Dual Inhibition Of EGFR Research Articles

Design, synthesis and bioevaluation of dual EGFR-PI3Kα inhibitors for potential treatment of NSCLC

Aberrant activation or mutation of the EGFR-PI3K-Akt-mTOR signaling pathway has been implicated in a wide range of human cancers, especially non-small-cell lung cancer (NSCLC). Thus, dual inhibition of EGFR and PI3K has been investigated as a promising strategy to address acquired drug resistance resulting from the use of tyrosine kinase inhibitors. A series of dual EGFR/PI3Kα inhibitors was synthesized using pharmacophore hybridization of the third-generation EGFR inhibitor olmutinib and the PI3Kα selective inhibitor TAK-117. The optimal compound 30k showed potent kinase inhibitory activities with IC50 values of 3.6 and 30.0 nM against EGFRL858R/T790M and PI3Kα, respectively. Compound 30k exhibited a significant antiproliferative effect in NCI-H1975 cells with a higher selectivity profile than olmutinib. The potential antitumor mechanism, molecular binding modes, and in vitro metabolic stability of compound 30k were also clarified.

Dual inhibition of EGFR and PI3K with a single drug.

Dual inhibition of EGFR and PI3K with a single drug.

Probing Dual Covalent Irreversible Inhibition of EGFR/FGFR4 by Electrophilic-Based Natural Compounds to Overcome Resistance and Enhance Combination Therapeutic Potentials and Management of Hepatocellular Carcinoma (HCC).

Hepatocellular carcinoma (HCC) is one of the most prevalent cancer types in the world and accounts for the majority of cases of primary liver cancer. A crucial part of the carcinogenesis of HCC involves aberrant stimulation of the FGF19-FGFR4 signaling pathway. Therefore, FGFR4 inhibition has become a strategic therapeutic approach for the treatment of HCC. However, the clinical treatment procedure is significantly hampered by the prevalence of kinase inhibitors resistance. It was recently established that the activation of EGFR signaling was found to be one of the primary mechanisms mediating the acquired resistance to FGFR4 inhibitors, moreover, sensitivity to FGFR4 inhibitors was effectively restored by inhibiting EGFR. These results provide compelling evidence that dual inhibition of EGFR and FGFR4 could represent a viable therapeutic approach to overcome resistance, hence enhanced management of HCC. To this end, we proposed a dual irreversible inhibition strategy through covalent binding by naturally occurring electrophilic warhead-bearing compounds (curcumin, deoxyelephantopin, eupalmerin acetate, syringolin A and andrographolide) to covalently target both EGFR and FGFR4 through cysteine residues, Cys797 and Cys552, respectively. Covalent docking and covalent molecular dynamics (MM/MDcov) simulations combined with thermodynamic binding free energy calculations were performed, and the results were compared against known potent and selective covalent EGFR and FGFR4 inhibitors with available X-ray crystal structures, Afatinib and BLU9931, respectively. Curcumin, deoxyelephantopin, eupalmerin acetate, syringolin A, and andrographolide showed relative binding free energies of -22.85, -17.14, -12.98, -21.81, and - 19.00kcal/mol against EGFR and - 41.06, -29.45, -24.76, -40.11, and - 37.55kcal/mol against FGFR4, respectively. The mechanisms of binding were emphasized by hydrogen bonding and binding forces analysis as well as active site physicochemical profiling. The findings of this study identified that curcumin, syringolin A and andrographolide-but not eupalmerin acetate or deoxyelephantopin -could be viable dual EGFR and FGFR4 covalent irreversible inhibitors and could be implemented in HCC combination therapy protocols alone or in conjunction with other chemotherapeutic agents. Investigations of this study conclusively indicate dual blockade of EGFR and FGFR4 may be a promising future therapeutic strategy for enhanced management of HCC.

Design, synthesis, and mechanistic evaluation of novel benzimidazole-hydrazone compounds as dual inhibitors of EGFR and HER2: Promising candidates for anticancer therapy

A series of twelve benzimidazole-hydrazone compounds were synthesized and evaluated for their cytotoxicity on three cancer cell lines (HepG-2, MCF-7, and HCT-116) using the MTT assay. The compounds' IC50 values were compared to the anticancer drug sunitinib. Additionally, we investigated the dual inhibition of compounds 6e and 6j on EGFR and HER2 tyrosine kinase enzymes. Flow cytometry analysis revealed that compound 6e induced cell cycle arrest and apoptosis in HepG-2 cells. Molecular docking analysis showed that compound 6e interacted with the ATP binding pocket of EGFR and HER2, exhibiting hydrophobic interactions and distinct hydrogen bonding patterns. Notably, compounds 6e, 6g, 6h, 6j, and 6l exhibited potent cytotoxicity comparable to sunitinib against all tested cancer cell lines. Compound 6e demonstrated dual inhibition of EGFR and HER2 and displayed favorable druglike properties. These findings suggest that compound 6e has potential as a lead compound for targeting the EGFR and HER2 pathways and warrant further optimization and development as a potential anticancer therapeutic agent.

Benzimidazole-Based Derivatives as Apoptotic Antiproliferative Agents: Design, Synthesis, Docking, and Mechanistic Studies.

A new class of benzimidazole-based derivatives (4a-j, 5, and 6) with potential dual inhibition of EGFR and BRAFV600E has been developed. The newly synthesized compounds were submitted for testing for antiproliferative activity against the NCI-60 cell line. All newly synthesized compounds 4a-j, 5, and 6 were selected for testing against a panel of sixty cancer cell lines at a single concentration of 10 µM. Some compounds tested demonstrated remarkable antiproliferative activity against the cell lines tested. Compounds 4c, 4e, and 4g were chosen for five-dose testing against 60 human tumor cell lines. Compound 4c demonstrated strong selectivity against the leukemia subpanel, with a selectivity ratio of 5.96 at the GI50 level. The most effective in vitro anti-cancer assay derivatives (4c, 4d, 4e, 4g, and 4h) were tested for EGFR and BRAFV600E inhibition as potential targets for antiproliferative action. The results revealed that compounds 4c and 4e have significant antiproliferative activity as dual EGFR/BRAFV600E inhibitors. Compounds 4c and 4e induced apoptosis by increasing caspase-3, caspase-8, and Bax levels while decreasing the anti-apoptotic Bcl2 protein. Moreover, molecular docking studies confirmed the potential of compounds 4c and 4e to act as dual EGFR/BRAFV600E inhibitors.

Development of new thieno[2,3-d]pyrimidines as dual EGFR and STAT3 inhibitors endowed with anticancer and pro-apoptotic activities

In part due to the resilience of cellular feedback pathways that develop therapeutic resistance to targeting the EGFR alone, using EGFR inhibitors alone was demonstrated to be unsuccessful in clinical trials. The over-activation of the signal transducer/activator of transcription 3 (STAT3) during the administration of an EGFR inhibitor is expected to play a substantial part in the failure and resistance of EGFR inhibitor treatment. Therein, we proposed a hypothesis that induced STAT3-mediated resistance to EGFR inhibition therapy could be addressed by a dual inhibition of EGFR and STAT3 method. To this end, we tried to discover new thieno[2,3-d]pyrimidine derivatives “5a-o”. Results from the screening on A549 and MCF7 cancer cell lines revealed that compounds 5j and 5k showed two-digit nanomolar with appropriate safety towards the WI-38 cell line. The best molecules, 5j and 5k, were subjected to γ-radiation, and their cytotoxic efficacy didn’t change after irradiation, demonstrating that not having to use it avoided its side effects. Compounds 5j and 5k demonstrated the highest inhibition when their potency was tested as dual inhibitors on EGFR 67 and 41 nM, respectively, and STAT3 5.52 and 3.34 nM, respectively, proved with in silico molecular docking and dynamic simulation. In light of the results presented above, the capacity of both powerful compounds to alter the cell cycle and initiate the apoptotic process in breast cancer MCF7 cells was investigated. Caspase-8, Bcl-2, Bax and Caspase-9 apoptotic indicators were studied.

Design, Synthesis, Antiproliferative Actions, and DFT Studies of New Bis-Pyrazoline Derivatives as Dual EGFR/BRAFV600E Inhibitors.

Some new Bis-pyrazoline hybrids 8-17 with dual EGFR and BRAFV600E inhibitors have been developed. The target compounds were synthesized and tested in vitro against four cancer cell lines. Compounds 12, 15, and 17 demonstrated strong antiproliferative activity with GI50 values of 1.05 µM, 1.50 µM, and 1.20 µM, respectively. Hybrids showed dual inhibition of EGFR and BRAFV600E. Compounds 12, 15, and 17 inhibited EGFR-like erlotinib and exhibited promising anticancer activity. Compound 12 is the most potent inhibitor of cancer cell proliferation and BRAFV600E. Compounds 12 and 17 induced apoptosis by increasing caspase 3, 8, and Bax levels, and resulted in the downregulation of the antiapoptotic Bcl2. The molecular docking studies verified that compounds 12, 15, and 17 have the potential to be dual EGFR/BRAFV600E inhibitors. Additionally, in silico ADMET prediction revealed that most synthesized bis-pyrazoline hybrids have low toxicity and adverse effects. DFT studies for the two most active compounds, 12 and 15, were also carried out. The values of the HOMO and LUMO energies, as well as softness and hardness, were computationally investigated using the DFT method. These findings agreed well with those of the in vitro research and molecular docking study.

Brief Report: Combination of Osimertinib and Dacomitinib to Mitigate Primary and Acquired Resistance in EGFR-Mutant Lung Adenocarcinomas.

Primary and acquired resistance to osimertinib remain significant challenges for patients with EGFR-mutant lung cancers. Acquired EGFR alterations such as EGFR T790M or C797S mediate resistance to EGFR tyrosine kinase inhibitors (TKI) and combination therapy with dual EGFR TKIs may prevent or reverse on-target resistance. We conducted two prospective, phase I/II trials assessing combination osimertinib and dacomitinib to address on-target resistance in the primary and acquired resistance settings. In the initial therapy study, patients received dacomitinib and osimertinib in combination as initial therapy. In the acquired resistance trial, dacomitinib with or without osimertinib was administered to patients with EGFR-mutant lung cancers with disease progression on osimertinib alone and evidence of an acquired EGFR second-site mutation. Cutaneous toxicities occurred in 93% (any grade) of patients and diarrhea in 72% (any grade) with the combination. As initial therapy, the overall response to the combination was 73% [95% confidence interval (CI), 50%-88%]. No acquired secondary alterations in EGFR were observed in any patients at progression. In the acquired resistance setting, the overall response was 14% (95% CI, 1%-58%). We observed no acquired secondary EGFR alterations with dual inhibition of EGFR as up-front treatment, but this regimen was associated with greater toxicity. The combination was not effective in reversing acquired resistance after development of a second-site acquired EGFR alteration. Our study highlights the need to develop better strategies to address on-target resistance in patients with EGFR-mutant lung cancers.

Dual EGFR-VEGF Pathway Inhibition: A Promising Strategy for Patients With EGFR-Mutant NSCLC

The VEGF pathway has been recognized as a key mediator of angiogenesis to support tumorigenesis. Multiple therapeutic agents targeting VEGF and VEGF receptors have been developed and approved for use in NSCLCs. Preclinical studies have found that the VEGF and EGFR pathways share common downstream signaling, and these pathways can function exclusively of one another during oncogenesis. In EGFR-mutant NSCLCs, up-regulated EGFR signaling increases VEGF through hypoxia-independent mechanisms, and elevated VEGF, in turn, contributes to the emergence of resistance to EGFR tyrosine kinase inhibitors (TKIs). In clinical trials, the addition of anti-VEGF therapy to EGFR TKIs considerably improved clinical outcomes. In recently reported large randomized studies, the addition of bevacizumab or ramucirumab to EGFR TKIs substantially improved progression-free survival in patients with TKI-naive EGFR-mutant NSCLC. This article reviews the preclinical and clinical data supporting dual inhibition of EGFR and VEGF in EGFR-mutant NSCLC as a way to improve patient outcomes.

Concurrent osimertinib plus gefitinib for first-line treatment of EGFR-mutated non-small cell lung cancer (NSCLC).

9507 Background: First-line treatment with an EGFR tyrosine kinase inhibitor (TKI) is standard of care for patients (pts) with EGFR-mutated NSCLC. The EGFR TKI osimertinib is active against the acquired gefitinib-resistant mutation EGFR T790M, as is gefitinib against the osimertinib-resistant EGFR C797S. Preclinical evidence suggests dual EGFR inhibition with gefitinib + osimertinib may delay emergence of acquired resistance. Methods: This ongoing phase I/II study enrolled pts with stage IV EGFR-mutated (L858R or del19) NSCLC, without prior therapy for metastatic disease. Treatment in dose escalation (n = 6): concurrent osimertinib 40 mg or 80 mg + gefitinib 250 mg daily. In dose expansion (n = 21): osimertinib + gefitinib at the maximum tolerated dose (MTD). Prior to protocol amendment 6 pts received alternating monthly cycles of TKI monotherapy and were excluded from this analysis. The primary endpoints in the dose escalation and expansion phases were, respectively, identification of the MTD and feasibility, defined as receipt of combination therapy for ≥ 6 four-week cycles. Secondary endpoints included overall response rate (ORR), survival outcomes, plasma EGFR mutation clearance (cell free DNA by droplet digital PCR (ddPCR)), and mechanisms of acquired resistance. Results: From May 2017 to July 2019 27 pts were enrolled and evaluable for the primary endpoints. The MTD was osimertinib 80 mg plus gefitinib 250 mg orally daily. In feasibility analysis, 81.5% completed ≥6 cycles combination therapy (1 pt discontinued for progression, 4 for toxicity). The ORR was 85.2% (95% CI 67.5%-94.1%). Best response: 85.2% partial response, 14.8% stable disease. The most common treatment-related adverse effects (TRAEs) (% any grade, % grade 3) were rash (96.3%, 3.7%), diarrhea (85.2%, 11.1%) and dry skin (70.4%, 0%). Plasma ddPCR (n = 25 pts) detected the driver EGFR mutation at baseline in 68% of pts. In these pts, plasma EGFR cleared to undetectable at 2 weeks treatment in 82.4%. At 14.8 months median follow up the median progression free survival was not yet reached. Conclusions: Combination therapy with osimertinib and gefitinib is tolerable for first-line treatment of EGFR-mutated NSCLC and resulted in rapid plasma clearance of the EGFR mutation. The observed ORR is consistent with previously reported first-line response rates to osimertinib. Analysis of survival outcomes and acquired resistance mechanisms are pending data maturity and will facilitate understanding of the role of first-line dual EGFR TKI therapy for this pt population. Clinical trial information: NCT03122717 .

Combined targeting of EGFR and HER2 against prostate cancer stem cells

ABSTRACTProgression of prostate cancer has been associated with EGFR and HER2 activation and to tumor-initiating cells contribution toward chemotherapy resistance. We investigated the efficacy of a dual intervention against EGFR and HER2 to deplete the tumor-initiating cells, optimize the chemotherapy management and prevent the progression of castration-resistant prostate cancer (CRPC) cells. Using DU145, PC3, and 22Rv1 CRPC cell lines, biochemical analysis revealed activation of EGFR, HER2, MAPK, and STAT3 in DU145 and 22Rv1, and AKT and SRC in DU145 and PC-3. pSTAT3 nuclear staining was observed in DU145 xenografts and in 12 out of 14 CRPC specimens. The in vivo dual targeting of ErbB receptors with Cetuximab and Trastuzumab combined with chemotherapy caused an effective antitumor response in DU145 xenografted mice displaying STAT3 activation; conversely PC-3 bearing mice experienced tumor relapse. The potentiating of in vivo cytotoxic effect in DU145 model was accompanied by a significant decrease of prostatosphere-forming capacity assessed in vitro on residual tumor cells. Additionally, combined treatment in vitro with Cetuximab, Trastuzumab and chemotherapy negatively affected DU145 and 22Rv1 sphere formation, suggesting the critical function of ErbB receptors for tumor-initiating cells proliferation; no effect on PC-3 clonogenic potential was observed, indicating that other receptors than EGFR and HER2 may sustain PC3 tumor-initiating cells. These findings provided the preclinical evidence that the dual inhibition of EGFR and HER2 by targeting tumor-initiating cells may improve the efficacy of the current chemotherapy regimen, bringing benefits especially to castration-resistant patients with activated STAT3, and preventing disease progression.

Senesce to Survive: YAP-Mediated Dormancy Escapes EGFR/MEK Inhibition

Therapeutic resistance is a major challenge in cancer treatment. In this issue of Cancer Cell, Kurppa etal. demonstrated that a senescence-like state enables lung cancer cells to survive dual inhibition of EGFR and MEK. This was mediated by the YAP/TEAD pathway, which drives epigenomic reprogramming and EMT to counteract apoptosis.

1,2,4-Oxadiazole derivatives targeting EGFR and c-Met degradation in TKI resistant NSCLC

Development of small-molecule agents with the ability to facilitate oncoprotein degradation has emerged as a promising strategy for cancer therapy. Since EGFR and c-Met are both implicated in oncogenesis and tumor progression, we initiated a screening program by using an in-house library to identify agents capable of inducing the concomitant suppression of EGFR and c-Met expression, which led to the identification of compound 1, a 1,2,4-oxadiazole derivative. Based on the scaffold of 1, we developed a series of derivatives to assess their efficacies in facilitating the downregulation of EGFR and c-Met, among which compound 48 represented the optimal agent. 48 showed equipotent antiproliferative activity against a panel of five NSCLC cell lines with different EGFR mutational status (IC50 = 0.2–0.6 μM), while the same panel exhibited differential sensitivity to different EGFR kinase inhibitors tested. Cell cycle analysis indicated that the antiproliferative activity of 48 was associated with its ability to cause G2/M arrest and, to a lesser extent, apoptosis. Western blot and RT-PCR analyses revealed that 48 facilitated the downregulation of EGFR and c-Met at the protein level. In vivo data showed that oral administration of 48 was effective in suppressing gefitinib-resistant H1975 xenograft tumor growth in nude mice, and at a suboptimal dose, could sensitize H1975 tumors to gefitinib. Based on these findings, 48 represents a promising candidate for further development to target EGFR TKI-resistant NSCLC via dual inhibition of EGFR and c-Met oncoproteins.

Licochalcone B inhibits growth and induces apoptosis of human non-small-cell lung cancer cells by dual targeting of EGFR and MET

BackgroundEpidermal growth factor receptor (EGFR) gene alterations are associated with sensitization to tyrosine kinase inhibitors such as gefitinib in lung cancer. Some patients suffering from non-small cell lung cancer (NSCLC) have difficulty in treating the cancer due to resistance acquired to gefitinib with MET amplification. Therefore EGFR and MET may be attractive targets for lung cancer therapy. PurposeThis study aimed to investigate the anti-cancer activity of Licochalcone (LC)B extracted from Glycyrrhiza inflata, in gefitinib-sensitive or gefitinib-resistant NSCLC cells, and to define its mechanisms. Study designWe investigated the mechanism of action of LCB by targeting EGFR and MET in human NSCLC cells. MethodsWe used the HCC827 and HCC827GR lines as gefitinib-sensitive and –resistant cells respectively, and determined the effects of LCB on both, by performing cell proliferation assay, flow cytometry analysis and Western blotting. Targets of LCB were identified by pull-down/kinase assay and molecular docking simulation. ResultsLCB inhibited both EGFR and MET kinase activity by directly binding to their ATP-binding pockets. The ability of this interaction was verified by computational docking and molecular dynamics simulations. LCB suppressed viability and colony formation of both HCC827 and HCC827GR cells while exhibiting no cytotoxicity to normal cells. The induction of G2/M cell-cycle arrest and apoptosis by LCB was confirmed by Annexin V/7-AAD double staining, ER stress and reactive oxygen species induction, mitochondrial membrane potential loss and caspase activation as well as related-proteins regulation. Inhibition of EGFR and MET by LCB decreased ERBB3 and AKT axis activation. ConclusionWe provide insights into the LCB-mediated mechanisms involved in reducing cell proliferation and inducing apoptosis in NSCLC cells. This occurs through dual inhibition of EGFR and MET in NSCLC cells regardless of their sensitivity or resistance to gefitinib. LCB may be a promising novel therapeutic medicine for gefitinib-sensitive or resistant NSCLC treatment.

Phase II randomized trial of afatinib with or without cetuximab as first-line treatment for EGFR mutated non-small cell lung cancer (NSCLC) patients (IFCT-1503 ACE-Lung).

9079 Background: First-line treatment of metastatic EGFR-mutated NSCLC relies on EGFR-TKIs. However, all patients (pts) eventually develop progression. Dual inhibition of EGFR with afatinib (A), an irreversible pan-erbB TKI, and cetuximab (C), an EGFR monoclonal antibody, has shown activity in EGFR-mutated pts with acquired resistance to TKIs, regardless of the T790M status. Methods: We conducted a phase II randomized trial in advanced NSCLC pts harboring an activating EGFR mutation, who had not received prior therapy. Pts were treated with A (40 mg/d) until progression alone or with C 500 mg/m² every 2 weeks during 6 months (mos) (beginning at D15 at 250 mg/m²). Primary endpoint was time-to-treatment failure (TTF) at 9 mos for pts with del19 and L858R mutations. Secondary endpoints include safety, progression-free survival (PFS), overall survival (OS). Prospective monitoring of the T790M mutation was performed on circulating tumoral DNA (ctDNA) by digital PCR. Results: Trial was stopped early due to futility analysis after 118 pts were enrolled (59 in each arm). Baseline characteristics were balanced between the 2 arms, and especially for the types of EGFR mutation (del19, 55.9 vs 50.8%; L858R, 39 vs 40.7%; others, 5.1 vs 8.5% in AC and A arms, respectively). Treatment-related AEs of any grades were similar, although there was an excess of grade 3 AEs in the AC arm (50 vs 37.3%), but no of grade 5. The excess in grade 3-5 AEs was essentially due to cutaneous (96.6 vs 81.4%), eyes (32.8 vs 27.1%), hematological (22.4 vs 15.3%) but not to digestive toxicities (89.7 vs 98.3%). Among the 117 pts included in the efficacy analysis, 9-months TTF was 63.3% (47.5-75.6) in arm A and 65.8% (50.1-77.66) in arm AC. Median TTF was 11.1 mos (8.3-not reached [NR]) and 10.8 mos (9.2-13.7) in arms A and AC, respectively. Median PFS was 11.1 mos (8.3-NR) and 12.8 mos (9.2-13.7), respectively. Median OS was 20.8 mos (17.5-NR) and NR (17-NR), respectively. Conclusions: Efficacy of AC was similar to that of A alone. These results don’t support further evaluation of this combination in this setting. Results of ctDNA monitoring will be reported during the meeting. Clinical trial information: NCT02716311.

Novel pyrazolo[3,4-d]pyrimidines: design, synthesis, anticancer activity, dual EGFR/ErbB2 receptor tyrosine kinases inhibitory activity, effects on cell cycle profile and caspase-3-mediated apoptosis

A series of novel pyrazolo[3,4-d]pyrimidines was synthesised. Twelve synthesised compounds were evaluated for their anticancer activity against 60 human tumour cell lines by NCI (USA). Compound 7d proved prominent anticancer activity. It showed 1.6-fold more potent anti-proliferative activity against OVCAR-4 cell line with IC50 = 1.74 μM. It also exhibited promising potent anticancer activity against ACHN cell line with IC50 value 5.53 μM, representing 2.2-fold more potency than Erlotinib. Regarding NCI-H460 cell line, compound 7d (IC50 = 4.44 μM) was 1.9-fold more potent than Erlotinib. It inhibited EGFR and ErbB2 kinases at sub-micromolar level (IC50 = 0.18 and 0.25 µM, respectively). Dual inhibition of EGFR and ErbB2 caused induction of apoptosis which was confirmed by a significant increase in the level of active caspase-3 (11-fold). It showed accumulation of cells in pre-G1 phase and cell cycle arrest at G2/M phase.

Arsenic promotes the COX2/PGE2-SOX2 axis to increase the malignant stemness properties of urothelial cells.

Chronic arsenic exposure is associated with the development of urothelial carcinoma of the bladder (UCB). To elucidate the contribution of arsenic exposure to urothelial cancer stem cell (CSC) generation, we established an in vitro stepwise malignant model transformed by chronically exposing human urothelial cells to arsenic. Using this model, we found that chronic arsenic exposure endows urothelial cells with malignant stemness properties including increased expression of stemness-related factors such as SOX2, sphere formation, self-renewal, invasion and chemoresistance. SOX2 was gradually and irreversibly overexpressed in line with acquired sphere-forming and self-renewal abilities. Following gene set enrichment analyses of arsenic-exposed and arsenic-unexposed cells, we found COX2 as an enriched gene for oncogenic signature. Mechanistically, arsenic-induced COX2/PGE2 increases SOX2 expression that eventually promotes malignant stem cell generation and repopulation. In urine samples from 90 subjects exposed to arsenic and 91 control subjects, we found a significant linear correlation between SOX2 and COX2 expression and the potential of SOX2 and COX2 expression as urinary markers to detect subjects exposed to arsenic. Furthermore, the combination marker yielded a high sensitivity for UCB detection in a separate cohort. Finally, our in vitro model exhibits basal-type molecular features and dual inhibition of EGFR and COX2 attenuated stem cell enrichment more efficiently than an EGFR inhibitor alone. In conclusion, the COX2/PGE2-SOX2 axis promotes arsenic-induced malignant stem cell transformation. In addition, our findings indicate the possible use of SOX2 and COX2 expression as urinary markers for the risk stratification and detection of UCB.

Strong radioprotective FGF1 signaling down-regulates proliferative and metastatic capabilities of the angiosarcoma cell line, ISOS-1, through the dual inhibition of EGFR and VEGFR pathways

Background and purposeAngiosarcoma is associated with a poor prognosis and is treated with radiotherapy. Although FGF1 is a potential radioprotector, the influence of FGF1 on the malignancy of angiosarcoma remains unknown. Materials and methodsHighly stable FGF1 mutants, which exhibit stronger mitogenic activity than wild-type FGF1, were examined as strong radioprotectors and signaling agonists to clarify the effects of FGF1 on the murine angiosarcoma cell line ISOS-1. ResultsFGF1 mutants reduced colony formation by and the in vitro invasion and migration of ISOS-1 cells, in addition to an increase in radiosensitivity to X-rays. In contrast, an FGFR inhibitor blocked the inhibitory effects of FGF1 mutants on colony formation, invasion, and migration. siRNA targeting the Fgfr1 gene showed that strong FGFR1 signaling reduced colony formation by ISOS-1 cells. However, the FGF1 mutant reduced the activation of VEGFRs and EGFRs in ISOS-1 cells more strongly than wild-type FGF1. Moreover, the inhibition of VEGFRs and EGFRs synergistically reduced colony formation by and invasion and migration of ISOS-1 cells. ConclusionThese results suggest that strong FGF1 signaling exerts not only radioprotective effects, but also inhibitory effects on proliferative and metastatic capacities of angiosarcoma through the dual inhibition of EGFR and VEGFR pathways.

Dual EGFR and BRAF blockade overcomes resistance to vemurafenib in BRAF mutated thyroid carcinoma cells

BackgroundBRAF inhibitors are effective anticancer agents in BRAF-mutated melanomas. By contrast, evidences about sensitivity of thyroid carcinomas to BRAF inhibition are conflicting and it has been proposed that BRAF V600E thyroid carcinoma cells are less sensitive to BRAF inhibitors due to activation of parallel signaling pathways. This study evaluated the hypothesis that feedback activation of EGFR signaling counteracts the cytostatic activity of vemurafenib (PLX4032) in BRAF V600E thyroid carcinoma cells.MethodsCell proliferation, cell cycle distribution, induction of apoptosis and EGFR and AKT signaling were evaluated in thyroid carcinoma cell lines bearing the BRAF V600E mutation in response to PLX4032.ResultsA partial and transient cytostatic response to PLX4032 was observed in thyroid carcinoma cell lines bearing the BRAF V600E mutation, with lack of full inhibition of ERK pathway. Interestingly, the exposure of thyroid carcinoma cells to PLX4032 resulted in a rapid feedback activation of EGFR signaling with parallel activation of AKT phosphorylation. Consistently, the dual inhibition of EGFR and BRAF, through combination therapy with PLX4032 and gefitinib, resulted in prevention of EGFR phosphorylation and sustained inhibition of ERK and AKT signaling and cell proliferation. Of note, the combined treatment with gefitinib and vemurafenib or the exposure of EGFR-silenced thyroid carcinoma cells to vemurafenib induced synthetic lethality compared to single agents.ConclusionsThese data suggest that the dual EGFR and BRAF blockade represents a strategy to by-pass resistance to BRAF inhibitors in thyroid carcinoma cells.

Abstract 35: The dual inhibition of Met and EGFR by ME22S, a novel Met/EGFR bispecific monoclonal antibody, suppresses the proliferation and invasion of laryngeal cancer

Abstract Background and purpose: It has been reported that the abnormal activation of receptor tyrosine kinases is associated with the development of many human carcinomas and the high activation of EGFR and Met mediates the tumorigenicity of laryngeal carcinoma. In this study, we have done the therapeutic efficacy of ME22S, a novel EGFR/Met bispecific antibody, in laryngeal carcinoma in vitro and in vivo was thoroughly evaluated. Methods: The effects of ME22S on cell viability was assessed through MTT assays, then Western blotting and immunocytochemistry were used to determine the expression of EGFR and Met. Also, wound healing and invasion assays were performed to observe the inhibitory effects of ME22S. Results: We found the ability of ME22S reducing the expression of both EGFR and Met and significantly inhibiting the cell migration, invasion, and proliferation of SNU899 and HN3 in vitro. Also the notably reduced levels of p-Met, p-ERK, and p-AKT were found when the cells were treated with only ME22S alone or with HGF together. Meanwhile, ME22S, interestingly enough, caused caspase-3-dependent apoptotic cell death when HN3 cells were treated with ME22S for 72h, decreased the HGF-induced Slug expression, and also inhibited the tumor growth of HN3 cells in a xenograft model in vivo. Conclusion: Taken together, our findings suggest that the dual inhibition of EGFR and Met through ME22S largely suppresses the invasion and growth of laryngeal carcinoma both in vitro and in vivo, hence, can be a practical approach as a novel therapeutic strategy for the treatment of laryngeal carcinoma. Citation Format: Bok-Soon Lee, Dae Ho Kim, Yeon Soo Kim, Jae Won Chang, Bon Seok Koo, Ho Ryun Won, Haeng-Jun Kim, Hyun-Young Cha, Ji Min Lee, Kyung-Ah Kim, Chul-Ho Kim. The dual inhibition of Met and EGFR by ME22S, a novel Met/EGFR bispecific monoclonal antibody, suppresses the proliferation and invasion of laryngeal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 35. doi:10.1158/1538-7445.AM2017-35