EGFR Tyrosine Kinase Domain Research Articles

Anticancer activity, DFT calculation, ADMET study and molecular docking of synthetic Chalcone (2E)-1-(4-Aminophenyl)-3-(3,4,5-trimethoxyphenyl)-2- propen-1-one in breast cancer cell line (MCF-7)

The (2E)-1-(4-Aminophenyl)-3-(3,4,5-trimethoxyphenyl)-2-propen-1-one was prepared by the reaction between 4-aminoacetophenone and 3,4,5, trimethoxy benzaldehyde in ethanolic NaOH. Then in vitro anticancer activity was evaluated against the MCF-7 breast cell line. The molecular properties like bond length, bond angle, molecular electrostatic potential surface (MEPs), Mullikan charge, HOMO-LUMO, and NBO analysis data obtained by DFT/B3LYP method using 6- 31+G(d,p) basis set to get electronic states and molecular parameters of the title molecule. Because of their advantageous pharmacokinetic properties, the synthesized compound was shown to have moderate anticancer activity and could advance to a higher level of the drug development process. Molecular docking studies were utilized to foresee the binding interactions of the title compound with the EGFR tyrosine kinase domain (PDB ID: 1M17). In silico ADME study is also performed to predict the pharmacokinetic profile which expressed good oral drug-like behavior.

In-silico molecular docking, ADME study, and molecular dynamic simulation of new azetidin-2-one derivatives with antiproliferative activity

Over the past two decades, protein kinase has been a heavily studied target in the development of new anti-proliferative medications. Heterocyclic systems have been identified as the preferred scaffold because of their wide range of biological properties. In this research, the objective was to design and develop fifteen novel azetidin-2-one derivatives and assess their cytotoxic potential as inhibitors of the epidermal growth factor receptor, which is considered one of the key factors influencing cell growth and proliferation. The crystal structure of inactive EGFR tyrosine kinase domain ligand erlotinib from protein data bank was retrieved in order to be docked with our proposed azetidine-2-one derivatives to evaluate their activity as anti-proliferative agents. In this article, an in-silico molecular docking approach proposes that these azetidine-2-one derivatives have satisfactory binding contact with the erlotinib binding site. Although, three compounds have been identified as the most powerful as they have PLP fitness scores of (77.79, 76.68 and 71.46), respectively, while the reference ligand’s fitness score was (71.94). Additionally, all of our derivatives have satisfied the Swiss-ADME parameters, indicating that they may be orally active compounds. In conclusion, two compounds (A-2 and A-8) have better PLP fitness, and one (A-14) has a comparable score in comparison to the reference ligand, at the active site of epidermal growth factor receptor. indicating that the novel azetidine-2-one derivatives have shown interesting results and could be used as model compounds to create novel anti-proliferative drugs. However, more pharmacological evaluation is needed.

Design, synthesis, in-silico studies and apoptotic activity of novel amide enriched 2-(1H)- quinazolinone derivatives

Cancer is a broad classification of diseases that can affect any organ or body tissue due to aberrant cellular proliferation for unknown reasons. Many present chemotherapeutic drugs are highly toxic and have little selectivity. Additionally, they lead to the development of medication resistance. Therefore, developing tailored chemotherapeutic drugs with minimal side effects and good selectivity is crucial for cancer treatment. 2-(1H)-Quinazolinone is one of the vital scaffold and anticancer activity is one of the prominent biological activities of this class. Here we report the novel set of amide-enriched 2-(1H)-quinazolinone derivatives (7a-j) and their apoptotic activity with the help of MTT assay method against four human cancer cell lines: PC3 (prostate cancer), DU-145 (prostate cancer), A549 (lung cancer), and MCF7 (breast cancer). When compared to etoposide, every synthetic test compound (7a-j) exhibited moderate to excellent activity. The IC50 values of the new amide derivatives (7a-j) varied from 0.07 ± 0.0061 μM to 10.8 ± 0.69 μM. While the positive control, etoposide, exhibited 1.97 ± 0.45 μM to 3.08 ± 0.135 μM range. Among the novel amide derivatives (7a-j), in particular, 7i and 7j showed strong apoptotic activity against MCF7; 7h showed against PC3, and 7g showed against DU-145. Molecular docking studies of test compounds (7a-j) with the EGFR tyrosine kinase domain (PDB ID: 1M17) protein provided the significant docking scores for each test compound (7a-j) (−9.00 to −9.67 kcal/mol). Additionally, DFT investigations and MD simulations validated the predictions of molecular docking. According to the findings of the ADME analysis, oral absorption by humans is anticipated to be higher than 85 % for all test compounds.

Docking, Synthesis and Evaluation of 4-hydroxy-1-phenyl-2(1H)-quinolone Derivatives as Anticancer Agents.

The estimated number of cancer cases in India for the year 2022 was found to be 14,61,427. The development of chemotherapeutic agents has reduced the mortality rate, however, they have high toxicity which is a disadvantage. Many researchers have found out that quinolin-2- one possesses anticancer activity, with this background we thought of synthesizing the quinolin-2-one derivatives. This study aimed to carry out docking, synthesis, characterization, and evaluation of 2-(2- (4-Hydroxy-2-oxoquinolin-1(2H)-yl)phenyl/ substituted phenyl)-3-(phenylamino) thiazolidon-4-one derivatives (IVa-g) as an anticancer agent. Diphenylamine and malonic acid treated with phosphoryl chloride gave compound I, which on formylation afforded compound II, which on reaction with various substituted aromatic phenylhydrazine derivatives gave compounds IIIa-g, which on further reaction with thioglycolic acid and anhydrous zinc chloride yielded the compounds IVa-g. Among all the synthesized novel derivatives, compounds IV a-d showed 50% lysis in the IC50 range of 25-50μg for the A549 cell line, and compounds IVa, and IVb showed 50% lysis in the IC50 range of 25-50μg for the MDA-MB cell line. The compound, 3-((4-fluorophenyl)amino)-2-(2-(4- hydroxy-2-oxoquinolin-1(2H)-yl)phenyl)thiazolidin-4-one (IVg) was found to be the most active against both the cell line, A549 and MDA-MB with IC50 value of 0.0298μmol and 0.0338μmol respectively. The docking results revealed that the synthesized compounds exhibited well-conserved hydrogen bonding with one or more amino acid residues in the active pocket of EGFR tyrosine kinase domain with 4-anilinoquinazoline inhibitor erlotinib (PDB ID:1M17). Compound IVg showed the highest MolDock score of -137.813 compared to the standard drug Imatinib having a MolDock score of -119.354. Compound IVg showed the highest MolDock score and was also found to be most potent against both the cell line, A549, and MDA-MB.

Schizandrin A induces non-small cell lung cancer apoptosis by suppressing the epidermal growth factor receptor activation.

The purpose of this study is to explore the biological mechanism of Schizandrin A (SchA) inducing non-small cell lung cancer (NSCLC) apoptosis. The reverse molecular docking tool "Swiss Target Prediction" was used to predict the targets of SchA. Protein-protein interaction analysis was performed on potential targets using the String database. Functional enrichment analyses of potential targets were performed with Gene Ontology and Kyoto Encyclopedia of Genes and Genomes. The conformation of SchA binding to target was simulated by chemical-protein interactomics and molecular docking. The effect of SchA on the expression and phosphorylation level of EGFR was detected by Western blot. Lipofectamine 3000 and EGFR plasmids were used to overexpress EGFR. Apoptosis was tested with Annexin V-FITC and propidium iodide staining, and cell cycle was detected by propidium iodide staining. The "Swiss Target Prediction" database predicted 112 and 111 targets based on the 2D and 3D structures of SchA, respectively, of which kinases accounted for the most, accounting for 24%. Protein interaction network analyses showed that molecular targets such as ERBB family and SRC were at the center of the network. Functional enrichment analyses indicated that ERBB-related signaling pathways were enriched. Compound-protein interactomics and molecular docking revealed that SchA could bind to the ATP-active pocket of the EGFR tyrosine kinase domain. Laboratory results showed that SchA inhibited the phosphorylation of EGFR. Insulin could counteract the cytotoxic effect of SchA. EGFR overexpression and excess EGF or IGF-1 had limited impacts on the cytotoxicity of SchA. Network pharmacology analyses suggested that ERBB family members may be the targets of SchA. SchA can inhibit NSCLC at least in part by inhibiting EGFR phosphorylation, and activating the EGFR bypass can neutralize the cytotoxicity of SchA.

Novel 5-bromoindole-2-carboxylic Acid Derivatives as EGFR Inhibitors: Synthesis, Docking Study, and Structure Activity Relationship.

The indole backbone is encountered in a class of N-heterocyclic compounds with physiological and pharmacological effects such as anti-cancer, anti-diabetic, and anti-HIV. These compounds are becoming increasingly popular in organic, medicinal, and pharmaceutical research. Nitrogen compounds' hydrogen bonding, dipole- dipole interactions, hydrophobic effects, Van der Waals forces, and stacking interactions have increased their relevance in pharmaceutical chemistry due to their improved solubility. Indole derivatives, such as carbothioamide, oxadiazole, and triazole, have been reported to act as anti-cancer drugs due to their ability to disrupt the mitotic spindle and prevent human cancer cell proliferation, expansion, and invasion. To synthesize new 5-bromoindole-2-carboxylic acid derivatives that function as EGFR tyrosine kinase inhibitors as deduced through molecular docking studies. Different derivatives of indole (carbothioamide, oxadiazole, tetrahydro pyridazine-3,6-dione, and triazole) were synthesized and evaluated through different chemical, spectroscopic methods (IR, 1HNMR, 13CNMR, and MS) and assessed in silico and in vitro for their antiproliferative activities against A549, HepG2, and MCF-7 cancer cell lines. According to molecular docking analyses, compounds 3a, 3b, 3f, and 7 exhibited the strongest EGFR tyrosine kinase domain binding energies. In comparison to erlotinib, which displayed some hepatotoxicity, all of the evaluated ligands displayed good in silico absorption levels, did not appear to be cytochrome P450 inhibitors, and were not hepatotoxic. The new indole derivatives were found to decrease cell growth of three different types of human cancer cell lines (HepG2, A549, and MCF-7), with compound 3a being the most powerful while still being cancer-specific. Cell cycle arrest and the activation of apoptosis were the results of compound 3a's inhibition of EGFR tyrosine kinase activity. The novel indole derivatives, compound 3a in particular, are promising anti-cancer agents which inhibit cell proliferation by inhibiting EGFR tyrosine kinase activity.

Synthesis and docking studies of new 5-bromoindole-2-carboxylic acid oxadiazole derivatives as EGFR inhibitors

The current study reports the design and synthesis of novel oxadiazole derivatives of 5-bromoindole-2-carboxylic acid and their molecular docking properties. In order to determine the structure of the new oxadiazole derivatives (3, 4a, 4b), a number of spectroscopic techniques (IR and 1HNMR) were employed. Molecular docking analysis indicated that compounds 3, 4a, 4b showed favorable binding free energy against the EGFR tyrosine kinase domain. None of these compounds appeared to suppress cytochrome P450, and all of them showed appropriate absorption levels. Moreover, they did not exhibit any hepatotoxicity when tested in vitro. Compound 4a Reported to have the highest stability, with a good net binding energy, and an excellent binding energy with hot amino acids. Dipole moment is also rather high. This result is an indication of compound 4a is with superior capacity to create hydrogen bonds over erlotinib. Likewise, compound 4a was found to be the most stable in its interaction with EGFR tyrosine kinase. Finally, molecular dynamic simulation revealed excellent outcomes for compound 4a.

Efficacy and safety study of osimertinib in advanced non-small cell lung cancer (NSCLC) with EGFR mutation.

e21150 Background: The frequency of transmission on EGFR is more frequent in women and non-smokers. It is characteristic of the Asian population (10% in the Caucasian race). EGFR tyrosine kinase domain inhibitors (EGFR-TKIs) are the standard first-line therapy in non-small cell lung cancer (NSCLC) with EGFR mutations. The efficacy and safety of osimertinib (third generation TKI-EGFR) were evaluated in the FLAURA phase III clinical trial, comparing it against first and second generation TKI-EGFR (erlotinib and gefitinib) in patients with advanced NSCLC. The objective is to evaluate the efficacy and safety of osimertinib in routine clinical practice at the Juan Ramon Jimenez Hospital. Methods: Descriptive observational, and retrospective study of patients with advanced NSCLC and EGFR + treated TKI-EGFR in the period 2015-2022. We analyzed (IBM SPSS Statistics 22) sociodemographic and clinical variables, toxicity profile, objective response rate (ORR), treatment response and progression-free survival (PFS) with Kappan-Meier curves. Results: We included 41 patients (p). 24 women (58.5%). The median age 66 years. 61% had a smoking habit (39% active smokers and 22% ex-smokers). 1p South American race, rest Caucasians. Present histologies: adenocarcinoma (87.8%), squamous cell (4.9%) and signet ring cells (2.4%). EGFR mutations: 53.7% exon 19 deletion and 36.6% exon 20 mutation and 9.8% were not specified. In 10 p the T790 mutation was analyzed, positive in 14.6%. Stage at diagnosis: IV (87.8%); III (12.2%). Symptoms at diagnosis: persistent cough (22%), pain (22%), dyspnea (19.5%) and constitutional syndrome (12.2%). Status Performance: 0-1 (85.4%). Pulmonary (48.8%), bone (36.6%), lymph node (29.3%), liver (22%), brain (22%), and adrenal (14.6%) metastases. 58.5% received osimertinib in the first line, 17.1% in the second. ORR: 77.4% partial response, 9.7% complete response and 12.9% disease progression. Toxicity (T) 90.9%. Digestive T: Diarrhea: grade G1 36.7%, G2 13.3%, G3 6.7%; nausea: G1 3.3%; hyporexia: G1 25.8%; hypertransaminasaemia: G1 3.2%. Dermal T: skin dryness: G1 20.7%, G2 10.3%, G3 3.4%; onycholysis: G1 19.4%. Asthenia: G1 25.8%, G2 12.9%, G3 12.9%. Pneumonitis: G1 3.2%, G2 3.2%, G3 6.5%. Hematological T: anemia: G1 6.5%, G2 16.1%, G3 3.2%. Dose reduction to 40 mg (37.5%). 12.5% discontinued osimertinib. mPFS in 1st line (L): 14 m (m). mPFS in 2nd L: 8 m. 95% CI (0-31.5). 95% CI (9.8- 18.1. mOS: 21 m (95% CI: 17.73-24.26). Conclusions: Osimertinib presents an adequate safety profile for patients with G1, G2 toxicities, generally easy to manage in clinical practice. A very low percentage of patients had to discontinue treatment. No fatal adverse event was reported. Our results in OS and PFS, although favorable, are somewhat lower than those of the FLAURA clinical trial, possibly due to dealing with real-life patients and/or the size of the sample, which is not entirely sufficiently representative.

Development and validation of a new LC–MS/MS method for the determination of mefatinib in human plasma and its first application in pharmacokinetic studies

Mefatinib (MET306) is a novel second-generation epidermal growth factor receptor-tyrosine kinase inhibitor (EGFR-TKI) designed to address the highly unmet clinical need of gefitinib-induced resistance and irreversibly bind to mutated tyrosine kinase domain of EGFR and human epidermal growth factor receptor 2 (HER2). In this study, a liquid chromatography–tandem mass spectrometry method was established and validated for determining MET306 in non-small cell lung cancer patients and a backpropagation artificial neural network was developed and constructed to predict the pharmacokinetic process. The mobile phase was water containing 5 mM ammonium acetate and acetonitrile at a flow rate of 0.3 mL min−1, within a 4.5 min run time. MET306 was separated on a Hypersil Gold-C18 at 40 °C and subjected to mass analysis using positive electrospray ionization. A total of 524 data were used as development groups and 145 data were used as testing groups. The final established Northern Goshawk Optimization-Backpropagation Artificial Neural Network (NGO-BPANN) model consisted of one input layer with 6 neurons, 1 hidden layer with 10 nodes, and 1 output layer with one node processed by MATLAB2021a.The calibration range of MET306 was 0.5–200 ng mL−1 with the correlation coefficient r ≥ 0.99. Accuracies ranged from 97.20 to 110.80% and the inter- and intra-assay precision were less than 15%. The ranges of extraction recoveries were 104.95% to 112.09% for analyte and internal standard and there was no significant matrix effect. The storage stability under different conditions was in accordance with the bioanalytical guidelines. The time-concentration profiles of the measured and predicted concentrations of MET306 by NGO-BPANN agree well. An NGO-BPANN model was developed to predict the plasma concentration and pharmacokinetic parameters of MET306 in the first time.

Computational Investigation of 1, 3, 4 Oxadiazole Derivatives as Lead Inhibitors of VEGFR 2 in Comparison with EGFR: Density Functional Theory, Molecular Docking and Molecular Dynamics Simulation Studies

Vascular endothelial growth factor (VEGF) is an angiogenic factor involved in tumor growth and metastasis. Gremlin has been proposed as a novel therapeutic pathway for the treatment of renal inflammatory diseases, acting via VEGFR 2 receptor. To date, most FDA-approved tyrosine kinase (TK) inhibitors have been reported as dual inhibitors of EGFR and VEGFR 2. The aim of the present study was to find the potent and selective inhibitor of VEGFR 2 specifically for the treatment of renal cancer. Fourteen previously identified anti-inflammatory compounds i.e., 1, 3, 4 oxadiazoles derivatives by our own group were selected for their anti-cancer potential, targeting the tyrosine kinase (TK) domain of VEGFR2 and EGFR. A detailed virtual screening-based study was designed viz density functional theory (DFT) study to find the compounds' stability and reactivity, molecular docking for estimating binding affinity, SeeSAR analysis and molecular dynamic simulations to confirm protein ligand complex stability and ADMET properties to find the pharmacokinetic profile of all compounds. The DFT results suggested that among all the derivatives, the 7g, 7j, and 7l were chemically reactive and stable derivatives. The optimized structures obtained from the DFTs were further selected for molecular docking, and the results suggested that 7g, 7j and 7l derivatives as the best inhibitors of VEGFR 2 with binding energy values -46.32, -48.89 and -45.01 kJ/mol. The Estimated inhibition constant (IC50) of hit compound 7j (0.009 µM) and simulation studies of its complexes confirms its high potency and best inhibitor of VEGFR2. All the derivatives were also docked with EGFR, where they showed weak binding energies and poor interactions, important compound 7g, 7j and 7i exhibited binding energy of -31.01, -33.23 and -34.19 kJ/mol respectively. Furthermore, the anticancer potential of the derivatives was confirmed by cell viability (MTT) assay using breast cancer and cervical cancer cell lines. At the end, the results of ADMET studies confirmed these derivatives as drug like candidates. Conclusively, the current study suggested substituted oxadiazoles as the potential anticancer compounds which exhibited more selectivity towards VEGFR2 in comparison to EGFR. Therefore, the identified lead molecules can be used for the synthesis of more potent derivatives of VEGFR2, along with extensive in vitro and in vivo experiments, that can be used to treat various cancers, especially renal cancers, and to prevent angiogenesis due to aberrant expression of VEGFR2.

Spectrum of EGFR mutation and its relation with high-risk predictors in thyroid cancer in Kashmiri population: 2 years prospective study at a tertiary care hospital

BackgroundEGFR mutation has not been extensively studied in thyroid cancer. This study was conducted to study spectrum of EGFR mutation in thyroid cancer in Kashmiri population for possible therapeutic purpose.MethodsIt was 2 years prospective cross-sectional study conducted at a tertiary care center in which histologically confirmed, untreated thyroid cancers were included. These specimens were subjected to EGFR mutation analysis by AS-PCR method.ResultsThere were a total 60 patients with preponderance of females [44(73%) vs 16(27%)]. Most were in the age group of less than 45 years (75%). Most of these patients were non-smokers [50(83.3%) vs 10 (17.3%)]. Papillary thyroid carcinoma (PTC) was the commonest type 48(80%), rest was follicular type (FTC) 12(20%). Well-differentiated carcinoma (WDC) was common than poorly differentiated (PDC) [41(68.4%) vs 19 (31.6%)]. Lymph node metastasis and vascular invasion were present in 32 (53.4%) and 17 (28.4%) respectively. Thirty-two (53.3%) patients were having 15 bp deletion in exon 19 of EGFR. These deletions were common in PTC than FTC, 29(60.5%) vs 3(25%) which was statistically significant (p = 0.04, CI = 0.2). The total mutational rate of T790M in EGFR tyrosine kinase domain (exon 20) was found to be only 8.4% (5 of 60). Only 4 (8.3%) of these mutations were detected in PTC and rest in FTC (1 of 12). Twenty-six (43.3%) of exon 21 were positive for L858R mutation in EGFR tyrosine kinase domain. Married persons and PDC were significant predictors of L858R mutation in EGFR tyrosine kinase domain in thyroid cancer as this was statistically significant in them with p = 0.04, 0.03 respectively.ConclusionIn our population, PTC is common in females with half of population harboring EGFR mutation and it is statistically significant in poorly differentiated carcinoma and in married individuals.It implies that EGFR may be used in thyroid cancer as a possible therapeutic agent in our set of population.

EP16.03-038 Single-cell Analyses Reveal Tumor Microenvironment Differences between EGFR 19del and L858R mutations in Lung Adenocarcinoma

Lung cancer is the leading cause of cancer-related mortality among malignant tumors worldwide. Lung adenocarcinoma (LUAD) accounts for 40% of all cases and is the most common histological subtype of non-small cell lung cancer (NSCLC). Activating somatic mutations, mostly identified as 19del and L858R in EGFR tyrosine kinase domain, may predict response to EGFR tyrosine kinase inhibitors (TKIs) in NSCLC patients. Although EGFR 19del and L858R patients are sensitive to TKIs, they showed different clinical benefits such as prolonged PFS in 19del patients.

Guidelines for HTRF technology in EGFR kinase assay

AbstractEpidermal growth factor receptor (EGFR) is among the first discovered transmembrane receptor tyrosine kinases. Ligand binding to the extracellular domain of EGFR results in the formation of homo‐dimeric or hetero‐dimeric complexes, activation, and auto‐phosphorylation in the intracellular tyrosine kinase domain, then regulates the cell's proliferation, differentiation, migration, and invasion. It has been reported that EGFR is overexpressed, amplified, or mutated in several epithelial tumors. EGFR tyrosine kinase inhibitors (EGFR‐TKIs) specifically bind to the tyrosine kinase domain of EGFR. Homogeneous time‐resolved fluorescence (HTRF) assay is successfully used in kinase assays and is a crucial tool in the drug discovery of various kinases. Moreover, the HTRF assay has been determined to exhibit many advantages, such as being easy to miniaturize, simple to read, and excellent to repeat. In these guidelines, the theory, materials, and reagent preparation of HTRF kinase assay are described and emphasized. Furthermore, the protocol and optimization of the assay are determined to establish an HTRF screening platform, then an optimized HTRF kinase assay is performed to test the compounds for screening potential EGFR‐TKIs. In addition, these guidelines can contribute to verifying the high‐throughput screening results of HTRF and help to reveal the underlying mechanism of the EGFR kinase and tested compounds.

Mutations in the helix αC of the catalytic domain from the EGFR affect its activity in cervical cancer cell lines.

The EGFR is a protein that belongs to the ErbB family of tyrosine kinase receptors. The EGFR is often overexpressed in human carcinomas. Amplification of the EGFR gene and mutations in the EGFR tyrosine kinase domain occur in patients with cancer. In cervical cancer, the expression level of the EGFR protein appears to directly associate with human papillomavirus infection. Our previous research demonstrated that in the cervical cancer cell lines, CALO and INBL, the EGFR is non-phosphorylated. The aim of the current study was to analyze the catalytic activity of the isolated EGFR and the presence of mutations in the control region αC. Catalytic activity was assessed by a universal in vitro kinase assay using polyGluTyr as a substrate, and the proteins were visualized by western blotting. For mutation analysis, DNA from CALO and INBL cell lines was isolated, and PCR was used to amplify the exons corresponding to the helix αC in the EGFR. The PCR products were visualized by agarose gel electrophoresis. The bands were isolated using a Zymoclean Gel DNA Recovery kit and directly sequenced. The EGFR, which was isolated and analyzed using the in vitro kinase assay, had catalytic activity. The receptor contained some mutations in the helix αC of the catalytic domain in both cell lines. The observed changes in the amino acid sequence may induce a different spatial arrangement and, therefore, a different conformation, which may confer different activities to this receptor. Thus, it was concluded that non-phosphorylated EGFR has catalytic activity, and it bears some amino acid changes in the helix αC of the catalytic domain in the CALO and INBL cells. These results suggest that the EGFR may function as an activator of other ErbB family receptors in these cervical cancer cells.

Identification of tripeptides against tyrosine kinase domain of EGFR for lung cancer cell inhibition by in silico and in vitro studies

Epidermal growth factor receptor tyrosine kinase domain (EGFR-TK) has been one of the prominent targets for therapeutics of several human cancers, in particular non-small cell lung cancer. Although several small chemical compounds targeting EGFR-TK have been approved by FDA for treatment of such a cancer, the discovery of a new class of EGFR-TK inhibitors, for example, small peptides, is still desired. In this study, using molecular docking-based virtual screening, we selected five small peptides with high docking scores from eight thousand peptides as candidate compounds against EGFR-TK. Among five, the tripeptide WFF had the most potency to suppress the survival of non-small cell lung cancer cells but had the least toxicity to human liver cancer cells. Our in vitro kinase assays showed that WFF exhibited much lower inhibitory activity against purified EGFR-TK than the drug erlotinib (i.e., IC50 values of≈0.62μM vs≈7.57nM, respectively). The relative free binding energies estimated from molecular dynamic simulations were consistent with the in vitro experiments in which the WFF bound had a lower affinity than erlotinib bound to EGFR-TK (i.e., ΔGbind values of -20.3kJ/mol vs≈-126.8kJ/mol, respectively). In addition, the simulation analyses demonstrated the difference in EGFR binding preference between the drug and tripeptide in which erlotinib was stably bound in the ATP-binding pocket for 4-anilinoquinazoline class of inhibitors, while WFF moved out of that pocket to interact with polar amino acid residues on the αC-helix, activation loop, and substrate-binding region. Our findings suggest preferable interactions of the potential tripeptide on enzyme inhibition that are useful for further development of a new class of inhibitors targeting EGFR-TK.

EGFR Aggregation in the Brain

Recent findings showed that preformed fibrils (PFFs) of misfolded proteins, including α-synuclein (α-syn) and amyloid-β (Aβ), activate EGFR in cell cultures and animal models of amyloid propagation. Comparing these supramolecular structures to normal EGFR ligands such as EGF and HB-EGF suggests that these PFFs might trigger the formation of high order clustering of EGFR that stimulates the aggregation of EGFR tyrosine kinase domain (EGFR-TKD) which is known to form fibrils. Subsequently, self-assembled fibril of EGFR-TKDs itself can serve as a seed to induce aggregation of monomeric forms of misfolded proteins in cytoplasm or endosomes. In this model, EGFR serves as an amyloidogenic receptor to facilitate (1) cellular uptake of exogenous PFFs and (2) seeding of endogenous misfolded proteins.

Pyrimidine-based EGFR TK inhibitors in targeted cancer therapy.

Despite significant improvements of new treatment options, cancer continues to represent as one of the most common and fatal disease. The EGFR signaling pathway is considered as a significant approach in targeted therapy of cancers. Blocking the EGFR-driven pathway by inhibiting the intracellular tyrosine kinase domain of EGFR have shown considerable improvement in cancer therapy. In an effort to identify EGFR tyrosine kinase inhibitors (TKI), several small molecules especially pyrimidine containing derivatives have been designed by applying molecular simulation and evaluated the emergence of epigenetic mutation and resistance problems restricted the long-term effectiveness of such medication and explained the need for further investigations in this field. In recent years, the studies have been focused on genetic alterations on EGFR tyrosine kinase domain, which led to the design and synthesis of more selective and effective inhibitors. Herein, we give an overview of the importance and status of EGFR inhibitors in cancer therapy. In addition, we provide an update of the recent advances in design, discovery and development of novel pyrimidine containing compounds as promising selective EGFR TK inhibitors.

PINK1-Mediated Inhibition of EGFR Dimerization and Activation Impedes EGFR-Driven Lung Tumorigenesis.

EGFR is established as a driver of lung cancer, yet the regulatory machinery underlying its oncogenic activity is not fully understood. PTEN-induced kinase 1 (PINK1) kinase is a key player in mitochondrial quality control, although its role in lung cancer and EGFR regulation is unclear. In this study, we show that PINK1 physically directly interacts with EGFR via the PINK1 C-terminal domain (PINK1-CTD) and the EGFR tyrosine kinase domain. This interaction constituted an endogenous steric hindrance to receptor dimerization and inhibited EGFR-mediated lung carcinogenesis. Depletion of PINK1 from lung cancer cells promoted EGFR dimerization, receptor activation, EGFR downstream signaling, and tumor growth. In contrast, overexpression of PINK1 or PINK1-CTD suppressed EGFR dimerization, activation, downstream signaling, and tumor growth. These findings identify key elements in the EGFR regulatory cascade and illustrate a new direction for the development of anti-EGFR therapeutics, suggesting translational potential of the PINK1-CTD in lung cancer. SIGNIFICANCE: This study identifies PINK1 as a critical tumor suppressor that impedes EGFR dimerization and highlights PINK1-CTD as a potential therapeutic agent in EGFR-driven lung cancer.

Synthesis and anticancer potential of novel 5,6-oxygenated and/or halogenated steroidal d-homo lactones

A series of 5,6-modified steroidal d-homo lactones, comprising of halogenated and/or oxygenated derivatives, was synthesized and evaluated for potential anticancer properties. Preparation of many of these compounds involved investigating alternative synthetic pathways. In silico ADME testing was performed for both novel and some previously synthesized compounds. Calculated physicochemical properties were in accordance with the Lipinski, Veber, Egan, Ghose and Muegge criteria, suggesting the potential of these molecules as orally active agents. Cytotoxicity of the synthesized steroid derivatives was tested on six tumor and one normal human cell line. None of the investigated derivatives was toxic to non-cancerous MRC-5 control cells. Most of the compounds showed significant cytotoxicity against the treated cancer cell lines. Most notably, the 3β,5α,6β-trihydroxy derivative exhibited strong cytotoxicity against multiple cell lines (MCF-7, MDA-MB-231 and HT-29), with the highest effect observed for lung adenocarcinoma (A549) cells, for which this steroid was more cytotoxic than all of the three commercial chemotherapeutic agents used as reference compounds. Molecular docking suggests the 3β,5α,6β-trihydroxy derivative could bind the EGFR tyrosine kinase domain with high affinity, providing a potential mechanism for its cytotoxicity via inhibition of EGFR signaling. The most active compounds were further studied for their potential to induce apoptosis by the double-staining fluorescence method; where the 5α,6β-dibromide, 5α,6β-dichloride and 3β,5α,6β-triol induced apoptotic changes in all three treated cell lines: MDA-MB-231, HT-29 and A549. To predict interactions with nuclear steroidal receptors, affinity for the ligand binding domains of ERα, ERβ and AR was measured using a yeast-based fluorescence assay. The 5β,6β-epoxide, dibromide and 5α-hydroxy-3,6-dioxo derivatives showed affinity for ERα, while the 5α-fluoro-6β-hydroxy and 3β-acetoxy-5α,6β-dihydroxy derivatives were identified as ERβ ligands. None of the tested compounds showed affinity for AR. Structure-activity relationships of selected compounds were also examined.

Caractérisation moléculaire de l’EGFR dans les cancers bronchiques non à petites cellules : étude prospective comparative des technologies NGS et automate Idylla

Detection of genetic alterations in the EGFR tyrosine kinase domain is a major concern in the management of non-small cell lung cancer because it conditions access to tyrosine kinase inhibitors. In practice, it is possible to characterize only well-documented mutations or to sequence all relevant EGFR exons and also other targets of theranostic interest. This prospective study compares the targeted EGFR characterization on Idylla platform (Biocartis) and a more extensive one by next generation sequencing using Ion Torrent technology. A total of 100 formalin-fixed paraffin-embedded tumour samples were tested simultaneously by both techniques under the conditions recommended by the suppliers. The comparison covered all technical and practical aspects of the laboratory. At least one EGFR mutation of interest for tyrosine kinase inhibitors for 9 and 7 samples was detected respectively by sequencing and by the Idylla system. For three samples, EGFR sensitive mutations to tyrosine kinase inhibitors were detected only by next-generation sequencing. In addition, for 37 samples, mutations of clinical interest outside EGFR were characterized by sequencing and communicated to the prescriber. Idylla technology allows the rapid characterization of a majority of EGFR variants. The result can be optimized by careful analysis of the amplification curves with the Idylla Explore tool or by increasing the amount of initial material. A complementary new generation sequencing analysis for non-contributory results by Idylla should also be recommended.