Identification of Enriched Driver Gene Alterations in Subgroups of Non-Small Cell Lung Cancer Patients Based on Histology and Smoking Status

BACKGROUND: Appropriate patient selection is needed for targeted therapies that are efficacious only in patients with specific genetic alterations. We aimed to define subgroups of patients with candidate driver genes in patients with non-small cell lung cancer. METHODS: Patients with primary lung cancer who underwent clinical genetic tests at Guangdong General Hospital were enrolled. Driver genes were detected by sequencing, high-resolution melt analysis, qPCR, or multiple PCR and RACE methods. RESULTS: 524 patients were enrolled in this study, and the differences in driver gene alterations among subgroups were analyzed based on histology and smoking status. In a subgroup of non-smokers with adenocarcinoma, EGFR was the most frequently altered gene, with a mutation rate of 49.8%, followed by EML4-ALK (9.3%), PTEN (9.1%), PIK3CA (5.2%), c-Met (4.8%), KRAS (4.5%), STK11 (2.7%), and BRAF (1.9%). The three most frequently altered genes in a subgroup of smokers with adenocarcinoma were EGFR (22.0%), STK11 (19.0%), and KRAS (12.0%). We only found EGFR (8.0%), c-Met (2.8%), and PIK3CA (2.6%) alterations in the non-smoker with squamous cell carcinoma (SCC) subgroup. PTEN (16.1%), STK11 (8.3%), and PIK3CA (7.2%) were the three most frequently enriched genes in smokers with SCC. DDR2 and FGFR2 only presented in smokers with SCC (4.4% and 2.2%, respectively). Among these four subgroups, the differences in EGFR, KRAS, and PTEN mutations were statistically significant. CONCLUSION: The distinct features of driver gene alterations in different subgroups based on histology and smoking status were helpful in defining patients for future clinical trials that target these genes. This study also suggests that we may consider patients with infrequent alterations of driver genes as having rare or orphan diseases that should be managed with special molecularly targeted therapies

Outer Membrane Vesicles Derived from Escherichia coli Induce Systemic Inflammatory Response Syndrome

Sepsis, characterized by a systemic inflammatory state that is usually related to Gram-negative bacterial infection, is a leading cause of death worldwide. Although the annual incidence of sepsis is still rising, the exact cause of Gram-negative bacteria-associated sepsis is not clear. Outer membrane vesicles (OMVs), constitutively secreted from Gram-negative bacteria, are nano-sized spherical bilayered proteolipids. Using a mouse model, we showed that intraperitoneal injection of OMVs derived from intestinal Escherichia coli induced lethality. Furthermore, OMVs induced host responses which resemble a clinically relevant condition like sepsis that was characterized by piloerection, eye exudates, hypothermia, tachypnea, leukopenia, disseminated intravascular coagulation, dysfunction of the lungs, hypotension, and systemic induction of tumor necrosis factor-α and interleukin-6. Our study revealed a previously unidentified causative microbial signal in the pathogenesis of sepsis, suggesting OMVs as a new therapeutic target to prevent and/or treat severe sepsis caused by Gram-negative bacterial infection

Heterodimerization of Glycosylated Insulin-Like Growth Factor-1 Receptors and Insulin Receptors in Cancer Cells Sensitive to Anti-IGF1R Antibody

Identification of predictive biomarkers is essential for the successful development of targeted therapy. Insulin-like growth factor 1 receptor (IGF1R) has been examined as a potential therapeutic target for various cancers. However, recent clinical trials showed that anti-IGF1R antibody and chemotherapy are not effective for treating lung cancer.In order to define biomarkers for predicting successful IGF1R targeted therapy, we evaluated the anti-proliferation effect of figitumumab (CP-751,871), a humanized anti-IGF1R antibody, against nine gastric and eight hepatocellular cancer cell lines. Out of 17 cancer cell lines, figitumumab effectively inhibited the growth of three cell lines (SNU719, HepG2, and SNU368), decreased p-AKT and p-STAT3 levels, and induced G 1 arrest in a dose-dependent manner. Interestingly, these cells showed co-overexpression and altered mobility of the IGF1R and insulin receptor (IR). Immunoprecipitaion (IP) assays and ELISA confirmed the presence of IGF1R/IR heterodimeric receptors in figitumumab-sensitive cells. Treatment with figitumumab led to the dissociation of IGF1-dependent heterodimeric receptors and inhibited tumor growth with decreased levels of heterodimeric receptors in a mouse xenograft model. We next found that both IGF1R and IR were N-linked glyosylated in figitumumab-sensitive cells. In particular, mass spectrometry showed that IGF1R had N-linked glycans at N913 in three figitumumab-sensitive cell lines. We observed that an absence of N-linked glycosylation at N913 led to a lack of membranous localization of IGF1R and figitumumab insensitivity.The data suggest that the level of N-linked glycosylated IGF1R/IR heterodimeric receptor is highly associated with sensitivity to anti-IGF1R antibody in cancer cells

High Content Screening Identifies Decaprenyl-Phosphoribose 2′ Epimerase as a Target for Intracellular Antimycobacterial Inhibitors

A critical feature of Mycobacterium tuberculosis, the causative agent of human tuberculosis (TB), is its ability to survive and multiply within macrophages, making these host cells an ideal niche for persisting microbes. Killing the intracellular tubercle bacilli is a key requirement for efficient tuberculosis treatment, yet identifying potent inhibitors has been hampered by labor-intensive techniques and lack of validated targets. Here, we present the development of a phenotypic cell-based assay that uses automated confocal fluorescence microscopy for high throughput screening of chemicals that interfere with the replication of M. tuberculosis within macrophages. Screening a library of 57,000 small molecules led to the identification of 135 active compounds with potent intracellular anti-mycobacterial efficacy and no host cell toxicity. Among these, the dinitrobenzamide derivatives (DNB) showed high activity against M. tuberculosis, including extensively drug resistant (XDR) strains. More importantly, we demonstrate that incubation of M. tuberculosis with DNB inhibited the formation of both lipoarabinomannan and arabinogalactan, attributable to the inhibition of decaprenyl-phospho-arabinose synthesis catalyzed by the decaprenyl-phosphoribose 2′ epimerase DprE1/DprE2. Inhibition of this new target will likely contribute to new therapeutic solutions against emerging XDR-TB. Beyond validating the high throughput/content screening approach, our results open new avenues for finding the next generation of antimicrobials

Proteomic Analysis of Fusarium solani Isolated from the Asian Longhorned Beetle, Anoplophora glabripennis

Wood is a highly intractable food source, yet many insects successfully colonize and thrive in this challenging niche. Overcoming the lignin barrier of wood is a key challenge in nutrient acquisition, but full depolymerization of intact lignin polymers has only been conclusively demonstrated in fungi and is not known to occur by enzymes produced by insects or bacteria. Previous research validated that lignocellulose and hemicellulose degradation occur within the gut of the wood boring insect, Anoplophora glabripennis (Asian longhorned beetle), and that a fungal species, Fusarium solani (ATCC MYA 4552), is consistently associated with the larval stage. While the nature of this relationship is unresolved, we sought to assess this fungal isolate's ability to degrade lignocellulose and cell wall polysaccharides and to extract nutrients from woody tissue. This gut-derived fungal isolate was inoculated onto a wood-based substrate and shotgun proteomics using Multidimensional Protein Identification Technology (MudPIT) was employed to identify 400 expressed proteins. Through this approach, we detected proteins responsible for plant cell wall polysaccharide degradation, including proteins belonging to 28 glycosyl hydrolase families and several cutinases, esterases, lipases, pectate lyases, and polysaccharide deacetylases. Proteinases with broad substrate specificities and ureases were observed, indicating that this isolate has the capability to digest plant cell wall proteins and recycle nitrogenous waste under periods of nutrient limitation. Additionally, several laccases, peroxidases, and enzymes involved in extracellular hydrogen peroxide production previously implicated in lignin depolymerization were detected. In vitro biochemical assays were conducted to corroborate MudPIT results and confirmed that cellulases, glycosyl hydrolases, xylanases, laccases, and Mn- independent peroxidases were active in culture; however, lignin- and Mn- dependent peroxidase activities were not detected While little is known about the role of filamentous fungi and their associations with insects, these findings suggest that this isolate has the endogenous potential to degrade lignocellulose and extract nutrients from woody tissue

Bioaccumulation and ecotoxicity of carbon nanotubes

Carbon nanotubes (CNT) have numerous industrial applications and may be released to the environment. In the aquatic environment, pristine or functionalized CNT have different dispersion behavior, potentially leading to different risks of exposure along the water column. Data included in this review indicate that CNT do not cross biological barriers readily. When internalized, only a minimal fraction of CNT translocate into organism body compartments. The reported CNT toxicity depends on exposure conditions, model organism, CNT-type, dispersion state and concentration. In the ecotoxicological tests, the aquatic organisms were generally found to be more sensitive than terrestrial organisms. Invertebrates were more sensitive than vertebrates. Single-walled CNT were found to be more toxic than double-/multi-walled CNT. Generally, the effect concentrations documented in literature were above current modeled average environmental concentrations. Measurement data are needed for estimation of environmental no-effect concentrations. Future studies with benchmark materials are needed to generate comparable results. Studies have to include better characterization of the starting materials, of the dispersions and of the biological fate, to obtain better knowledge of the exposure/effect relationships