EGFR/ErbB receptor Tyrosine Kinase Superfamily and Ligand-stimulated EGFR endocytosis

EGFR/ErbB receptors is Tyrosine Kinase Superfamily.belong to ERK superfamily.

There are 8 ligands:TNF-al;pha, EGF, transforming growth factor-alpha (TGFA), heparin-binding EGF-like growth factor (HBEGF), betacellulin (BTC), amphiregulin (AREG), epiregulin (EREG), and epigen (EPGN). 

ligand-stimulated phospholation and endocytosis, binding to DNA leads to enhanced uncontrolled proliferation

Receptor Tyrosine kinase inhibitors:vitamin C,EGCG,curcumin,Dehydroascorbic acid (DHA) directly inhibits IκBα kinase β (IKKβ) and IKKα enzymatic activity

vitamin c block TNF-alpha signaling, thus

curcumin is a TKI

APMG Lung Molecular Pathways
http://www.apmggroup.net/innovation/molecular_testing/Lung_Pathways/lung.html

IJMS | Free Full-Text | Epidermal Growth Factor Receptor Transactivation Is Required for Mitogen-Activated Protein Kinase Activation by Muscarinic Acetylcholine Receptors in HaCaT Keratinocytes | HTML
https://www.mdpi.com/1422-0067/15/11/21433/htm

https://www.researchgate.net/figure/The-nuclear-accumulation-and-receptor-internalization-of-EGFR-family-members-Upon-signal_fig3_51994766

The ErbB family of proteins contains four receptor tyrosine kinases, structurally related to the epidermal growth factor receptor (EGFR), its first discovered member. In humans, the family includes Her1 (EGFR, ErbB1), Her2 (Neu, ErbB2), Her3 (ErbB3), and Her4 (ErbB4). May 13 2019
ErbB - Wikipedia
en.wikipedia.org/wiki/ErbB

Given that ligand binding is essential for the rapid internalization of epidermal growth factor receptor (EGFR), the events induced by ligand binding probably contribute to the regulation of EGFR internalization. These events include receptor dimerization, activation of intrinsic tyrosine kinase activity and autophosphorylation.
Control of epidermal growth factor receptor endocytosis by ...
www.embopress.org/doi/full/10.1038/sj.embor.7400491

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Internalization and intracellular sorting of the EGF ...
https://jcs.biologists.org/content/122/19/3433

Oct 01, 2009 · The epidermal growth factor receptor (EGFR; also known as ErbB1) is so far the best characterized of the four ErbB-family members (EGFR, ErbB2, ErbB3 and ErbB4). EGFR binds to EGF with high affinity in a specific and saturable manner (for a review, see Carpenter and Cohen, 1979). This binding elicits the internalization and intracellular sorting of receptor-ligand complexes and activates …

Cited by: 151
Publish Year: 2009
Author: Inger Helene Madshus, Espen Stang
EGF receptor trafficking: consequences for signaling and ...
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3884125

Ligand-stimulated EGFR endocytosis: a positive and negative regulator of signaling. However, internalization of activated EGFR also enables specific signaling pathways from intracellular sites, and endocytic trafficking of EGFRs is required for optimal activation of a subset of signal transducers [2].

Cited by: 399
Publish Year: 2014
Author:

PLoS One. 2013;8(3):e58148. doi: 10.1371/journal.pone.0058148. Epub 2013 Mar 5.
Internalization mechanisms of the epidermal growth factor receptor after activation with different ligands.
Henriksen L1, Grandal MV, Knudsen SL, van Deurs B, Grøvdal LM.
Author information
1
Department of Cellular and Molecular Medicine, University of Copenhagen, Copenhagen, Denmark.
Abstract
The epidermal growth factor receptor (EGFR) regulates normal growth and differentiation, but dysregulation of the receptor or one of the EGFR ligands is involved in the pathogenesis of many cancers. There are eight ligands for EGFR, however most of the research into trafficking of the receptor after ligand activation focuses on the effect of epidermal growth factor (EGF) and transforming growth factor-α (TGF-α). For a long time it was believed that clathrin-mediated endocytosis was the major pathway for internalization of the receptor, but recent work suggests that different pathways exist. Here we show that clathrin ablation completely inhibits internalization of EGF- and TGF-α-stimulated receptor, however the inhibition of receptor internalization in cells treated with heparin-binding EGF-like growth factor (HB-EGF) or betacellulin (BTC) was only partial. In contrast, clathrin knockdown fully inhibits EGFR degradation after all ligands tested. Furthermore, inhibition of dynamin function blocked EGFR internalization after stimulation with all ligands. Knocking out a number of clathrin-independent dynamin-dependent pathways of internalization had no effect on the ligand-induced endocytosis of the EGFR. We suggest that EGF and TGF-α lead to EGFR endocytosis mainly via the clathrin-mediated pathway. Furthermore, we suggest that HB-EGF and BTC also lead to EGFR endocytosis via a clathrin-mediated pathway, but can additionally use an unidentified internalization pathway or better recruit the small amount of clathrin remaining after clathrin knockdown.

Internalization mechanisms of the epidermal growth factor receptor after activation with different ligands. - PubMed - NCBI
https://www.ncbi.nlm.nih.gov/pubmed/23472148

Ligand-activated epidermal growth factor receptor (EGFR) signaling governs endocytic trafficking of unliganded receptor monomers by non-canonical phosphorylation
Tomohiro Tanaka‡, Yue Zhou‡,§, Tatsuhiko Ozawa¶, Ryuya Okizono‡, Ayako Banba‡, Tomohiro Yamamura‡, Eiji Oga‡, Atsushi Muraguchi¶ and Hiroaki Sakurai‡1
- Author Affiliations

From the Departments of ‡Cancer Cell Biology and
¶Immunology, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Toyama 930-0194, Japan and
the §MOE Key Laboratory for Standardization of Chinese Medicines and the Shanghai Key Laboratory of Compound Chinese Medicines, Institute of Chinese Materia Medica, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China

Abstract
The canonical description of transmembrane receptor function is initial binding of ligand, followed by initiation of intracellular signaling and then internalization en route to degradation or recycling to the cell surface. It is known that low concentrations of extracellular ligand lead to a higher proportion of receptor that is recycled and that non-canonical mechanisms of receptor activation, including phosphorylation by the kinase p38, can induce internalization and recycling. However, no connections have been made between these pathways; i.e. it has yet to be established what happens to unbound receptors following stimulation with ligand. Here we demonstrate that a minimal level of activation of epidermal growth factor receptor (EGFR) tyrosine kinase by low levels of ligand is sufficient to fully activate downstream mitogen-activated protein kinase (MAPK) pathways, with most of the remaining unbound EGFR molecules being efficiently phosphorylated at intracellular serine/threonine residues by activated mitogen-activated protein kinase(MAPK). This non-canonical, p38-mediated phosphorylation of the C-tail of EGFR, near Ser-1015, induces the clathrin-mediated endocytosis of the unliganded EGFR monomers, which occurs slightly later than the canonical endocytosis of ligand-bound EGFR dimers via tyrosine autophosphorylation. EGFR endocytosed via the non-canonical pathway is largely recycled back to the plasma membrane as functional receptors, whereas p38-independent populations are mainly sorted for lysosomal degradation. Moreover, ligand concentrations balance these endocytic trafficking pathways. These results demonstrate that ligand-activated EGFR signaling controls unliganded receptors through feedback phosphorylation, identifying a dual-mode regulation of the endocytic trafficking dynamics of EGFR.

Ligand-activated epidermal growth factor receptor (EGFR) signaling governs endocytic trafficking of unliganded receptor monomers by non-canonical phosphorylation
http://www.jbc.org/content/293/7/2288.short

Targeting receptor tyrosine kinases for chemoprevention by green tea catechin, EGCG.

f2-ijms-9-6-1034: Schematic representation of the erbB family and the IGF/IGF-1R system. (A) The family of erbB receptors includes four members: EGFR (erbB1), HER2 (neu/erbB2), HER3 (erbB3), and HER4 (erbB4). All members have an extracellular ligand binding region (cysteine rich domain), a single membrane-spanning region, and a cytoplasmic tyrosine-kinase-containing domain. Ligand, such as TGF-α etc, binding to erbB receptors induces the formation of receptor homo- and heterodimers and the activation of the intrinsic kinase domain, thus resulting in phosphorylation on specific tyrosine residues within the cytoplasmic tail. These phosphorylated residues serve as docking sites for a range of proteins, the recruitment of which leads to the activation of intracellular signaling pathways, including the Ras/MAPK and PI3K/Akt pathways. (B) The IGF/IGF-1R system is composed of ligands (IGF-1 and IGF-2), receptor (IGF-1R), and ligand binding proteins (IGFBPs). IGF-1 and IGF-2 are found in the circulation complexed to IGFBPs, which serve to regulate the bioavailability of these ligands in the tissues. The IGF-1R contains two α (cysteine rich domain) and two β (tyrosine kinase domain) subunits which are joined by disulfide bridges to form a heterotetrameric receptor complex. The IGF/IGF-1R interaction results in phosphorylation of tyrosine residues in the tyrosine kinase domain. After autophosphorylation, the receptor kinase phosphorylates intracellular proteins, which enable activation of the PI3K/Akt and Ras/MAPK signaling pathways. A detailed description of the downstream signaling pathway is provided in “Figure 3”.

Abstract
Tea is one of the most popular beverages consumed worldwide. Epidemiologic studies show an inverse relationship between consumption of tea, especially green tea, and development of cancers. Numerous in vivo and in vitro studies indicate strong chemopreventive effects for green tea and its constituents against cancers of various organs. (-)-Epigallocatechin-3-gallate (EGCG), the major catechin in green tea, appears to be the most biologically active constituent in tea with respect to inhibiting cell proliferation and inducing apoptosis in cancer cells. Recent studies indicate that the receptor tyrosine kinases (RTKs) are one of the critical targets of EGCG to inhibit cancer cell growth. EGCG inhibits the activation of EGFR (erbB1), HER2 (neu/erbB2) and also HER3 (neu/erbB3), which belong to subclass I of the RTK superfamily, in various types of human cancer cells. The activation of IGF-1 and VEGF receptors, the other members of RTK family, is also inhibited by EGCG. In addition, EGCG alters membrane lipid organization and thus inhibits the dimerization and activation of EGFR. Therefore, EGCG inhibits the Ras/MAPK and PI3K/Akt signaling pathways, which are RTK-related cell signaling pathways, as well as the activation of AP-1 and NF-kappaB, thereby modulating the expression of target genes which are associated with induction of apoptosis and cell cycle arrest in cancer cells. These findings are significant because abnormalities in the expression and function of RTKs and their downstream effectors play a critical role in the development of several types of human malignancies. In this paper we review evidence indicating that EGCG exerts anticancer effects, at least in part, through inhibition of activation of the specific RTKs and conclude that targeting RTKs and related signaling pathway by tea catechins might be a promising strategy for the prevention of human cancers.

Bottom Line: In addition, EGCG alters membrane lipid organization and thus inhibits the dimerization and activation of EGFR.These findings are significant because abnormalities in the expression and function of RTKs and their downstream effectors play a critical role in the development of several types of human malignancies.In this paper we review evidence indicating that EGCG exerts anticancer effects, at least in part, through inhibition of activation of the specific RTKs and conclude that targeting RTKs and related signaling pathway by tea catechins might be a promising strategy for the prevention of human cancers.

Schematic representation of the erbB family and the IGF | Open-i
https://openi.nlm.nih.gov/detailedresult?img=PMC2658783_ijms-9-6-1034-f2&req=4

European Journal of Medicinal Chemistry
Volume 181, 1 November 2019, 111512
European Journal of Medicinal Chemistry
Review article
Curcumin as tyrosine kinase inhibitor in cancer treatment

Author links open overlay panelA.GolonkoaH.LewandowskabR.ŚwisłockacU.T.JasińskaaW.PriebedW.Lewandowskia
a
Institute of Agricultural and Food Biotechnology, Rakowiecka 36, 02-532, Warsaw, Poland
b
Institute of Nuclear Chemistry and Technology, Centre for Radiobiology and Biological Dosimetry, Dorodna 16, 03-195, Warsaw, Poland
c
Bialystok University of Technology, Faculty of Civil Engineering and Environmental Engineering, Department of Chemistry, Biology and Biotechnology, Wiejska 45E, 15-351, Bialystok, Poland
d
Department of Experimental Therapeutics, University of Texas MD, Anderson Cancer Center, Houston, TX, USA

Highlights
•
Our article summarizes knowledge about the mechanisms of curcumin activity directed to signaling pathways of tyrosine kinases.

•
We have attempted to explain how the antitumor activity can be enhanced by modifications in the structure of the molecule.

•
We have discussed the possibility of using curcumin in anti-cancer therapy and problems with its bioavailability.


Abstract
Curcumin is a natural substance known for ages, exhibiting a multidirectional effect in cancer prevention and adjuvant cancer therapies. The great advantage of using nutraceuticals of vegetable origin in comparison to popular cytostatic drugs is the minimized side effect and reduced toxicity. The targets in oncological therapy are, among others, tyrosine kinases, important mediators of signaling pathways whose impaired expression is observed in many types of cancer. Unfortunately, the hydrophobic nature of the curcumin molecule often limits its bioavailability, which is why many studies focus on the chemical modification of this compound. Current research is aimed at modifying structures that improve the pharmacokinetic parameters of curcumin, e.g. the formation of nanoparticles, complexes with metals or the synthesis of curcumin derivatives with functional substituents that allow tumor targeting. The article is a review and analysis of current literature on the properties of curcumin and its derivatives in the treatment of cancers directed to signaling pathways of tyrosine kinases and confronts the problem of low assimilation of curcumin with potential therapeutic effects.

Curcumin as tyrosine kinase inhibitor in cancer treatment - ScienceDirect
https://www.sciencedirect.com/science/article/pii/S0223523419306361

LUNG CARCINOMA

NORMAL MOLECULAR PATHWAYS AND ASSOCIATED MUTATIONS

The RAS-RAF-MEK-ERK signaling pathway (MAPK pathway) is a classical intracellular pathway that plays a crucial role in the homeostasis of normal cell turnover, cellular proliferation, differentiation, survival, and apoptosis. When activated aberrantly, this signaling pathway can induce tumorigenesis and has been associated with various malignancies.

MAPK Signaling Pathway

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EGFR

The Epidermal Growth Factor Receptor (EGFR), also known as HER1 and ERBB1, is an important trans-membrane tyrosine kinase receptor involved in the initiation of the MAPK pathway. Activating mutations in EGFR can lead to aberrant cellular proliferation, inhibition of apoptosis, angiogenesis, increased cell survival, and gene transcription.

Some patients with tumors carrying EGFR mutations may respond to selective anti-EGFR therapies. EGFR has an extracellular ligand-binding region and a cytoplasmic tyrosine kinase-containing domain. Currently, there are two major types of anti-EGFR drug therapies. The first drug type (cetuximab, panitumumab) is a monoclonal antibody that inhibits EGFR by binding to an extracellular portion of the EGFR protein. The second type (gefitinib, erlotinib) is an intracellular tyrosine kinase inhibitor that inhibits EGFR by crossing the cell membrane and blocking the receptor’s active site.


Monoclonal Anti-EGFR Therapy

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Tyrosine Kinase Inhibitor Therapy

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Studies show that patients whose lung cancers exhibit EGFR mutations may benefit from anti-EGFR therapies. Tumor subtypes most likely to respond to anti-EGFR therapies include adenocarcinoma, non-mucinous bronchioloalveolar carcinoma (BAC), and adenosquamous carcinoma. EGFR mutations are rare in other histologic types of lung cancer (small cell carcinoma, squamous cell carcinoma, and large cell carcinoma). Thus, anti-EGFR therapies are most commonly employed for treatment of adenocarcinoma. The two most common EGFR mutations, comprising about 90% of all EGFR mutations in lung adenocarcinomas, are on exon 19 and 21 (L858R). Patients with these mutations are likely to respond to anti-EGFR therapies. There are also less common mutations (exon 20 insertion), including those that actually cause poor response to anti-EGFR therapies, but these mutations are relatively uncommon.

KRAS
KRAS is a small G-protein that is important for EGFR signaling. KRAS mutations activate downstream signaling cascade despite the upstream EGFR regulation. Anti-EGFR therapies are ineffective in tumors with such activating KRAS mutations, because the KRAS mutations trigger the EGFR-signaling cascade at a point downstream of the therapy’s target. Therefore, both monoclonal antibodies and tyrosine kinase inhibitors may fail to respond in the presence of KRAS mutations.

KRAS mutations are seen in about 15-20% of non-small cell lung cancers and 30-50% of lung adenocarcinomas. EGFR and KRAS mutations are essentially mutually exclusive in lung adenocarcinomas. The presence of both EGFR and KRAS mutations in lung adenocarcinomas is rare, but if they do occur together, EGFR inhibitors are less likely to be effective. KRAS mutation is a poor prognostic indicator. The presence of KRAS mutation usually means failure to respond to anti-EGFR therapy. Anti-EGFR therapies are likely to be ineffective in patients with advanced or metastatic lung adenocarcinomas when a KRAS mutation is present.

In patients with lung adenocarcinoma, non-mucinous BAC, or adenosquamous carcinoma, EGFR mutation analysis by polymerase chain reaction (PCR) is recommended to detect those patients who may benefit from anti-EGFR therapy. Laboratories often offer EGFR mutation PCR test, with reflex to KRAS (if EGFR is negative), since the two mutations are mutually exclusive.

KRAS Mutations

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ALK

Non-small cell lung cancers have also been linked to rearrangements of the gene encoding anaplastic lymphoma kinase (ALK). The most common ALK rearrangement found in non-small cell lung cancers is the fusion between echinoderm microtubule-associated protein-like 4 (EML4) and ALK gene on chromosome 2p23. EML4-ALK non-small cell lung cancer is a unique subset of non-small cell lung cancer that is most commonly seen in adenocarcinomas of never or light smokers, whose tumors lack EGFR and KRAS mutations. Patients with ALK rearrangements tend to be younger than most patients with non-small cell lung cancer. These patients usually do not benefit from EGFR-specific therapy, but may benefit from specific ALK inhibitors. Crizotinib is an oral ALK tyrosine kinase inhibitor approved for advanced or metastatic ALK-positive non-small cell lung cancer (i.e patients who harbor the ALK gene rearrangement). FISH is the current standard method for determining the presence of the EML4-ALK gene translocation. EML4-ALK translocations, KRAS mutations, and EGFR mutations in lung cancer are almost always mutually exclusive.

ROS1

ROS1 is a receptor tyrosine kinase. ROS1 fusions are encountered in approximately 2% of non-small cell lung cancers and are identified as a potential driver mutation in non-small cell lung cancers. ROS1 rearrangement-positive lung cancer is a distinct subset of lung cancer with clinical characteristics similar to ALK-rearranged lung cancer. Similar to ALK rearrangements, ROS1 fusions are more commonly seen in light or never smokers. They are also associated with younger age and adenocarcinomas. ROS1 fusion-positive cancers are reportedly sensitive to tyrosine kinase inhibitors that inhibit ROS1. Crizotinib has shown early evidence of clinical response in ROS1 fusion-positive patients. FISH is the current standard method for detecting ROS1 rearrangement. ROS1 mutations are non-overlapping with other oncogenic mutations found in non-small cell lung cancer such as ALK, EGFR, and KRAS mutations.

CLINICAL INDICATIONS FOR EGFR/KRAS/ALK/ROS1 TESTING

Testing should be performed on:
Patients with metastatic adenocarcinoma
Patients with unresectable tumors
Patients with recurrent tumors
Patients who cannot tolerate conventional chemotherapy
Patients with mixed lung cancers with any adenocarcinoma component in a lung resection specimen. For patients without any adenocarcinoma component by histology or IHC (i.e. pure SCC or small cell carcinoma) EGFR and ALK testing is not recommended
Patients with limited specimens (i.e. core biopsy or cytology) where an adenocarcinoma component cannot be excluded and the clinical presentation is suspicious for adenocarcinoma (i.e. young age or absence of smoking history).
Patients with multiple separate primary lung adenocarcinomas. Each separate primary may be tested, however, testing of different areas within the same primary adenocarcinoma is not recommended.
Testing may be useful at the time of the initial resection specimen even in patients who do not have advanced metastatic disease, in order to prevent difficulties/delays in obtaining archival material for molecular testing at the time of recurrence (sometimes resulting in patients receiving additional procedures)

Acceptable specimens usually include:
Formalin-fixed paraffin-embedded tissue
Fresh snap-frozen tissue
Fine needle aspiration cell block

PROPOSED ALGORITHM FOR EGFR/KRAS/ALK/ROS1 TESTING

Testing for EGFR, ALK, KRAS, and ROS1 mutations may all be ordered separately at the time of the resection. Alternatively, since all these mutations are essentially mutually exclusive, KRAS and ROS1 testing are often ordered as reflex, if EGFR or ALK1 is negative, respectively. I am recommending (as available) automatic reflex to KRAS and ROS1 when EGFR and ALK1 are negative. See algorithm below.

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**Although it is counter-intuitive to pursue further testing in a patient who is EGFR- negative, determining the presence of KRAS mutation is still useful information because patients who are EGFR-mutation negative often are still candidates for anti-EGFR therapy as a second-line agent. Studies have shown that anti-EGFR therapy may be useful in patients with NSCLC as second-line, third-line, or as maintenance therapy, including those patients who are EGFR mutation-negative.

The above information is largely based upon the National Comprehensive Cancer Network (NCCN) recommendations and guidelines.

TEST COMMENTS

EGFR mutation analysis by PCR is available upon request. This test is intended to detect EGFR mutation-positive patients. Pulmonary non-small cell carcinomas, including adenocarcinomas, non-mucinous bronchioloalveolar carcinomas, and adenosquamous carcinomas with certain EGFR mutations may respond to anti-EGFR therapies. Alternatively, rare EGFR mutations predict failure to respond to anti-EGFR therapies.

EML4-ALK gene translocation and ROS1 fusion analysis by FISH are also available upon request. These tests are intended to detect patients who may respond to anti-ALK therapy.

KRAS mutation analysis by PCR is also available upon request. This test is useful in patients found to be negative for EGFR mutations. Patients with KRAS mutation may also show relative insensitivity to anti-EGFR therapies that target the RAS/RAF pathway.
TEST INTERPRETATION

PCR GENE MUTATION ANALYSIS INTERPRETATION:
EGFR Mutation Analysis:
Positive = Mutation Detected; specify genotype*
Negative = Mutation Not Detected

*When an EGFR mutation is detected, a specific genotype is reported. Although most EGFR mutations predict a favorable response to anti-EGFR therapies, rare mutations are associated with a lack of clinical response to anti-EGFR therapies.

KRAS Mutation Analysis:
Positive = Mutation Detected: Unlikely to respond to anti-EGFR therapy
Negative = Mutation Not Detected: May respond to anti-EGFR therapy


ALK Fusion Analysis:
Positive = Fusion Detected: May respond to anti-ALK therapy
Negative = Fusion Not Detected: Unlikely to respond to anti-ALK therapy

ROS1 Rearrangement Analysis:
Positive = Fusion Detected: May respond to anti-ALK therapy
Negative = Fusion Not Detected: Unlikely to respond to anti-ALK therapy
SUGGESTED READING


Ladanyi M, Pao W. Lung adenocarcinoma: guiding EGFR-targeted therapy and beyond. Modern Pathology 21: S16-S22, 2008.
http://www.nature.com/modpathol/journal/v21/n2s/full/3801018a.html

CAP/IASLC/AMP: Molecular Testing Guideline for the Selection of Lung Cancer Patients for EGFR and ALK Tyrosine Kinase Inhibitors. Archives of Pathology 2013, 137:828-860

CAP Lung Molecular Reporting Template

CAP Webinar on lung cancer testing can be viewed here:

CAP Resource Page on Molecular Testing in Lung Cancer

CAP Today article on lung cancer molecular testing.

CAP Patient Guide to Lung Cancer Testing.

CAP FAQs on Lung Cancer IHC Guidelines.

APMG Lung Molecular Pathways
http://www.apmggroup.net/innovation/molecular_testing/Lung_Pathways/lung.html

Linking tyrosine kinase inhibitor-mediated inflammation with normal epithelial cell homeostasis and tumor therapeutic responses
https://cdrjournal.com/article/view/2662