METTL14 enhances the m6A modification level of lncRNA MSTRG.292666.16 to promote the progression of non-small cell lung cancer.

BACKGROUND: m6A modification has close connection with the occurrence, development, and prognosis of tumors. This study aimed to explore the roles of m6A modification and its related mechanisms in non-small cell lung cancer (NSCLC). METHODS: NSCLC tissues and their corresponding para-cancerous tissues were collected to determine the m6A levels of total RNA/lncRNAs and the expression of m6A modification-related genes/lncRNAs. Then, A549 cells were transfected with si-METTL14 or oe-METTL14, and the cell transfection efficiency was assessed. Subsequently, the viability, apoptosis, cell colony formation, migration and invasion of the different cells were determined. Finally, the nude mouse tumorigenicity experiments were performed to observe the effects of METTL14 in vivo. RESULTS: Compared to the para-NSCLC tissues, the m6A level and METTL14 expression were both significantly increased in the NSCLC tissues (P < 0.05). Based on the expression of METTL14 in the different cell lines, A549 cells were chosen for further experiments. Then, the A549 cells with METTL14 knockdown and overexpression were successfully established, as well as it was found that METTL14 knockdown could inhibit the viability, colony formation, migration, and invasion of A549 cells, while facilitate their apoptosis. In vivo experiments also showed that METTL14 knockdown could inhibit tumor formation and growth. Additionally, the m6A level of MSTRG.292666.16 was higher in the NSCLC tissues; and after METTL14 knockdown, the expression and m6A level of MSTRG.292666.16 were both significantly reduced in A549 cells, and vice versa. CONCLUSION: METTL14 may promote the progression of NSCLC through up-regulating MSTRG.292666.16 and enhance its m6A modification level.

Exosomes derived from M2 type tumor-associated macrophages promote osimertinib resistance in non-small cell lung cancer through MSTRG.292666.16-miR-6836-5p-MAPK8IP3 axis.

BACKGROUND: Osimertinib resistance limits the treatment of epidermal growth factor receptor-(EGFR)-mutated non-small-cell lung carcinoma (NSCLC). The mechanisms of osimertinib resistance need to be elucidated to determine alternative treatment strategies. This study explores the role of M2 type tumor-associated macrophage (TAM)-derived exosomal MSTRG.292666.16 in osimertinib resistance, and its related competing endogenous RNA (ceRNA) mechanism. METHODS: M2 type TAMs were induced with 200 ng/mL phorbol 12-myristate 13-acetate, 20 ng/mL IL-4 and IL-13, and M2 type macrophage markers were measured by RT-qPCR. Next, the exosomes were isolated and characterized. Tumor formation in nude mice was conducted using H1975 cells under different treatment conditions. Small RNA sequencing was performed on exosomes derived from sensitive and resistant plasma, and ceRNA networks were constructed. Fluorescence in situ hybridization was used to observe the localization of MSTRG.292666.16, and a ceRNA network (MSTRG.292666.16-miR-6836-5p-MAPK8IP3) was selected for further validation. RESULTS: M2 type TAMs, and M2 type TAM-derived exosomes were successfully induced and isolated. Nude mice results showed that M2 type TAM-derived exosomes and MSTRG.292666.16 overexpression significantly increased tumor volume after administration of osimertinib for 4 weeks. M2 type TAMs were found in the resistant plasma, and MSTRG.292666.16 localized in the cytoplasm of H1975 cells. In addition, the genes in the ceRNA networks were significantly enriched in eight GO terms and seven KEGG pathways, including the MAPK signaling pathway. Subsequently, the levels of MSTRG.292666.16 and MAPK8IP3 significantly increased in both resistant plasma-derived exosomes and M2 type TAM-derived exosomes, while miR-6836-5p levels were significantly reduced. Finally, MSTRG.292666.16, miR-6836-5p, and MAPK8IP3 were part of the same network. CONCLUSIONS: M2 type TAM-derived exosomes promoted osimertinib resistance in NSCLC by regulating the MSTRG.292666.16/miR-6386-5p/MAPK8IP3 axis.

A novel virtual barcode strategy for accurate panel-wide variant calling in circulating tumor DNA.

BACKGROUND: Hybrid capture-based next-generation sequencing of DNA has been widely applied in the detection of circulating tumor DNA (ctDNA). Various methods have been proposed for ctDNA detection, but low-allelic-fraction (AF) variants are still a great challenge. In addition, no panel-wide calling algorithm is available, which hiders the full usage of ctDNA based 'liquid biopsy'. Thus, we developed the VBCALAVD (Virtual Barcode-based Calling Algorithm for Low Allelic Variant Detection) in silico to overcome these limitations. RESULTS: Based on the understanding of the nature of ctDNA fragmentation, a novel platform-independent virtual barcode strategy was established to eliminate random sequencing errors by clustering sequencing reads into virtual families. Stereotypical mutant-family-level background artifacts were polished by constructing AF distributions. Three additional robust fine-tuning filters were obtained to eliminate stochastic mutant-family-level noises. The performance of our algorithm was validated using cell-free DNA reference standard samples (cfDNA RSDs) and normal healthy cfDNA samples (cfDNA controls). For the RSDs with AFs of 0.1, 0.2, 0.5, 1 and 5%, the mean F1 scores were 0.43 (0.25~0.56), 0.77, 0.92, 0.926 (0.86~1.0) and 0.89 (0.75~1.0), respectively, which indicates that the proposed approach significantly outperforms the published algorithms. Among controls, no false positives were detected. Meanwhile, characteristics of mutant-family-level noise and quantitative determinants of divergence between mutant-family-level noises from controls and RSDs were clearly depicted. CONCLUSIONS: Due to its good performance in the detection of low-AF variants, our algorithm will greatly facilitate the noninvasive panel-wide detection of ctDNA in research and clinical settings. The whole pipeline is available at https://github.com/zhaodalv/VBCALAVD.

SPOP promotes SIRT2 degradation and suppresses non-small cell lung cancer cell growth.

SIRT2 is a NAD-dependent deacetylase and inhibition of SIRT2 has a broad anticancer activity. Here we report that SPOP binds to SIRT2 and mediates its degradation by the 26S proteasome, which can be blocked by MG132 treatment. We also found that the levels of SPOP significantly decreased, while the levels of SIRT2 significantly increased in non-small cell lung cancer (NSCLC) cell lines, compared to normal bronchial epithelial cell line and NSCLC specimens, compared to the paired non-tumor lung tissue. Furthermore, SPOP can suppress NSCLC cell growth. Notably, mutations in NSCLC inhibit the abilities of SPOP to degrade SIRT2 and suppress NSCLC cell growth. These results reveal a novel regulation of SIRT2 by SPOP mediated degradation, which is important for the growth of lung tumor cells.

Overexpression of miR-100 inhibits cancer growth, migration, and chemosensitivity in human NSCLC cells through fibroblast growth factor receptor 3.

Nonsmall cell lung cancer (NSCLC) is a commonly occurring lung cancer. A combination of molecular biological treatments with regular chemotherapy may result in improved therapeutic outcome. Here, we reported significantly higher levels of fibroblast growth factor receptor 3 (FGFR3) and significantly lower levels of miR-100 in the NSCLC specimen, compared to the paired NSCLC-adjacent normal lung tissues. Moreover, the levels of FGFR3 and miR-100 were inversely correlated. Bioinformatics analyses followed by luciferase reporter assay showed that miR-100 bound to the 3'-UTR of FGFR3 messenger RNA (mRNA) to inhibit its translation. Overexpression of miR-100 in NSCLC cells decreased FGFR3 protein levels, whereas inhibition of miR-100 increased FGFR3 protein levels, without affecting FGFR3 mRNA levels. Furthermore, overexpression of miR-100 suppressed cancer growth, migration, and chemosensitivity in NSCLC cells, while inhibition of miR-100 significantly facilitated them. Taken together, our data demonstrate that miR-100 may inhibit NSCLC through FGFR3.

[Interventional therapy for lung cancer patients with superior vena cava syndrome].

OBJECTIVE: To investigate the method, therapeutic effect and safety of interventional therapy for lung cancer patients with superior vena cava syndrome (SVCS). METHODS: Fifty-two cases of lung cancer with SVCS who received interventional therapy in our hospital between Jan to Dec 2011 were included in this study. Of the 52 cases, 50 cases had successfully carried out superior vena cava stent implantation. The distal venous pressure was measured before and after angioplasty, and the results were assessed by Wilcoxon matched-pairs test. In addition, the 50 patients were followed up and the therapeutic effect and postoperative survival rate were evaluated. RESULTS: The mean distal venous pressure in the 50 patients was significantly decreased from preoperative (28.2 ± 1.9)cm H2O to postoperative (8.7 ± 0.5)cm H2O (P = 0.0085). The efficacy of the treatment was as follows: complete remission (20/52, 38.5%), partial remission (28/52, 53.8%), ineffective 4 (4/52, 7.7%), and total effective rate 92.3%. The complications after angioplasty and stent implantation included chest pain (12 cases, 23.1%), hematoma at the puncture site (5 cases, 9.6%), and fever (2 cases, 3.8%). No serious complications such as massive hemorrhage, pulmonary embolism and stent migration into the cardiac atrium were observed. The rate of postoperative restenosis was low (2/52, 3.8%). For the SCLC group, the objective effective rate was 74.1% and 1-year survival rate was 21.0%. For the NSCLC group, the objective effective rate was 21.7% and 1-year survival rate was 35.0%. CONCLUSIONS: For lung cancer patients with SVCS, interventional therapy may relief obstruction effectively, promote blood flow recovery, and relieve clinical symptoms. Interventional therapy with endovascular angioplasty and stenting may be highly recommended as the first choice for palliative treatment of SVCS. It is an effective initial palliative treatment. However, subsequent comprehensive anti-tumor treatment is necessary.

Temporal genomic heterogeneity guiding individualized therapy in recurrent non-small cell lung cancer.

Introduction: Despite the benefit of adjuvant systemic therapy for patients with resected non-small cell lung cancer (NSCLC), the risk of postoperative recurrence remains high. Our objective was to characterize temporal genetic heterogeneity between primary resected and recurrent tumors, and its impact on treatment outcomes. Methods: In this study, next-generation sequencing (NGS) testing was performed on tissue specimens and circulating tumor DNA (ctDNA) collected at postoperative recurrence, and results were compared to the genotypes of initial surgical specimens. Results: Of forty-five patients with matched primary and post-operative recurrent tumors, EGFR status switched in 17 patients (37.8%) at post-operative recurrence and 28 patients (62.2%) had no genotype change (17 mutant, 11 wild-type). Based on the changes of EGFR status, patients were divided into 4 groups. Following subsequent treatment with EGFR TKI o chemotherapy: In group A, with sustained sensitive mutation, the percentage achieving partial response (PR) was the highest, at 72.2%, the median progression-free survival (PFS) was 17 months, and the median overall survival (OS) was 44.0 months respectively; In group B, with genotype changed from wild-type to mutant, 50% achieved PR, PFS was 10 months, and OS was 35 months; In group C, in which mutant status shifted to wild-type or new co-mutation emerged, the percentage achieving PR was 30%, PFS was 9 months, and OS was 35 months. In group D, with sustained wild type, the percentage achieving PR was 27.3%, PFS was 8 months, and OS was 22 months. Discussion: Genotypic shift between paired primary and post-operative recurrent tumors was not infrequent, and this temporal genomic heterogeneity substantially impacted subsequent treatment outcomes.

DNMT3A governs tyrosine kinase inhibitors responses through IAPs and in a cell senescence-dependent manner in non-small cell lung cancer.

Patients with non-small cell lung cancer (NSCLC) treated with tyrosine kinase inhibitors (TKIs) inevitably exhibit drug resistance, which diminishes therapeutic effects. Nonetheless, the molecular mechanisms of TKI resistance in NSCLC remain obscure. In this study, data from clinical and TCGA databases revealed an increase in DNMT3A expression, which was correlated with a poor prognosis. Using NSCLC organoid models, we observed that high DNMT3A levels reduced TKI susceptibility of NSCLC cells via upregulating inhibitor of apoptosis proteins (IAPs). Simultaneously, the DNMT3Ahigh subset, which escaped apoptosis, underwent an early senescent-like state in a CDKN1A-dependent manner. Furthermore, the cellular senescence induced by TKIs was observed to be reversible, whereas DNMT3Ahigh cells reacquired their proliferative characteristics in the absence of TKIs, resulting in subsequent tumour recurrence and growth. Notably, the blockade of DNMT3A/IAPs signals enhanced the efficacy of TKIs in DNMT3Ahigh tumour-bearing mice, which represented a promising strategy for the effective treatment of NSCLC.

Individualized Chemotherapy in Advanced NSCLC Patients Based on mRNA Levels of BRCA1 and RRM1.

OBJECTIVE: Experimental evidence suggests that the overexpression of breast cancer-specific tumor suppressor protein 1 (BRCA1) gene enhances sensitivity to docetaxel and resistance to cisplatin and ribonucleotide reductase M1 (RRM1) gene overexpression enhances resistance to gemcitabine. To further examine the effect of BRCA1 and RRM1 mRNA levels on outcome in advanced non-small cell lung cancer (NSCLC), we performed this non-randomized phase II clinical trial which tested the hypothesis that customized therapy would confer improved outcome over non-customized therapy. METHODS: RNA was isolated from fresh tumor tissue. Patients received chemotherapy regimen based on their BRCA1 and RRM1 mRNA levels: both low-cisplatin plus gemcitabine (GP); both high-vinorelbine plus cisplatin (NP); BRCA1 low and RRM1 high-cisplatin plus docetaxel (TP); BRCA1 high and RRM1 low-vinorelbine plus gemcitabine (GN). RESULTS: From Dec 2005 to Nov 2008, 94 metastatic and locally advanced NSCLC patients from our institute were enrolled in this study. The median age was 58 years old. Among them, 21 patients received GP, 30 patients received TP and 43 patients received NP chemotherapy. GP group had a higher response rate, and longer median time to progression (TTP) and median overall survival (OS) time than the other 2 groups. The response rates in the GP, TP and NP groups were 42.9%, 36.7% and 27.9%, respectively (P=0.568). The median TTP was 5.6, 5.0, 4.8 months (P=0.975), respectively, and the median OS time was 12.5, 11.0, 9.7 months (P=0.808), respectively. CONCLUSION: Chemotherapy customized according to BRCA1 and RRM1 expression levels is associated with higher response rate and longer TTP and OS time in the GP group. This suggests that BRCA1 and RRM1 mRNA levels could be used as biomarkers in individual therapy in NSCLC.

Erlotinib versus chemotherapy as first-line treatment for patients with advanced EGFR mutation-positive non-small-cell lung cancer (OPTIMAL, CTONG-0802): a multicentre, open-label, randomised, phase 3 study.

BACKGROUND: Activating mutations in EGFR are important markers of response to tyrosine kinase inhibitor (TKI) therapy in non-small-cell lung cancer (NSCLC). The OPTIMAL study compared efficacy and tolerability of the TKI erlotinib versus standard chemotherapy in the first-line treatment of patients with advanced EGFR mutation-positive NSCLC. METHODS: We undertook an open-label, randomised, phase 3 trial at 22 centres in China. Patients older than 18 years with histologically confirmed stage IIIB or IV NSCLC and a confirmed activating mutation of EGFR (exon 19 deletion or exon 21 L858R point mutation) received either oral erlotinib (150 mg/day) until disease progression or unacceptable toxic effects, or up to four cycles of gemcitabine plus carboplatin. Patients were randomly assigned (1:1) with a minimisation procedure and were stratified according to EGFR mutation type, histological subtype (adenocarcinoma vs non-adenocarcinoma), and smoking status. The primary outcome was progression-free survival, analysed in patients with confirmed disease who received at least one dose of study treatment. The trial is registered at ClinicalTrials.gov, number NCT00874419, and has completed enrolment; patients are still in follow-up. FINDINGS: 83 patients were randomly assigned to receive erlotinib and 82 to receive gemcitabine plus carboplatin; 82 in the erlotinib group and 72 in the chemotherapy group were included in analysis of the primary endpoint. Median progression-free survival was significantly longer in erlotinib-treated patients than in those on chemotherapy (13.1 [95% CI 10.58-16.53] vs 4.6 [4.21-5.42] months; hazard ratio 0.16, 95% CI 0.10-0.26; p<0.0001). Chemotherapy was associated with more grade 3 or 4 toxic effects than was erlotinib (including neutropenia in 30 [42%] of 72 patients and thrombocytopenia in 29 [40%] patients on chemotherapy vs no patients with either event on erlotinib); the most common grade 3 or 4 toxic effects with erlotinib were increased alanine aminotransferase concentrations (three [4%] of 83 patients) and skin rash (two [2%] patients). Chemotherapy was also associated with increased treatment-related serious adverse events (ten [14%] of 72 patients [decreased platelet count, n=8; decreased neutrophil count, n=1; hepatic dysfunction, n=1] vs two [2%] of 83 patients [both hepatic dysfunction]). INTERPRETATION: Compared with standard chemotherapy, erlotinib conferred a significant progression-free survival benefit in patients with advanced EGFR mutation-positive NSCLC and was associated with more favourable tolerability. These findings suggest that erlotinib is important for first-line treatment of patients with advanced EGFR mutation-positive NSCLC. FUNDING: F Hoffmann-La Roche Ltd (China); Science and Technology Commission of Shanghai Municipality.

High-level mRNA of excision repair cross-complementation group 1 gene is associated with poor outcome of platinum-based doublet chemotherapy of advanced nonsmall cell lung cancer patients.

BACKGROUND: DNA excision repair gene expression plays a pivotal role in the resistance of platinum-based doublet chemotherapy of nonsmall cell lung cancer (NSCLC) in clinical practice. The aim of this study was to investigate the relationship of the excision repair cross-complementation group 1 (ERCC1) mRNA level in fresh tumor tissue and the efficacy of platinum-based chemotherapy of NSCLC. PATIENTS AND METHODS: 100 patients diagnosed with NSCLC, including stage IIIB with malignant pleural effusion, stage IV, and recurrent disease, were enrolled in this study. Before clinical treatment, tumor biopsy specimens were collected, and total RNA was purified to analyze ERCC1 mRNA level by real-time polymerase chain reaction assay. All patients were treated with platinum-based third-generation doublet chemotherapy. RESULTS: Patient median age was 60 years. Forty-seven patients had NSCLC with low expression of ERCC1 mRNA, and 53 patients had high expression of ERCC1 mRNA. Although the ERCC1 mRNA level was not correlated with the response rate (p = .665) and progression-free survival (median, 6.4 months vs. 5.5 months; p = .446), the high level of ERCC1 mRNA demonstrated a significant association with poor overall survival (median, 11 months vs. 17 months; p = .02). High level of ERCC1 mRNA was an independent prognostic factor for poor overall survival (p < .001) along with lack of disease control (p < .001). CONCLUSIONS: High level of ERCC1 mRNA may serve as a useful prognostic factor for poor outcome in advanced NSCLC patients treated with platinum-based third-generation doublet chemotherapy and may provide important information to guide tailored therapy of NSCLC patients.

Predictive effects of ERCC1 and XRCC3 SNP on efficacy of platinum-based chemotherapy in advanced NSCLC patients.

OBJECTIVE: The purpose of the study was to investigate whether single-nucleotide polymorphisms of deoxyribonucleic acid repair gene excision repair cross-complementing group 1 at codon 118 and X-ray repair cross-complementing group 3 at codon 241 affected clinical outcomes in advanced non-small cell lung cancer patients receiving first-line platinum-based chemotherapy. METHODS: A total of 130 patients treated with platinum-based doublets were examined for genotyping of excision repair cross-complementing group 1 118 and X-ray repair cross-complementing group 3 241 in peripheral blood lymphocytes with the method of the TaqMan assay plus the real-time polymerase chain reaction method. Multivariate logistic or Cox's regression analyses were used to adjust for possible confounding variables. RESULTS: There were no differences in clinical characteristics among the different single-nucleotide polymorphisms. Overall response rate in the 130 patients was 20% with 85.4% of disease control rate. Followed up to 31 March 2008, there were 47 patients still alive. Overall survival was 15 months. No relationship was found between excision repair cross-complementing group 1 or X-ray repair cross-complementing group 3 single-nucleotide polymorphisms and tumor response to platinum-based chemotherapy. A significant correlation was found between excision repair cross-complementing group 1 118 C/T single-nucleotide polymorphisms and survival (P = 0.003). In the multivariate model, the survival was highly related with excision repair cross-complementing group 1 118 C/T or T/T genotypes and tumor response to chemotherapy. CONCLUSIONS: Overall survival was significantly improved in the patients with excision repair cross-complementing group 1 118 T/T or C/T treated by platinum-based chemotherapy.

Exosomal long non-coding RNA MSTRG.292666.16 is associated with osimertinib (AZD9291) resistance in non-small cell lung cancer.

Acquired resistance of osimertinib is encountered in clinic treatment of non-small cell lung cancer (NSCLC). However, the molecular mechanisms of osimertinib resistance are not fully revealed. This study aimed to investigate the roles of exosomes in delivering osimertinib resistance in NSCLC. Exosomes were successfully isolated. LncRNA sequencing identified a total of 123 differentially expressed lncRNAs, including 45 upregulated lncRNAs and 78 downregulated lncRNAs. The relative expression level of lncRNA MSTRG.292666.16 was significantly upregulated in osimertinib-resistant plasma, osimertinib-resistant H1975R cells and their derived exosomes, compared with those in osimertinib- sensitive plasma, H1975 cells and exosomes (P < 0.05). Besides, osimertinib-resistant exosomes could regulate gene expressions induced by osimertinib, including miRNA-21, miRNA-125b, TGFß, ARF6 and c-Kit. Osimertinib-resistant exosomes could be taken up by osimertinib-sensitive H1975 cells and resulting in osimertinib-resistance in vivo. Knockdown of lncRNA MSTRG.292666.16 decreased osimertinib resistance of H1975R cells. Our results suggest that exosomal lncRNA MSTRG.292666.16 might be associated with osimertinib resistance in NSCLC.

Cyclooxygenase-2 mediates gefitinib resistance in non-small cell lung cancer through the EGFR/PI3K/AKT axis.

Gefitinib is a potent inhibitor of EGFR and represents the front-line treatment for non-small cell lung cancer (NSCLC) therapeutics. However, NSCLC patients are prone to develop acquired resistance through as yet, undefined mechanisms of resistance. Here, we investigated the role of COX-2 during gefitinib resistance in NSCLC cells and revealed its underlying mechanism(s) of action. We report the upregulation of COX-2 in gefitinib-resistant NSCLC tissues and cells, which is associated with poor prognosis. In vitro assays in NSCLC cells (PC9/GR) showed that COX-2 facilitates gefitinib resistance in NSCLC cells through its effects on P-gp, MRP1, and BCRP, and cancer cell migration and invasion. In vivo, COX-2 silencing could repress tumor growth. We found that the overexpression of COX-2 enhances the transcription of MMP-2, MMP-7, and MMP-9 which mediates PI3K-AKT activation. In summary, we demonstrate that COX-2 mediates the gefitinib resistance of NSCLC cells through its interaction with EGFR and the PI3K-AKT axis. This highlights COX-2 as a novel molecular target for NSCLC.

Predictive value of unmethylated RASSF1A on disease progression in non-small cell lung cancer patients receiving pemetrexed-based chemotherapy.

BACKGROUND AND OBJECTIVE: Chemotherapy remains the basis of the treatment of lung cancer, and screening biomarkers with predictive value for chemotherapy is of great interest. The present study focused on status of genes methylation in NSCLC patients receiving pemetrexed- or gemcitabine-based chemotherapy. PATIENTS AND METHODS: Promoter methylation of Ras association domain family (RASSF1A) and short stature homeobox 2 (SHOX2) was examined in bronchoalveolar lavage (BAL) from 117 NSCLC patients treated with chemotherapy. Multivariate analysis was used to identify the predictive value of gene methylation. Progression-free survival (PFS) rather than overall survival (OS) was used as the clinical outcome to minimize the impact of chemotherapy on gene methylation. RESULTS: The methylation of RASSF1A and SHOX2 was significantly associated with shorter PFS (RASSF1A: HR = 2.355, 95% CI: 1.533-3.617, P< 0.0001; SHOX2: HR = 2.123, 95% CI: 1.392-3.236, P= 0.0004). After adjusting for confounding factors, RASSF1A methylation was still a predictive factor for PFS (HR = 1.765, 95% CI: 1.064-2.928, P= 0.0278). In the pemetrexed group, unmethylated RASSF1A could be used to predict longer PFS (P= 0.0001), and no predictive value was found in the gemcitabine group. CONCLUSION: Unmethylated RASSF1A is a favorable prognostic indicator for patients receiving pemetrexed doublets. Because of the promoting effect of most chemotherapeutic drugs on gene methylation, unmethylated RASSF1A is not suitable as a predictor for gemcitabine doublets.

Detection of plasma EGFR mutations for personalized treatment of lung cancer patients without pathologic diagnosis.

INTRODUCTION: Next-generation sequencing (NGS) and digital polymerase chain reaction (PCR) based platforms have been used to detect EGFR mutations in plasma circulating tumor DNA (ctDNA) with high accuracy. Generally, molecular testing is performed after histopathological analysis. However, many patients with suspected advanced nonsmall cell lung cancer are unable to undergo biopsy thus forgoing potential treatment with highly effective tyrosine kinase inhibitors (TKIs) in patients with sensitizing EGFR mutations. We examined the utility of ctDNA testing to detect EGFR mutations in patients' plasma, where tissue biopsy is not feasible. METHODS: We conducted a single-center, prospective study of 30 Chinese patients with suspected advanced lung cancer, who were unable to undergo a biopsy for initial diagnosis due to comorbidities or poor performance status. Patients with plasma EGFR sensitizing mutations were treated with first-generation EGFR TKIs. RESULTS: Twenty of 30 patients enrolled had sensitizing EGFR mutations in ctDNA and were started on EGFR TKIs. After a median follow-up of 12 months, median progression-free survival (PFS) was 10 months and median overall survival (OS) was not reached. The median OS for the 10 untreated patients was 3 months. CONCLUSIONS: In our study, patients with plasma EGFR mutations treated with TKIs showed disease control rate (DCR) and PFS similar to historical controls that were treated based on tissue testing. This is the first prospective study showing that ctDNA genotyping provides a feasible diagnostic approach for frail lung cancer patients who are unable to undergo biopsy, which subsequently leads to EGFR-targeted therapy, and improved outcomes in this subgroup of patients.

Clinical efficacy of erlotinib in patients previously treated for advanced non-small cell lung cancer.

BACKGROUND AND OBJECTIVE: Erlotinib is one of the standard second/third line treatments for patients with advanced non-small cell lung cancer (NSCLC). This study investigated the efficacy of erlotinib in a Chinese population with advanced NSCLC and compared the predictive value of serum vascular endothelial growth factor (VEGF) and transforming growth factor (TGF)-alpha for the efficacy of erlotinib. METHODS: Patients with advanced, previously treated NSCLC received 150 mg of erlotinib once daily orally until disease progression or intolerable toxicity. Serum levels of VEGF and TGF-alpha were measured by ELISA at baseline and 1 month after treatment commenced. RESULTS: There were 112 patients enrolled during the period October 2005 to February 2008 and followed until July 2008. Serum samples were available in 50 patients. Tumour response to erlotinib was partial in 35.7% of patients and 41.1% of patients had stable disease. The severity of skin rash (P < 0.001) had a significant positive correlation with the response to erlotinib. Median progression-free survival (PFS) and overall survival were 6.3 months and 12.5 months, respectively. After erlotinib treatment, serum VEGF levels did not change significantly, while serum TGF-alpha levels increased in patients who had partial response (P = 0.075) or stable disease (P = 0.055), but not in patients with progressive disease (P = 0.155). In patients with measurable serum TGF-alpha levels at baseline the PFS and median survival were 5 and 9.9 months respectively, and in patients with no measurable TGF-alpha at baseline the PFS and median survival were 11 and 21 months, respectively. Overall survival was significantly longer in patients with negative baseline serum TGF-alpha (P = 0.002). CONCLUSIONS: Oral erlotinib was effective as a second/third line treatment for patients with advanced NSCLC. Baseline serum TGF-alpha levels may be a predictor for the efficacy of erlotinib treatment.

[Impact of erlotinib treatment on symptoms and quality of life in patients with advanced non-small-cell lung cancer].

OBJECTIVE: To investigate the impact of erlotinib as a second or third line treatment on the symptoms and quality of life (QOL) in patients with advanced non-small cell lung cancer (NSCLC). METHODS: Fifty patients with stage III b and IV NSCLC, treated previously with at least one regimen of platinum-based chemotherapy, received 150 mg of erlotinib orally, once a day till disease progression. QOL was assessed by European Organization for Research and Treatment of Cancer QLQ-C30 and the lung cancer module (QLQ-LC13). The primary end points for QOL analysis were time to deterioration of three common lung cancer symptoms: cough, dyspnea and pain. RESULTS: Among 47 evaluable cases, there were partial remission (PR) in 18 cases, stable disease (SD) in 21 cases, and progressive disease (PD) in 8 cases. After two cycles of treatment, the mean scores of global QOL and all 5 functioning scales except the cognitive function increased significantly (P < 0.05). Mean scores of major general symptoms, hypodynamia and anorexia, and disease-related symptoms alleviated significantly. Both response rates of five functioning and global QOL were more than 44% after erlotinib treatment. Response rates of major general symptoms and disease-related symptoms varied from 14% to 76%. Patients with complete or partial response likely had improvement in the QOL response (P < 0.05), and the time to major symptom deterioration in those were significantly longer (P < 0.001) than that in patients with stable or even progressive disease. CONCLUSION: Erlotinib is effective to improve not only survival, but also tumor-related symptoms and quality of life in patients with advanced NSCLC previously treated with cisplatin-contained regimens. The improvement in the quality of life is positively correlated with objective tumor response.

Engineering docetaxel-loaded micelles for non-small cell lung cancer: a comparative study of microfluidic and bulk nanoparticle preparation.

Bulk preparation of micelles has the drawbacks of facile formation of large aggregates and heterogeneous particle size distribution. Microfluidic technology has shown clear potential to address these challenges for robust nanomedicine applications. In this study, docetaxel-loaded PLGA-PEG-Mal-based micelles were prepared by microfluidics and dialysis methods and their physicochemical properties were analyzed. The biological behaviors of these micelles were also investigated in the non-small cell lung cancer (NSCLC) cell line A549 in vitro as well as in vivo. Encouragingly, the mean particle size of the micelles prepared by microfluidics (DMM) was smaller, with an average size of 72 ± 1 nm and a narrow size distribution with a polydispersity index (PDI) of 0.072; meanwhile, micelles prepared by the dialysis method (DMD) had larger particle sizes (range, 102 to 144 nm) and PDIs (up to 0.390). More importantly, significantly high drug loading was achieved using the microfluidic process. The IC50 value of DMM was lower than that of DMD. Whole-body fluorescence imaging of live mice showed that DMM achieved higher accumulation in tumors compared with DMD. DMM showed superior antitumor efficacy, with a tumor inhibition rate of 91.5%. Moreover, pathological histology analysis revealed that no evident biological toxicity was caused by the micelles. In addition, Arg-Gly-Asp (RGD) was employed as a targeting agent on the basis of DMM to prepare targeting micelles, and the targeting micelles exhibited stronger cytotoxicity and obvious antitumor efficacy. In conclusion, DMM may have obvious clinical advantages for the treatment of NSCLC due to its optimized physiochemical properties. Therefore, microfluidic technology-based micelles are a promising platform as an effective drug delivery system for incorporating anticancer agents.