ErbB/HER receptor activation and preclinical efficacy of lapatinib in vestibular schwannoma

Vestibular schwannomas (VS) arising sporadically or in patients with neurofibromatosis type 2 (NF2) consistently lack expression of Merlin, a tumor suppressor. Conventional treatment options include surgery and radiotherapy but there is no validated medical option. Recent evidence suggests that Merlin deficiency may result in abnormal activation of receptor tyrosine kinases (RTKs) and downstream signaling, promoting tumor growth. Although small-molecule RTK inhibitors are widely available for clinical use, no such therapy has been validated in patients with VS. To screen for RTK activation, surgical VS specimens from patients with and without NF2 were analyzed by phospho-RTK profiling arrays. Downstream signaling pathway activation was analyzed by phospho-MAPK arrays. Activated RTKs and downstream kinases were validated immunohistochemically in corresponding formalin-fixed, paraffin-embedded tissues. Phospho-RTK arrays and immunohistochemistry showed consistent overexpression and activation of EGFR family receptors and evidence of ERK1/2 downstream signaling was observed in all samples analyzed (n = 11). Based on the findings, the small-molecule EGFR/ErbB2 kinase inhibitor lapatinib was selected for evaluation of target inhibition and treatment efficacy in our in vitro human schwannoma model. EGFR/ErbB2 targeted therapy with lapatinib inhibited ErbB2 phosphorylation and survivin upregulation, as well as downstream ERK1/2 and AKT activation, resulting in decreased proliferation. We conclude that EGFR family receptor activation is a consistent feature of both sporadic and NF2-related VS. Molecular targeted therapy with lapatinib downregulates survivin and has antiproliferative activity in a preclinical VS model. Based on these findings, a clinical trial with lapatinib for the treatment of VS is currently underway.     

ErbB receptor tyrosine kinase network inhibition radiosensitizes carcinoma cells

PURPOSE: The expression of epidermal growth factor receptor (EGFR)-CD533, a truncation mutant of the wild-type EGFR, radiosensitizes carcinoma and malignant glioma cell lines. This deletion mutant disrupts EGFR activation and downstream signaling through the formation of inhibitory dimerizations. In this study, the effects of EGFR-CD533 on other ErbB receptor tyrosine kinase (RTK) family members were quantified to better understand the mechanism of EGFR-CD533-mediated radiosensitization. METHODS AND MATERIALS: Breast carcinoma cell lines with different ErbB RTK expression profiles were transduced with EGFR or ErbB2 deletion mutants (EGFR-CD533 and ErbB2-CD572) using an adenoviral vector. ErbB RTK activation, mitogen activated protein kinase (MAPK) and phosphatidylinositol-3-kinase (PI3K)/p70S6K signaling, and clonogenic survival were determined for expression of each deletion mutant. RESULTS: EGFR-CD533 radiosensitizes carcinoma cells with either high EGFR expression (MDA-MB231) or low EGFR expression (T47D) through significant blockade of the ErbB RTK network. Analysis of clonogenic survival demonstrate significant enhancement of the alpha/beta ratios, as determined by the linear-quadratic model. Split-dose survival experiments confirm that EGFR-CD533 reduces the repair of cellular damage after ionizing radiation. CONCLUSION: Expression of EGFR-CD533 inhibits the ErbB RTK network and radiosensitizes carcinoma cells irrespective of the ErbB RTK expression patterns, and ErbB2-CD572 does not radiosensitize cells with low EGFR expression. These studies demonstrate that the mechanism of action for EGFR-CD533-mediated radiosensitization is inhibition of the ErbB RTK network, and is an advantage for radiosensitizing multiple malignant cell types.     

Epidermal growth factor ligand/receptor loop and downstream signaling activation pattern in completely resected nonsmall cell lung cancer

BACKGROUND: In recent years, molecular insights shed light on the role of the epidermal growth factor receptor (EGFR) in nonsmall cell lung cancer (NSCLC), and new therapeutic agents, such as the EGFR tyrosine kinase inhibitors, were tested successfully, with responsiveness to those agents more likely in those patients with specific EGFR gene alterations. The objective of the current study was to investigate the protein profiles of EGFR, c-erb-B2, transforming growth factor alpha (TGF-alpha) (one of the EGFR ligands commonly expressed in NSCLC), and some downstream molecules, potentially to detect a subset of tumors with an activated autocrine loop that is responsible for higher intracellular signaling. METHODS: One hundred twelve consecutive patients with resected NSCLC were analyzed by immunohistochemistry for EGFR, the c-erb-B2 receptor, TGF-alpha, and pivotal molecules downstream from EGFR activation. Statistical correlations between the investigated molecular expression profiles and clinicopathologic data were performed. RESULTS: EGFR, c-erb-B2, TGF-alpha and downstream molecule expression, per se, was not correlated significantly with any clinicopathologic variables, with the exception of a significant correlation between squamous histology and EGFR and between adenocarcinoma and TGF-alpha. However, nearly 30% of NSCLCs demonstrated coexpression of both TGF-alpha and EGFR, and this molecular status was associated positively with a statistically significant expression of phosphatidylinositol 3 kinase and an inversely with mitogen-activated protein kinase expression. CONCLUSIONS: The presence of a subgroup of NSCLCs with an activated autocrine loop may help to explain the mechanisms that lead to the relative ineffectiveness of the EGFR tyrosine kinase inhibitor and may support new clinical trials to define whether the subgroup of patients with these tumors reasonably may benefit from higher doses of such inhibitors or from the simultaneous inhibition of EGFR downstream signaling targets.     

Expression and mutation analysis of epidermal growth factor receptor in head and neck squamous cell carcinoma

The epidermal growth factor receptor (EGFR)–RAS–RAF–mitogen-activated protein kinase signaling cascade is an important pathway in cancer development and recent reports show that EGFR and its downstream signaling molecules are mutated in a number of cancers. We have analyzed 91 Japanese head and neck squamous cell carcinomas (HNSCC) and 12 HNSCC cell lines for mutations in EGFR , ErbB2 , and K-ras . Exons encoding the hot-spot regions in the tyrosine kinase domain of both EGFR (exons 18, 19, and 21) and ErbB2 (exons 18–23), as well as exons 1 and 2 of K-ras were amplified by polymerase chain reaction and sequenced directly. EGFR expression was also analyzed in 65 HNSCC patients using immunohistochemistry. Only one silent mutation, C836T, was found in exon 21 of EGFR in the UT-SCC-16A cell line and its corresponding metastasic cell line UT-SCC-16B. No other mutation was found in EGFR , ErbB2 , or K-ras . All tumors showed EGFR expression. In 21 (32%) tumors, EGFR was expressed weakly (+1). In 27 (42%) tumors it was expressed (+2) moderately, and in 17 (26%) tumors high expression (+3) was detected. Overexpression (+2, +3) was found in 44 tumors (68%). A worse tumor differentiation and a positive nodal stage were significantly associated with EGFR overexpression ( P  = 0.02, P  = 0.032, respectively). Similar to patients from western ethnicity, mutations are absent or rare in Japanese HNSCC. Protein overexpression rather than mutation might be responsible for activation of the EGFR pathway in HNSCC. ( Cancer Sci 2008; 99: 1589–1594)     

Noninvasive imaging and quantification of epidermal growth factor receptor kinase activation in vivo

Epidermal growth factor receptor (EGFR) is a receptor tyrosine kinase (RTK) critical in tumor growth and a major target for anticancer drug development. However, thus far, there is no effective system to monitor its activities in vivo. Here, we report a novel approach to monitor EGFR activation based on the bifragment luciferase reconstitution system. The EGFR receptor and its interacting partner proteins (EGFR, growth factor receptor binding protein 2, and Src homology 2 domain-containing) were fused to NH(2) terminal and COOH terminal fragments of the firefly luciferase. After establishing tumor xenograft from cells transduced with the reporter genes, we show that the activation of EGFR and its downstream factors could be quantified through optical imaging of reconstituted luciferase. Changes in EGFR activation could be visualized after radiotherapy or EGFR inhibitor treatment. Rapid and sustained radiation-induced EGFR activation and inhibitor-mediated signal suppression were observed in the same xenograft tumors over a period of weeks. Our data therefore suggest a new methodology where activities of RTKs can be imaged and quantified optically in mice. This approach should be generally applicable to study biological regulation of RTK, as well as to develop and evaluate novel RTK-targeted therapeutics.     

PDGFRA,PDGFRB, EGFR,and downstream signaling activation in malignant peripheral nerve sheath tumor

We investigated the activation of platelet-derived growth factor (PDGF) receptor A (PDGFRA), PDGF receptor B (PDGFRB), epidermal growth factor receptor (EGFR), and their downstream pathways in malignant peripheral nerve sheath tumors (MPNSTs). PDGFRA, PDGFRB, and EGFR were immunohistochemically, biochemically, cytogenetically, and mutationally analyzed along with the detection of their cognate ligands in 16 neurofibromatosis type 1 (NF1)-related and 11 sporadic MPNSTs. The activation of the downstream receptor pathways was also studied by means of v-akt murine thymoma viral oncogene homolog (AKT), extracellular signal-regulated kinase (ERK), and mammalian target of rapamycin (mTOR) Western blotting experiments, as well as rat sarcoma viral oncogene homolog (RAS), v-raf murine sarcoma viral oncogene homolog B1 (BRAF), phosphoinositide-3-kinase, catalytic, alpha polypeptide (PI3KCA), and phosphatase and tensin homolog deleted on chromosome ten (PTEN) mutational analysis and fluorescence in situ hybridization. PDGFRA, PDGFRB, and EGFR were expressed/activated, with higher levels of EGFR expression/phosphorylation paralleling increasing EGFR gene copy numbers in the NF1-related cases (71%). Autocrine loop activation of these receptors along with their coactivation were suggested by the expression of the cognate ligands in the absence of mutations and the presence of receptor tyrosine kinase (RTK) heterodimers, respectively. Both MPNST groups showed AKT, ERK, and mTOR expression/phosphorylation. No BRAF, PI3KCA, or PTEN mutations were found in either group of MPNSTs, but 18% of the sporadic MPNSTs showed RAS mutations. PTEN monosomy segregated with the NF1-related cases (50%, p = 0.018), but PTEN protein was expressed in all but two cases. In conclusion, PDGFRA, PDGFRB, and EGFR seem to be promising molecular targets for tailored treatments in MPNST. In particular, the ligand- and heterodimerization-dependent RTK activation/expression coupled with a downstream signaling phosphorylation, mediated by the upstream receptors or RAS activation, may provide a rationale to apply combined RTK and mTOR inhibitor treatments both to sporadic and NF1-related cases.     

Erlotinib is a viable treatment for tumors with acquired resistance to cetuximab

The epidermal growth factor receptor (EGFR) is a ubiquitously expressed receptor tyrosine kinase (RTK) and is recognized as a key mediator of tumorigenesis in many human tumors. Currently there are five EGFR inhibitors used in oncology, two monoclonal antibodies (panitumumab, and cetuximab) and three tyrosine kinase inhibitors (erlotinib, gefitinib, and lapatinib). Both strategies of EGFR inhibition have demonstrated clinical successes, however many tumors remain non-responsive or acquire resistance during therapy. To explore potential molecular mechanisms of acquired resistance to cetuximab we previously established a series of cetuximab-resistant clones by chronically exposing the NCI-H226 NSCLC cell line to escalating doses of cetuximab. Cetuximab-resistant clones exhibited a dramatic increase in steady-state expression of EGFR, HER2, and HER3 receptors as well as increased signaling through the MAPK and AKT pathways. RNAi studies demonstrated dependence of cetuximab-resistant clones on the EGFR signaling network. These findings prompted investigation on whether or not cells with acquired resistance to cetuximab would be sensitive to the EGFR targeted TKI erlotinib. In vitro, erlotinib was able to decrease signaling through the EGFR axis, decrease cellular proliferation, and induce apoptosis. To determine if erlotinib could have therapeutic benefit in vivo, we established cetuximab-resistant NCI-H226 mouse xenografts, and subsequently treated them with erlotinib. Mice harboring cetuximab-resistant tumors treated with erlotinib exhibited either a tumor regression or growth delay as compared to vehicle controls. Analysis of the erlotinib treated tumors demonstrated a decrease in cell proliferation and increase rates of apoptosis. The work presented herein suggests that 1) cells with acquired resistance to cetuximab maintain their dependence on EGFR and 2) tumors developing resistance to cetuximab can benefit from subsequent treatment with erlotinib, providing rationale for its use in the setting of cetuximab resistance.     

Targeted Inhibition of Multiple Receptor Tyrosine Kinases in Mesothelioma

The receptor tyrosine kinases (RTKs) epidermal growth factor receptor (EGFR) and MET are activated in subsets of mesothelioma, suggesting that these kinases might represent novel therapeutic targets in this notoriously chemotherapy-resistant cancer. However, clinical trials have shown little activity for EGFR inhibitors in mesothelioma. Despite the evidence for RTK activation in mesothelioma pathogenesis, it is unclear whether transforming activity is dependent on an individual kinase oncoprotein or the coordinated activity of multiple kinases. Using phospho-RTK and immunoblot assays, we herein demonstrate activation of multiple RTKs (EGFR, MET, AXL, and ERBB3) in individual mesothelioma cell lines but not in normal mesothelioma cells. Inhibition of mesothelioma multi-RTK signaling was accomplished using combinations of RTK direct inhibitors or by inhibition of the RTK chaperone, heat shock protein 90 (HSP90). Multi-RTK inhibition by the HSP90 inhibitor 17-allyloamino-17-demethoxygeldanamycin (17-AAG) had a substantially greater effect on mesothelioma proliferation and survival compared with inhibition of individual activated RTKs. HSP90 inhibition also suppressed phosphorylation of downstream signaling intermediates (AKT, mitogen-activated protein kinase, and S6); upregulated the p53, p21, and p27 cell cycle checkpoints; induced G₂ phase arrest; induced caspase 3/7 activity; and led to an increase in the sub-G₁ apoptotic population. These compelling proapoptotic and antiproliferative responses indicate that HSP90 inhibition warrants clinical evaluation as a novel therapeutic strategy in mesothelioma.     

The Plasticity of Oncogene Addiction: Implications for Targeted Therapies Directed to Receptor Tyrosine Kinases

A common mutation of the epidermal growth factor receptor (EGFR) in glioblastoma multiforme (GBM) is an extracellular truncation known as the de2-7 EGFR (or EGFRvIII). Hepatocyte growth factor (HGF) is the ligand for the receptor tyrosine kinase (RTK) c-Met, and this signaling axis is often active in GBM. The expression of the HGF/c-Met axis or de2-7 EGFR independently enhances GBMgrowth and invasiveness, particularly through the phosphatidylinositol-3 kinase/pAkt pathway. Using RTK arrays, we show that expression of de2-7 EGFR in U87MG GBM cells leads to the coactivation of several RTKs, including platelet-derived growth factor receptor β and c-Met. A neutralizing antibody to HGF (AMG102) did not inhibit de2-7 EGFR-mediated activation of c-Met, demonstrating that it is ligand-independent. Therapy for parental U87MG xenografts with AMG 102 resulted in significant inhibition of tumor growth, whereas U87MG.Δ2-7 xenografts were profoundly resistant. Treatment of U87MG.Δ2-7 xenografts with panitumumab, an anti-EGFR antibody, only partially inhibited tumor growth as xenografts rapidly reverted to the HGF/c-Met signaling pathway. Cotreatment with panitumumab and AMG 102 prevented this escape leading to significant tumor inhibition through an apoptotic mechanism, consistent with the induction of oncogenic shock. This observation provides a rationale for using panitumumab and AMG 102 in combination for the treatment of GBM patients. These results illustrate that GBM cells can rapidly change the RTK driving their oncogene addiction if the alternate RTK signals through the same downstream pathway. Consequently, inhibition of a dominant oncogene by targeted therapy can alter the hierarchy of RTKs resulting in rapid therapeutic resistance.     

The tyrosine phosphatase PTPRO sensitizes colon cancer cells to anti-EGFR therapy through activation of SRC-mediated EGFR signaling

Inappropriate activation of epidermal growth factor receptor (EGFR) plays a causal role in many cancers including colon cancer. The activation of EGFR by phosphorylation is balanced by receptor kinase and protein tyrosine phosphatase activities. However, the mechanisms of negative EGFR regulation by tyrosine phosphatases remain largely unexplored. Our previous results indicate that protein tyrosine phosphatase receptor type O (PTPRO) is down-regulated in a subset of colorectal cancer (CRC) patients with a poor prognosis. Here we identified PTPRO as a phosphatase that negatively regulates SRC by directly dephosphorylating Y416 phosphorylation site. SRC activation triggered by PTPRO down-regulation induces phosphorylation of both EGFR at Y845 and the c-CBL ubiquitin ligase at Y731. Increased EGFR phosphorylation at Y845 promotes its receptor activity, whereas enhanced phosphorylation of c-CBL triggers its degradation promoting EGFR stability. Importantly, hyperactivation of SRC/EGFR signaling triggered by loss of PTPRO leads to high resistance of colon cancer to EGFR inhibitors. Our results not only highlight the PTPRO contribution in negative regulation of SRC/EGFR signaling but also suggest that tumors with low PTPRO expression may be therapeutically targetable by anti-SRC therapies.     

Tyrosine kinase inhibitors-ZD1839 (Iressa).

Several epithelial tumors display epidermal growth factor receptor (EGFR) overexpression (with or without EGFR gene amplification) that is often associated with increased production of EGFR ligands. This permits the activation of endogenous tumor EGFR via autocrine mechanisms, resulting in cellular proliferation and tumor growth. Interruption of receptor signaling with bivalent EGFR antibodies or with small molecule inhibitors of the EGFR tyrosine kinase results in inhibition of tumor cell proliferation or viability in vitro and in vivo. One small molecule currently undergoing preclinical and clinical investigation is ZD1839 (Iressa), a synthetic anilinoquinazoline capable of inhibiting EGFR tyrosine kinase in vitro. The early results of clinical trials indicate this drug possesses antitumor activity in certain malignancies of the upper aerodigestive tract.     

Molecular mechanisms of epidermal growth factor receptor (EGFR) activation and response to gefitinib and other EGFR-targeting drugs.

The epidermal growth factor receptor (EGFR) family of receptor tyrosine kinases, including EGFR, HER2/erbB2, and HER3/erbB3, is an attractive target for antitumor strategies. Aberrant EGFR signaling is correlated with progression of various malignancies, and somatic tyrosine kinase domain mutations in the EGFR gene have been discovered in patients with non-small cell lung cancer responding to EGFR-targeting small molecular agents, such as gefitinib and erlotinib. EGFR overexpression is thought to be the principal mechanism of activation in various malignant tumors. Moreover, an increased EGFR copy number is associated with improved survival in non-small cell lung cancer patients, suggesting that increased expression of mutant and/or wild-type EGFR molecules could be molecular determinants of responses to gefitinib. However, as EGFR mutations and/or gene gains are not observed in all patients who respond partially to treatment, alternative mechanisms might confer sensitivity to EGFR-targeting agents. Preclinical studies showed that sensitivity to EGFR tyrosine kinase inhibitors depends on how closely cell survival and growth signalings are coupled with EGFR, and also with HER2 and HER3, in each cancer. This review also describes a possible association between EGFR phosphorylation and drug sensitivity in cancer cells, as well as discussing the antiangiogenic effect of gefitinib in association with EGFR activation and phosphatidylinositol 3-kinase/Akt activation in vascular endothelial cells.     

Antagonistic and agonistic effects of quinazoline tyrosine kinase inhibitors on mutant EGF receptor function

A mutated form of the EGF receptor (EGFRvIII), resulting from deletion of exons 2-7, is an oncogenic protein that is expressed in multiple human tumors. This mutation induces ligand-independent activation of the EGFR tyrosine kinase and thereby can initiate unregulated cell growth and tumorigenesis. Thus, inhibition of the kinase activity of EGFRvIII is a potential means of suppressing its oncogenic properties. Certain tyrosine kinase inhibitors (tyrphostins) specifically inhibit the wild-type EGFR and thereby inhibit tumor growth both in vitro and in vivo. We demonstrate that the quinazoline tyrphostins AG 1478 and AG 1517 can suppress morphologic transformation of cell lines by EGFRvIII. Quinazolines were found to inhibit receptor autophosphorylation and signaling through MAP kinase, but had minimal effects on association of EGFRvIII with Grb2/SOS. Low concentrations of quinazoline also increased receptor dimerization and phosphotyrosine content. This was associated with increases in colony formation in soft agar and increased invasion through matrigel for AG 1478. Thus, both AG 1478 and AG 1517 can inhibit multiple EGFRvIII signaling pathways, but at low concentrations AG 1478 can enhance colony formation, presumably related to augmented homodimerization of the receptor and activation of downstream signaling.     

Flavopiridol Synergizes with Sorafenib to Induce Cytotoxicity and Potentiate Antitumorigenic Activity in EGFR/HER-2 and Mutant RAS/RAF Breast Cancer Model Systems

Oncogenic receptor tyrosine kinase (RTK) signaling through the Ras-Raf-Mek-Erk (Ras-MAPK) pathway is implicated in a wide array of carcinomas, including those of the breast. The cyclin-dependent kinases (CDKs) are implicated in regulating proliferative and survival signaling downstream of this pathway. Here, we show that CDK inhibitors exhibit an order of magnitude greater cytotoxic potency than a suite of inhibitors targeting RTK and Ras-MAPK signaling in cell lines representative of clinically recognized breast cancer (BC) subtypes. Drug combination studies show that the pan-CDK inhibitor, flavopiridol (FPD), synergistically potentiated cytotoxicity induced by the Raf inhibitor, sorafenib (SFN). This synergy was most pronounced at sub-EC₅₀ SFN concentrations in MDA-MB-231 (KRAS-G13D and BRAF-G464V mutations), MDA-MB-468 [epidermal growth factor receptor (EGFR) overexpression], and SKBR3 [ErbB2/EGFR2 (HER-2) overexpression] cells but not in hormone-dependent MCF-7 and T47D cells. Potentiation of SFN cytotoxicity by FPD correlated with enhanced apoptosis, suppression of retinoblastoma (Rb) signaling, and reduced Mcl-1 expression. SFN and FPD were also tested in an MDA-MB-231 mammary fat pad engraftment model of tumorigenesis. Mice treated with both drugs exhibited reduced primary tumor growth rates and metastatic tumor load in the lungs compared to treatment with either drug alone, and this correlated with greater reductions in Rb signaling and Mcl-1 expression in resected tumors. These findings support the development of CDK and Raf co-targeting strategies in EGFR/HER-2-overexpressing or RAS/RAF mutant BCs.     

Co-expression of epidermal growth factor receptor and transforming growth factor-alpha is independent of ras mutations in lung adenocarcinoma

The interaction of epidermal growth factor receptor (EGFR) and its ligand transforming growth factor-alpha (TGF-alpha) leads to an autocrine activation of the ras signaling pathway and putatively its oncogenic activity. It is thus hypothesized that the co-overexpression of EGFR-TGFalpha will be redundant hence rare in tumors with oncogenic ras mutations. To test this hypothesis, we studied by immunohistochemistry the expression of EGFR and TGF-alpha in primary non small cell lung cancers. Such putative EGFR autocrine loop activation was found in 73% of squamous cell carcinomas that rarely develop ras mutations. In contrast, EGFR-TGFalpha co-expression occurred with equal frequency in adenocarcinomas irrespective of their ras genotype. The results indicate that EGFR autocrine loop activity in adenocarcinoma may have alternative signaling activities aside from the activation of ras-MAP kinase pathway.     

Inhibition of the Growth of Patient-Derived Pancreatic Cancer Xenografts with the MEK Inhibitor Trametinib Is Augmented by Combined Treatment with the Epidermal Growth Factor Receptor/HER2 Inhibitor Lapatinib

Mutations of the oncogene KRAS are important drivers of pancreatic cancer progression. Activation of epidermal growth factor receptor (EGFR) and human EGFR2 (HER2) is observed frequent in pancreatic adenocarcinomas. Because of co-activation of these two signaling pathways, we assessed the efficacy of inhibition of EGFR/HER2 receptors and the downstream KRAS effector, mitogen-activated protein kinase/extracellular-signal regulated kinase (ERK) kinase 1 and 2 (MEK1/2), on pancreatic cancer proliferation in vitro and in a murine orthotopic xenograft model. Treatment of established and patient-derived pancreatic cancer cell lines with the MEK1/2 inhibitor trametinib (GSK1120212) inhibited proliferation, and addition of the EGFR/HER2 inhibitor lapatinib enhanced the inhibition elicited by trametinib in three of eight cell lines. Importantly, in the orthotopic xenograft model, treatment with lapatinib and trametinib resulted in significantly enhanced inhibition of tumor growth relative to trametinib treatment alone in four of five patient-derived tumors tested and was, in all cases, significantly more effective in reducing the size of established tumors than treatment with lapatinib or trametinib alone. Acute treatment of established tumors with trametinib resulted in an increase in AKT2 phosphorylation that was blunted in mice treated with both trametinib and lapatinib. These data indicate that inhibition of the EGFR family receptor signaling may contribute to the effectiveness of MEK1/2 inhibition of tumor growth possibly through the inhibition of feedback activation of receptor tyrosine kinases in response to inhibition of the RAS-RAF-MEK-ERK pathway. These studies provide a rationale for assessing the co-inhibition of these pathways in the treatment of pancreatic cancer patients.     

Acetylsalicylic acid enhances antiproliferative effects of the EGFR inhibitor gefitinib in the absence of activating mutations in gastric cancer

The epidermal growth factor receptor (EGFR) is highly expressed in gastric cancer indicating its suitability as a target for receptor tyrosine kinase (RTK) inhibitors. In the current study we explored the role of EGFR and its potential use as a therapeutic target in gastric cancer. First we analyzed 66 gastric cancer samples of Asian and Caucasian patients for the presence of EGFR mutations. No activating EGFR mutations were found and gefitinib alone was only weakly effective in gastric cancer cell lines. However, acetylsalicylic acid (ASA) significantly enhanced the inhibitory effects of gefitinib indicating synergistic action. Whole genome expression profiling indicated significant regulation of 120 genes in the case of co-administration of gefitinib and ASA (32 induced, 88 repressed) in gastric adenocarcinoma cells. Further analyses indicated that several important signalling pathways were effectively inhibited by simultaneous exposure to gefitinib and ASA. Our findings indicate that although gastric cancer does not seem to harbour mutations which render the cancer cells constitutively susceptible to gefitinib, the co-administration of ASA can strengthen RTK inhibitor activity in adenocarcinoma cells by EGFR activation. This is the first report of effective modulation of EGFR-inhibition activity in cancer.