East Asian winter monsoon intensification over the Northwest Pacific Ocean driven by late Miocene atmospheric CO2 decline.

East Asian winter monsoon (EAWM) activity has had profound effects on environmental change throughout East Asia and the western Pacific. Much attention has been paid to Quaternary EAWM evolution, while long-term EAWM fluctuation characteristics and drivers remain unclear, particularly during the late Miocene when marked global climate and Asian paleogeographic changes occurred. To clarify understanding of late Miocene EAWM evolution, we developed a high-precision 9-million-year-long stacked EAWM record from Northwest Pacific Ocean abyssal sediments based on environmental magnetism, sedimentology, and geochemistry, which reveals a strengthened late Miocene EAWM. Our paleoclimate simulations also indicate that atmospheric CO2 decline played a vital role in this EAWM intensification over the Northwest Pacific Ocean compared to other factors, including central Asian orogenic belt and northeastern Tibetan Plateau uplift and Antarctic ice-sheet expansion. Our results expand understanding of EAWM evolution from inland areas to the open ocean and indicate the importance of atmospheric CO2 fluctuations on past EAWM variability over large spatial scales.

Association of integrin beta1 and c-MET in mediating EGFR TKI gefitinib resistance in non-small cell lung cancer.

Although some patients are initially sensitive to epidermal growth factor receptor tyrosine kinase inhibitors (EGFR TKIs), resistance invariably develops. Therefore, it's very important to study the molecular mechanism of this resistance. In our previous study we found that integrin beta1 can induce EGFR TKIs resistance in non-small cell lung cancer (NSCLC) cells. Here we analyzed the association of integrin beta1 and c-MET that is a recognized mechanism of EGFR TKIs resistance in NSCLC to demonstrate the mechanism of integrin beta1 related EGFR TKIs resistance. We found that the ligands of integrin beta1 and c-MET could synergistically promote cell proliferation and their inhibitors could synergistically improve the sensitivity to gfitinib, increase apoptosis, and inhibit the downstream signal transduction: focal adhesion kinase (FAK) and AKT. On the other hand, ligand-dependent activation of integrin beta1 could induce EGFR TKIs resistance through activating c-MET and its downstream signals. Thus, it can be concluded that there is crosstalk between integrin beta1 and c-MET and integrin beta1 mediates EGFR TKI resistance associating with c-MET signaling pathway in non-small cell lung cancer.

Survival, Chemotherapy and Chemosensitivity Predicted by CTC Cultured In Vitro of SCLC Patients.

PURPOSE: The prognosis for small cell lung cancer (SCLC) patients receiving later-line treatment is very poor and there is still no standard treatments after the second-line setting. Analyzing the susceptibility of chemotherapeutic drugs with circulating tumor cells (CTCs) cultured in vitro may contribute to optimize the therapeutic regimen. However, so far CTCs have been barely used for studying their chemosensitivity due to the lack of technology to obtain wholly intact and viable CTCs. METHODS: Based on a retrospective study of the therapeutic response of 99 patients with unresectable SCLC, the CTC count in 14 SCLC patients was detected before and after chemotherapy to evaluate its role as a potential marker of response. Furthermore, the drug susceptibility of CTCs cultured in vitro obtained from ClearCell FX® System was tested and the therapy response was evaluated. RESULTS: All of the 99 patients received the first-line chemotherapy and the objective response rate (ORR) was 74.7%. A total of 36 patients received the second-line therapy and the average duration was 2.6 months, and only 11 cases out of them received the third-line therapy but no one responded. The change of CTC counts was identified to be correlated with therapy response. However all the five SCLC patients who were administered with the drugs according to the drug susceptibility test of CTCs for two cycles underwent progression of disease. CONCLUSION: The results showed that the responses of chemotherapy are very poor in later lines and the drug susceptibility test using CTCs primary cultured in vitro may not benefit the improvement of therapeutic regimen of SCLC patients.

Efficacy of the Xpert Mycobacterium tuberculosis/rifampicin assay for diagnosing sputum-smear negative or sputum-scarce pulmonary tuberculosis in bronchoalveolar lavage fluid.

OBJECTIVE: To evaluate he diagnostic performance of the Xpert Mycobacterium tuberculosis/Rifampin (MTB/RIF) assay in bronchoalveolar lavage fluid (BALF). METHODS: We retrospectively reviewed the clinical data from 671 sputum-smear negative or sputum-scarce adult patients with suspected pulmonary tuberculosis (PTB) who had an Xpert MTB/RIF assay performed on BALF. The diagnostic performance of the Xpert MTB/RIF assay, smear microscopy (SM) and MTB culture was evaluated using MTB culture or final clinical diagnosis as the reference standard. RESULTS: Compared with MTB culture, the sensitivity and specificity were 87.8% and 72.7% for the Xpert MTB/RIF assay and 11.0% and 99.2% for SM, respectively. Compared with final diagnosis, diagnostic performance was 58.9% and 83.9% for the Xpert MTB/RIF assay, 5.0% and 98.3% for SM, and 43.3% and 100% for culture, for sensitivity and specificity respectively. The Xpert MTB/RIF assay had low specificity and high sensitivity. When very low results were re-evaluated and considered MTB-negative, the specificity increased significantly. The sensitivity remained higher than SM and was similar to that of culture. CONCLUSIONS: The Xpert MTB/RIF assay adds microbiologic evidence to clinical decisions; however, close attention should be paid to very low semi-quantitative positive results.

Integrin beta1 over-expression associates with resistance to tyrosine kinase inhibitor gefitinib in non-small cell lung cancer.

The epidermal growth factor receptor tyrosine kinase inhibitors (EGFR TKIs) such as gefitinib and erlotinib have been widely used in treating patients with advanced non-small cell lung cancer (NSCLC). However, acquired resistance to EGFR TKI almost occurs in every patient eventually. To identify its potential mechanism, we established a human NSCLC cell line PC9/AB2 which was 576-fold decrease in gefitinib sensitivity compared with its parental PC9 cell lines. No EGFR-T790M mutation or abnormal expression of c-Met protein was found in PC9/AB2 cells. Over-expression of integrin ß1 was found, accompanied with increase of the cells' adhesion and migration. To further confirm the role of integrin ß1 in gefitinib acquired resistance, we transferred its siRNA-expressing plasmid and its whole cDNA expressing plasmid into PC9/AB2 and into PC9 cells, respectively. The sensitivity of NSCLC cells to gefitinib was negatively correlated with integrin ß1 expression levels. All these data suggest that up-regulation of integrin ß1 might be an important factor for gefitinib resistance in NSCLC cell line PC9/AB2.

[Drug resistance mechanism of non small cell lung cancer PC9/AB2 cell line with acquired drug resistance to gefitinib].

OBJECTIVE: To study the drug resistance mechanism of non-small cell lung cancer (NSCLC) cell line PC9/AB2 with acquired drug resistance to gefitinib. METHODS: The human lung adenocarcinoma cell line PC9 was cultured in vitro, and was induced by MNNG to obtain the cell line PC9/AB2 with acquired drug resistance to gefitinib. The sensitivity of the cell line PC9 and PC9/AB2 to gefitinib was determined by MTT assay. The effects of gefitinib on cell apoptosis of the 2 cell lines were determined by flow cytometry. The genomic DNA of the 2 cell lines were extracted, and then the exons 19-21 of EGFR gene were amplified by PCR and sequenced. The protein expression of c-MET and integrin beta1 in the 2 cell lines was determined by Western blot method. The adhesion ability and migration ability of the 2 cell lines were determined by adhesion test and scratch assay. RESULTS: (1) The data form MTT and apoptosis detection showed that the IC50 of PC9/AB2 cells was (24.2+/-5.5) micromol/L, 576 times higher than PC9 cells [IC50 (0.04+/-0.01) micromol/L]. Given the same concentration of gefitinib, the apoptosis rate of PC9 cells was 38.48%, while that of PC9/AB2 cells was 2.2%. (2) The results of gene sequencing showed that there was a deletion of 15 bp in both exon 19 of the 2 cell lines, while no T790M mutation occurred. (3) The results from Western blot showed that there was no significant difference in protein expression of c-MET between the 2 cell lines, while the protein expression of integrin beta1 in PC9/AB2 cells was significantly higher than that of the PC9 cells. (4) The result from adhesion test and scratch assay showed that the adhesion ability and migration ability of the PC9/AB2 cells was significantly higher that those of PC9 cells. CONCLUSION: The high expression of integrin beta1 may be associated with acquired drug resistance of NSCLC cell line PC9/AB2 to gefitinib.

Mechanism of intrinsic resistance of lung squamous cell carcinoma to epithelial growth factor receptor-tyrosine kinase inhibitors revealed by high-throughput RNA interference screening.

Although targeted therapy has achieved a great breakthrough in the treatment of lung adenocarcinoma, there are still no effective targeted drugs for lung squamous cell carcinoma (SqCC). In addition, as immunotherapy can only prolong the overall survival (OS) of lung SqCC by ≤5 months, chemotherapy and radiotherapy are still the main types of therapy for advanced SqCC. The expression level of epithelial growth factor receptor (EGFR) in patients with lung SqCC is higher compared with those with adenocarcinoma, but the former group is intrinsically resistant to EGFR-tyrosine kinase inhibitors (EGFR-TKIs). Therefore, if the drug resistance in patients with lung SqCC could be reversed, the majority of patients may benefit from EGFR-TKIs. In the present study, the high-throughput RNA interference technology was used to screen the genes involved in the EGFR-TKI erlotinib resistance of lung SqCCs, and integrin-linked kinase (ILK) was identified to be the most effective. The role of ILK in erlotinib resistance was further studied in cell lines, and the expression of ILK was analyzed in patients with SqCC and adenocarcinoma. Finally, the mechanism of ILK in EGFR-TKIs resistance was analyzed using Kyoto Encyclopedia of Genes and Genomes (KEGG), Gene Ontology (GO) and ingenuity pathway analysis (IPA). The results demonstrated that the ILK gene knockdown could overcome erlotinib resistance by inhibiting cell proliferation, inducing apoptosis and blocking the cell cycle at the G2/M phase. The expression of ILK in patients with SqCC was significantly higher compared with those with adenocarcinoma with sensitizing EGFR mutations. In addition, the cell cycle pathway 'G2/M DNA damage and checkpoint regulation' was identified to be significantly inhibited by ILK knockdown in IPA, KEGG and GO analysis. The results of the present study may improve the understanding of EGFR-TKI resistance in lung SqCCs, thus promoting the development of potential targeted therapies for lung SqCCs.

A lung squamous carcinoma patient with ROS1 rearrangement sensitive to crizotinib.

It is widely known that ROS1 rearrangement mostly occurs in the adenocarcinoma subtype of non-small-cell lung cancer. Patients with squamous cell carcinoma harboring ROS1 rearrangement are extremely rare. This is a case report of a squamous cell carcinoma patient with ROS1 rearrangement. An 84-year-old Chinese woman with a about 6.6 cm mass in the right middle lobe and right pleural effusion, enlarged mediastinal and hilar lymph nodes was diagnosed with stage IIIB lung squamous cell carcinoma harboring ROS1 rearrangement is highly sensitive to crizotinib in the first treatment setting. This is the first time to report lung squamous carcinoma patient with ROS1 rearrangement sensitive to crizotinib. We hope that oncologists will notice this phenomenon and it will help the lung squamous cell carcinoma patient to get longer overall survival time.

MicroRNA Signature of Lung Adenocarcinoma with EGFR Exon 19 Deletion.

The findings of EGFR mutations and the development of targeted therapies have significantly improved the overall survival of lung cancer patients. Still, the prognosis remains poor, so we need to know more about the genetic alterations in lung cancer. MicroRNAs are dysregulated in lung cancer, and some of them can regulate EGFR. So it is very important to predict the candidate microRNAs that target mutated EGFR and to investigate the role of these candidate microRNAs in lung cancer. In this study, we investigated the difference of microRNAs expression between lung adenocarcinoma cell lines with EGFR exon 19 deletion (H1650 and PC9) and wild-type (H1299 and A549) using the Phalanx Human Whole Genome Microarray. Then the expression of individual microRNAs was validated by qRT-PCR assays. Moreover, we detected the microRNAs expression in plasma of lung adenocarcinoma patients with EGFR exon 19 deletion and wild-type. Lastly, we explored the function of the positive microRNA in EGFR tyrosine kinase inhibitors (EGFR-TKIs ) resistance using MTT and Annexin V-APC assays. The expression of 1,732 microRNAs was evaluated, and we found that microRNAs expression was different between these two groups. Hsa-miR-141-3p, hsa-miR-200c-3p, hsa-miR-203, hsa-miR-3182, hsa-miR-934 were up-regulated and hsa-miR-3196 was down-regulated in the EGFR exon 19 deletion group compared with wild-type group. The detection of circulating microRNAs showed that miR-3196 was down-regulated in lung adenocarcinoma patients with EGFR exon 19 deletion compared with wild-type. And then the MTT assay results showed that miR-3196 had no effect on the sensitivity of erlotinib. The results of apoptosis analysis showed that inhibition of miR-3196 and erlotinib induced more apoptosis in H1299 cells than erlotinib alone, and overexpressed miR-3196 and erlotinib induced less apoptosis in PC9 cells than erlotinib alone (P<0.05). It is suggested that microRNAs associate with EGFR exon 19 deletion and miR-3196 may be further explored as a potential predictor and targeted biomarker when it is difficult to get the tumors.

Isolated brain metastasis as a late recurrence of completely resected non-small cell lung cancer.

The brain is one of the most common sites for non-small cell lung cancer (NSCLC) metastasis; however, late isolated brain metastasis as a recurrence of NSCLC is rare. The present study describes a case of isolated solitary brain metastasis as a late recurrence of NSCLC, which occurred >2 years following the successful resection of the primary tumor, and was identified by magnetic resonance imaging. To the best of our knowledge, this is the first report of isolated brain metastasis as a postoperative recurrence of NSCLC. The aim of the present study was to highlight that, despite its rarity, such recurrence should be considered possible, and particular attention to the treatment of such patients should be paid.

EGFR, KRAS and ROS1 variants coexist in a lung adenocarcinoma patient.

The c-ros oncogene 1 (ROS1) fusion is almost mutually exclusive to epidermal growth factor receptor (EGFR), anaplastic lymphoma kinase (ALK) or Kirsten rat sarcoma viral oncogene homolog (KRAS) mutation in non-small cell lung cancer (NSCLC), and it is not seen in the literature for patients to exhibit three mutations. The present study reported a case of a 53-year-old male diagnosed with adenocarcinoma, exhibiting combined EGFR, KRAS mutations and ROS1 rearrangement. At the first line therapy, the patient was treated with crizotinib because of the KRAS mutation that is a known resistant factor of EGFR-TKI resistance, but no responsive. At the second line therapy, EGFR-TKI Icotinib revealed a good response until now. To the best of to our knowledge, this is the first case report of a patient with concurrent EGFR, KRAS mutations and ROS1 fusion. This patient had an excellent response to Icotinib but not crizotinib, suggesting that the EGFR mutation was the oncogenic driver but ROS1 fusion and KRAS mutation not.

Integrin beta 1 enhances the epithelial-mesenchymal transition in association with gefitinib resistance of non-small cell lung cancer.

We have previously shown that integrinß1 associates with gefitinib resistance. As epithelial-mesenchymal transition (EMT) also induces gefitinib resistance in vitro, we wished to determine the relation of them in gefitinib resistance. In this study, we show that integrinß1 induced epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) resistance in xenograft tumors and gefitinib-resistant NSCLC tumors acquired EMT phenotype. Furthermore, inhibition of integrinß1 reverses EMT, meanwhile overexpression and activation of integrinß1 aggravates EMT. Lastly, we further identified that integrinß1 enhanced EMT via FAK-AKT signaling pathway. These findings highlight a novel relation of integrinß1 and EMT in EGFR TKI resistant NSCLC.

RGD-targeted paramagnetic liposomes for early detection of tumor: in vitro and in vivo studies.

Magnetic resonance molecular imaging has emerged as a potential approach for tumor diagnosis in the last few decades. This approach consists of the delivery of MR contrast agents to the tumor by specific targeted carriers. For this purpose, a lipopeptide was constructed by using a cyclic RGD peptide headgroup coupled to palmitic acid anchors via a KGG tripeptide spacer. Targeted paramagnetic liposomes were then prepared by the incorporation of RGD-coupled-lipopeptides into lipid bilayers for specific bounding to tumor. In vitro, study demonstrated that RGD-targeted liposomes exhibited a better binding affinity to targeted cells than non-targeted liposomes. MR imaging of mice bearing A549 tumors with the RGD-targeted paramagnetic liposomes also resulted in a greater signal enhancement of tumor compared to non-targeted liposomes and pure contrast agents groups. In addition, biodistribution study also showed specific tumor targeting of RGD-targeted paramagnetic liposomes in vivo. Therefore, RGD-targeted paramagnetic liposomes prepared in the present study may be a more promising method for early tumor diagnosis.