A Novel Bispecific Tetravalent Antibody Fusion Protein to Target Costimulatory Activity for T-cell Activation to Tumor Cells Overexpressing ErbB2/HER2

Biburger, Markus; Weth, Robert; Wels, Winfried S.

doi:10.1016/j.jmb.2004.12.052

Cited by 20 publications

(8 citation statements)

References 67 publications

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“…However, this was the case only under the condition that both proteins were linked to the same stimulator cells. Similar observations were reported for other systems (25)(26)(27). Also, for specific antigenpresenting cells, signals 1 and 2 have to be expressed on the same cell in order to achieve optimal effects (28,29).…”

Section: Discussionsupporting

confidence: 91%

Optimization studies for the coupling of bispecific antibodies to viral anchor molecules of a tumor vaccine

Fournier

Aigner²,

Schirrmacher³

2010

Int J Oncol

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Abstract. Tumor vaccines have to provide several signals for T cell activation. Among them, signal 1 (through TCR/ CD3) and signal 2 (through CD28) are the most important. We herein describe a procedure to introduce anti-CD3 and anti-CD28 signals into any tumor cell which is susceptible to infection by Newcastle disease virus (NDV). We developed the ATV-NDV tumor vaccine which consists of patientderived tumor cells (ATV) modified through infection by NDV. We tested for further improvement of vaccine efficiency the addition of two bispecific single-chain antibodies. They bind with one arm to the viral hemagglutininneuraminidase (HN) or fusion (F) protein of NDV expressed at the surface of the vaccine cells while the second arm is directed either against CD3 or CD28 of T cells. The aim of this study was to optimize the coupling of these new reagents to the tumor vaccine. When anti-CD3 and anti-CD28 molecules bind to the same anchoring viral molecule (e.g. HN), competition for binding could occur under certain conditions. This was not the case when the bispecific reagents bound to separate viral molecules (HN or F, respectively). When using transfectants expressing HN and F either separately or on the same cell, we show that T cell activation works best when anti-CD3 and anti-CD28 are attached to the same stimulator cell. The clinical application of such a combined therapy with ATV-NDV vaccine cells and bi-specific antibodies allows to modify the strength of signal 1 and 2 in a quantitative and predictable way according to the immune status of the T cells and the requirements of the patients' immune system.

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Section: Discussionsupporting

confidence: 91%

Optimization studies for the coupling of bispecific antibodies to viral anchor molecules of a tumor vaccine

Fournier

Aigner²,

Schirrmacher³

2010

Int J Oncol

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“…Proteins can be efficiently secreted into the culture supernatant, which aids proper folding and allows posttranslational modification such as glycosylation [24], [32], [33], [34]. Nevertheless, protein expression levels comparable to those of bacteria can often not be achieved.…”

Section: Resultsmentioning

confidence: 99%

Maltose-Binding Protein Enhances Secretion of Recombinant Human Granzyme B Accompanied by In Vivo Processing of a Precursor MBP Fusion Protein

et al. 2010

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BackgroundThe apoptosis-inducing serine protease granzyme B (GrB) is an important factor contributing to lysis of target cells by cytotoxic lymphocytes. Expression of enzymatically active GrB in recombinant form is a prerequisite for functional analysis and application of GrB for therapeutic purposes.Methods and FindingsWe investigated the influence of bacterial maltose-binding protein (MBP) fused to GrB via a synthetic furin recognition motif on the expression of the MBP fusion protein also containing an N-terminal α-factor signal peptide in the yeast Pichia pastoris. MBP markedly enhanced the amount of GrB secreted into culture supernatant, which was not the case when GrB was fused to GST. MBP-GrB fusion protein was cleaved during secretion by an endogenous furin-like proteolytic activity in vivo, liberating enzymatically active GrB without the need of subsequent in vitro processing. Similar results were obtained upon expression of a recombinant fragment of the ErbB2/HER2 receptor protein or GST as MBP fusions.ConclusionsOur results demonstrate that combination of MBP as a solubility enhancer with specific in vivo cleavage augments secretion of processed and functionally active proteins from yeast. This strategy may be generally applicable to improve folding and increase yields of recombinant proteins.

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“…ScFv(FRP5) was, in another approach, also assembled into a bispecific tetravalent construct containing an immunoglobulin variable domain (IgV)-like CD28 binding domain of the T cell costimulatory molecule CD86. This CD86(111)-IgG-scFv(FRP5) molecule is able to bind two ErbB2 targets on tumor cells and two CD28 proteins on T cells, providing a strong costimulatory signal in addition to mediating T cell-target interaction [170]. Along similar principles, Shi et al [171] have recently reported about an anti-ErbB2 scFv-Fc-IL2 fusion protein that was shown to mediate ADCC in vitro, and exhibited significant antitumor activity in vivo.…”

Section: Other Erbb2 Specific Antibodies and Antibody Derivativesmentioning

confidence: 97%

ErbB-directed immunotherapy: Antibodies in current practice and promising new agents

Friedländer¹,

Barok²,

Szöllősi³

et al. 2008

Immunology Letters

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A Novel Bispecific Tetravalent Antibody Fusion Protein to Target Costimulatory Activity for T-cell Activation to Tumor Cells Overexpressing ErbB2/HER2

Cited by 20 publications

References 67 publications

Optimization studies for the coupling of bispecific antibodies to viral anchor molecules of a tumor vaccine

Optimization studies for the coupling of bispecific antibodies to viral anchor molecules of a tumor vaccine

Maltose-Binding Protein Enhances Secretion of Recombinant Human Granzyme B Accompanied by In Vivo Processing of a Precursor MBP Fusion Protein

ErbB-directed immunotherapy: Antibodies in current practice and promising new agents

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