Breakthroughs and Views
The ErbB/HER receptor protein-tyrosine kinases and cancer

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Abstract

The ErbB/HER protein-tyrosine kinases, which include the epidermal growth factor receptor, consist of a growth-factor-binding ectodomain, a single transmembrane segment, an intracellular protein-tyrosine kinase catalytic domain, and a tyrosine-containing cytoplasmic tail. The genes for the four members of this family, ErbB1–ErbB4, are found on different human chromosomes. Null mutations of any of the ErbB family members result in embryonic lethality. ErbB1 and ErbB2 are overexpressed in a wide variety of tumors including breast, colorectal, ovarian, and non-small cell lung cancers. The structures of the ectodomains of the ErbB receptors in their active and inactive conformation have shed light on the mechanism of receptor activation. The extracellular component of the ErbB proteins consists of domains I–IV. The activating growth factor, which binds to domains I and III, selects and stabilizes a conformation that allows a dimerization arm to extend from domain II to interact with an ErbB dimer partner. As a result of dimerization, protein kinase activation, trans-autophosphorylation, and initiation of signaling occur. The conversion of the inactive to active receptor involves a major rotation of the ectodomain. The ErbB receptors are targets for anticancer drugs. Two strategies for blocking the action of these proteins include antibodies directed against the ectodomain and drugs that inhibit protein-tyrosine kinase activity. A reversible ATP competitive inhibitor of ErbB1 (ZD1839, or Iressa) and an ErbB1 ectodomain directed antibody (IMC-C225, or Erbitux) have been approved for the treatment of non-small cell lung cancer and colorectal cancer, respectively. An ErbB2/HER2 ectodomain directed antibody (trastuzumab, or Herceptin) has also been approved for the treatment of breast cancer. Current research promises to produce additional agents based upon these approaches.

Section snippets

Protein-tyrosine kinases and cancer

Many protein kinases are implicated in the mechanisms leading to malignancies [9]. In 1980, Hunter and Sefton [8] demonstrated that the Rous sarcoma virus (v-src) oncogene product is a protein-tyrosine kinase. Later work showed that v-src (the viral protein) and c-src (the cellular homologue) are non-receptor protein-tyrosine kinases.

Cohen and co-workers discovered that the receptor for epidermal growth factor is a protein-tyrosine kinase, the first receptor protein-tyrosine kinase to be

The ErbB/HER growth factor ligands

The EGF family of ligands, which consists of about a dozen members, has an EGF-like domain and three disulfide-bonded intramolecular loops [16]. These peptide ligands are expressed in the extracellular domain of transmembrane proteins and are generated by regulated proteolysis to yield growth factors that contain 49–85 amino acids.

The EGR receptor family has four members [3]. The first family member is the EGF receptor [17]; its human gene is designated ErbB1 after the v-erbB oncogene of avian

The ErbB family of receptor protein-tyrosine kinases

The various ErbB growth factors have different specificities for the receptor family members (Fig. 1). For example, EGF and TGF-α have high affinity for the EGF receptor, ErbB1. Moreover, the transforming potential and signaling pathways activated by different dimers are distinctive. For example, ErbB1–ErbB2 heterodimers are associated with a more robust signal then ErbB1–ErbB1 homodimers [19]. All four members of the ErbB family have the potential to stimulate the Raf-MEK-ERK protein kinase

Ligand binding to the ErbB ectodomains and receptor activation

The stoichiometry for epidermal growth factor binding to activated receptor is 2 mol EGF to 2 mol EGFR [24]. This finding is consistent with a mechanism involving each of two EGF molecules spanning the EGF receptor dimer interface (a divalent EGF binding simultaneously to two EGF receptors) or with EGF binding to a single receptor and promoting receptor dimerization (a monovalent EGF binding to a single EGF receptor). The extracellular component of the four family members consists of domains I–IV

Structure of the ErbB2 ectodomain

The second member of the family, ErbB2/HER2, has several unique properties [3]. First, ErbB2 lacks a known direct ligand; in order to function, it must work as a co-receptor, or heterodimerization partner, for other ErbB receptors that possess stimulatory ligands [15], [19], [20]. Second, unlike other ErbB receptors, ErbB2 overexpression can cause malignant transformation without the expression of a growth factor. This observation suggests that ErbB2 has a high level of constitutive

Therapeutic antibodies directed to ErbB ectodomains

A search for genetic alterations in breast cancers showed that ErbB2, or HER2 (human EGF receptor 2), is amplified up to 100-fold in tumor cells from about 25% to 30% of people with invasive breast cancer. A significant clinical correlation exists between HER2 overexpression and the severity of the malignancy [14].

Mouse chimeric humanized monoclonal antibodies targeted against the extracellular component of ErbB2 are clinically useful [31]. Trastuzumab (Herceptin from Genentech) is a humanized

Structure of the epidermal growth factor receptor protein-tyrosine kinase domain

The EGF receptor protein-tyrosine kinase domain has the characteristic bilobed architecture observed in all protein kinases (Fig. 3) as predicted by Taylor and co-workers [35]. Residues 685–769 make up the amino-terminal lobe of the kinase, and residues 773–953 make up the carboxyterminal lobe. The smaller lobe has a predominantly antiparallel β-sheet structure. It contains the glycine-rich (GSGAFG) ATP-phosphate binding loop composed of residues 695–700. The large lobe is predominantly

The activation loop

The conformation of the activation loop differs between activate and dormant kinases [40], [41]. The activation loop of nearly all protein kinases begins with DFG and ends with APE. That of the EGF receptor kinase begins with DFG (831–833), but ends in ALE (858–860). In protein kinases that are dormant, the activation loop has a compact conformation that inhibits the binding of protein substrates (and ATP in some kinases). In most protein-tyrosine kinases, a tyrosine residue in the activation

The protein kinase inhibitory ectodomain of ErbB proteins

Two lines of evidence indicate that the ectodomain constrains EGF receptor kinase activity. First, the v-erb murine oncoprotein, homologous to the EGF receptor, is a constitutively active protein kinase that consists of a short ectodomain, a transmembrane segment, and an intracellular catalytic domain [13]. The ectodomain does not bind ligand. Furthermore, the human oncoprotein (EGFRvIII) that lacks residues 6–273 is unable to bind EGF and is constitutively active [17]. These findings indicate

Inhibition of the EGF receptor protein-tyrosine kinase by anilinoquinazolines

Inhibition of the EGF receptor kinase has been proposed as a rational approach to cancer therapy for about 15 years. The identification of 4-anilinoquinazolines ushered in a new era owing to the potency for this enzyme [43]. These compounds are reversible competitive inhibitors with respect to ATP.

Two drugs that have emerged as therapeutic EGF receptor kinase inhibitors include ZD1839 and OSI-774 (Fig. 4), both of which are active in humans when given orally. The antineoplastic effects are

Binding of OSI-774 to the EGF receptor protein kinase domain

The X-ray crystallographic structure of the catalytic domain of the human EGF receptor protein-tyrosine kinase domain has been solved in the presence and absence of OSI-774 [4]. The drug lies with the N1 and C8-containing edge of the quinazoline directed toward the peptide segment connecting the N-terminal and C-terminal lobes of the kinase. The N1 of the quinazoline accepts a hydrogen bond from the Met769 amide nitrogen (Fig. 5); this is how the N1 of ATP is inferred to bind to the enzyme.

ErbB glycoprotein synthesis and degradation

Each of the ErbB proteins is an N-linked glycoprotein; about 20% of the mass of these proteins is carbohydrate. When glycosylation of the EGF receptor is blocked by tunicamycin during synthesis in cell culture, the EGF receptor protein kinase activity is only 30% of that of the control receptor [47]. This treatment inhibits the attachment of carbohydrate, and the molecular weight is decreased from 170,000 to 140,000. When carbohydrate is cleaved enzymatically from the mature receptor, protein

Epilogue

The EGF receptor is one of the first proteins, if not the first, that was implicated in the production of human malignancies. Neoplasms that are often associated with dysregulation of this receptor include cancers of the lung, colon, and breast. These neoplasms are among the most prevalent malignancies in the US. If inhibitors of the EGF receptor were efficacious in only 10% of these disorders, the number of new treatable cases would be more than 50,000 annually in the US. A major difficulty

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    Abbreviations: AR, amphiregulin; BTC, betacellulin; EGF; epidermal growth factor; EGFR, epidermal growth factor receptor; EPR, epiregulin; HB-EGF, heparin-binding epidermal growth factor; NRG, neuregulin/heregulin/neu differentiation factor; PTB, phosphotyrosine binding; SH2, Src homology 2; TGF-α, transforming growth factor-α.

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