Dual Inhibition Of EGFR Research Articles

KRAS mutant lung cancer cells are differentially responsive to MEK inhibitor due to AKT or STAT3 activation: Implication for combinatorial approach

KRAS is frequently mutated in nonsmall cell lung cancer (NSCLC), resulting in the activation of the MAPK/ERK kinase (MEK)/ERK pathway. High-throughput mutation profile has shown that lung cancer frequently harbors comutation of cancer-related genes. Therefore, given that cancer cells have multiple genetic alterations, combinatorial therapeutic strategy is demanded for effective cancer therapy. To address this, we first characterized MEK dependence in four NSCLC cells. Two cells (H358, A549) carried KRAS mutation only, and the other two (H23, H157) harbored comutation of KRAS/PTEN. H358 cells with KRAS mutation only were sensitive to MEK inhibition. However, the other KRAS mutant A549 cells were resistant to MEK inhibition. Previously, we have shown that dual inhibition of EGFR and MEK signaling shows a synergistic effect on KRAS mutant gastric cancer cells by suppressing compensatory activation of AKT. Here we also observed that this combination was effective in KRAS mutant A549 cells. However, the combination was ineffective in H23 and 157 cells with comutation of KRAS/PTEN. Compared to KRAS mutant/PTEN wild-type cells, signal transducer and activator of transcription 3 (STAT3) was significantly activated following MEK inhibition in KRAS/PTEN comutant cells. Combined STAT3 inhibition by a JAK2 inhibitor or gene knockdown with MEK inhibition blocked STAT3 activation, synergistically suppressed cell growth, and induced apoptosis in comutant cells. Taken together, our study provides molecular insights that help explain the heterogeneous response to MEK inhibition in KRAS mutant lung cancers, and presents a rationale for the clinical investigation of combination of MEK and EGFR inhibitor or MEK and JAK2 inhibitor depending on PTEN status.

Synergistic interaction between trastuzumab and EGFR/HER-2 tyrosine kinase inhibitors in HER-2 positive breast cancer cells

Overexpression of HER-2 in breast cancer is frequently associated with expression of EGFR, and EGFR expression influences response to HER-2 inhibition. The aim of this study was to examine the effects of combining dual inhibition of EGFR and HER-2, using trastuzumab, gefitinib and lapatinib, in HER-2 overexpressing breast cancer cells. Combination proliferation assays were performed in two HER-2 positive breast cancer cell lines, SKBR-3 and BT-474. Trastuzumab combined with lapatinib was also tested in BT-474 xenografts. In proliferation assays, dual targeting with trastuzumab and gefitinib or lapatinib showed synergy or additivity in both SKBR-3 and BT-474 cells. Trastuzumab (10 nM) or gefitinib (5 µM) alone did not induce significant apoptosis, whereas lapatinib (0.75 µM) induced significant apoptosis in SKBR-3 cells. Trastuzumab combined with lapatinib further enhanced apoptosis induction. Trastuzumab (10 nM) and gefitinib (5 µM) induced apoptosis comparable to lapatinib alone (0.75 µM), suggesting that inhibition of both EGFR and HER-2 may be required to induce apoptosis in these cells. Pre-treatment with trastuzumab and gefitinib or lapatinib enhanced response to chemotherapy in vitro. The combination of trastuzumab and lapatinib also effectively blocked tumour growth in vivo. Dual targeting of EGFR and HER-2, by combining trastuzumab with EGFR/HER-2 tyrosine kinase inhibitors, may improve response in HER-2 overexpressing breast cancer cells that also express EGFR.

Abstract A75: Dual inhibition of EGFR and ErbB2 during androgen ablation prevent the progression of prostate cancer cells to androgen independence

Abstract Introduction: Patients with metastatic prostate cancer are treated with androgen ablation therapy which is initially effective, but most patients on this therapy ultimately become refractory to this treatment. Studies show that androgen withdrawal does not result in cell death, but simply cell growth arrest, whereas the development of androgen independence allows them to come out of the growth arrest and undergo cell cycle progression. Hence, induction of apoptosis during androgen ablation will prevent progression to androgen independence. The goal of the present project is to determine whether dual inhibition of EGFR and ErbB2 in prostate cancer cells during androgen withdrawal prevent their progression to androgen independence. EGFR, ErbB2, ErbB3 and ErbB4 constitute a family of receptor tyrosine kinases that require dimerization for complete activation. Experimental Procedures: Androgen dependent LNCaP prostate cancer cells and its androgen-independent subline LNCaP-AI were treated with combinations of the EGFR inhibitors AG1478 or Erlotinib (Tarceva), and the ErbB2 inhibitors AG879 or Trastuzumab (Herceptin), either in the presence of androgens, or together with androgen withdrawal. These experiments were also conducteds in androgen receptor (AR)-null pRNS-1-1 cells transfected with vector only, wild type AR or mutant AR(T877A). Results: Dual inhibition of EGFR and ErbB2 was more effective in inhibiting cell growth compared to single inhibition of either kinase. This effect was more pronounced in the absence of androgens compared to its presence, and was also more effective in androgen-dependent than in androgen-independent cells. Investigation of the mechanism of this effect demonstrated that dual inhibition of EGFR and ErbB2 also inhibited ErbB3 phosphorylation, likely by preventing its dimerization. Androgen withdrawal results in increased Akt phosphorylation, and we show here that dual EGFR/ErbB2 inhibition, but not single inhibition, prevented this effect and induced apoptosis. This effect is most pronounced in androgen sensitive cells undergoing androgen ablation, whereas in androgen-independent cells where Akt phosphorylation is already increased, the effect of the dual inhibition was less pronounced. Conclusions: Our results indicate that cell survival during androgen withdrawal caused by an increase in Akt phosphorylation may be prevented by dual use of EGFR and ErbB2 inhibitors. If the cells can be killed during androgen ablation, then no cells will remain to progress to androgen independence. Hence, the effect of the dual inhibition prevents the development of androgen-independence in prostate cancer cells. The significance of these results is that they suggest that the use of dual EGFR/ErbB2 inhibitors during androgen ablation may prevent the development of androgen independence in patients undergoing androgen ablation therapy. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):A75.

Abstract A252: KRAS mutant lung cancer cells are differentially responsive to MEK inhibition due to AKT or STAT3 activation: Implication for combinatorial approach

Abstract KRAS is frequently mutated in non-small cell lung cancer (NSCLC), resulting in the activation of the MEK/ERK pathway. High-throughput mutation profile has shown that lung cancer frequently harbors co-mutation of cancer related genes. Therefore, given that cancer cells have multiple genetic alterations, combinatorial therapeutic strategy is demanded for effective cancer therapy. To address this, we first characterized MEK dependence in four NSCLC cells. Two cells (H358, A549) carried KRAS mutation only, and the other two (H23, H157) harbored co-mutation of KRAS/PTEN. H358 cells with KRAS mutation only were sensitive to MEK inhibition. However, the other KRAS mutant A549 cells were resistant to MEK inhibition. Previously, we have shown that dual inhibition of EGFR and MEK signaling shows a synergistic effect on KRAS mutant gastric cancer cells by suppressing compensatory activation of AKT. Here we also observed that this combination was effective in KRAS mutant A549 cells. However, the combination was ineffective in H23 and 157 cells with co-mutation of KRAS/PTEN. Compared to KRAS mutant/PTEN wild-type cells, STAT3 was significantly activated following MEK inhibition in KRAS/PTEN co-mutant cells. Combined STAT3 inhibition by a JAK2 inhibitor or gene knockdown with MEK inhibition blocked STAT3 activation, synergistically suppressed cell growth, and induced apoptosis in co-mutant cells. Taken together, our study provides molecular insights that help explain the heterogeneous response to MEK inhibition in KRAS mutant lung cancers, and presents a rationale for the clinical investigation of combination of MEK and EGFR inhibitor or MEK and STAT3 inhibitor depending on PTEN status. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):A252.

Lapatinib, a Dual EGFR and HER2 Tyrosine Kinase Inhibitor, Downregulates Thymidylate Synthase by Inhibiting the Nuclear Translocation of EGFR and HER2

BackgroundEpidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI) has been shown to exert a synergistic antitumor effect when combined with fluoropyrimidine. This synergy may be attributable to the downregulation of thymidylate synthase (TS), which is frequently overexpressed in fluoropyrimidine-resistant cancer cells. However, the molecular mechanism underlying the downregulation of TS has yet to be clearly elucidated.Methodology and Principal FindingsIn this study, we demonstrate that lapatinib, a dual TKI of EGFR and HER2 downregulates TS via inhibition of the nuclear translocation of EGFR and HER2. From our cDNA microarray experiments, we determined that a variety of nucleotide synthesis-related genes, including TS, were downregulated with lapatinib, and this was apparent in HER2-amplified cells. Targeted and pharmacologic inhibition assays confirmed that the dual inhibition of EGFR and HER2 is required for the more effective reduction of TS as compared to what was observed with gefitinib or trasutuzumab alone. Additionally, we determined that co-transfected EGFR and HER2 activate the TS gene promoter more profoundly than do either EGFR or HER2 alone. The translocation of EGFR and HER2 into the nucleus and the subsequent activation of the TS promoter were inhibited by lapatinib.Conclusions and SignificanceThese results demonstrate that lapatinib inhibits the nuclear translocation of EGFR and HER2 and downregulates TS, thus sensitizing cancer cells to fluoropyrimidine.

Dual Inhibition of the Epidermal Growth Factor Receptor Pathway with Cetuximab and Erlotinib: A Phase I Study in Patients with Advanced Solid Malignancies

To determine the optimal dose of the antiepidermal growth factor receptor (EGFR) monoclonal antibody cetuximab that can be safely administered in combination with a standard daily dose of erlotinib in patients with advanced solid malignancies. Patients with advanced solid malignancies who had failed standard chemotherapies received escalating doses of cetuximab without a loading dose (100, 200, 250 mg/m(2) i.v. weekly) in combination with a fixed dose of erlotinib (150 mg daily orally) until disease progression or unacceptable toxicity. Twenty-two patients were treated, including 14 patients (64%) with non-small cell lung cancer. Twenty patients received combination treatment at the highest dose level for a median of 5.5 weeks (range, 1-31 weeks). One dose-limiting toxicity was observed: grade 3 skin rash. Overall, the most common adverse events (any grade, grade 3/4) were consistent with the safety profiles of the individual drugs: acneform rash (100%, 9%), diarrhea (77%, 5%), and hypomagnesemia (59%, 12%). Seven of 18 evaluable patients (38.9%) had stable disease lasting for a median of 16.6 weeks (range, 6.1-25.1 weeks). Dual EGFR inhibition with cetuximab and erlotinib is feasible; the observed toxicities were manageable and consistent with the safety profiles of the individual drugs. The recommended doses for phase II studies are 250 mg/m(2) i.v. weekly for cetuximab and 150 mg daily orally for erlotinib.

Dual Epidermal Growth Factor Receptor and Vascular Endothelial Growth Factor Receptor Inhibition with NVP-AEE788 for the Treatment of Aggressive Follicular Thyroid Cancer

Patients with radioiodine-resistant follicular thyroid cancer (FTC) have a poor prognosis, if metastasized, with currently available treatment modalities. Epidermal growth factor (EGF) and vascular endothelial growth factor (VEGF) and their receptors (EGFR and VEGFR) have been reported to be overexpressed in FTC and have been implicated in FTC development. We hypothesized that inhibiting the phosphorylation of EGFR and VEGFR by treatment with NVP-AEE788 (AEE788), a novel dual specific EGFR and VEGFR inhibitor, either alone or in combination with paclitaxel, would inhibit the growth of FTC xenografts in an orthotopic nude mouse model. To confirm previous reports, EGF and EGFR expression and vascularity were analyzed in human samples of FTC, Hürthle cell carcinoma, and normal thyroid tissues. EGFR expression in four FTC cell lines was measured using Western blotting. The antitumor effect of AEE788 on FTC cells in vitro was evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays and Western blotting. The effect of AEE788, alone and in combination with paclitaxel, on FTC tumor growth in an orthotopic nude mouse model was also investigated. Immunohistochemical analysis of EGFR and VEGFR signaling status, cell proliferation, apoptosis, and microvessel density was done. EGF, EGFR, and vascularity were increased in human thyroid tumor samples and EGFR was increased in FTC cells. AEE788 inhibited FTC cell growth in vitro and reduced the phosphorylation status of EGFR, VEGFR, and two downstream targets, AKT and mitogen-activated protein kinase, in FTC cells. AEE788 alone and, to a greater extent, AEE788 plus paclitaxel suppressed FTC tumor growth in the thyroids of nude mice. Dual inhibition of EGFR and VEGFR by AEE788 could represent a novel approach to the treatment of radioiodine-resistant FTC.