Structure of the Complex between HER2 and an Antibody Paratope Formed by Side Chains from Tryptophan and Serine

Fisher, Robert D.; Ultsch, Mark; Lingel, Andreas; Schaefer, Gabriele; Shao, Lily; Birtalan, Sara; Sidhu, Sachdev S.; Eigenbrot, Charles

doi:10.1016/j.jmb.2010.07.027

Cited by 44 publications

(44 citation statements)

References 50 publications

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“…In addition, unlike recombinant proteins expressed in E. coli , recombinant proteins expressed in S2 cells will form intramolecular disulfide bonds. This is particularly important for the HER2 DIV, which contains intramolecular disulfide bridges and is structurally divided into seven modules ( 25-27 ) ( Figure 1 ) that are difficult to reconstitute from a recombinant protein expressed by E. coli . As a result of these advantages, S2 cells are being used for a variety of purposes, including the production of vaccine antigens and other heterologous proteins.…”

Section: Introductionmentioning

confidence: 99%

Expression and purification of HER2 extracellular domain proteins in Schneider2 insect cells

Kanthala

Mill

Riese

et al. 2016

Protein Expression and Purification

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Overexpression of human epidermal growth factor receptor 2 (HER2/ErbB2/Neu) results in ligand independent activation of kinase signaling and is found in about 30% of human breast cancers, and is correlated with a more aggressive tumor phenotype. The HER2 extracellular domain (ECD) consists of four domains − I, II, III and IV. Although the role of each domain in the dimerization and activation of the receptor has been extensively studied, the role of domain IV (DIV) is not clearly understood yet. In our previous studies, we reported peptidomimetic molecules inhibit HER2:HER3 heterodimerization. In order to study the binding interactions of peptidomimetics with HER2 DIV in detail, properly folded recombinant HER2 protein in pure form is important. We have expressed and purified HER2 ECD and DIV proteins in the Drosophila melanogaster Schneider2 (S2) cell line. Using the commercial Drosophila expression system (DES), we transfected S2 cells with plasmids designed to direct the expression of secreted recombinant HER2 ECD and DIV proteins. The secreted proteins were purified from the conditioned medium by filtration, ultrafiltration, dialysis and nickel affinity chromatography techniques. The purified HER2 proteins were then analyzed using western blot, mass spectrometry and circular dichroism (CD) spectroscopy.

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Section: Introductionmentioning

confidence: 99%

Expression and purification of HER2 extracellular domain proteins in Schneider2 insect cells

Kanthala

Mill

Riese

et al. 2016

Protein Expression and Purification

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“…The final dynamics structures were represented using PyMol software. The crystal structure of HER2 ECD was downloaded from the Protein Data Bank (PDB ID 3N85)[13, 49]. Polar hydrogens were added.…”

Section: Methodsmentioning

confidence: 99%

Design of a doxorubicin-peptidomimetic conjugate that targets HER2-positive cancer cells

Pallerla

Gauthier

Sable

et al. 2017

European Journal of Medicinal Chemistry

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Doxorubicin (DOX) belongs to the anthracycline class of drugs that are used in the treatment of various cancers. It has limited cystostatic effects in therapeutic doses, but higher doses can cause cardiotoxicity. In the current approach, we conjugated a peptidomimetic (Arg-aminonaphthylpropionic acid-Phe, compound 5) known to bind to HER2 protein to DOX via a glutaric anhydride linker. Antiproliferative assays suggest that the DOX-peptidomimetic conjugate has activity in the lower micromolar range. The conjugate exhibited higher toxicity in HER2-overexpressed cells than in MCF-7 and MCF-10A cells that do not overexpress HER2 protein. Cellular uptake studies using confocal microscope experiments showed that the conjugate binds to HER2-overexpressed cells and DOX is taken up into the cells in 4 h compared to conjugate in MCF-7 cells. Binding studies using surface plasmon resonance indicated that the conjugate binds to the HER2 extracellular domain with high affinity compared to compound 5 or DOX alone. The conjugate was stable in the presence of cells with a half-life of nearly 4 h and 1 h in human serum. DOX is released from the conjugate and internalized into the cells in 4 h, causing cellular toxicity. These results suggest that this conjugate can be used to target DOX to HER2-overexpressing cells and can improve the therapeutic index of DOX for HER2-positive cancer.

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“…The region of PPI and its inhibition is marked by an oval shape. The model of EGFR:HER2 heterodimer was generated [108,109] using the crystal structures of EGFR (PDB ID 3NJP) [110] and HER2 (PDB ID) [111] with a homodimer of EGFR as a template (PDB ID 3NJP). EGFR: EGF receptor; PPI: Protein–protein interaction.…”

Section: Executive Summarymentioning

confidence: 99%

Current and future targeted therapies for non-small-cell lung cancers with aberrant EGF receptors

2015

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Expression of the EGF receptors (EGFRs) is abnormally high in many types of cancer, including 25% of lung cancers. Successful treatments target mutations in the EGFR tyrosine kinase domain with EGFR tyrosine kinase inhibitors (TKIs). However, almost all patients develop resistance to this treatment, and acquired resistance to first-generation TKI has prompted the clinical development of a second generation of EGFR TKI. Because of the development of resistance to treatment of TKIs, there is a need to collect genomic information about EGFR levels in non-small-cell lung cancer patients. Herein, we focus on current molecular targets that have therapies available as well as other targets for which therapies will be available in the near future.

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Structure of the Complex between HER2 and an Antibody Paratope Formed by Side Chains from Tryptophan and Serine

Cited by 44 publications

References 50 publications

Expression and purification of HER2 extracellular domain proteins in Schneider2 insect cells

Expression and purification of HER2 extracellular domain proteins in Schneider2 insect cells

Design of a doxorubicin-peptidomimetic conjugate that targets HER2-positive cancer cells

Current and future targeted therapies for non-small-cell lung cancers with aberrant EGF receptors

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