Kye Young Lee

Afatinib versus cisplatin plus gemcitabine for first-line treatment of Asian patients with advanced non-small-cell lung cancer harbouring EGFR mutations (LUX-Lung 6): an open-label, randomised phase 3 trial

BACKGROUND Afatinib-an oral irreversible ErbB family blocker-improves progression-free survival compared with pemetrexed and cisplatin for first-line treatment of patients with EGFR mutation-positive advanced non-small-cell lung cancer (NSCLC). We compared afatinib with gemcitabine and cisplatin-a chemotherapy regimen widely used in Asia-for first-line treatment of Asian patients with EGFR mutation-positive advanced NSCLC. METHODS This open-label, randomised phase 3 trial was done at 36 centres in China, Thailand, and South Korea. After central testing for EGFR mutations, treatment-naive patients (stage IIIB or IV cancer [American Joint Committee on Cancer version 6], performance status 0-1) were randomly assigned (2:1) to receive either oral afatinib (40 mg per day) or intravenous gemcitabine 1000 mg/m(2) on day 1 and day 8 plus cisplatin 75 mg/m(2) on day 1 of a 3-week schedule for up to six cycles. Randomisation was done centrally with a random number-generating system and an interactive internet and voice-response system. Randomisation was stratified by EGFR mutation (Leu858Arg, exon 19 deletions, or other; block size three). Clinicians and patients were not masked to treatment assignment, but the independent central imaging review group were. Treatment continued until disease progression, intolerable toxic effects, or withdrawal of consent. The primary endpoint was progression-free survival assessed by independent central review (intention-to-treat population). This study is registered with ClinicalTrials.gov, NCT01121393. FINDINGS 910 patients were screened and 364 were randomly assigned (242 to afatinib, 122 to gemcitabine and cisplatin). Median progression-free survival was significantly longer in the afatinib group (11·0 months, 95% CI 9·7-13·7) than in the gemcitabine and cisplatin group (5·6 months, 5·1-6·7; hazard ratio 0·28, 95% CI 0·20-0·39; p<0·0001). The most common treatment-related grade 3 or 4 adverse events in the afatinib group were rash or acne (35 [14·6%] of 239 patients), diarrhoea (13 [5·4%]), and stomatitis or mucositis (13 [5·4%]), compared with neutropenia (30 [26·5%] of 113 patients), vomiting (22 [19·5%]), and leucopenia (17 [15·0%]) in the gemcitabine and cisplatin group. Treatment-related serious adverse events occurred in 15 (6·3%) patients in the afatinib group and nine (8·0%) patients in the gemcitabine and cisplatin group. INTERPRETATION First-line afatinib significantly improves progression-free survival with a tolerable and manageable safety profile in Asian patients with EGFR mutation-positive advanced lung NSCLC. Afatinib should be considered as a first-line treatment option for this patient population. FUNDING Boehringer Ingelheim.

Afatinib versus cisplatin-based chemotherapy for EGFR mutation-positive lung adenocarcinoma (LUX-Lung 3 and LUX-Lung 6): analysis of overall survival data from two randomised, phase 3 trials

BACKGROUND We aimed to assess the effect of afatinib on overall survival of patients with EGFR mutation-positive lung adenocarcinoma through an analysis of data from two open-label, randomised, phase 3 trials. METHODS Previously untreated patients with EGFR mutation-positive stage IIIB or IV lung adenocarcinoma were enrolled in LUX-Lung 3 (n=345) and LUX-Lung 6 (n=364). These patients were randomly assigned in a 2:1 ratio to receive afatinib or chemotherapy (pemetrexed-cisplatin [LUX-Lung 3] or gemcitabine-cisplatin [LUX-Lung 6]), stratified by EGFR mutation (exon 19 deletion [del19], Leu858Arg, or other) and ethnic origin (LUX-Lung 3 only). We planned analyses of mature overall survival data in the intention-to-treat population after 209 (LUX-Lung 3) and 237 (LUX-Lung 6) deaths. These ongoing studies are registered with ClinicalTrials.gov, numbers NCT00949650 and NCT01121393. FINDINGS Median follow-up in LUX-Lung 3 was 41 months (IQR 35-44); 213 (62%) of 345 patients had died. Median follow-up in LUX-Lung 6 was 33 months (IQR 31-37); 246 (68%) of 364 patients had died. In LUX-Lung 3, median overall survival was 28.2 months (95% CI 24.6-33.6) in the afatinib group and 28.2 months (20.7-33.2) in the pemetrexed-cisplatin group (HR 0.88, 95% CI 0.66-1.17, p=0.39). In LUX-Lung 6, median overall survival was 23.1 months (95% CI 20.4-27.3) in the afatinib group and 23.5 months (18.0-25.6) in the gemcitabine-cisplatin group (HR 0.93, 95% CI 0.72-1.22, p=0.61). However, in preplanned analyses, overall survival was significantly longer for patients with del19-positive tumours in the afatinib group than in the chemotherapy group in both trials: in LUX-Lung 3, median overall survival was 33.3 months (95% CI 26.8-41.5) in the afatinib group versus 21.1 months (16.3-30.7) in the chemotherapy group (HR 0.54, 95% CI 0.36-0.79, p=0.0015); in LUX-Lung 6, it was 31.4 months (95% CI 24.2-35.3) versus 18.4 months (14.6-25.6), respectively (HR 0.64, 95% CI 0.44-0.94, p=0.023). By contrast, there were no significant differences by treatment group for patients with EGFR Leu858Arg-positive tumours in either trial: in LUX-Lung 3, median overall survival was 27.6 months (19.8-41.7) in the afatinib group versus 40.3 months (24.3-not estimable) in the chemotherapy group (HR 1.30, 95% CI 0.80-2.11, p=0.29); in LUX-Lung 6, it was 19.6 months (95% CI 17.0-22.1) versus 24.3 months (19.0-27.0), respectively (HR 1.22, 95% CI 0.81-1.83, p=0.34). In both trials, the most common afatinib-related grade 3-4 adverse events were rash or acne (37 [16%] of 229 patients in LUX-Lung 3 and 35 [15%] of 239 patients in LUX-Lung 6), diarrhoea (33 [14%] and 13 [5%]), paronychia (26 [11%] in LUX-Lung 3 only), and stomatitis or mucositis (13 [5%] in LUX-Lung 6 only). In LUX-Lung 3, neutropenia (20 [18%] of 111 patients), fatigue (14 [13%]) and leucopenia (nine [8%]) were the most common chemotherapy-related grade 3-4 adverse events, while in LUX-Lung 6, the most common chemotherapy-related grade 3-4 adverse events were neutropenia (30 [27%] of 113 patients), vomiting (22 [19%]), and leucopenia (17 [15%]). INTERPRETATION Although afatinib did not improve overall survival in the whole population of either trial, overall survival was improved with the drug for patients with del19 EGFR mutations. The absence of an effect in patients with Leu858Arg EGFR mutations suggests that EGFR del19-positive disease might be distinct from Leu858Arg-positive disease and that these subgroups should be analysed separately in future trials. FUNDING Boehringer Ingelheim.

Detection and comparison of peptide nucleic acid-mediated real-time polymerase chain reaction clamping and direct gene sequencing for epidermal growth factor receptor mutations in patients with non-small cell lung cancer

EGFR tyrosine kinase inhibitors (EGFR-TKIs) are recommended as first-line therapy in patients with advanced, recurrent, or metastatic non-squamous non-small cell lung cancer (NSCLC) that have active EGFR mutations. The importance of rapid and sensitive methods for the detection of EGFR mutations is emphasized. The aim of this study is to examine the EGFR mutational status by both direct DNA sequencing and peptide nucleic acid (PNA)-mediated real-time PCR clamping and to evaluate the correlation between the EGFR mutational status and the clinical response to EGFR-tyrosine kinase inhibitors. Clinical specimens from 240 NSCLC patients were analyzed for EGFR mutations in exons 18, 19, 20 and 21. All clinical data and tumor specimens were obtained from 8 centers of the Korean Molecular Lung Cancer Group (KMLCG). After genomic DNA was extracted from paraffin-embedded tissue specimens, we performed PNA-mediated real-time PCR clamping and direct DNA sequencing for the detection of EGFR mutations. Of 240 tumor samples, PNA-mediated PCR clamping was used to detect genomic alterations in 83 (34.6%) samples, including 61 identified by sequencing and 22 additional samples (10 in exon 19, 9 in exon 21, and 3 in both exons); direct DNA sequencing was used to identify a total of 63 (26.3%) mutations that contained 40 deletion mutations in exon 19 (63.5%) and 18 substitution mutations (28.6%) in exon 21. PNA-mediated PCR clamping was used to identify more mutations than clinical direct sequencing, whereas clinical outcomes were not significantly different between the groups harboring activating mutations detected by each method. These data suggest that PNA-mediated real-time PCR clamping exhibits high sensitivity and is a simple procedure relative to direct DNA sequencing that is a useful screening tool for the detection of EGFR mutations in clinical settings.

Identification and characterization of proteins isolated from microvesicles derived from human lung cancer pleural effusions

Microvesicles (MVs, also known as exosomes, ectosomes, microparticles) are released by various cancer cells, including lung, colorectal, and prostate carcinoma cells. MVs released from tumor cells and other sources accumulate in the circulation and in pleural effusion. Although recent studies have shown that MVs play multiple roles in tumor progression, the potential pathological roles of MV in pleural effusion, and their protein composition, are still unknown. In this study, we report the first global proteomic analysis of highly purified MVs derived from human nonsmall cell lung cancer (NSCLC) pleural effusion. Using nano‐LC–MS/MS following 1D SDS‐PAGE separation, we identified a total of 912 MV proteins with high confidence. Three independent experiments on three patients showed that MV proteins from PE were distinct from MV obtained from other malignancies. Bioinformatics analyses of the MS data identified pathologically relevant proteins and potential diagnostic makers for NSCLC, including lung‐enriched surface antigens and proteins related to epidermal growth factor receptor signaling. These findings provide new insight into the diverse functions of MVs in cancer progression and will aid in the development of novel diagnostic tools for NSCLC.

IFNα sensitizes ME‐180 human cervical cancer cells to TNFα‐induced apoptosis by inhibiting cytoprotective NF‐κB activation

Tumor necrosis factor α (TNFα) induces apoptosis of a variety of tumor cell types. The anti‐tumor effect of TNFα is often augmented by interferon (IFN) γ. We hypothesized that IFNα, which shares many biological activities with IFNγ, might also synergize with TNFα for the induction of tumor cell death. We tested our hypothesis using ME‐180 human cervical cancer cells exposed to either IFNα or TNFα alone or both. We analyzed the death of ME‐180 cells by biochemical and cytological means, and investigated the molecular mechanism underlying cytotoxic synergism between the two cytokines. We found that (i) IFNα/TNFα synergistically induced apoptosis of ME‐180 cells, which was accompanied by activation of caspases‐3 and ‐8; (ii) IFNα induced signal transducer and activator of transcription (STAT) 1 phosphorylation, and transfection of phosphorylation‐defective STAT1 dominant‐negative mutant inhibited IFNα/TNFα‐induced apoptosis; (iii) inhibition of nuclear factor κB (NF‐κB) by proteasome inhibitor MG‐132 sensitized ME‐180 cells to TNFα alone; (iv) IFNα treatment attenuated TNFα‐induced NF‐κB reporter activity, while it did not inhibit DNA binding of NF‐κB. Taken collectively, our results indicate that IFNα sensitizes ME‐180 cells to TNFα‐induced apoptosis by inhibiting TNFα‐mediated cytoprotective NF‐κB activation, and this sensitizing effect of IFNα is mediated through a STAT1‐dependent pathway.

A promoter SNP rs4073T>A in the common allele of the interleukin 8 gene is associated with the development of idiopathic pulmonary fibrosis via the IL-8 protein enhancing mode

BackgroundInterleukin-8 (IL-8) is a potent chemo-attractant cytokine responsible for neutrophil infiltration in lungs with idiopathic pulmonary fibrosis (IPF). The IL-8 protein and mRNA expression are increased in the lung with IPF. We evaluated the effect of single nucleotide polymorphisms (SNPs) of the IL-8 gene on the risk of IPF.MethodsOne promoter (rs4073T>A) and two intronic SNPs (rs2227307T>G and rs2227306C>T) of the IL-8 genes were genotyped in 237 subjects with IPF and 456 normal controls. Logistic regression analysis was applied to evaluate the association of these SNPs with IPF. IL-8 in BAL fluids was measured using a quantitative sandwich enzyme immunoassay, and promoter activity was assessed using the luciferase reporter assay.ResultsThe minor allele frequencies of rs4073T>A and rs2227307T>G were significantly lower in the 162 subjects with surgical biopsy-proven IPF and 75 subjects with clinical IPF compared with normal controls in the recessive model (OR = 0.46 and 0.48, p = 0.006 and 0.007, respectively). The IL-8 protein concentration in BAL fluids significantly increased in 24 subjects with IPF compared with 14 controls (p = 0.009). Nine IPF subjects homozygous for the rs4073 T>A common allele exhibited higher levels of the IL-8 protein compared with six subjects homozygous for the minor allele (p = 0.024). The luciferase activity of the rs4073T>A common allele was significantly higher than that of the rs4073T>A minor allele (p = 0.002).ConclusionThe common allele of a promoter SNP, rs4073T>A, may increase susceptibility to the development of IPF via up-regulation of IL-8.

Symptom and Quality of Life Improvement in LUX-Lung 6: An Open-Label Phase III Study of Afatinib Versus Cisplatin/Gemcitabine in Asian Patients With EGFR Mutation-Positive Advanced Non–small-cell Lung Cancer

Introduction: In the phase III, LUX-Lung 6 trial, afatinib prolonged progression-free survival (PFS) versus cisplatin/gemcitabine in Asian patients with epidermal growth factor receptor (EGFR) mutation-positive non–small-cell lung cancer (NSCLC). This article provides detailed assessments of patient-reported outcomes (PROs), a LUX-Lung 6 secondary end point, and explores the relationship between PFS and health-related quality of life (QoL) in these patients. Methods: Patients (n = 364) were randomized (2:1) to oral afatinib (40 mg/day) or up to six cycles of cisplatin/gemcitabine (21-day cycle; cisplatin 75 mg/m2 [d1]; gemcitabine 1000 mg/m2 [d1,8]). QoL was assessed using the European Organization for Research and Treatment of Cancer Quality of Life Questionnaire and its lung cancer-specific module. The relationship between PFS (investigator assessment and independent review) and QoL was evaluated using analysis of covariance and a longitudinal model. Results: More patients treated with afatinib versus cisplatin/gemcitabine showed improvements in global health status/QoL (p < 0.0001) and physical (p < 0.0001), role (p = 0.013), and social (p < 0.001) functioning scales. Delayed symptom deterioration and better QoL over time was also observed with afatinib. QoL measured before tumor assessment was considerably poorer for patients with progression than those without progression, with significant differences in mean scores at multiple assessment time points. Results from the longitudinal analysis consistently demonstrated a significant negative impact of progression on QoL (p < 0.0001). Conclusion: Afatinib improved PFS and PROs versus chemotherapy in EGFR mutation-positive NSCLC patients. Progression was associated with statistically significant worsening in QoL measured before tumor assessment, underscoring the value of PFS as a clinically relevant end point.

Extracellular vesicles shed from gefitinib-resistant nonsmall cell lung cancer regulate the tumor microenvironment.

Epidermal growth factor receptor (EGFR)‐tyrosine kinase inhibitors (TKIs), including gefitinib, are the first‐line treatment of choice for nonsmall cell lung cancer patients who harbor activating EGFR mutations, however, acquired resistance to EGFR‐TKIs is inevitable. The main objective of this study was to identify informative protein signatures of extracellular vesicles (EV) derived from gefitinib‐resistant nonsmall cell lung cancer cells using proteomics analysis. Nano‐LC–MS/MS analysis identified with high confidence (false discovery rate < 0.05, fold change ≥2) 664 EV proteins enriched in PC9R cells, which are resistant to gefitinib due to EGFR T790M mutation. Computational analyses suggested components of several signal transduction mechanisms including the AKT (also PKB, protein kinase B)/mTOR (mechanistic target of rapamycin) pathway are overrepresented in EV from PC9R cells. Treatment of recipient cells with EV harvested from PC9R cells increased phosphorylation of signaling molecules, and enhanced proliferation, invasion, and drug resistance to gefitinib‐induced apoptosis. Dose‐ and time‐dependent pharmaceutical inhibition of AKT/mTOR pathway overcame drug resistance of PC9R cells and those of H1975 exhibiting EGFR T790M mutation. Our findings provide new insight into an oncogenic EV protein signature regulating tumor microenvironment, and will aid in the development of novel diagnostic strategies for prediction and assessment of gefitinib resistance.

The performance of a novel amino acid multivariate index for detecting lung cancer: A case control study in Korea

INTRODUCTION Previous studies have shown that plasma free amino acid (PFAA) profiles are altered in cancer patients compared with healthy controls. A multivariate index based on PFAAs was generated from a Japanese dataset and has been previously demonstrated to be clinically valuable for discriminating patients in the early stages of lung cancer. However, it remains unclear whether similar PFAA profile changes occur in cancer patients from other populations. Therefore, this study aimed to validate the performance of this index in discriminating lung cancer patients from controls in the Korean population. METHODS Samples were collected from a total of 142 Korean subjects (72 lung cancer/70 controls) for this study. PFAAs were quantified by high-performance liquid chromatography-electrospray ionization-mass spectrometry, and the clinical performance characteristics of the amino acid multivariate index were evaluated across cancer stages and histological types. RESULTS The concentrations of several PFAAs were significantly decreased in the Korean lung cancer patients compared with the controls. Significant decreases in threonine, citrulline, histidine and tryptophan and increases in proline, isoleucine, phenylalanine and ornithine were observed, which are similar to the PFAA changes reported by a previous Japanese study. The area under the receiver-operator characteristic curve (AUC of the ROC) for the index was 0.80, and similar performances were demonstrated for the different histological types. CONCLUSIONS These results suggest that the amino acid multivariate index previously developed from a Japanese dataset has the potential to aid in the early detection of lung cancers of different histological types in Korean patients.

Treatment of Non-small Cell Lung Carcinoma after Failure of Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitor

Since the first description of non-small cell lung cancer (NSCLC) with activating epidermal growth factor receptor (EGFR) mutation as a distinct clinical entity, studies have proved EGFR tyrosine kinase inhibitors (TKIs) as a first choice of treatment. The median response duration of TKIs as a first-line treatment for EGFR mutant tumors ranges from 11 to 14 months. However, acquired resistance to EGFR-TKIs is inevitable due to various mechanisms, such as T790M, c-Met amplification, activation of alternative pathways (IGF-1, HGF, PI3CA, AXL), transformation to mesenchymal cell or small cell features, and tumor heterogeneity. Until development of a successful treatment strategy to overcome such acquired resistance, few options are currently available. Here we provide a summary of the therapeutic options after failure of first line EGFR-TKI treatment for NSCLC.