Jan G J van de Winkel

Publications (111)689.73 Total impact

• Article: Efficient generation of stable bispecific IgG1 by controlled Fab-arm exchange.

  Aran F Labrijn, Joyce I Meesters, Bart E C G de Goeij, Ewald T J van den Bremer, Joost Neijssen, Muriel D van Kampen, Kristin Strumane, Sandra Verploegen, Amitava Kundu, Michael J Gramer, Patrick H C van Berkel, Jan G J van de Winkel, Janine Schuurman, Paul W H I Parren
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  ABSTRACT: The promise of bispecific antibodies (bsAbs) to yield more effective therapeutics is well recognized; however, the generation of bsAbs in a practical and cost-effective manner has been a formidable challenge. Here we present a technology for the efficient generation of bsAbs with normal IgG structures that is amenable to both antibody drug discovery and development. The process involves separate expression of two parental antibodies, each containing single matched point mutations in the CH3 domains. The parental antibodies are mixed and subjected to controlled reducing conditions in vitro that separate the antibodies into HL half-molecules and allow reassembly and reoxidation to form highly pure bsAbs. The technology is compatible with standard large-scale antibody manufacturing and ensures bsAbs with Fc-mediated effector functions and in vivo stability typical of IgG1 antibodies. Proof-of-concept studies with HER2×CD3 (T-cell recruitment) and HER2×HER2 (dual epitope targeting) bsAbs demonstrate superior in vivo activity compared with parental antibody pairs.
  Proceedings of the National Academy of Sciences 03/2013; · 9.68 Impact Factor
• Article: Mimicking an Induced Self Phenotype by Coating Lymphomas with the NKp30 Ligand B7-H6 Promotes NK Cell Cytotoxicity.

  Christian Kellner, Tina Maurer, Daniela Hallack, Roland Repp, Jan G J van de Winkel, Paul W H I Parren, Thomas Valerius, Andreas Humpe, Martin Gramatzki, Matthias Peipp
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  ABSTRACT: Induced self expression of the NKp30 ligand B7-H6 facilitates NK cell-mediated elimination of stressed cells. A fusion protein consisting of the ectodomain of B7-H6 and the CD20 single-chain fragment variable 7D8 was generated to mimic an induced self phenotype required for NK cell-mediated target cell elimination. B7-H6:7D8 had bifunctional properties as reflected by its ability to simultaneously bind to the CD20 Ag and to the NKp30 receptor. B7-H6:7D8 bound by CD20(+) lymphoma cells activated human NK cells and triggered degranulation. Consequently, the immunoligand B7-H6:7D8 induced killing of lymphoma-derived cell lines as well as fresh tumor cells from chronic lymphocytic leukemia or lymphoma patients. B7-H6:7D8 was active at nanomolar concentrations in a strictly Ag-specific manner and required interaction with both CD20 and NKp30. Remarkably, NK cell cytotoxicity was further augmented by concomitant activation of Fcγ receptor IIIa or NK group 2 member D. Thus, B7-H6:7D8 acted synergistically with the CD20 Ab rituximab and the immunoligand ULBP2:7D8, which was similarly designed as B7-H6:7D8 but engaging the NK group 2 member D receptor. In conclusion, to our knowledge, B7-H6:7D8 represents the first Ab-based molecule stimulating NKp30-mediated NK cell cytotoxicity for therapeutic purposes and provides proof of concept that Ag-specific NKp30 engagement may represent an innovative strategy to enhance antitumoral NK cell cytotoxicity.
  The Journal of Immunology 10/2012; · 5.79 Impact Factor
• Source

  Available from: Jeanette H W Leusen

  Article: Patients with unstable angina pectoris show an increased frequency of the Fc gamma RIIa R131 allele.

  Dorette Raaz-Schrauder, Arif B Ekici, Luis E Munoz, Lutz Klinghammer, Reinhard E Voll, Jeanette H W Leusen, Jan G J van de Winkel, André Reis, Georg Schett, Christoph D Garlichs, Martin Herrmann
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  ABSTRACT: Patients with Systemic Lupus Erythematosus (SLE) carry an increased risk for the development of coronary artery disease (CAD). The R131 allele of the Fc gamma receptor IIa (FcγRIIa) is associated with SLE incidence and disease severity but also with CAD. Compared to stable angina pectoris (SAP) the unstable angina (UAP), as a manifestation of destabilizing CAD, is associated with increased risk of persistent instability, myocardial infarction, and death. Identification of clinically relevant determinants for unstable angina promises reduction of UAP-associated mortality in patients with SLE. We conducted a clinical study among 553 consecutive patients with stable angina pectoris (n = 330) and unstable angina pectoris (n = 223). All patients were genotyped for a frequent functional variant at position 131 of the mature FcγRIIa. UAP, but not SAP was significantly associated with the R/R131 genotype (P < 0.001). In troponin-negative patients with angina carrying the R/R131 genotype the odds ratio for suffering from UAP was 4.02 (95% confidence interval, 2.52-6.41) compared to those with non-R/R131 genotypes. In a multivariable analysis, the R/R131 genotype independently predicted the risk for development of UAP in a model adjusted for classical atherogenic risk factors. Our data imply that risk stratification of SLE- and other high risk patients with troponin-negative angina could be significantly improved by FcγRIIa genotyping.
  Autoimmunity 05/2012; 45(7):556-64. · 2.47 Impact Factor
• Article: Enhanced FcαRI-mediated neutrophil migration towards tumour colonies in the presence of endothelial cells.

  Marielle A Otten, Jantine E Bakema, Cornelis W Tuk, Martin J Glennie, Alison L Tutt, Robert H J Beelen, Jan G J van de Winkel, Marjolein van Egmond
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  ABSTRACT: Neutrophils potently kill tumour cells in the presence of anti-tumour antibodies in vitro. However, for in vivo targeting, the neutrophils need to extravasate from the circulation by passing through endothelial barriers. To study neutrophil migration in the presence of endothelial cells in vitro, we established a three-dimensional collagen culture in which SK-BR-3 tumour colonies were grown in the presence or absence of an endothelial barrier. We demonstrated that - in contrast to targeting FcγR on neutrophils with mAbs - targeting the immunoglobulin A Fc receptor (FcαRI) instead triggered neutrophil migration and degranulation leading to tumour destruction, which coincided with release of the pro-inflammatory cytokines interleukin (IL)-1β and tumour necrosis factor (TNF)-α. Interestingly, neutrophil migration was enhanced in the presence of endothelial cells, which coincided with production of significant levels of the neutrophil chemokine IL-8. This supports the idea that stimulation of neutrophil FcαRI, but not FcγR, initiates cross-talk between neutrophils and endothelial cells, leading to enhanced neutrophil migration towards tumour colonies and subsequent tumour killing.
  European Journal of Immunology 04/2012; 42(7):1815-21. · 5.10 Impact Factor
• Article: Retraction for Lammerts van Bueren et al. The antibody zalutumumab inhibits epidermal growth factor receptor signaling by limiting intra- and intermolecular flexibility.

  Jeroen J Lammerts van Bueren, Wim K Bleeker, Annika Brännström, Magnus Jansson, Matthias Peipp, Tanja Schneider-Merck, Thomas Valerius, Jan G J van de Winkel, Paul W H I Parren
  Proceedings of the National Academy of Sciences 04/2012; 109(14):5548. · 9.68 Impact Factor
• Article: Quantitative analysis of the interaction strength and dynamics of human IgG4 half molecules by native mass spectrometry.

  Rebecca J Rose, Aran F Labrijn, Ewald T J van den Bremer, Stefan Loverix, Ignace Lasters, Patrick H C van Berkel, Jan G J van de Winkel, Janine Schuurman, Paul W H I Parren, Albert J R Heck
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  ABSTRACT: Native mass spectrometry (MS) is a powerful technique for studying noncovalent protein-protein interactions. Here, native MS was employed to examine the noncovalent interactions involved in homodimerization of antibody half molecules (HL) in hinge-deleted human IgG4 (IgG4Δhinge). By analyzing the concentration dependence of the relative distribution of monomer HL and dimer (HL)(2) species, the apparent dissociation constant (K(D)) for this interaction was determined. In combination with site-directed mutagenesis, the relative contributions of residues at the CH3-CH3 interface to this interaction could be characterized and corresponding K(D) values quantified over a range of 10(-10)-10(-4) M. The critical importance of this noncovalent interaction in maintaining the intact dimeric structure was also proven for the full-length IgG4 backbone. Using time-resolved MS, the kinetics of the interaction could be measured, reflecting the dynamics of IgG4 HL exchange. Hence, native MS has provided a quantitative view of local structural features that define biological properties of IgG4.
  Structure 09/2011; 19(9):1274-82. · 6.35 Impact Factor
• Source

  Available from: Jeanette H W Leusen

  Article: The in vivo mechanism of action of CD20 monoclonal antibodies depends on local tumor burden.

  Peter Boross, J H Marco Jansen, Simone de Haij, Frank J Beurskens, Cees E van der Poel, Lisette Bevaart, Maaike Nederend, Josée Golay, Jan G J van de Winkel, Paul W H I Parren, Jeanette H W Leusen
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  ABSTRACT: CD20 monoclonal antibodies are widely used in clinical practice. Antibody-dependent cellular cytotoxicity, complement-dependent cytotoxicity and direct cell death have been suggested to be important effector functions for CD20 antibodies. However, their specific contributions to the in vivo mechanism of action of CD20 immunotherapy have not been well defined. Here we studied the in vivo mechanism of action of type I (rituximab and ofatumumab) and type II (HuMab-11B8) CD20 antibodies in a peritoneal, syngeneic, mouse model with EL4-CD20 cells using low and high tumor burden. Interestingly, we observed striking differences in the in vivo mechanism of action of CD20 antibodies dependent on tumor load. In conditions of low tumor burden, complement was sufficient for tumor killing both for type I and type II CD20 antibodies. In contrast, in conditions of high tumor burden, activating FcγR (specifically FcγRIII), active complement and complement receptor 3 were all essential for tumor killing. Our data suggest that complement-enhanced antibody-dependent cellular cytotoxicity may critically affect tumor killing by CD20 antibodies in vivo. The type II CD20 antibody 11B8, which is a poor inducer of complement activation, was ineffective against high tumor burden. Tumor burden affects the in vivo mechanism of action of CD20 antibodies. Low tumor load can be eliminated by complement alone, whereas elimination of high tumor load requires multiple effector mechanisms.
  Haematologica 08/2011; 96(12):1822-30. · 6.42 Impact Factor
• Article: Loss of CD20 and bound CD20 antibody from opsonized B cells occurs more rapidly because of trogocytosis mediated by Fc receptor-expressing effector cells than direct internalization by the B cells.

  Paul V Beum, Elizabeth M Peek, Margaret A Lindorfer, Frank J Beurskens, Patrick J Engelberts, Paul W H I Parren, Jan G J van de Winkel, Ronald P Taylor
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  ABSTRACT: We previously reported that 1 h after infusion of CD20 mAb rituximab in patients with chronic lymphocytic leukemia (CLL), >80% of CD20 was removed from circulating B cells, and we replicated this finding, based on in vitro models. This reaction occurs via an endocytic process called shaving/trogocytosis, mediated by FcγR on acceptor cells including monocytes/macrophages, which remove and internalize rituximab-CD20 immune complexes from B cells. Beers et al. reported that CD20 mAb-induced antigenic modulation occurs as a result of internalization of B cell-bound mAb-CD20 complexes by the B cells themselves, with internalization of ∼40% observed after 2 h at 37°C. These findings raise fundamental questions regarding the relative importance of shaving versus internalization in promoting CD20 loss and have substantial implications for the design of mAb-based cancer therapies. Therefore, we performed direct comparisons, based on flow cytometry, to determine the relative rates and extent of shaving versus internalization. B cells, from cell lines, from patients with CLL, and from normal donors, were opsonized with CD20 mAbs rituximab or ofatumumab and incubated for varying times and then reacted with acceptor THP-1 monocytes to promote shaving. We find that shaving induces considerably greater loss of CD20 and bound mAb from opsonized B cells in much shorter time periods (75-90% in <45 min) than is observed for internalization. Both shaving/trogocytosis and internalization could contribute to CD20 loss when CLL patients receive rituximab therapy, but shaving should occur more rapidly and is most likely to be the key mechanism of CD20 loss.
  The Journal of Immunology 08/2011; 187(6):3438-47. · 5.79 Impact Factor
• Article: Species-specific determinants in the IgG CH3 domain enable Fab-arm exchange by affecting the noncovalent CH3-CH3 interaction strength.

  Aran F Labrijn, Theo Rispens, Joyce Meesters, Rebecca J Rose, Tamara H den Bleker, Stefan Loverix, Ewald T J van den Bremer, Joost Neijssen, Tom Vink, Ignace Lasters, Rob C Aalberse, Albert J R Heck, Jan G J van de Winkel, Janine Schuurman, Paul W H I Parren
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  ABSTRACT: A distinctive feature of human IgG4 is its ability to recombine half molecules (H chain and attached L chain) through a dynamic process termed Fab-arm exchange, which results in bispecific Abs. It is becoming evident that the process of Fab-arm exchange is conserved in several mammalian species, and thereby represents a mechanism that impacts humoral immunity more generally than previously thought. In humans, Fab-arm exchange has been attributed to the IgG4 core-hinge sequence (226-CPSCP-230) in combination with unknown determinants in the third constant H chain domain (CH3). In this study, we investigated the role of the CH3 domain in the mechanism of Fab-arm exchange, and thus identified amino acid position 409 as the critical CH3 determinant in human IgG, with R409 resulting in exchange and K409 resulting in stable IgG. Interestingly, studies with IgG from various species showed that Fab-arm exchange could not be assigned to a common CH3 domain amino acid motif. Accordingly, in rhesus monkeys (Macaca mulatta), aa 405 was identified as the CH3 determinant responsible (in combination with 226-CPACP-230). Using native mass spectrometry, we demonstrated that the ability to exchange Fab-arms correlated with the CH3-CH3 dissociation constant. Species-specific adaptations in the CH3 domain thus enable Fab-arm exchange by affecting the inter-CH3 domain interaction strength. The redistribution of Ag-binding domains between molecules may constitute a general immunological and evolutionary advantage. The current insights impact our view of humoral immunity and should furthermore be considered in the design and evaluation of Ab-based studies and therapeutics.
  The Journal of Immunology 08/2011; 187(6):3238-46. · 5.79 Impact Factor
• Article: Epidermal growth factor receptor (EGFR) antibody-induced antibody-dependent cellular cytotoxicity plays a prominent role in inhibiting tumorigenesis, even of tumor cells insensitive to EGFR signaling inhibition.

  Marije B Overdijk, Sandra Verploegen, Jeroen H van den Brakel, Jeroen J Lammerts van Bueren, Tom Vink, Jan G J van de Winkel, Paul W H I Parren, Wim K Bleeker
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  ABSTRACT: Ab-dependent cellular cytotoxicity (ADCC) is recognized as a prominent cytotoxic mechanism for therapeutic mAbs in vitro. However, the contribution of ADCC to in vivo efficacy, particularly for treatment of solid tumors, is still poorly understood. For zalutumumab, a therapeutic epidermal growth factor receptor (EGFR)-specific mAb currently in clinical development, previous studies have indicated signaling inhibition and ADCC induction as important therapeutic mechanisms of action. To investigate the in vivo role of ADCC, a panel of EGFR-specific mAbs lacking specific functionalities was generated. By comparing zalutumumab with mAb 018, an EGFR-specific mAb that induced ADCC with similar potency, but did not inhibit signaling, we observed that ADCC alone was insufficient for efficacy against established A431 xenografts. Interestingly, however, both zalutumumab and mAb 018 prevented tumor formation upon early treatment in this model. Zalutumumab and mAb 018 also completely prevented outgrowth of lung metastases, in A431 and MDA-MB-231-luc-D3H2LN experimental metastasis models, already when given at nonsaturating doses. Finally, tumor growth of mutant KRAS-expressing A431 tumor cells, which were resistant to EGFR signaling inhibition, was completely prevented by early treatment with zalutumumab and mAb 018, whereas ADCC-crippled N297Q-mutated variants of both mAbs did not show any inhibitory effects. In conclusion, ADCC induction by EGFR-specific mAbs represents an important mechanism of action in preventing tumor outgrowth or metastasis in vivo, even of cancers insensitive to EGFR signaling inhibition.
  The Journal of Immunology 08/2011; 187(6):3383-90. · 5.79 Impact Factor
• Article: Combined Fc-protein- and Fc-glyco-engineering of scFv-Fc fusion proteins synergistically enhances CD16a binding but does not further enhance NK-cell mediated ADCC.

  Roland Repp, Christian Kellner, Anja Muskulus, Matthias Staudinger, Sahar Mohseni Nodehi, Pia Glorius, Dalia Akramiene, Michael Dechant, Georg H Fey, Patrick H C van Berkel, Jan G J van de Winkel, Paul W H I Parren, Thomas Valerius, Martin Gramatzki, Matthias Peipp
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  ABSTRACT: Protein- or glyco-engineering of antibody molecules can be used to enhance Fc-mediated effector functions. ScFv-Fc fusion proteins (scFv-Fc) represent interesting antibody derivatives due to their relatively simple design and increased tissue penetration. Here, the impact of protein- and glyco-engineering on ADCC potency of a panel of human IgG1-based scFv-Fc was tested. Three matched sets of scFv-Fc variants targeting CD7, CD20 or HLA class II and optimized for CD16a binding by mutagenesis, lack of core-fucose, or their combination, were generated and functionally tested in comparison to the corresponding wild type scFv-Fc. Antigen binding activity was not compromised by altered glycosylation or Fc mutagenesis, whereas Fc binding to CD16a was significantly enhanced in the order: non-core fucosylated/Fc-mutated double-engineered≫Fc-mutated≥non-core-fucosylated>wild-type IgG1-Fc. All engineered variants triggered potent ADCC with up to 100-fold reduced EC50 values compared to non-engineered variants. Interestingly, double-engineered variants were similarly effective in triggering ADCC compared to single-engineered variants irrespective of their 1 log greater CD16a binding affinity. Thus, these data demonstrate that protein- and glyco-engineering enhances NK-cell mediated ADCC of scFv-Fc similarly and show that enhancing CD16a affinity beyond a certain threshold does not result in a further increase of NK-cell mediated ADCC.
  Journal of immunological methods 08/2011; 373(1-2):67-78. · 2.35 Impact Factor
• Article: Complement‐mediated tumor‐specific cell lysis by antibody combinations targeting epidermal growth factor receptor (EGFR) and its variant III (EGFRvIII)

  Katja Klausz, Sven Berger, Jeroen J. Lammerts van Bueren, Stefanie Derer, Stefan Lohse, Michael Dechant, Jan G. J. van de Winkel, Matthias Peipp, Paul W. H. I. Parren, Thomas Valerius
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  ABSTRACT: Monoclonal antibodies (mAb) against variant III of epidermal growth factor receptor (EGFRvIII) hold promise for improving tumor selectivity of EGFR-targeted therapy. Here, we compared Fc-mediated effector functions of three mAb against EGFRvIII (MR1-1, ch806, 13.1.2) with those of zalutumumab, a high affinity EGFR mAb in advanced clinical trials. MR1-1 and ch806 demonstrated preferential and 13.1.2 exclusive binding to EGFRvIII, in contrast to zalutumumab, which bound both wild-type and EGFRvIII. All four human IgG1κ mAb mediated antibody-dependent cellular cytotoxicity (ADCC) of EGFRvIII-expressing cells with mononuclear cells and isolated monocytes, while only zalutumumab in addition triggered ADCC by polymorphonuclear cells. Interestingly, combinations of zalutumumab and EGFRvIII mAb specifically mediated complement-dependent cytotoxicity (CDC) of EGFRvIII-transfected but not wild-type cells. Moreover, EGFRvIII-specific CDC was significantly enhanced when zalutumumab was combined with a Fc-engineered variant of MR1-1 (K326A/E333A). These observations confirm the immunotherapeutic potential of antibody combinations against EGFR, and demonstrate that tumor selectivity can be improved by combining therapeutic EGFR mAb with an antibody against EGFRvIII. (Cancer Sci 2011; 102: 1761–1768)
  Cancer Science 08/2011; 102(10):1761 - 1768. · 3.33 Impact Factor
• Article: Penetration of antibody-opsonized cells by the membrane attack complex of complement promotes Ca(2+) influx and induces streamers.

  Paul V Beum, Margaret A Lindorfer, Elizabeth M Peek, P Todd Stukenberg, Michel de Weers, Frank J Beurskens, Paul W H I Parren, Jan G J van de Winkel, Ronald P Taylor
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  ABSTRACT: We have reported that during complement-mediated cytolysis of B cells promoted by the CD20 mAbs rituximab or ofatumumab (OFA), long, thin structures that we call streamers (≥ 3 cell diameters) are rapidly generated and grow out from the cell surface. Streamers appear before cells are killed and contain opsonizing mAbs and membrane lipids. By exploiting the differential Ca(2+) requirements of discrete steps in the complement cascade, we determined that mAb-opsonized cells first tagged with C3b using C5-depleted serum are killed on addition of serum and EDTA, but the cells do not produce streamers. Also, cells first opsonized with OFA are lysed in serum containing Mg-EGTA by the alternative complement pathway but streamers are not produced. These findings indicate that Ca(2+) influx is necessary for streamer formation. Other mAbs that promote complement-mediated cytolysis also induce streamers on target cells. Streamer-like structures called nanotubes have been reported in several cellular systems, and are thought to promote intercellular communication/signaling. We tested whether this signaling could influence the susceptibility of neighboring cells contacted by streamers to complement attack and found that complement-mediated cytolysis of OFA-opsonized cells increases the resistance of unopsonized indicator cell populations to subsequent lysis when these cells are exposed to OFA and complement.
  European Journal of Immunology 06/2011; 41(8):2436-46. · 5.10 Impact Factor
• Article: Molecular engineering to improve antibodies' anti-lymphoma activity.

  Matthias Peipp, Jan G J van de Winkel, Thomas Valerius
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  ABSTRACT: The human/mouse chimeric CD20 antibody rituximab has significantly improved the survival of lymphoma patients. However, translational research into pharmacology and effector mechanisms of rituximab has identified several limitations of this prototypic antibody. For example, humanized or fully human next-generation antibodies demonstrated reduced immunogenicity, which may translate into improved applicability in certain patient populations. Furthermore, novel technologies of antibody engineering offer the potential to tailor antibody effector functions. Here, glyco- or protein engineering of antibodies' Fc region has demonstrated promising activity in preclinical models. However, these novel molecules are still in early phases of clinical development, and data from on-going and future studies will determine whether promising preclinical results will indeed translate into improved drugs for the treatment of lymphoma patients.
  Best practice & research. Clinical haematology 06/2011; 24(2):217-29. · 3.13 Impact Factor
• Article: Functional characteristics of the high affinity IgG receptor, FcγRI.

  Cees E van der Poel, Robbert M Spaapen, Jan G J van de Winkel, Jeanette H W Leusen
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  ABSTRACT: IgG FcRs are important mediators of immunity and play a key role during Ab-based immunotherapy. Within the leukocyte IgG receptor family, only FcγRI is capable of IgG binding with high affinity. FcγRI exists as a complex of a ligand binding α-chain and an FcR γ-chain. The receptors' α-chain can, furthermore, elicit several functions independent of the ITAM-bearing FcR γ-chain. Functional implications of high-affinity IgG binding and mechanisms underlying FcR γ-chain-independent signaling remain unclear to this day. In this paper, we provide an overview of past literature on FcγRI and address the implications of recently described interactions between cytosolic proteins and the FcγRI α-chain, as well as cytokine-enhanced FcγRI immune complex binding. Furthermore, an analysis of potential polymorphisms within the FCGR1A gene is provided.
  The Journal of Immunology 03/2011; 186(5):2699-704. · 5.79 Impact Factor
• Article: Daratumumab, a novel therapeutic human CD38 monoclonal antibody, induces killing of multiple myeloma and other hematological tumors.

  Michel de Weers, Yu-Tzu Tai, Michael S van der Veer, Joost M Bakker, Tom Vink, Daniëlle C H Jacobs, Lukas A Oomen, Matthias Peipp, Thomas Valerius, Jerry W Slootstra, Tuna Mutis, Wim K Bleeker, Kenneth C Anderson, Henk M Lokhorst, Jan G J van de Winkel, Paul W H I Parren
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  ABSTRACT: CD38, a type II transmembrane glycoprotein highly expressed in hematological malignancies including multiple myeloma (MM), represents a promising target for mAb-based immunotherapy. In this study, we describe the cytotoxic mechanisms of action of daratumumab, a novel, high-affinity, therapeutic human mAb against a unique CD38 epitope. Daratumumab induced potent Ab-dependent cellular cytotoxicity in CD38-expressing lymphoma- and MM-derived cell lines as well as in patient MM cells, both with autologous and allogeneic effector cells. Daratumumab stood out from other CD38 mAbs in its strong ability to induce complement-dependent cytotoxicity in patient MM cells. Importantly, daratumumab-induced Ab-dependent cellular cytotoxicity and complement-dependent cytotoxicity were not affected by the presence of bone marrow stromal cells, indicating that daratumumab can effectively kill MM tumor cells in a tumor-preserving bone marrow microenvironment. In vivo, daratumumab was highly active and interrupted xenograft tumor growth at low dosing. Collectively, our results show the versatility of daratumumab to effectively kill CD38-expressing tumor cells, including patient MM cells, via diverse cytotoxic mechanisms. These findings support clinical development of daratumumab for the treatment of CD38-positive MM tumors.
  The Journal of Immunology 02/2011; 186(3):1840-8. · 5.79 Impact Factor
• Article: Recombinant dimeric IgA antibodies against the epidermal growth factor receptor mediate effective tumor cell killing.

  Stefan Lohse, Stefanie Derer, Thomas Beyer, Katja Klausz, Matthias Peipp, Jeanette H W Leusen, Jan G J van de Winkel, Michael Dechant, Thomas Valerius
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  ABSTRACT: Dimeric IgA Abs contribute significantly to the humoral part of the mucosal immune system. However, their potential as immunotherapeutic agent has hardly been explored. In this article, we describe the production, purification, and functional evaluation of recombinant dimeric IgA against the epidermal growth factor receptor. Human joining chain-containing IgA was produced by nonadherent Chinese hamster ovarian (CHO)-K1 cells under serum-free conditions. Purification by anti-human κ and anti-His-tag affinity, as well as size exclusion chromatography, resulted in a homogenous preparation of highly pure IgA dimers. Functional studies demonstrated dimeric IgA to be at least as effective as monomeric IgA in triggering Ab-dependent cellular cytotoxicity by isolated monocytes or polymorphonuclear cell and in human whole-blood assays. Importantly, dimeric IgA was more effective in F(ab)-mediated killing mechanisms, such as inhibition of ligand binding, receptor downmodulation, and growth inhibition. Furthermore, only dimeric but not monomeric IgA or IgG was directionally transported by the polymeric Ig receptor through an epithelial cell monolayer. Together, these studies demonstrate that recombinant dimeric IgA Abs recruit a distinct repertoire of effector functions compared with monomeric IgA or IgG1 Abs.
  The Journal of Immunology 02/2011; 186(6):3770-8. · 5.79 Impact Factor
• Article: Anti-galactose-α-1,3-galactose IgE from allergic patients does not bind α-galactosylated glycans on intact therapeutic antibody Fc domains.

  Jeroen J Lammerts van Bueren, Theo Rispens, Sandra Verploegen, Tjitske van der Palen-Merkus, Steven Stapel, Lisa J Workman, Hayley James, Patrick H C van Berkel, Jan G J van de Winkel, Thomas A E Platts-Mills, Paul W H I Parren
  Nature Biotechnology 01/2011; 29(7):574-6. · 29.50 Impact Factor
• Article: Anti-galactose-α-1,3-galactose IgE from allergic patients does not bind α-galactosylated glycans on intact therapeutic antibody Fc domains. Jeroen J Lammerts van Bueren, Theo Rispens, Sandra Verploegen, Tjitske van der Palen-Merkus, Steven Stapel, Lisa J Workman, Hayley James, Patrick H C van Berkel, Jan G J van de Winkel, Thomas A E Platts-Mills, Paul W H I Parren Nature Biotechnology01/2011; 29(7):574-6

  Jeroen J Lammerts van Bueren, Theo Rispens, Sandra Verploegen, Tjitske van der Palen-Merkus, Steven Stapel, Lisa J Workman, Hayley James, Patrick H C van Berkel, Jan G J van de Winkel, Thomas A E Platts-Mills, Paul W H I Parren
  Nature Biotechnology 01/2011; 29(7):4-6. · 23.27 Impact Factor
• Article: Cytokine-induced immune complex binding to the high-affinity IgG receptor, FcγRI, in the presence of monomeric IgG.

  Cees E van der Poel, Roos A Karssemeijer, Peter Boross, Joke A van der Linden, Miranda Blokland, Jan G J van de Winkel, Jeanette H W Leusen
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  ABSTRACT: FcγRI is the sole high-affinity immunoglobulin G (IgG) receptor on leukocytes. Its role in immunity and the clearance of opsonized particles has been challenged, as the receptor function may well be hindered by serum IgG. Here, we document immune complex binding by FcγRI to be readily enhanced by cytokine stimulation, whereas binding of monomeric IgG only modestly increased. Enhanced immune complex binding was independent of FcγRI surface expression levels. FcγRI, saturated with prebound IgG, was found capable of effective immune complex binding upon cytokine stimulation. Cytokine-enhanced binding was observed across a variety of immune complexes, including huIgG3- or mIgG2a-opsonized red blood cells, rituximab- or ofatumumab-opsonized B-cell lymphoma, and cetuximab-opsonized glioblastoma cells. This study contributes to our understanding of how FcγRI can participate in the clearance of opsonized particles despite saturation by monomeric IgG.
  Blood 12/2010; 116(24):5327-33. · 9.90 Impact Factor