To EGFR-tyrosine Kinase Inhibitors Research Articles

The content of mutant EGFR DNA correlates with response to EGFR-TKIs in lung adenocarcinoma patients with common EGFR mutations.

This study aimed to elucidate the association of the content of mutant epidermal growth factor receptor (EGFR) deoxyribonucleic acid (DNA) with the treatment response to EGFR-tyrosine kinase inhibitor (TKI) and survival in patients with lung cancer.This retrospective cohort study included 77 lung adenocarcinoma patients with common EGFR mutations from December 2012 to February 2015. The content of mutant EGFR DNA in lung cancer tissues was determined using an Amplification Refractory Mutation System. The association of the amount of mutant EGFR DNA with treatment response, the clinical variables, and the progression-free survival (PFS) after EGFR-TKI therapy were evaluated.Using the amount of mutant EGR DNA above 4.77% as the cut-off value, the sensitivity to predict EGFR-TKI responder is 82.0% and the specificity is 75.0% (area under the curve [AUC]: 0.734, P = 0.003). The high content of mutant EGFR DNA is an independent factor associated with the response to EGFR-TKIs (odds ratio: 13.07, 95% confidence interval [CI]: 3.23–52.11, P = 0.0003). A significantly longer PFS was observed in the group with the high content of mutant EGFR DNA (26.3 months, 95% CI: 12.2–26.3) compared with the low content of mutant EGFR DNA groups (12.3 months, 95% CI: 5.7–14.8, P = 0.0155). A better predictive value of the content of mutant EGFR DNA was noted in patients with exon 19 deletions (AUC: 0.892, P < 0.0001) than exon 21 L858R mutations (AUC: 0.675, P = 0.0856).Our results show that the content of mutant EGFR DNA is associated with the clinical response to EGFR-TKIs, especially in patients with exon 19 deletions mutation.

Development of an efficient scoring system for detection of EGFR-T790M mutation in non-small cell lung cancer patients treated with EGFR-tyrosine kinase inhibitors.

e20599Background: Secondary T790M mutation of epidermal growth factor receptor (EGFR) accounts for approximately half of all cases of acquired resistance to EGFR-tyrosine kinase inhibitors (TKIs) i...

Ganoderma lucidum Combined with the EGFR Tyrosine Kinase Inhibitor, Erlotinib Synergize to Reduce Inflammatory Breast Cancer Progression.

The high incidence of resistance to Tyrosine Kinase Inhibitors (TKIs) targeted against EGFR and downstream pathways has increased the necessity to identify agents that may be combined with these therapies to provide a sustained response for breast cancer patients. Here, we investigate the therapeutic potential of Ganoderma lucidum extract (GLE) in breast cancer, focusing on the regulation of the EGFR signaling cascade when treated with the EGFR TKI, Erlotinib. SUM-149, or intrinsic Erlotinib resistant MDA-MB-231 cells, and a successfully developed Erlotinib resistant cell line, rSUM-149 were treated with increasing concentrations of Erlotinib, GLE, or their combination (Erlotinib/GLE) for 72h. Treatment effects were tested on cell viability, cell proliferation, cell migration and invasion. To determine tumor progression, severe combined immunodeficient mice were injected with SUM-149 cells and then treated with Erlotinib/GLE or Erlotinib for 13 weeks. We assessed the protein expression of ERK1/2 and AKT in in vitro and in vivo models. Our results show that GLE synergizes with Erlotinib to sensitize SUM-149 cells to drug treatment, and overcomes intrinsic and developed Erlotinib resistance. Also, Erlotinib/GLE decreases SUM-149 cell viability, proliferation, migration and invasion. GLE increases Erlotinib sensitivity by inactivating AKT and ERK signaling pathways in our models. We conclude that a combinatorial therapeutic approach may be the best way to increase prognosis in breast cancer patients with EGFR overexpressing tumors.

Baseline, Trend, and Normalization of Carcinoembryonic Antigen as Prognostic Factors in Epidermal Growth Factor Receptor-Mutant Nonsmall Cell Lung Cancer Patients Treated With First-Line Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitors.

Among epidermal growth factor receptor (EGFR) mutation status unknown nonsmall cell lung cancer (NSCLC) patients, those with higher carcinoembryonic antigen (CEA) level are more likely to response to EGFR-tyrosine kinase inhibitors (TKIs) because they tend to have mutant epidermal growth factor receptor (EGFR). However, patients with higher CEA also have more tumor burden. With the above paradoxical evidence, it is prudent to understand the prognostic significance of baseline CEA in patients with EGFR-mutant NSCLC treated with first-line EGFR-TKIs. The clinical significance of the trend in CEA after treatment and the impact of CEA normalization during EGFR-TKI therapy are also unknown and potentially important.A total of 241 patients who received first-line EGFR-TKIs were included. As to baseline CEA, patients were divided into normal, low, and high baseline CEA by cut point determined by receiver operating characteristic curves. As to CEA responses, patients were divided into 3 groups accordingly to their amount of CEA change after taking TKIs. In group A, 1-month follow-up CEA level decreased more than 35% with nadir CEA normalization; in group B, 1-month follow-up CEA level decreased more than 35% without nadir CEA normalization; and in group C, 1-month follow-up CEA level decreased less than 35% or increased.Patients with higher baseline CEA levels had shorter progression-free survival (PFS) and overall survival (OS) (CEA > 32 vs 5–32 vs <5 ng/mL, PFS = 8.8 vs 11.3 vs 14.4 months, respectively, P < 0.001; OS = 17.8 vs 22.0 vs 27.9 months, respectively, P = 0.01). For trend and CEA normalization in groups A, B, and C, PFS was 14.3, 10.6, and 7.1 months, respectively (P < 0.001); OS was 29.7, 20.0, and 16.2 months, respectively (P < 0.001).Baseline, trend, and normalization of CEA levels are potential prognostic markers for patients with EGFR-mutant advanced NSCLC treated with first line EGFR-TKIs.

Next-generation sequencing of tyrosine kinase inhibitor-resistant non-small-cell lung cancers in patients harboring epidermal growth factor-activating mutations.

BackgroundThe aim of this study was to detect the epidermal growth factor receptor (EGFR)-activating mutations and other oncogene alterations in patients with non-small-cell lung cancers (NSCLC) who experienced a treatment failure in response to EGFR-tyrosine kinase inhibitors (TKIs) with a next generation sequencer.MethodsFifteen patients with advanced NSCLC previously treated with EGFR-TKIs were examined between August 2005 and October 2014. For each case, new biopsies were performed, followed by DNA sequencing on an Ion Torrent Personal Genome Machine (PGM) system using the Ion AmpliSeq Cancer Hotspot Panel version 2.ResultsAll 15 patients were diagnosed with NSCLC harboring EGFR-activating mutations (seven cases of exon 19 deletion, seven cases of L858R in exon 21, and one case of L861Q in exon 21). Of the 15 cases, acquired T790M resistance mutations were detected in 9 (60.0 %) patients. In addition, other mutations were identified outside of EGFR, including 13 cases (86.7 %) exhibiting TP53 P72R mutations, 5 cases (33.3 %) of KDR Q472H, and 2 cases (13.3 %) of KIT M541L.ConclusionsHere, we showed that next-generation sequencing (NGS) is able to detect EGFR T790M mutations in cases not readily diagnosed by other conventional methods. Significant differences in the degree of EGFR T790M and other EGFR-activating mutations may be indicative of the heterogeneity of disease phenotype evident within these patients. The co-existence of known oncogenic mutations within each of these patients may play a role in acquired EGFR-TKIs resistance, suggesting the need for alternative treatment strategies, with PCR-based NGS playing an important role in disease diagnosis.

Spatiotemporal T790M Heterogeneity in Individual Patients with EGFR-Mutant Non–Small-Cell Lung Cancer after Acquired Resistance to EGFR-TKI

Epidermal growth factor receptor (EGFR) mutation T790M accounts for approximately half of acquired resistances to EGFR-tyrosine kinase inhibitor (TKI). Because T790M is mediated by TKI exposure, its penetration and "on-off" may affect T790M status. We retrospectively reviewed T790M status and clinical course of patients who had undergone multiple rebiopsies after acquired resistance to EGFR-TKI. Of 145 patients with EGFR-mutant NSCLC receiving rebiopsy after acquired resistance, 30 underwent multiple site rebiopsies, and 24 received repeated rebiopsies at the same lesion. In 22 patients who underwent rebiopsies from both central nervous system (CNS; 20 cerebrospinal fluids [CSF] and 2 brain tumoral tissues) and thoracic lesions (7 lung tissues, 14 pleural effusions, and 1 lymph node), 12 were thoracic-T790M-positive. Of these 12 patients, 10 were CNS-T790M-negative, despite exhibiting thoracic-T790M-positive. All 10 thoracic-T790M-negatives were CNS-T790M-negative. Three patients revealed a spatial heterogeneous T790M status among their thoracic lesions. In 24 patients receiving repeated rebiopsies at the same lesion (12 lung tissues, 6 CSFs, and 6 pleural effusions), T790M status of lung lesions varied in five patients after TKI-free interval. In all five patients whose T790M status changed from positive to negative, EGFR-TKI rechallenge was effective. In three of these five patients, after further TKI exposure, T790M status changed from negative to positive again. There was also a patient whose CSF T790M status changed from negative to positive after high-dose erlotinib therapy. T790M status in an individual patient can be spatiotemporally heterogeneous because of selective pressure from EGFR-TKI.

Reduced expression of EI24 confers resistance to gefitinib through IGF-1R signaling in PC9 NSCLC cells.

Lung cancer is the commonly diagnosed cancer and is the leading cause of cancer-related mortality worldwide. The most prevalent form of lung cancer is NSCLC, comprising 80% of all lung cancer cases, and epidermal growth factor receptor (EGFR) is frequently mutated in NSCLC. EI24 is a p53-responsive gene and plays an important role in tumor suppression. In the current study, we found that reduced expression of EI24 conferred resistance to EGFR-tyrosine-kinase inhibitor (TKI) in NSCLC cells. The correlation between EI24 expression and EGFR-TKI drug resistance in EGFR-driven tumors were determined from microarray datasets. The phospho-protein expression profiles of receptor tyrosine kinases and protein kinases were examined using antibody arrays method in PC9 cells expressing shRNAs targeting EI24 and gefitinib-resistant PC9-GR cells expressing exogenous EI24. The EGFR-TKI resistant clones had reduced expression of EI24 mRNA compared to the sensitive clones, and EI24 knockdown rendered sensitive cells resistant to EGFR-TKI. Receptor tyrosine kinase screening revealed the involvement of a kinase switch in EI24-mediated regulation of drug sensitivity. We found that EI24 modulates the insulin growth factor-1 receptor (IGF-1R) pathway through the induction of IGF-1. Combination treatment with EGFR and IGF-1R inhibitors significantly reduced the viability of EI24 knockdown-induced resistant cell lines compared to single-agent treatments. We also showed that low EI24 and high IGF-1R expressions in lung cancer patients were correlated with reduced overall survival. Taken together, these results suggest a potential role for EI24 as a biomarker of drug resistance, and indicate that combination therapy with EGFR and IGF-1R inhibitors would be effective in NSCLC patients with low EI24 expression.

Abstract 5358: The hypoxia-activated EGFR-TKI TH-4000 overcomes erlotinib-resistance in preclinical NSCLC models at plasma levels achieved in a Phase 1 clinical trial

Abstract Clinical data indicate that mutant EGFR NSCLC is often heterozygous (PLoS ONE 2013; 8: e54170; PLoS ONE 2009; 4: e7464) and the presence of wild type (WT) EGFR allele is associated with limited response to EGFR-tyrosine kinase inhibitor (TKI) therapy (Cancer Sci 2008; 99:929). Tumor hypoxia upregulates WT EGFR protein and its cognate ligand TGFα via several HIF-dependent mechanisms (reviewed in: Curr Pharm Des 2013; 19:907). NSCLC is known to be a hypoxic tumor, and thus hyperactivation of WT EGFR may be an important cause of resistance to EGFR-TKI therapy. TH-4000 (formerly called PR610) is a clinical-stage hypoxia-activated prodrug that releases an irreversible EGFR-TKI under hypoxic conditions and may overcome resistance to conventional TKI therapy. We tested this hypothesis using the heterozygous WT/Δ19 EGFR PC9 tumor model and found it to be resistant to clinically relevant doses of the EGFR-TKI erlotinib; 100% of tumors progressed during treatment with human matched plasma PK exposures of erlotinib. In contrast, the homozygous Δ19 mutant EGFR tumor HCC827 was readily controlled by erlotinib (100% tumor regression). TH-4000 (15 mg/kg) produced 100% tumor regressions in both models. In vitro, PC9 cells exposed to hypoxia had elevated EGFR protein and were more resistant to erlotinib as measured by EGFR phosphorylation. In nude mice, single-dose administration of 15 mg/kg TH-4000 achieved a plasma AUC equivalent to 32 mg/m2 in human subjects, one-fifth of the maximum tolerated dose (MTD) defined in the Phase 1 trial (MTD = 150 mg/m2/week; NCT01631279). A single dose of TH-4000 (15 mg/kg) cleared rapidly from mouse plasma (T½ = 0.37 h) but had durable residency in PC9 tumors (T½ = 39 h), releasing TKI above efficacious levels for 7 days (T½β = 84 h). Consistent with these PK properties, tumor shutdown of EGFR signalling was durable, with no recovery by day 7. To confirm the mechanism of action, TH-4000 was shown to be metabolized efficiently under hypoxia using a panel of human NSCLC cell lines (rate of TKI release 0.4-2.1 nmol/hr/106 cells), a process that was inhibited by oxygen (TKI release &amp;lt; 0.002 nmol/hr/106 cells). Cellular anti-proliferative and receptor phosphorylation assays demonstrated a 14-80 fold reduction of TH-4000 activity relative to TKI. Using PC9 tumors, hyperbaric oxygen breathing suppressed release of TKI from TH-4000 by &amp;gt;80% (538 vs 99 nmol/kg; p &amp;lt;0.01) compared to air breathing controls. Collectively, these data further validate that TH-4000 is a hypoxia-activated irreversible EGFR-TKI, and show that TH-4000 has greater activity compared with erlotinib in a heterozygous xenograft model of mutant EGFR NSCLC. Thus, TH-4000 may overcome hypoxia-induced resistance to erlotinib at plasma exposures readily achieved in human subjects. Citation Format: Adam V. Patterson, Shevan Silva, Christopher Guise, Maria Abbattista, Matthew Bull, Huai-Ling Hsu, Charles Hart, Jessica Sun, Angus Grey, Amir Ashoorzadeh, Robert Anderson, Jeff B. Smaill. The hypoxia-activated EGFR-TKI TH-4000 overcomes erlotinib-resistance in preclinical NSCLC models at plasma levels achieved in a Phase 1 clinical trial. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5358. doi:10.1158/1538-7445.AM2015-5358

Abstract 5248: Non-invasive EGFR T790M detection using droplet digital PCR system

Abstract Background: Non-small cell lung cancers (NSCLCs) harboring activating epidermal growth factor receptor (EGFR) mutations show significant response to EGFR-tyrosine kinase inhibitors (TKIs). However, most of them acquire resistance to EGFR-TKIs, and a half of the resistance is caused by EGFR T790M mutation. Recently, the third generation-TKIs which are effective to lung cancer harboring T790M mutation have been developed, thus the importance of T790M mutation detection is increasing in NSCLC treatment. Nowadays, the highly sensitive assays such as PNA-LNA PCR clamp, Scorpion ARMS have been developed to detect T790M mutation. In this study, we newly established non-invasive assay for T790M mutation detection from circulating cell free DNA (ccfDNA) using droplet digital PCR system (ddPCR). Materials and Methods: The sensitivity of ddPCR was determined using variable proportion of mixture DNAs of human bronchial epithelial cell with wild-type EGFR and NCI-H1975 with EGFR L858R and T790M mutations. We collected ccfDNA extracted from the 39 plasma samples of 22 NSCLC patients, who had been treated with EGFR-TKI and examined EGFR T790M mutational status by re-biopsy. The T790M mutational status of the re-biopsy samples were determined using PNA-LNA PCR clamp method. The circulating T790M alleles in ccfDNAs were detected using ddPCR, and compared with the T790M mutational status of re-biopsy samples. Results: In 8 of the 39 plasma samples, T790M alleles were detected by ddPCR. As 10 of 39 plasma samples were collected from the patient who determined T790M positive by re-biopsy after acquiring resistance to EGFR -TKI, the sensitivity of ddPCR was 80%. In 3 plasma samples, T790M alleles were detected by ddPCR although PNA-LNA PCR clamp method could not detect T790M mutation from the re-biopsy tissues of the patients who provided their plasmas. The specificity of ddPCR was determined as 89.7% although we have to consider about the difference between the timings of plasma collection and re-biopsy, and tumors’ heterogeneity. Conclusions: The result of circulating T790M allele detection using ddPCR was highly concordant with the result of T790M detection in re-biopsy samples using PNA-LNA PCR clamp method. The ddPCR is promising method for non-invasive T790M mutation detection. Citation Format: Shinsuke Hashida, Kadoaki Ohashi, Takehiro Matsubara, Tomoaki Ohtsuka, Mototsugu Watanabe, Ken Suzawa, Yuho Maki, Hiromasa Yamamoto, Junichi Soh, Hiroaki Asano, Kazunori Tsukuda, Shinichiro Miyoshi, Katsuyuki Kiura, Shinichi Toyooka. Non-invasive EGFR T790M detection using droplet digital PCR system. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5248. doi:10.1158/1538-7445.AM2015-5248

Phase II study of a combination regimen of gefitinib and pemetrexed as first-line treatment in patients with advanced non-small cell lung cancer harboring a sensitive EGFR mutation

Patients with advanced non-small cell lung cancer (NSCLC) harboring a sensitive epidermal growth factor receptor (EGFR) mutation have been shown to exhibit a marked response to EGFR-tyrosine kinase inhibitor (TKI) treatment. Pemetrexed and gefitinib were reported to have a schedule-dependent cytotoxic synergism. We evaluated the efficacy and safety of a combination regimen of gefitinib and pemetrexed as first-line chemotherapy in EGFR-mutated NSCLC patients. Systemic therapy-naïve patients with advanced non-squamous NSCLC harboring a sensitive EGFR mutation were included in this study. Pemetrexed was administered on day 1 at a dose of 500 mg/m(2), and gefitinib was sequentially administered on days 2-16. This treatment regimen was repeated every 3 weeks until disease progression. Twenty-six patients were enrolled in this study. The median number of treatment cycles was 16 (range, 1-35). The overall response rate (ORR) was 84.6% (95% confidence interval [CI], 70.7-98.5%), and the disease control rate (DCR) was 96.2% (95% CI, 88.9-100%). Grade 3/4 hematological toxicities included neutropenia (15.4%), leukopenia (7.7%), and anemia (3.8%). No grade 4 non-hematological toxicities were observed. The main grade 3 non-hematological toxicities were infection (11.5%), increased alanine aminotransferase (11.5%) and aspartate aminotransferase (7.7%) levels, fatigue (3.8%), diarrhea (3.8%), and pneumonitis (3.8%). We observed a median progression-free survival (PFS) of 18.0 months (95% CI, 15.0-21.0 months) and a median survival time (MST) of 32.0 months (95% CI, 28.5-35.5 months). There were no treatment-related deaths. The combination regimen used in this study showed a high ORR, long median PFS, and acceptable toxicity. A future randomized trial on pemetrexed plus gefitinib compared with gefitinib alone is warranted.

Rebiopsy rate for non-small cell lung cancer patients after EGFR-TKI failure in clinical setting: Toward the third-generation EGFR-TKI era.

e19080 Background: The secondary epidermal growth factor receptor (EGFR) mutation Thr790Met (T790M) accounts for approximately half of acquired resistances to EGFR-tyrosine kinase inhibitors (TKIs)...

Epidermal growth factor receptor variant III mutation in Chinese patients with squamous cell cancer of the lung.

BackgroundThe aim of this study was to investigate the distribution of epidermal growth factor receptor (EGFR)vIII mutation in Chinese non-small cell lung cancer (NSCLC) patients and to explore the likely relationship between EGFRvIII mutation and response to EGFR-tyrosine kinase inhibitors (TKIs) in squamous cell carcinoma (SCC).MethodsSamples were derived from two patient cohorts: (i) 114 early-stage NSCLC who received surgical resection; and (ii) 31 advanced-stage SCC who received EGFR-TKI EGFRvIII. EGFR and V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) mutations were detected by reverse transcription polymerase chain reaction (RT-PCR), denaturing high-performance liquid chromatography, and PCR-restriction fragment length polymorphism, respectively. The associations of EGFRvIII, EGFR, and KRAS mutations with clinical outcome of EGFR-TKI treatment were evaluated using the Kaplan-Meier method, descriptive analysis, and multi-variable Cox regression analysis.ResultsIn the first cohort, EGFRvIII mutation was detected in eight of 114 (7.0%) patients, including 11.1% (6/54) SCC and 3.6% (2/55) adenocarcinomas (ADC) (P = 0.269). In the second cohort, five (16.1%) and 10 out of 31 advanced SCC presented EGFRvIII and EGFR mutations, respectively. No appreciable discrepancy of progression-free survival or disease control rate was detected between the patients with and without EGFRvIII mutation (P > 0.05). However, longer median overall survival (OS) was observed in patients harboring EGFRvIII compared to those without EGFRvIII, although the difference did not reach statistical significance.ConclusionThe frequency of EGFRvIII mutation in SCC was higher than in ADC. SCC patients harboring EGFRvIII mutations had a tendency for prolonged OS.

A Case Report of a Metastatic Adenocarcinoma of Lung with Dual Positivity for EGFR Mutation and ALK Fusion

Non-small cell lung cancer ranks among the most lethal cancers worldwide. The rate of epidermal growth factor receptor (EGFR) mutations and echinoderm microtubuleassociated protein-like 4-anaplastic lymphoma kinase (EML4-ALK) gene fusion is the most common in younger age, non-smoking Asian adenocarcinoma lung cancer patients. EGFR mutations and ALK gene rearrangements are known to be mutually exclusive and as mutual causes of resistance to EGFR-tyrosine kinase inhibitors (TKIs) or ALK-TKIs. However, rarely such co-alterations do co-exist in some clinical cases. Here we report a 62 year old male, heavy smoker with cough, hemoptysis and fatigue. Histopathological examination of bronchoscopic guided biopsy showed adenocarcinoma histology. Staging evaluation, he was found to have stage IV disease on positron emission tomography-computed tomography scan. The biopsy blocks tested positive for both EGFR mutation and ALK fusion. Patient was initiated on tablet Gefitinib 250 mg once daily. To the best of our knowledge, there has been no report of dual EGFR mutation and ALK fusion positivity from India.

Survival Outcome Assessed According to Tumor Burden and Progression Patterns in Patients With Epidermal Growth Factor Receptor Mutant Lung Adenocarcinoma Undergoing Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitor Therapy

BackgroundMutations in the epidermal growth factor receptor (EGFR) have been associated with a marked therapeutic response to EGFR-tyrosine kinase inhibitors (TKIs) in patients with advanced non–small-cell lung cancer (NSCLC). However, the clinical predictors of the survival benefit of EGFR-TKI therapy for NSCLC with EGFR-activating mutations have not been well elucidated. Therefore, the present study evaluated the clinical predictors of survival outcome in patients with EGFR-mutant NSCLC who had been treated with EGFR-TKIs. Material and MethodsThe data from 224 patients with EGFR-mutant lung adenocarcinoma treated with EGFR-TKIs were retrospectively reviewed. The treatment outcomes were evaluated according to the clinical factors, number of metastasis sites, and progression patterns. ResultsThe clinical factors associated with reduced progression-free survival (PFS) and overall survival (OS) on univariate analysis were Eastern Cooperative Oncology Group (ECOG) performance status (PS) ≥ 2, intra- and extrathoracic metastasis, extrathoracic metastasis, a high number of metastatic sites, metastasis to the liver or adrenal gland at baseline, and rapid progression at the diagnosis of progressive disease (PD). On multivariate analysis, the factors that remained significantly associated with a shorter PFS were ECOG PS ≥ 2 (odds ratio [OR], 2.189; 95% confidence interval [CI], 1.374-3.437; P < .001) and rapid tumor progression at PD (OR, 1.800; 95% CI, 1.059-3.058; P = .030). ConclusionThus, the tumor burden, expressed as the number of metastatic sites at EGFR-TKI treatment, and rapid tumor progression at PD were predictive of inferior survival in patients with lung adenocarcinoma with activating EGFR mutations.

O2. Frequency of EGFR gene mutations in patients with NSCLC in Czech Republic

Background: Worldwide, lung cancer is the most common cause of cancer-related death, and was responsible for 1.56 million deaths annually, as of 2012. The situation in the Czech Republic is identical, lung cancer is the leading cause of mortality due to malignant tumours, with 5,572 cancer related deaths in 2010. The detection of epidermal growth factor receptor (EGFR) gene mutations can guide targeted therapies for non-small cell lung cancer (NSCLC), because most of these mutations are strongly associated with clinical response to EGFR tyrosine kinase inhibitors. The aim of this study was to verify the frequency of EGFR mutations in a large group of Caucasians patients with advanced stage non-squamous NSCLC. Methods: Analysed data were collected in the period from November 15, 2011 to June 30, 2013 in three pneumo-oncological centres. Totally, 486 patients diagnosed with adenocarcinoma (459 cases) or NSCLC-NOS (27 cases) were tested for EGFR mutations, consisting of 284 males and 202 females, with median age of 66.5 years. Results: EGFR mutations were detected in 74 patients (15.2%). Mutations were shown in 25 (8.8%) men vs. 49 (24.3%) women (P<0.001), in 12 (6.7%) smokers vs. 20 (11.2%) ex-smokers vs. 38 (37.3%) non-smokers (P<0.001), and in 32 (15.7%) patients under 65 years of age vs. 42 (14.9%) patients over 65 years of age (P=0.898). Conclusions: The authors showed that the overall frequency of EGFR mutations in patients with non-squamous NSCLC was 15.2%, most commonly in female patients and non-smokers. The study results are in concordance with recent studies in which the frequency of EGFR mutations varies between 8-23% in Caucasian population. This information can help us to estimate number of patients who will need EGFR tyrosine kinase inhibitors in the first line of treatment.

AXL inhibition sensitizes mesenchymal cancer cells to antimitotic drugs.

Molecularly targeted drug therapies have revolutionized cancer treatment; however, resistance remains a major limitation to their overall efficacy. Epithelial-to-mesenchymal transition (EMT) has been linked to acquired resistance to tyrosine kinase inhibitors (TKI), independent of mutational resistance mechanisms. AXL is a receptor tyrosine kinase associated with EMT that has been implicated in drug resistance and has emerged as a candidate therapeutic target. Across 643 human cancer cell lines that were analyzed, elevated AXL was strongly associated with a mesenchymal phenotype, particularly in triple-negative breast cancer and non-small cell lung cancer. In an unbiased screen of small-molecule inhibitors of cancer-relevant processes, we discovered that AXL inhibition was specifically synergistic with antimitotic agents in killing cancer cells that had undergone EMT and demonstrated associated TKI resistance. However, we did not find that AXL inhibition alone could overcome acquired resistance to EGFR TKIs in the EMT setting, as previously reported. These findings reveal a novel cotreatment strategy for tumors displaying mesenchymal features that otherwise render them treatment refractory.

O3-12-3 - Survival Outcome Assessed According to Tumor Response and Shrinkage Pattern in Egfr(+)Nsclc Treated with Egfr-Tki

Introduction: Somatic mutations in the epidermal growth factor receptor gene (EGFR) are associated with a marked therapeutic response to EGFR-tyrosine kinase inhibitors (TKIs) in patients with advanced non-small cell lung cancer (NSCLC). Clinical indicators of the likely survival benefit of EGFR-TKI treatment in NSCLC patients with EGFR mutations have not been identified, however. We therefore evaluated progression-free survival (PFS) and overall survival (OS) according to tumor response and tumor shrinkage pattern in such patients.Methods: Among 145 EGFR mutation-positive NSCLC patients treated with EGFR-TKIs, 68 individuals were selected for analysis.Results: Of the 68 selected patients, 6 achieved a complete response (CR), 42 a partial response (PR), and 14 stable disease (SD). Both PFS and OS were significantly longer in patients who achieved a CR or PR than in those who experienced SD. Multivariate analysis showed that a response (CR or PR) to EGFR-TKIs was significantly associated with both PFS and OS. Among the CR/PR group, the median maximal tumor shrinkage relative to baseline was 56%, and the median time to response (TTR) was 4.2 weeks. The subsets of these patients who experienced rapid tumor regression (TTR of =< 4.2 weeks) or a high degree of tumor shrinkage (=> 56%) did not show a more favorable PFS or OS compared with those who experienced slow tumor regression or a low degree of tumor shrinkage.Conclusion: Response (CR or PR) may represent the optimal surrogate for efficacy among EGFR mutation positive NSCLC patients treated with EGFR-TKIs.

Abstract 1834: The molecular characters of acquired resistant non-small cell lung cancer cells to afatinib

Abstract [Background] The non-small cell lung carcinomas (NSCLCs) with activating epidermal growth factor receptor (EGFR) gene mutation show significant response to EGFR-tyrosine kinase inhibitors (TKIs). However, almost all of them acquire resistance to EGFR-TKIs due to various mechanisms, such as T790M mutation or MET amplification. Afatinib is an irreversible TKI for EGFR and HER2 and known to be effective to the EGFR T790M variant. However, a half of the patient responded afatinib acquire resistance by 12 months in clinical use. Thus, to unravel the mechanisms of the resistances and to overcome them are still important issues of EGFR-TKI treatment. In this study, we established various kinds of afatinib resistant cell lines with changing drug exposing condition as we previously reported (Shien K. et al Cancer Res. 2013). We also investigated these cell lines to understand mechanisms of afatinib resistances in NSCLC. [Materials and Methods] Afatinib resistant cell lines were established using four NSCLC cell lines with activating EGFR mutations by exposing afatinib with different procedures, escalation from 1 nM, escalation from 10 nM, intermittent 2 µM and continuous 2 µM procedures. These cell lines were investigated about previously reported resistant related features, T790M mutation, MET amplification, and epithelial to mesenchymal transition (EMT). ALDH1A1 and ABCB1 expression, which were putative stem cell markers, were also investigated. Proliferative inhibitions with various drugs were evaluated using MTS assay. [Results] There were no T790M mutations in all afatinib resistant cell lines. HCC827 sublines exposed afatinib with escalation from 1 nM and intermittent 2 µM, and HCC4011 sublines showed MET amplification. HCC827 sublines exposed afatinib with escalation from 10 nM and continuous 2 µM, and HCC4006 sublines showed EMT features. Furthermore, ALDH1A1 and ABCB1 expression were up-regulated in HCC827 afatinib resistant sublines with EMT features. Next, we examined the effect of afatinib or crizotinib for the afatinib resistant cell line with MET amplification. We found that combination therapy with afatinib and crizotinib was effective to the MET amplified cell. The combination exposure was continued to establish a resistant subline (HCC827-ACR) to combination therapy. The HCC827-ACR was also showed EMT features and up-regulation of stem cell markers. [Conclusions] As in the cases of reversible EGFR-TKI, MET amplification, EMT, and stem cell-like features were emerged in afatinib resistant cells. To develop strategies to resistant cells with stem cell-like property may be critical part to overcome EGFR-TKI resistance. Citation Format: Shinsuke Hashida, Shinichi Toyooka, Tomoaki Ohtsuka, Ken Suzawa, Kazuhiko Shien, Hiromasa Yamamoto, Junichi Soh, Hiroaki Asano, Kazunori Tsukuda, Shinichiro Miyoshi. The molecular characters of acquired resistant non-small cell lung cancer cells to afatinib. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1834. doi:10.1158/1538-7445.AM2014-1834

Impact of EGFR mutation detection methods on the efficacy of erlotinib in patients with advanced EGFR-wild type lung adenocarcinoma.

IntroductionMethods used for epidermal growth factor receptor (EGFR) mutation testing vary widely. The impact of detection methods on the rates of response to EGFR-tyrosine kinase inhibitors (TKIs) in EGFR-wild type (wt) lung adenocarcinoma patients is unknown.MethodsWe recruited the Group-I patients to evaluate the efficacy of erlotinib in patients with EGFR-wt lung adenocarcinoma by either direct sequencing (DS) or mutant type-specific sensitive (MtS) methods in six medical centers in Taiwan. Cross recheck of EGFR mutations was performed in patients who achieved objective response to erlotinib and had adequate specimens. The independent Group-II lung adenocarcinoma patients whose EGFR mutation status determined by DS were recruited to evaluate the potential limitations of three MtS methods.ResultsIn Group-I analysis, 38 of 261 EGFR-wt patients (14.6%) achieved partial response to erlotinib treatment. Nineteen patients (50.0%) had adequate specimens for cross recheck of EGFR mutations and 10 of them (52.6%) had changes in EGFR mutation status, 5 in 10 by DS and 5 in 9 by MtS methods originally. In Group-II analysis, 598 of 996 lung adenocarcinoma patients (60.0%) had detectable EGFR mutations. The accuracy rates of the three MtS methods, MALDI-TOF MS, Scorpions ARMS and Cobas, were 87.8%, 86.8% and 85.8%, respectively.ConclusionsA significant portion of the erlotinib responses in EGFR-wt lung adenocarcinoma patients were related to the limitations of detection methods, not only DS but also MtS methods with similar percentages. Prospective studies are needed to define the proper strategy for EGFR mutation testing.

Sequential treatment of icotinib after first-line pemetrexed in advanced lung adenocarcinoma with unknown EGFR gene status.

In non-small cell lung cancer (NSCLC), the well-developed epidermal growth factor receptor (EGFR) is an important therapeutic target. EGFR activating gene mutations have been proved strongly predictive of response to EGFR-tyrosine kinase inhibitors (TKI) in NSCLC. However, both in daily clinical practice and clinical trials, patients with unknown EGFR gene status (UN-EGFR-GS) are very common. In this study, we assessed efficacy and tolerability of sequential treatment of first-line pemetrexed followed by icotinib in Chinese advanced lung adenocarcinoma with UN-EGFR-GS. We analyzed 38 patients with advanced lung adenocarcinoma with UN-EGFR-GS treated with first-line pemetrexed-based chemotherapy followed by icotinib as maintenance or second-line therapy. The response rates to pemetrexed and icotinib were 21.1% and 42.1%, respectively. The median overall survival was 27.0 months (95% CI, 19.7-34.2 months). The 12-month overall survival probability was 68.4%. The most common toxicities observed in icotinib phase were rashes, diarrheas, and elevated aminotransferase. Subgroup analysis indicated that the overall survival is correlated with response to icotinib. The sequence of first-line pemetrexed-based chemotherapy followed by icotinib treatment is a promising option for advanced lung adenocarcinoma with UN-EGFR-GS in China.