NK cells. Thus, NK cells could exercise cytolytic activity against ICAM-1 deficient tumor cells in the absence of proinflammatory cytokines, emphasizing the importance of NK cells in tumor-specific immunity at early stages of cancer.
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Affiliation(s)
- Nadia Anikeeva
- Department of Microbiology and Immunology, Kimmel Cancer Center, Thomas Jefferson University, Philadelphia, PA, USA
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Wang C, Thudium KB, Han M, Wang XT, Huang H, Feingersh D, Garcia C, Wu Y, Kuhne M, Srinivasan M, Singh S, Wong S, Garner N, Leblanc H, Bunch RT, Blanset D, Selby MJ, Korman AJ. In vitro characterization of the anti-PD-1 antibody nivolumab, BMS-936558, and in vivo toxicology in non-human primates. Cancer Immunol Res 2014; 2:846-56. [PMID: 24872026 DOI: 10.1158/2326-6066.cir-14-0040] [Citation(s) in RCA: 478] [Impact Index Per Article: 43.5] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
The programmed death-1 (PD-1) receptor serves as an immunologic checkpoint, limiting bystander tissue damage and preventing the development of autoimmunity during inflammatory responses. PD-1 is expressed by activated T cells and downmodulates T-cell effector functions upon binding to its ligands, PD-L1 and PD-L2, on antigen-presenting cells. In patients with cancer, the expression of PD-1 on tumor-infiltrating lymphocytes and its interaction with the ligands on tumor and immune cells in the tumor microenvironment undermine antitumor immunity and support its rationale for PD-1 blockade in cancer immunotherapy. This report details the development and characterization of nivolumab, a fully human IgG4 (S228P) anti-PD-1 receptor-blocking monoclonal antibody. Nivolumab binds to PD-1 with high affinity and specificity, and effectively inhibits the interaction between PD-1 and its ligands. In vitro assays demonstrated the ability of nivolumab to potently enhance T-cell responses and cytokine production in the mixed lymphocyte reaction and superantigen or cytomegalovirus stimulation assays. No in vitro antibody-dependent cell-mediated or complement-dependent cytotoxicity was observed with the use of nivolumab and activated T cells as targets. Nivolumab treatment did not induce adverse immune-related events when given to cynomolgus macaques at high concentrations, independent of circulating anti-nivolumab antibodies where observed. These data provide a comprehensive preclinical characterization of nivolumab, for which antitumor activity and safety have been demonstrated in human clinical trials in various solid tumors.
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Affiliation(s)
- Changyu Wang
- Biologics Discovery California, Bristol-Myers Squibb Company, Redwood City, California
| | - Kent B Thudium
- Biologics Discovery California, Bristol-Myers Squibb Company, Redwood City, California
| | - Minhua Han
- Biologics Discovery California, Bristol-Myers Squibb Company, Redwood City, California
| | - Xi-Tao Wang
- Biologics Discovery California, Bristol-Myers Squibb Company, Redwood City, California
| | - Haichun Huang
- Biologics Discovery California, Bristol-Myers Squibb Company, Redwood City, California
| | - Diane Feingersh
- Biologics Discovery California, Bristol-Myers Squibb Company, Redwood City, California
| | - Candy Garcia
- Biologics Discovery California, Bristol-Myers Squibb Company, Redwood City, California
| | - Yi Wu
- Biologics Discovery California, Bristol-Myers Squibb Company, Redwood City, California
| | - Michelle Kuhne
- Biologics Discovery California, Bristol-Myers Squibb Company, Redwood City, California
| | - Mohan Srinivasan
- Biologics Discovery California, Bristol-Myers Squibb Company, Redwood City, California
| | - Sujata Singh
- Biologics Discovery California, Bristol-Myers Squibb Company, Redwood City, California
| | - Susan Wong
- Biologics Discovery California, Bristol-Myers Squibb Company, Redwood City, California
| | - Neysa Garner
- Biologics Discovery California, Bristol-Myers Squibb Company, Redwood City, California
| | - Heidi Leblanc
- Biologics Discovery California, Bristol-Myers Squibb Company, Redwood City, California
| | - R Todd Bunch
- Bristol-Myers Squibb Company, Mount Vernon, Indiana; and
| | | | - Mark J Selby
- Biologics Discovery California, Bristol-Myers Squibb Company, Redwood City, California
| | - Alan J Korman
- Biologics Discovery California, Bristol-Myers Squibb Company, Redwood City, California;
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Lauc G, Krištić J, Zoldoš V. Glycans - the third revolution in evolution. Front Genet 2014; 5:145. [PMID: 24904645 PMCID: PMC4033155 DOI: 10.3389/fgene.2014.00145] [Citation(s) in RCA: 45] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/06/2014] [Accepted: 05/04/2014] [Indexed: 11/13/2022] Open
Abstract
The development and maintenance of a complex organism composed of trillions of cells is an extremely complex task. At the molecular level every process requires a specific molecular structures to perform it, thus it is difficult to imagine how less than tenfold increase in the number of genes between simple bacteria and higher eukaryotes enabled this quantum leap in complexity. In this perspective article we present the hypothesis that the invention of glycans was the third revolution in evolution (the appearance of nucleic acids and proteins being the first two), which enabled the creation of novel molecular entities that do not require a direct genetic template. Contrary to proteins and nucleic acids, which are made from a direct DNA template, glycans are product of a complex biosynthetic pathway affected by hundreds of genetic and environmental factors. Therefore glycans enable adaptive response to environmental changes and, unlike other epiproteomic modifications, which act as off/on switches, glycosylation significantly contributes to protein structure and enables novel functions. The importance of glycosylation is evident from the fact that nearly all proteins invented after the appearance of multicellular life are composed of both polypeptide and glycan parts.
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Affiliation(s)
- Gordan Lauc
- Department of Biochemistry and Molecular Biology, Faculty of Pharmacy and Biochemistry, University of Zagreb Zagreb, Croatia ; Genos Glycoscience Zagreb, Croatia
| | | | - Vlatka Zoldoš
- Department of Molecular Biology, Faculty of Science, University of Zagreb Zagreb, Croatia
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Costa AR, Rodrigues ME, Henriques M, Oliveira R, Azeredo J. Glycosylation: impact, control and improvement during therapeutic protein production. Crit Rev Biotechnol 2013; 34:281-99. [PMID: 23919242 DOI: 10.3109/07388551.2013.793649] [Citation(s) in RCA: 110] [Impact Index Per Article: 9.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022]
Abstract
The emergence of the biopharmaceutical industry represented a major revolution for modern medicine, through the development of recombinant therapeutic proteins that brought new hope for many patients with previously untreatable diseases. There is a ever-growing demand for these therapeutics that forces a constant technological evolution to increase product yields while simultaneously reducing costs. However, the process changes made for this purpose may also affect the quality of the product, a factor that was initially overlooked but which is now a major focus of concern. Of the many properties determining product quality, glycosylation is regarded as one of the most important, influencing, for example, the biological activity, serum half-life and immunogenicity of the protein. Consequently, monitoring and control of glycosylation is now critical in biopharmaceutical manufacturing and a requirement of regulatory agencies. A rapid evolution is being observed in this context, concerning the influence of glycosylation in the efficacy of different therapeutic proteins, the impact on glycosylation of a diversity of parameters/processes involved in therapeutic protein production, the analytical methodologies employed for glycosylation monitoring and control, as well as strategies that are being explored to use this property to improve therapeutic protein efficacy (glycoengineering). This work reviews the main findings on these subjects, providing an up-to-date source of information to support further studies.
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Affiliation(s)
- Ana Rita Costa
- IBB - Institute for Biotechnology and Bioengineering, Centre of Biological Engineering, University of Minho, Campus de Gualtar , Braga , Portugal
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50
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Kellner C, Derer S, Valerius T, Peipp M. Boosting ADCC and CDC activity by Fc engineering and evaluation of antibody effector functions. Methods 2013; 65:105-13. [PMID: 23851282 DOI: 10.1016/j.ymeth.2013.06.036] [Citation(s) in RCA: 53] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/03/2013] [Revised: 06/26/2013] [Accepted: 06/27/2013] [Indexed: 01/18/2023] Open
Abstract
In recent years, therapy with monoclonal antibodies has become standard of care in various clinical applications. Despite obvious clinical activity, not all patients respond and benefit from this generally well tolerated treatment option. Therefore, rational optimization of antibody therapy represents a major area of interest in translational research. Animal models and clinical data suggested important roles of Fc-mediated effector mechanisms such as antibody dependent cell-mediated cytotoxicity (ADCC) or complement dependent cytotoxicity (CDC) in antibody therapy. These novel insights into the mechanisms of action mediated by monoclonal antibodies inspired the development of different engineering approaches to enhance/optimize antibodies' effector functions. Fc-engineering approaches by altering the Fc-bound glycosylation profile or by exchanging amino acids in the protein backbone have been intensively studied. Here, advanced and emerging technologies in Fc-engineering resulting in altered ADCC and CDC activity are summarized and experimental strategies to evaluate antibodies' effector functions are discussed.
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Affiliation(s)
- Christian Kellner
- Division of Stem Cell Transplantation and Immunotherapy, 2nd Department of Medicine, Christian-Albrechts-University Kiel, Germany
| | - Stefanie Derer
- Division of Stem Cell Transplantation and Immunotherapy, 2nd Department of Medicine, Christian-Albrechts-University Kiel, Germany
| | - Thomas Valerius
- Division of Stem Cell Transplantation and Immunotherapy, 2nd Department of Medicine, Christian-Albrechts-University Kiel, Germany
| | - Matthias Peipp
- Division of Stem Cell Transplantation and Immunotherapy, 2nd Department of Medicine, Christian-Albrechts-University Kiel, Germany.
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