A Case of Tension Subcutaneous Emphysema Treated With Minimally Invasive Open-Window Thoracostomy Using a Wound Protector/Retractor and Three-Sided Taping.

Subcutaneous emphysema is a common complication of thoracic surgery. Tension subcutaneous emphysema that causes airway obstruction is rare but life-threatening. This report presents a patient who developed tension subcutaneous emphysema after recurrent secondary pneumothorax surgery which was treated with minimally invasive open-window thoracostomy. A wound protector/retractor and three-sided taping were successfully used to prevent air from entering the subcutaneous space via the wound while draining trapped air without creating an open pneumothorax. This approach is an option for managing subcutaneous and intrathoracic air leakage in emergency situations.

A Case of Tension Pneumomediastinum Treated With Mediastinal Drainage Using a Semi-flexible Fiberscope via a Subxiphoid Approach.

Tension pneumomediastinum with hemodynamic failure is a rare but life-threatening condition. Rapid decompression of the mediastinum by drainage is essential to save the patient's life. This report presents a case of tension pneumomediastinum that developed during conservative management of a pneumomediastinum associated with idiopathic pulmonary fibrosis. Endoscopically guided mediastinal drainage was successfully performed in the emergency situation of tension pneumomediastinum. Using the semi-flexible fiberscope inserted through a subxiphoid approach, the drainage catheter was easily and safely placed at the appropriate site in the mediastinum. Good mediastinal decompression was achieved, and the patient was out of this critical condition.

Advances in the Evaluation of Gastrointestinal Absorption Considering the Mucus Layer.

Because of the increasing sophistication of formulation technology and the increasing polymerization of compounds directed toward undruggable drug targets, the influence of the mucus layer on gastrointestinal drug absorption has received renewed attention. Therefore, understanding the complex structure of the mucus layer containing highly glycosylated glycoprotein mucins, lipids bound to the mucins, and water held by glycans interacting with each other is critical. Recent advances in cell culture and engineering techniques have led to the development of evaluation systems that closely mimic the ecological environment and have been applied to the evaluation of gastrointestinal drug absorption while considering the mucus layer. This review provides a better understanding of the mucus layer components and the gastrointestinal tract's biological defense barrier, selects an assessment system for drug absorption in the mucus layer based on evaluation objectives, and discusses the overview and features of each assessment system.

Liver metastases 2 years after resection of a very-low-risk duodenal gastrointestinal stromal tumor: a case report.

BACKGROUND: Gastrointestinal stromal tumors (GISTs) are rare mesenchymal tumors, but are the most common mesenchymal tumors of the gastrointestinal tract. The risk classification of GISTs is based on the tumor size, mitotic index, tumor site, and presence of tumor rupture. Recurrence in the very-low-risk group is extremely rare. We herein report a case of liver metastases 2 years after resection of a very-low-risk duodenal GIST. CASE PRESENTATION: A 57-year-old woman presented to the hospital for evaluation of melena. Esophagogastroduodenoscopy showed bleeding from the exposed blood vessels at the top of a submucosal tumor approximately 20 mm in size located in the second (descending) part of the duodenum, and the bleeding was controlled with electrocoagulation. A GIST was suspected, and the patient underwent wedge resection of the duodenum. The resected specimen contained a 16- × 12-mm (< 20-mm) white submucosal tumor composed of spindle cells with a mitotic count of 4 per 50 high-power fields, and a histologically negative margin was achieved. Immunochemical analysis revealed positive tumor staining for c-kit protein and alpha-smooth muscle actin and negative staining for CD34, desmin, and S-100 protein. Therefore, the tumor was diagnosed as a very-low-risk duodenal GIST based on the Fletcher classification and modified Fletcher classification (Joensuu classification). The postoperative course was uneventful, and the patient was discharged on postoperative day 11. At the follow-up visit 2 years postoperatively, contrast-enhanced computed tomography revealed liver tumors in S8 and S6 measuring 26 × 24 and 10 × 10 mm, respectively. Both lesions showed peripheral dominant hyperenhancement with hypoenhancement inside, indicating tissue degeneration within the tumors. These imaging findings closely resembled those of the duodenal GIST. Hence, the patient was diagnosed with liver metastases of GIST 2 years postoperatively. She was subsequently started on treatment with 400 mg of imatinib. At the time of this writing (2 months after diagnosis), the patient was clinically well and asymptomatic and was continuing imatinib therapy. CONCLUSIONS: Recurrence of very-low-risk GISTs is extremely rare. Even a small GIST with low mitotic activity can never be considered completely benign, and long-term follow-up is necessary.

CD44 Facilitates Epithelial-to-Mesenchymal Transition Phenotypic Change at Acquisition of Resistance to EGFR Kinase Inhibitors in Lung Cancer.

Epithelial-to-mesenchymal transition (EMT) is one of the acquired resistance mechanisms to EGFR tyrosine kinase inhibitors (TKI) in lung cancers. Because EMT is related to tumor invasion, metastases, and resistance to various treatments, it is important to prevent the emergence of EMT. However, molecular mechanism(s) underlying EMT phenotypic changes, as well as biomarker(s) that predict the emergence of EMT in EGFR-mutated lung cancers, are unclear to date. Through the comparison of expression data between isogenic lung cancer cell lines that acquired resistance to EGFR-TKI(s), we identified that high CD44 expression is related to a mesenchymal phenotype and that shRNA-mediated knockdown of CD44 reversed the EMT change. High membranous CD44 expression was identified in lesions with mesenchymal phenotype that were obtained from lung cancer patients who developed acquired resistance to gefitinib or afatinib, whereas isogenic lesions without EMT change showed negative/weak staining for CD44. Immunohistochemistry for treatment-naïve lung cancer cell lines with EGFR mutations found those that acquire resistance to EGFR-TKIs via EMT (HCC4006 and H1975 cells) had strong membranous CD44 expression compared with non-EMT-transforming lines which demonstrated negative or weak staining (Fisher exact test P value = 0.036). shRNA-mediated CD44 knockdown in HCC4006 cells prevented the emergence of EMT after chronic exposure to osimertinib. These results suggest that upregulation of CD44 facilitates EMT-phenotypic change in lung cancers with EGFR mutations when treated with EGFR-TKIs. In addition, our results suggest that CD44 can be a useful biomarker to predict the emergence of EMT upon EGFR-TKI monotherapy. Mol Cancer Ther; 17(10); 2257-65. ©2018 AACR.

Effect of dasatinib on EMT-mediated-mechanism of resistance against EGFR inhibitors in lung cancer cells.

OBJECTIVE: The epithelial to mesenchymal transition (EMT) is associated with acquired resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) in certain non-small cell lung cancers that harbor EGFR mutations. Because no currently available drugs specifically kill cancer cells via EMT, novel treatment strategies that overcome or prevent EMT are needed. A recent report suggested that dasatinib (an ABL/Src kinase inhibitor) inhibits EMT induced by transforming growth factor (TGF)-beta in lung cancer cells (Wilson et al., 2014). In this study, we analyzed effects of dasatinib on the resistance mechanism in HCC4006 cells, which tend to acquire resistance to EGFR-TKIs via EMT. MATERIALS AND METHODS: Sensitivity to dasatinib in HCC4006 and HCC4006 erlotinib-resistant (ER) cells with an EMT phenotype was analyzed. HCC4006 cells acquired resistance against the combination of erlotinib and dasatinib (HCC4006EDR) following chronic treatment with these drugs. The expression of EMT markers and the resistance mechanism were analyzed. RESULTS: Short-term or long-term treatment with dasatinib did not reverse EMT in HCC4006ER. In contrast, HCC4006EDR cells maintained an epithelial phenotype, and the mechanism underlying resistance to erlotinib plus dasatinib combination therapy was attributable to a T790M secondary mutation. HCC4006EDR cells, but not HCC4006ER cells, were highly sensitive to a third-generation EGFR-TKI, osimertinib. CONCLUSIONS: Although dasatinib monotherapy did not reverse EMT in HCC4006ER cells, preemptive combination treatment with erlotinib and dasatinib prevented the emergence of acquired resistance via EMT, and led to the emergence of T790M. Our results indicate that preemptive combination therapy may be a promising strategy to prevent the emergence of EMT-mediated resistance.

MEK inhibitors against MET-amplified non-small cell lung cancer.

Several receptor tyrosine kinases (RTKs) including EGFR, ALK, and MET have been identified as therapeutic targets in non-small cell lung cancer (NSCLC). Among the downstream pathways of RTKs, the MAPK pathway is particularly important for cancer cell proliferation, differentiation, and survival. In this study, the effects of MEK inhibitors (trametinib and PD0325901) in several NSCLC cell lines with driver gene alterations, especially RTK genes, were tested in vitro using an MTT assay, and a wide range of sensitivities was found. In particular, all the EGFR-mutated cell lines were resistant to MEK inhibitors, whereas all the MET-amplified cell lines were sensitive. A bioinformatics technique and western blot analyses showed that the PI3K/AKT pathway is more activated in EGFR-mutated NSCLC than in MET-amplified NSCLC, and a PI3K inhibitor enhanced the sensitivity to trametinib in the EGFR-mutated cell lines, suggesting that this pathway is associated with resistance to MEK inhibitors. Although the HCC827 cell line (EGFR mutation) was resistant to MEK inhibitors, the HCC827CNXR cell line, whose driver gene shifts from EGFR to MET, exhibited enhanced sensitivity to MEK inhibitors, indicating the biological importance of the MAPK pathway for MET-amplified NCSLC. Furthermore, a synergistic effect of crizotinib (a MET inhibitor) and trametinib was observed in MET-amplified NCLC cell lines. Our findings indicate that the MAPK pathway is biologically important for MET-amplified NSCLC and strongly encourage the development of combination therapy with a MET inhibitor and a MEK inhibitor against MET-amplified NSCLC.

Clinical, Pathological, and Molecular Features of Lung Adenocarcinomas with AXL Expression.

The receptor tyrosine kinase AXL is a member of the Tyro3-Axl-Mer receptor tyrosine kinase subfamily. AXL affects several cellular functions, including growth and migration. AXL aberration is reportedly a marker for poor prognosis and treatment resistance in various cancers. In this study, we analyzed clinical, pathological, and molecular features of AXL expression in lung adenocarcinomas (LADs). We examined 161 LAD specimens from patients who underwent pulmonary resections. When AXL protein expression was quantified (0, 1+, 2+, 3+) according to immunohistochemical staining intensity, results were 0: 35%; 1+: 20%; 2+: 37%; and 3+: 7% for the 161 samples. AXL expression status did not correlate with clinical features, including smoking status and pathological stage. However, patients whose specimens showed strong AXL expression (3+) had markedly poorer prognoses than other groups (P = 0.0033). Strong AXL expression was also significantly associated with downregulation of E-cadherin (P = 0.025) and CD44 (P = 0.0010). In addition, 9 of 12 specimens with strong AXL expression had driver gene mutations (6 with EGFR, 2 with KRAS, 1 with ALK). In conclusion, we found that strong AXL expression in surgically resected LADs was a predictor of poor prognosis. LADs with strong AXL expression were characterized by mesenchymal status, higher expression of stem-cell-like markers, and frequent driver gene mutations.

Oncogene swap as a novel mechanism of acquired resistance to epidermal growth factor receptor-tyrosine kinase inhibitor in lung cancer.

Mutant selective epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs), such as rociletinib and AZD9291, are effective for tumors with T790M secondary mutation that become refractory to first-generation EGFR-TKI. However, acquired resistance to these prospective drugs is anticipated considering the high adaptability of cancer cells and the mechanisms remain largely obscure. Here, CNX-2006 (tool compound of rociletinib) resistant sublines were established by chronic exposure of HCC827EPR cells harboring exon 19 deletion and T790M to CNX-2006. Through the analyses of these resistant subclones, we identified two resistant mechanisms accompanied by MET amplification. One was bypass signaling by MET amplification in addition to T790M, which was inhibited by the combination of CNX-2006 and MET-TKI. Another was loss of amplified EGFR mutant allele including T790M while acquiring MET amplification. Interestingly, MET-TKI alone was able to overcome this resistance, suggesting that oncogenic dependence completely shifted from EGFR to MET. We propose describing this phenomenon as an "oncogene swap." Furthermore, we analyzed multiple lesions from a patient who died of acquired resistance to gefitinib, then found a clinical example of an oncogene swap in which the EGFR mutation was lost and a MET gene copy was gained. In conclusion, an "oncogene swap" from EGFR to MET is a novel resistant mechanism to the EGFR-TKI. This novel mechanism should be considered in order to avoid futile inhibition of the original oncogene.

Functional Analyses of Mutations in Receptor Tyrosine Kinase Genes in Non-Small Cell Lung Cancer: Double-Edged Sword of DDR2.

PURPOSE: This study investigated whether mutations of receptor tyrosine kinase (RTK) genes detected using next-generation sequencing (NGS) are suitable therapeutic targets. EXPERIMENTAL DESIGN: Fifty surgically resected non-small cell lung cancer (NSCLC) samples were target resequenced using NGS. We then investigated the functions of the identified RTK gene mutations, including their oncogenic potential, in vitro RESULTS: Mutations in RTK genes were found in 20 samples (EGFR, 15; ERBB4, 1; ALK, 1; DDR2, 2; FGFR1, 1), mutations in MAPK pathway genes were found in nine samples (KRAS, 7; NRAS, 1; BRAF, 2), and mutations in PI3K pathway genes were found in three samples (PIK3CA, 1; PTEN, 3). Among the mutations in RTKs, the functions of four mutations were unclear (ERBB4 D245G; DDR2 H246R and E655K; FGFR1 A263V). These mutations did not exhibit any transformational activities. Neither the phosphorylation nor the protein expressions of RTKs were changed by the DDR2 H246R, ERBB4 D245G, and FGFR1 A263V mutations, although the expression level of the DDR2 protein harboring the E655K mutation was particularly low. Collagen stimulation decreased cellular proliferation through p38 activation in the DDR2 wild-type-overexpressed cell lines, whereas the growth-suppressive effect was weakened in DDR2 E655K-overexpressed cell lines. Furthermore, the DDR2 E655K protein strongly bound to ubiquitin ligase E3 (Cbl-b), and the mutant protein expression was increased after treatment with a proteasome inhibitor. CONCLUSIONS: Our experimental findings suggest that RTK mutations are not always suitable as therapeutic targets. The DDR2 E655K mutation can play a role in cancer progression by reducing the growth-inhibitory effect of collagen. Clin Cancer Res; 22(14); 3663-71. ©2016 AACR.

Impact of bevacizumab in combination with erlotinib on EGFR-mutated non-small cell lung cancer xenograft models with T790M mutation or MET amplification.

Erlotinib (ERL), an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor, shows notable efficacy against non-small cell lung cancer (NSCLC) harboring EGFR mutations. Bevacizumab (BEV), a humanized monoclonal antibody to vascular endothelial cell growth factor (VEGF), in combination with ERL (BEV+ERL) significantly extended progression-free survival in patients with EGFR-mutated NSCLC compared with ERL alone. However, the efficacy of BEV+ERL against EGFR-mutated NSCLC harboring T790M mutation or MET amplification, is unclear. Here, we examined the antitumor activity of BEV+ERL in four xenograft models of EGFR-mutated NSCLC (three harboring ERL resistance mutations). In the HCC827 models (exon 19 deletion: DEL), ERL significantly inhibited tumor growth by blocking EGFR signal transduction. Although there was no difference between ERL and BEV+ERL in maximum tumor growth inhibition, BEV+ERL significantly suppressed tumor regrowth during a drug-cessation period. In the HCC827-EPR model (DEL+T790M) and HCC827-vTR model (DEL+MET amplification), ERL reduced EGFR signal transduction and showed less pronounced but still significant tumor growth inhibition than in the HCC827 model. In these models, tumor growth inhibition was significantly stronger with BEV+ERL than with each single agent. In the NCI-H1975 model (L858R+T790M), ERL did not inhibit growth or EGFR signal transduction, and BEV+ERL did not inhibit growth more than BEV. BEV alone significantly decreased microvessel density in each tumor. In conclusion, addition of BEV to ERL did not enhance antitumor activity in primarily ERL-resistant tumors with T790M mutation; however, BEV+ERL enhanced antitumor activity in T790M mutation- or MET amplification-positive tumors as long as their growth remained significantly suppressed by ERL.

Heterogeneity in resistance mechanisms causes shorter duration of epidermal growth factor receptor kinase inhibitor treatment in lung cancer.

OBJECTIVES: Epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors (TKIs) are used as a first line therapy for metastatic lung cancer harboring somatic EGFR mutation. However, acquisition of resistance to these drugs is almost inevitable. T790M (threonine to methionine substitution at codon 790 of the EGFR gene) and MET amplification are well-known resistance mechanisms, and we previously demonstrated that three of six autopsied patients showed inter-tumor heterogeneity in resistance mechanisms by analyzing T790M and MET gene copy number (Suda et al., 2010). To further elucidate the role of heterogeneity in acquired resistance, here we performed further analyses including additional five patients. MATERIALS AND METHODS: We analyzed somatic mutations in 50 cancer-related genes for 26 EGFR-TKI refractory lesions from four autopsied patients using target sequencing. MET and ERBB2 copy numbers were analyzed by real-time PCR. Data for additional one patient was obtained from our recent study (Suda et al., 2015). Relationship between heterogeneity in resistance mechanism(s) and time to treatment failure (TTF) of EGFR-TKI and post-progression survival (PPS) were analyzed. RESULTS AND CONCLUSION: We observed heterogeneity of resistance mechanisms in two of four patients analyzed (T790M+MET gene copy number gain, and mutant EGFR loss+unknown). We also identified quantitative heterogeneity in EGFR T790M mutation ratio among EGFR-TKI refractory lesions. In analyzing patient outcomes, we found that patients who developed multiple resistance mechanisms had shorter TTF compared with those who developed single resistance mechanism (p=0.022). PPS after EGFR-TKI treatment failure was compatible between these two groups (p=0.42). These findings further our understanding of acquired resistance mechanisms to EGFR-TKIs, and may lead to better treatment strategies after acquisition of resistance to first generation EGFR-TKIs in lung cancer patients with EGFR mutations.

MET gene exon 14 deletion created using the CRISPR/Cas9 system enhances cellular growth and sensitivity to a MET inhibitor.

BACKGROUND: MET splice site mutations resulting in an exon 14 deletion have been reported to be present in about 3% of all lung adenocarcinomas. Patients with lung adenocarcinoma and a MET splice site mutation who have responded to MET inhibitors have been reported. The CRISPR/Cas9 system is a recently developed genome-engineering tool that can easily and rapidly cause small insertions or deletions. MATERIALS AND METHODS: We created an in vitro model for MET exon 14 deletion using the CRISPR/Cas9 system and the HEK293 cell line. The phenotype, which included MET inhibitor sensitivity, was then investigated in vitro. Additionally, MET splice site mutations were analyzed in several cancers included in The Cancer Genome Atlas (TCGA) dataset. RESULTS: An HEK293 cell line with a MET exon 14 deletion was easily and rapidly created; this cell line had a higher MET protein expression level, enhanced MET phosphorylation, and prolonged MET activation. In addition, a direct comparison of phenotypes using this system demonstrated enhanced cellular growth, colony formation, and MET inhibitor sensitivity. In the TCGA dataset, lung adenocarcinomas had the highest incidence of MET exon 14 deletions, while other cancers rarely carried such mutations. Approximately 10% of the lung adenocarcinoma samples without any of driver gene alterations carried the MET exon 14 deletion. CONCLUSIONS: These findings suggested that this system may be useful for experiments requiring the creation of specific mutations, and the present experimental findings encourage the development of MET-targeted therapy against lung cancer carrying the MET exon 14 deletion.

Small cell lung cancer transformation and T790M mutation: complimentary roles in acquired resistance to kinase inhibitors in lung cancer.

Lung cancers often harbour a mutation in the epidermal growth factor receptor (EGFR) gene. Because proliferation and survival of lung cancers with EGFR mutation solely depend on aberrant signalling from the mutated EGFR, these tumours often show dramatic responses to EGFR tyrosine kinase inhibitors (TKIs). However, acquiring resistance to these drugs is almost inevitable, thus a better understanding of the underlying resistance mechanisms is critical. Small cell lung cancer (SCLC) transformation is a relatively rare acquired resistance mechanism that has lately attracted considerable attention. In the present study, through an in-depth analysis of multiple EGFR-TKI refractory lesions obtained from an autopsy case, we observed a complementary relationship between SCLC transformation and EGFR T790M secondary mutation (resistance mutation). We also identified analogies and differences in genetic aberration between a TKI-refractory lesion with SCLC transformation and one with EGFR T790M mutation. In particular, target sequencing revealed a TP53 P151S mutation in all pre- and post-treatment lesions. PTEN M264I mutation was identified only in a TKI-refractory lesion with SCLC transformation, while PIK3CA and RB1 mutations were identified only in pre-treatment primary tumour samples. These results provide the groundwork for understanding acquired resistance to EGFR-TKIs via SCLC transformation.

Solitary pulmonary metastasis from malignant melanoma of the bulbar conjunctiva presenting as a pulmonary ground glass nodule: Report of a case.

We herein report a case of solitary pulmonary metastasis from malignant melanoma that presented as a pulmonary ground glass nodule. A 57-year-old man who had undergone resection of a malignant melanoma of the right bulbar conjunctiva at the age of 51 was referred to our hospital for management of ground glass opacity in his left lung. Because radiological examination suggested the nodule was an adenocarcinoma in situ, computed tomography (CT) follow-up was planned. CT examination performed nine months later showed that the nodule had grown from 6 mm to 8 mm. Moreover, CT performed one and a half years after first detection revealed that the nodule had grown up to 10 mm. The patient, therefore, underwent partial resection of the lung for diagnosis and treatment. Pathological examination of the resected specimen revealed atypical cells with melanin granules proliferating in a lepidic-like fashion. The cells were positive on S-100 staining, indicating a pulmonary metastasis from malignant melanoma. Thus, metastatic tumors from malignant melanoma can present as ground glass opacities.

EGFR Exon 18 Mutations in Lung Cancer: Molecular Predictors of Augmented Sensitivity to Afatinib or Neratinib as Compared with First- or Third-Generation TKIs.

PURPOSE: Lung cancers harboring common EGFR mutations respond to EGFR tyrosine kinase inhibitors (TKI), whereas exon 20 insertions (Ins20) are resistant to them. However, little is known about mutations in exon 18. EXPERIMENTAL DESIGN: Mutational status of lung cancers between 2001 and 2015 was reviewed. Three representative mutations in exon 18, G719A, E709K, and exon 18 deletion (Del18: delE709_T710insD) were retrovirally introduced into Ba/F3 and NIH/3T3 cells. The 90% inhibitory concentrations (IC90s) of first-generation (1G; gefitinib and erlotinib), second-generation (2G; afatinib, dacomitinib, and neratinib), and third-generation TKIs (3G; AZD9291 and CO1686) were determined. RESULTS: Among 1,402 EGFR mutations, Del19, L858R, and Ins20 were detected in 40%, 47%, and 4%, respectively. Exon 18 mutations, including G719X, E709X, and Del18, were present in 3.2%. Transfected Ba/F3 cells grew in the absence of IL3, and NIH/3T3 cells formed foci with marked pile-up, indicating their oncogenic abilities. IC90s of 1G and 3G TKIs in G719A, E709K, and Del18 were much higher than those in Del19 (by >11-50-fold), whereas IC90s of afatinib were only 3- to 7-fold greater than those for Del19. Notably, cells transfected with G719A and E709K exhibited higher sensitivity to neratinib (by 5-25-fold) than those expressing Del19. Patients with lung cancers harboring G719X exhibited higher response rate to afatinib or neratinib (∼ 80%) than to 1G TKIs (35%-56%) by compilation of data in the literature. CONCLUSIONS: Lung cancers harboring exon 18 mutations should not be overlooked in clinical practice. These cases can be best treated with afatinib or neratinib, although the currently available in vitro diagnostic kits cannot detect all exon 18 mutations.

Collateral chemoresistance to anti-microtubule agents in a lung cancer cell line with acquired resistance to erlotinib.

Various alterations underlying acquired resistance to epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) have been described. Although treatment strategies specific for these mechanisms are under development, cytotoxic agents are currently employed to treat many patients following failure of EGFR-TKIs. However, the effect of TKI resistance on sensitivity to these cytotoxic agents is mostly unclear. This study investigated the sensitivity of erlotinib-resistant tumor cells to five cytotoxic agents using an in vitro EGFR-TKI-resistant model. Four erlotinib-sensitive lung adenocarcinoma cell lines and their resistant derivatives were tested. Of the resistant cell lines, all but one showed a similar sensitivity to the tested drugs as their parental cells. HCC4006ER cells with epithelial mesenchymal transition features acquired resistance to the three microtubule-targeting agents, docetaxel, paclitaxel and vinorelbine, but not to cisplatin and gemcitabine. Gene expression array and immunoblotting demonstrated that ATP-binding cassette subfamily B, member 1 (ABCB1) was up-regulated in HCC4006ER cells. ABCB1 knockdown by siRNA partially restored sensitivity to the anti-microtubule agents but not to erlotinib. Moreover, the histone deacetylase inhibitor entinostat sensitized HCC4006ER cells to anti-microtubule agents through ABCB1 suppression. Our study indicates that sensitivity of tumor cells to cytotoxic agents in general does not change before and after failure of EGFR-TKIs. However, we describe that two different molecular alterations confer acquired resistance to EGFR-TKIs and cytotoxic agents, respectively. This phenomenon should be kept in mind in selection of subsequent therapy after failure of EGFR-TKIs.

Recent evidence, advances, and current practices in surgical treatment of lung cancer.

In the last 10-15 years, strategies and modalities of lung cancer treatment have changed dramatically. Meanwhile, the treatment objectives, the lung cancers themselves, have also changed, probably owing to early detection by computed tomography and aging of the population. In particular, the proportions of smaller lung cancers, lung adenocarcinomas with ground-glass opacity, and lung cancers in older patients are increasing. Along with these changes, surgeons have innovated and evaluated novel procedures for pulmonary resection. These include the application of minimally invasive surgical techniques, such as video-assisted thoracoscopic surgery (VATS) and robotic surgery, and sub-lobar resection, such as wedge resection and segmentectomy, for small peripheral lung cancers. Currently, VATS has gained wide acceptance and several institutions in Japan have started using robotic surgery for lung cancers. Two important clinical trials of sub-lobar resection for small peripheral lung cancers are now underway in Japan. In addition, surgery itself is of growing importance in lung cancer treatment. In particular, recent evidence supports the use of surgery in strictly selected patients with locally advanced disease, lung cancers with N2 lymph node metastases, small cell lung cancers, recurrent oligo-metastasis after pulmonary resection, or relapsed tumors after drug treatment. Surgical treatment also provides abundant tumor samples for molecular analysis, which can be used for drug selection in the adjuvant setting or after disease relapse. In the era of personalized treatment, surgery is still one of the most important treatment modalities to combat lung cancer.

CRKL amplification is rare as a mechanism for acquired resistance to kinase inhibitors in lung cancers with epidermal growth factor receptor mutation.

OBJECTIVES: Epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors (TKIs) often provide dramatic responses in lung cancer patients with somatic EGFR mutation. However, acquired resistance to the drugs usually emerges within a few years. EGFR T790M secondary mutation, MET gene amplification, and transformation to small cell lung cancer are well-validated mechanisms that underlie acquisition of resistance to EGFR-TKIs. In addition, many molecular aberrations have been reported as candidates for mechanisms of acquired resistance to EGFR-TKIs. Amplification of the CRKL gene was reportedly observed in 1 of 11 lung cancer patients with EGFR mutations who acquired resistance to EGFR-TKI. This study is the first report, to our knowledge, that validated the role of CRKL gene amplification as a mechanism for acquisition of resistance to EGFR-TKIs. MATERIALS AND METHODS: We analyzed CRKL gene copy numbers, using a quantitative real-time PCR method, in 2 in vitro acquired-resistance cell-line models: 11 clinical samples from patients who developed acquired resistance to EGFR-TKIs, and 39 tumor specimens obtained from 7 autopsy patients whose cancers acquired resistance to EGFR-TKIs. Mutational status of EGFR codon 790 and copy numbers for the MET gene were also determined. RESULTS AND CONCLUSION: In analysis for in vitro models, CRKL gene copy numbers were identical between EGFR-TKI-sensitive parental cells and their acquired resistant descendant cells. In addition, we found no clinical tumor specimens with acquired EGFR-TKI resistance to harbor amplified CRKL genes. These results indicate that CRKL gene amplification is rare in acquisition of resistance to EGFR-TKIs in lung cancer patients with EGFR mutations.

The insulin-like growth factor 1 receptor causes acquired resistance to erlotinib in lung cancer cells with the wild-type epidermal growth factor receptor.

Epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitor (TKI) therapy often provides a dramatic response in lung cancer patients with EGFR mutations. In addition, moderate clinical efficacy of the EGFR-TKI, erlotinib, has been shown in lung cancer patients with the wild-type EGFR. Numerous molecular mechanisms that cause acquired resistance to EGFR-TKIs have been identified in lung cancers with the EGFR mutations; however, few have been reported in lung cancers with the wild-type EGFR. We used H358 lung adenocarcinoma cells lacking EGFR mutations that showed modest sensitivity to erlotinib. The H358 cells acquired resistance to erlotinib via chronic exposure to the drug. The H358 erlotinib-resistant (ER) cells do not have a secondary EGFR mutation, neither MET gene amplification nor PTEN downregulation; these have been identified in lung cancers with the EGFR mutations. From comprehensive screening of receptor tyrosine kinase phosphorylation, we observed increased phosphorylation of insulin-like growth factor 1 receptor (IGF1R) in H358ER cells compared with parental H358 cells. H358ER cells responded to combined therapy with erlotinib and NVP-AEW541, an IGF1R-TKI. Our results indicate that IGF1R activation is a molecular mechanism that confers acquired resistance to erlotinib in lung cancers with the wild-type EGFR.