Efficacy, long-term toxicity, and mechanistic studies of gold nanorods photothermal therapy of cancer in xenograft mice.

Gold nanorods (AuNRs)-assisted plasmonic photothermal therapy (AuNRs-PPTT) is a promising strategy for combating cancer in which AuNRs absorb near-infrared light and convert it into heat, causing cell death mainly by apoptosis and/or necrosis. Developing a valid PPTT that induces cancer cell apoptosis and avoids necrosis in vivo and exploring its molecular mechanism of action is of great importance. Furthermore, assessment of the long-term fate of the AuNRs after treatment is critical for clinical use. We first optimized the size, surface modification [rifampicin (RF) conjugation], and concentration (2.5 nM) of AuNRs and the PPTT laser power (2 W/cm2) to achieve maximal induction of apoptosis. Second, we studied the potential mechanism of action of AuNRs-PPTT using quantitative proteomic analysis in mouse tumor tissues. Several death pathways were identified, mainly involving apoptosis and cell death by releasing neutrophil extracellular traps (NETs) (NETosis), which were more obvious upon PPTT using RF-conjugated AuNRs (AuNRs@RF) than with polyethylene glycol thiol-conjugated AuNRs. Cytochrome c and p53-related apoptosis mechanisms were identified as contributing to the enhanced effect of PPTT with AuNRs@RF. Furthermore, Pin1 and IL18-related signaling contributed to the observed perturbation of the NETosis pathway by PPTT with AuNRs@RF. Third, we report a 15-month toxicity study that showed no long-term toxicity of AuNRs in vivo. Together, these data demonstrate that our AuNRs-PPTT platform is effective and safe for cancer therapy in mouse models. These findings provide a strong framework for the translation of PPTT to the clinic.

[Effects of ambient air pollution on acute respiratory diseases and symptoms among adults in three cities in the Yangtze River Delta in 2015].

OBJECTIVE: To understand the effects of ambient air pollution on the prevalence of acute respiratory diseases and symptoms among adults in three cities in the Yangtze River Delta. METHODS: During September to November 2015, 4144 permanent residents aged 18 years and above from four investigation sites in three cities in the Yangtze River Delta were randomly surveyed by questionnaire. Daily concentrations of air pollutants, including PM_(2. 5), NO_2、O_3、CO and SO_2 nearest to the investigation sites were collected from the department of environmental protection. Logistic regression was used to analyze the association between air pollution and acute respiratory diseases and symptoms after other risk factors were adjusted. RESULTS: The prevalence of acute respiratory diseases and symptoms in two weeks among adults were 0. 99% and 3. 88%, respectively. Chemicals related to air pollution(OR=2. 339, 95%CI 1. 156-4. 734) and allergy(OR=4. 857, 95%CI 2. 279-10. 350) were the risk factors of acute respiratory diseases in two weeks among adults while occupational hazards such as toxic chemicals and high temperature(OR=1. 796, 95%CI 1. 220-2. 644), family history of respiratory diseases(OR=2. 670, 95%CI 1. 865-3. 823) and allergy(OR=3. 703, 95%CI 2. 395-5. 725) were the risk factor of respiratory symptoms in two weeks among adults. In addition, the average exposure level of PM_(2. 5)in two weeks was associated with acute respiratory diseases(OR=1. 014, 95%CI 1. 000-1. 028) and symptoms(OR=1. 025, 95%CI 1. 018-1. 033) in two weeks among adults. CONCLUSION: The increases of the prevalence of acute respiratory diseases and symptoms among adults are associated with ambient air pollution in three cities in the Yangtze River Delta.

Quantitative Examination of the Active Targeting Effect: The Key Factor for Maximal Tumor Accumulation and Retention of Short-Circulated Biopolymeric Nanocarriers.

Targeted and nontargeted biopolymeric nanoparticles with identical hydrodynamic sizes and surface charges were quantitatively examined in terms of the pharmacokinetic and biodistribution differences in detail. In adding cancer cell targeting folate molecules to the surface of the heparin nanocarriers, the amount of drug delivered to the tumor is doubled, and tumor growth inhibition is significantly enhanced. The folate-targeted heparin particles offered similar therapeutic potentials compared to their synthetic long-circulating analogues, thus presenting a viable alternative for drug-delivery vehicle construction using biological polymers, which are easier for the body to eliminate.

Quantitative Analysis of Lipid-Rich Necrotic Core in Carotid Atherosclerotic Plaques by In Vivo Magnetic Resonance Imaging and Clinical Outcomes.

BACKGROUND The aim of this study was to explore the accuracy of in vivo magnetic resonance imaging (MRI) in the quantitative evaluation of lipid-rich necrotic core (LRNC) in carotid atherosclerotic plaques compared with histopathology, and to assess the association of LRNC size with cerebral ischemia symptoms. MATERIAL AND METHODS Thirty patients were enrolled and 19 patients (16 men and 3 women) were analyzed. All the patients were submitted to MRI on a Siemens Avanto (1.5-Tesla) device before carotid endarterectomy (CEA). The scanning protocol included three-dimensional time of flight (3D TOF), T1-weighted image (T1WI), T2-weighted image (T2WI), turbo spin-echo T2-weighted (T2-TSE), and contrast-enhanced T1-weighted image. MRI images were reviewed for quantitative measurements of LRNC areas. LRNC specimens were collected for histology. Percentages of LRNC area to total vessel area were assessed to determine the association of MRI with histological findings. RESULTS There were 151 pairs of matched MRI and pathological sections. LRNC area percentages (LRNC area/vessel area) measured by MRI and histology were 20.6±9.0% and 18.7±9.5%, respectively (r=0.69, p<0.001). Twelve out of 19 patients had symptoms (S-group; 3 had recent stroke, 3 had a recent stroke and a history of transient ischemic attack (TIA), and 6 had TIA); the remaining 7 subjects showed no symptoms (NS-group). LRNC area percentages in the S- and NS-groups were 22.2±5.8% and 12.6±10.7%, respectively (p<0.05). CONCLUSIONS MRI can quantitatively measure LRNC in carotid atherosclerotic plaques, and may be useful in predicting the rupture risk of plaques. These findings provide a basis for imaging use in individualized treatment plan.

Electrochemical Fabrication of Functional Gelatin-Based Bioelectronic Interface.

Gelatin remains one of the most important biopolymeric material platforms because of its availability, safety, biocompatibility, biodegradability, and stimuli-responsive properties. Here we report a simple, rapid, and reagentless anodic deposition method to assemble gelatin hydrogels from aqueous salt solutions onto an electrode surface. Results indicate that anodic reactions partially oxidize gelatin to yield a covalently cross-linked network that can perform multiple functions. First, anodically deposited gelatin remains activated, allowing covalent protein grafting and thus enabling biofunctionalization for electrochemical biosensing. Second, the anodically deposited gelatin retains its thermally responsive physical cross-linking properties that enable switching functions. Finally, the physical and chemical cross-linking mechanisms are reversible, which enables self-healing functions. Thus, anodic deposition provides a facile method to assemble gelatin-based multifunctional matrices for diverse applications in bioelectronics.

Tumor-expressed adrenomedullin accelerates breast cancer bone metastasis.

INTRODUCTION: Adrenomedullin (AM) is secreted by breast cancer cells and increased by hypoxia. It is a multifunctional peptide that stimulates angiogenesis and proliferation. The peptide is also a potent paracrine stimulator of osteoblasts and bone formation, suggesting a role in skeletal metastases-a major site of treatment-refractory tumor growth in patients with advanced disease. METHODS: The role of adrenomedullin in bone metastases was tested by stable overexpression in MDA-MB-231 breast cancer cells, which cause osteolytic bone metastases in a standard animal model. Cells with fivefold increased expression of AM were characterized in vitro, inoculated into immunodeficient mice and compared for their ability to form bone metastases versus control subclones. Bone destruction was monitored by X-ray, and tumor burden and osteoclast numbers were determined by quantitative histomorphometry. The effects of AM overexpression on tumor growth and angiogenesis in the mammary fat pad were determined. The effects of AM peptide on osteoclast-like multinucleated cell formation were tested in vitro. A small-molecule AM antagonist was tested for its effects on AM-stimulated ex vivo bone cell cultures and co-cultures with tumor cells, where responses of tumor and bone were distinguished by species-specific real-time PCR. RESULTS: Overexpression of AM mRNA did not alter cell proliferation in vitro, expression of tumor-secreted factors or cell cycle progression. AM-overexpressing cells caused osteolytic bone metastases to develop more rapidly, which was accompanied by decreased survival. In the mammary fat pad, tumors grew more rapidly with unchanged blood vessel formation. Tumor growth in the bone was also more rapid, and osteoclasts were increased. AM peptide potently stimulated bone cultures ex vivo; responses that were blocked by small-molecule adrenomedullin antagonists in the absence of cellular toxicity. Antagonist treatment dramatically suppressed tumor growth in bone and decreased markers of osteoclast activity. CONCLUSIONS: The results identify AM as a target for therapeutic intervention against bone metastases. Adrenomedullin potentiates osteolytic responses in bone to metastatic breast cancer cells. Small-molecule antagonists can effectively block bone-mediated responses to tumor-secreted adrenomedullin, and such agents warrant development for testing in vivo.

Prognostic value of pre-treatment PNH clone among the patients with aplastic anemia: a meta-analysis.

BACKGROUND: Paroxysmal nocturnal hemoglobinuria (PNH) clone can be detected in some patients with aplastic anemia (AA) before treatment. But the prognostic value of the presence of pre-treatment PNH clone for intensive immunosuppressive therapy (IIST) is controversial and no consensus on whether the occurrence of PNH/AA-PNH syndrome is related to pre-treatment PNH clone. OBJECTIVE: This study aims to summarize the prognostic value of the presence of pre-treatment PNH clone treated with IIST among the AA patients and to elucidate its relationship with the development of PNH / AA-PNH syndrome. METHODS: All published studies on the prognostic value of pre-treatment PNH clone among AA patients were retrieved. Pooled odds ratio (OR) was calculated to compare the rates, along with 95% confidence intervals (CI) and p value to assess whether the results were statistically significant. RESULTS: The meta-analysis consisted of 15 studies with a combined total of 1349 patients in the cohort. Pre-treatment PNH clone had a positive effect on AA patients 6-month (pooled OR = 1.49,95% Cl: 1.06-2.08, P = 0.020), 12-month (pooled OR = 3.10,95% Cl: 1.89-5.10, P = 0.000), and overall hematological response rate (pooled OR = 1.69,95% Cl: 1.07-2.68, P = 0.024) after IIST. Patients with pre-treatment PNH clone are more likely to develop PNH/AA-PNH syndrome after IIST(pooled OR = 2.78,95%Cl:1.21-6.39, P = 0.016). CONCLUSION: Patients with positive pre-treatment PNH clone had better hematological responses to IIST than negative. And, those patients are more likely to develop PNH/AA-PNH syndrome after IIST.

Facile and Scalable Synthesis and Self-Assembly of Chitosan Tartaric Sodium.

Chitosan-based nanostructures have been widely applied in biomineralization and biosensors owing to its polycationic properties. The creation of chitosan nanostructures with controllable morphology is highly desirable, but has met with limited success yet. Here, we report that nanostructured chitosan tartaric sodium (CS-TA-Na) is simply synthesized in large amounts from chitosan tartaric ester (CS-TA) hydrolyzed by NaOH solution, while the CS-TA is obtained by dehydration-caused crystallization. The structures and self-assembly properties of CS-TA-Na are carefully characterized by Fourier-transform infrared spectroscopy (FTIR), nuclear magnetic resonance spectroscopy (1H-NMR), X-ray diffraction (XRD), differential scanning calorimeter (DSC), transmission electron microscopy (TEM), a scanning electron microscope (SEM) and a polarizing optical microscope (POM). As a result, the acquired nanostructured CS-TA-Na, which is dispersed in an aqueous solution 20-50 nm in length and 10-15 nm in width, shows both the features of carboxyl and amino functional groups. Moreover, morphology regulation of the CS-TA-Na nanostructures can be easily achieved by adjusting the solvent evaporation temperature. When the evaporation temperature is increased from 4 °C to 60 °C, CS-TA-Na nanorods and nanosheets are obtained on the substrates, respectively. As far as we know, this is the first report on using a simple solvent evaporation method to prepare CS-TA-Na nanocrystals with controllable morphologies.

Facile Fabrication of a Functional Filter Tip for Highly Efficient Reduction of Nicotine Content in Mainstream Smoke.

The nicotine addiction problem is of great concern, particularly in adolescents. Notably, nicotine addiction drives humans to continue smoking. Notably, several diseases and disorders are caused by smoking. To date, various adsorbents have been proposed to develop a functionalization filter tip for reducing nicotine content in mainstream smoke. However, the nicotine adsorption efficiencies of most of the reported functionalization filter tips were not satisfactory, and their preparation process was complex and time-consuming. Herein, we demonstrate a highly active and adsorbing filter tip for cigarettes, fabricated by decorating polydopamine (PDA) on the surface of a commercial filter tip in situ. The PDA coating on the filter tip was obtained by the self-polymerization of dopamine (DA) within 16 h, which was quicker and easier than the preparation processes of other reported functionalized filter tips. Significantly, the PDA-decorated filter tip had a nicotine adsorption efficiency as high as ∼95%, which was much higher than most of the commercial filter tips.

Molecular imaging of pancreatic cancer in an animal model using targeted multifunctional nanoparticles.

BACKGROUND & AIMS: Identification of a ligand/receptor system that enables functionalized nanoparticles to efficiently target pancreatic cancer holds great promise for the development of novel approaches for the detection and treatment of pancreatic cancer. Urokinase plasminogen activator receptor (uPAR), a cellular receptor that is highly expressed in pancreatic cancer and tumor stromal cells, is an excellent surface molecule for receptor-targeted imaging of pancreatic cancer using multifunctional nanoparticles. METHODS: The uPAR-targeted dual-modality molecular imaging nanoparticle probe is designed and prepared by conjugating a near-infrared dye-labeled amino-terminal fragment of the receptor binding domain of urokinase plasminogen activator to the surface of functionalized magnetic iron oxide nanoparticles. RESULTS: We have shown that the systemic delivery of uPAR-targeted nanoparticles leads to their selective accumulation within tumors of orthotopically xenografted human pancreatic cancer in nude mice. The uPAR-targeted nanoparticle probe binds to and is subsequently internalized by uPAR-expressing tumor cells and tumor-associated stromal cells, which facilitates the intratumoral distribution of the nanoparticles and increases the amount and retention of the nanoparticles in a tumor mass. Imaging properties of the nanoparticles enable in vivo optical and magnetic resonance imaging of uPAR-elevated pancreatic cancer lesions. CONCLUSIONS: Targeting uPAR using biodegradable multifunctional nanoparticles allows for the selective delivery of the nanoparticles into primary and metastatic pancreatic cancer lesions. This novel receptor-targeted nanoparticle is a potential molecular imaging agent for the detection of pancreatic cancer.

Single chain epidermal growth factor receptor antibody conjugated nanoparticles for in vivo tumor targeting and imaging.

Epidermal growth factor receptor (EGFR) targeted nanoparticle are developed by conjugating a single-chain anti-EGFR antibody (ScFvEGFR) to surface functionalized quantum dots (QDs) or magnetic iron oxide (IO) nanoparticles. The results show that ScFvEGFR can be successfully conjugated to the nanoparticles, resulting in compact ScFvEGFR nanoparticles that specifically bind to and are internalized by EGFR-expressing cancer cells, thereby producing a fluorescent signal or magnetic resonance imaging (MRI) contrast. In vivo tumor targeting and uptake of the nanoparticles in human cancer cells is demonstrated after systemic delivery of ScFvEGFR-QDs or ScFvEGFR-IO nanoparticles into an orthotopic pancreatic cancer model. Therefore, ScFvEGFR nanoparticles have potential to be used as a molecular-targeted in vivo tumor imaging agent. Efficient internalization of ScFvEGFR nanoparticles into tumor cells after systemic delivery suggests that the EGFR-targeted nanoparticles can also be used for the targeted delivery of therapeutic agents.

Quantum dot-based quantification revealed differences in subcellular localization of EGFR and E-cadherin between EGFR-TKI sensitive and insensitive cancer cells.

Nanoparticle quantum dots (QDs) provide sharper and more photostable fluorescent signals than organic dyes, allowing quantification of multiple biomarkers simultaneously. In this study, we quantified the expression of epidermal growth factor receptor (EGFR) and E-cadherin (E-cad) in the same cells simultaneously by using secondary antibody-conjugated QDs with two different emission wavelengths (QD605 and QD565) and compared the cellular distribution of EGFR and E-cad between EGFR-tyrosine kinase inhibitor (TKI)-insensitive and -sensitive lung and head and neck cancer cell lines. Relocalization of EGFR and E-cad upon treatment with the EGFR-TKI erlotinib in the presence of EGF was visualized and analyzed quantitatively. Our results showed that QD-immunocytochemistry (ICC)-based technology can not only quantify basal levels of multiple biomarkers but also track the localization of the biomarkers upon biostimulation. With this new technology we found that in EGFR-TKI-insensitive cells, EGFR and E-cad were located mainly in the cytoplasm; while in sensitive cells, they were found mainly on the cell membrane. After induction with EGF, both EGFR and E-cad internalized to the cytoplasm, but the internalization capability in sensitive cells was greater than that in insensitive cells. Quantification also showed that inhibition of EGF-induced EGFR and E-cad internalization by erlotinib in the sensitive cells was stronger than that in the insensitive cells. These studies demonstrate substantial differences between EGFR-TKI-insensitive and -sensitive cancer cells in EGFR and E-cad expression and localization both at the basal level and in response to EGF and erlotinib. QD-based analysis facilitates the understanding of the features of EGFR-TKI-insensitive versus -sensitive cancer cells and may be used in the prediction of patient response to EGFR-targeted therapy.

Thorough survey and analysis of pulmonary lymphoepithelioma-like carcinoma in Macau and multimodality treatment for advanced disease.

OBJECTIVE: Pulmonary lymphoepithelioma-like carcinoma (LELC) is a rare type of non-small cell lung cancer. The clinical course and prognosis of advanced LELC are largely unknown. Few reports have discussed multimodality treatment for LELC. MATERIALS AND METHODS: This retrospective study identified records from 2007 to 2018 of pulmonary LELCs and other lung cancer subtypes from hospital information systems and collected demographic, treatment, and survival data. RESULTS: In this cohort of 69 LELCs (median age: 55.4), more female, non-smokers, and fewer right upper lobe tumors (4.3%) were observed in the LELC subgroup compared with others. The median overall survival (OS) of LELCs was 40 months, superior to other subtypes (p < 0.05), except adenocarcinoma (p = 0.062). Patients with early stage disease and primary tumor resection tended to have better OS in univariate analysis, but surgery was the independent predictor in multivariate analysis (0.042). The median OS of 52 advanced LELCs was 22.7 months. Platinum-based chemotherapy and radiotherapy with curative purpose were independent predictors for OS of advanced LELCs (p = 0.004 and 0.003, respectively). For patients who received multimodality treatment in advanced setting, the median line of treatments was two. The overall response and disease-control rates were 61.8% and 80.6%, respectively. There were no differences in response or survival between patients receiving taxane-combined and non-taxane-combined chemotherapy. However, patients treated with radiotherapy in upfront settings had significantly favorable response and progression-free survival compared with those without. One case with PD-L1 positivity had pembrolizumab in the 4th line and achieved tumor shrinkage and stable disease for 12 months. CONCLUSION: Patients who underwent radical resection of primary tumors had better prognoses. Patients with advanced LELC could achieve satisfactory survival by receiving multimodality treatment, including platinum-based chemotherapy and/or radiotherapy. Immune checkpoint inhibitors may be part of future therapies. A well-organized clinical trial should be performed to determine the optimal treatment regimen.

Ethanol-Precipitation-Assisted Highly Efficient Synthesis of Nitrogen-Doped Carbon Quantum Dots from Chitosan.

Nitrogen-doped carbon quantum dots (NCQDs) were prepared from chitosan through a hydrothermal reaction. When ethanol precipitation was used as the purification method, a high product yield of 85.3% was obtained. A strong blue fluorescence emission with a high quantum yield (QY) of 6.6% was observed from the NCQD aqueous solution. Physical and chemical characteristics of the NCQDs were carefully investigated by transmission electron microscopy (TEM), X-ray diffraction (XRD), Fourier transform infrared spectra (FTIR), Raman spectra, X-ray photoelectron spectroscopy (XPS), and transient fluorescence spectra. Experimental results showed that diameters of the NCQDs were in the range of 2-10 nm. The carbon quantum dots possess good water dispersibility and precipitation by ethanol. When used for metal ion detection, the detection limit of the NCQDs for Fe3+ was as low as 1.57 µM. This work proposed a facile method to synthesize NCQDs from chitosan with high yield and demonstrated that carbon quantum dots derived from chitosan were promising for ion detection.

Periampullary Metastases from Breast Cancer: A Case Report and Literature Review.

We presented a metastatic breast cancer case who was afflicted with obstructive jaundice caused by an ampullary neoplasm. Since jaundice due to periampullary metastasis from breast cancer was a rare entity, a literature review of similar cases through the PubMed database was done. A total of 23 additional cases were found. Among these 24 cases, 5 presented with periampullary metastasis synchronously with the diagnosis of breast cancer, while 19 had metachronous periampullary metastasis with an interval ranging between 1.3 and 23 years from the initial diagnosis of breast cancer to the emergence of jaundice. It is intriguing to establish a differential diagnosis for common bile tract stricture prior to tissue biopsy, even with diagnostic workups including serum tumor markers, MRI plus MRCP, ERCP with intraductal brushing, and endoscopic ultrasound, in that the clinical, radiological, and endoscopic findings of metastatic lesions overlapped extensively with those found with primary periampullary malignancies. An immunohistochemical portfolio including cytokeratin7/20 (CK7/20), homeobox protein CDX2, human epidermal growth factor receptor 2 (HER2/neu), estrogen receptor alfa (ERα), progesterone receptor (PgR), mammaglobin, gross cystic disease fluid protein-15 (GCDFP-15), and transacting T-cell-specific transcription factor (GATA-3) was helpful for differential diagnosis among cases with ambiguous microscopic features.

Synergy of Single-ion Conductive and Thermo-responsive Copolymer Hydrogels Achieving Anti-Arrhenius Ionic Conductivity.

Artificial intelligence sensations have aroused scientific interest from electronic conductors to bio-inspired ionic conductors. The conductivity of electrons decreases with increasing temperature, while the ionic conductivity agrees with an Arrhenius equation or a modified Vogel-Tammann-Fulcher (VTF) equation. Herein, thermo-responsive poly(N-isopropyl amide) (PNIPAm) and single-ion-conducting poly(2-acrylamido-2-methyl-1-propanesulfonic lithium salt) (PAMPSLi) were copolymerized via a facile radical polymerization to demonstrate a very intriguing anti-Arrhenius ionic conductivity behaviour during thermally induced volume-phase transition. These smart hydrogels presented very promising scaffolds for architecting flexible, wearable or advanced functional ionic devices.

Real-time detection of gene expression in cancer cells using molecular beacon imaging: new strategies for cancer research.

Development of novel approaches for quantitative analysis of gene expression in intact tumor cells should provide new means for cancer detection and for studying the response of cancer cells to biological and therapeutic reagents. We developed procedures for detecting the levels of expression of multiple genes in fixed as well as viable cells using molecular beacon imaging technology. We found that simultaneous delivery of molecular beacons targeting survivin and cyclin D1 mRNAs produced strong fluorescence in breast cancer but not in normal breast cells. Importantly, fluorescence intensity correlated well with the level of gene expression in the cells detected by real-time reverse transcription-PCR or Western blot analysis. We further show that molecular beacons can detect changes of survivin gene expression in viable cancer cells following epidermal growth factor stimulation, docetaxel treatment, and overexpression of p53 gene. Thus, molecular beacon imaging is a simple and specific method for detecting gene expression in cancer cells. It has great potential for cancer detection and drug development.

Phenformin enhances the therapeutic effect of selumetinib in KRAS-mutant non-small cell lung cancer irrespective of LKB1 status.

MEK inhibition is potentially valuable in targeting KRAS-mutant non-small cell lung cancer (NSCLC). Here, we analyzed whether concomitant LKB1 mutation alters sensitivity to the MEK inhibitor selumetinib, and whether the metabolism drug phenformin can enhance the therapeutic effect of selumetinib in isogenic cell lines with different LKB1 status. Isogenic pairs of KRAS-mutant NSCLC cell lines A549, H460 and H157, each with wild-type and null LKB1, as well as genetically engineered mouse-derived cell lines 634 (krasG12D/wt/p53-/-/lkb1wt/wt) and t2 (krasG12D/wt/p53-/-/lkb1-/-) were used in vitro to analyze the activities of selumetinib, phenformin and their combination. Synergy was measured and potential mechanisms investigated. The in vitro findings were then confirmed in vivo using xenograft models. The re-expression of wild type LKB1 increased phospho-ERK level, suggesting that restored dependency on MEK->ERK->MAPK signaling might have contributed to the enhanced sensitivity to selumetinib. In contrast, the loss of LKB1 sensitized cells to phenformin. At certain combination ratios, phenformin and selumetinib showed synergistic activity regardless of LKB1 status. Their combination reduced phospho-ERK and S6 levels and induced potent apoptosis, but was likely through different mechanisms in cells with different LKB1 status. Finally, in xenograft models bearing isogenic A549 cells, we confirmed that loss of LKB1 confers resistance to selumetinib, and phenformin significantly enhances the therapeutic effect of selumetinib. Irrespective of LKB1 status, phenformin may enhance the anti-tumor effect of selumetinib in KRAS-mutant NSCLC. The dual targeting of MEK and cancer metabolism may provide a useful strategy to treat this subset of lung cancer.

Elevated glucose activates protein synthesis in cultured cardiac myocytes.

Diabetes mellitus results in chronic hyperglycemia, a serious metabolic disorder associated with a markedly increased risk of cardiovascular disease. However, the effects of high glucose (HG) on cardiac myocyte growth have not been fully clarified. In this study, the effect of glucose on cardiac myocyte growth was examined using leucine incorporation as an index of protein synthesis. High glucose (HG, 25 mmol/L) increased leucine incorporation (167% +/- 0.2% over normal glucose, n=4, P<.01) compared with a physiological glucose concentration (5.5 mmol/L, normal glucose). The HG-induced increase in leucine incorporation was time- and dose-dependent and was not due to osmotic changes because 25 mmol/L mannitol did not change leucine incorporation. High glucose also significantly reduced elongation factor 2 phosphorylation, an effect known to result in increased protein synthesis at the elongation step. Western blot analysis showed that HG-activated protein kinase B (PKB), also called Akt (PKB/Akt), at 18 hours. High glucose-induced leucine incorporation was attenuated with phosphatidylinositol 3-kinase (PI3K) inhibition using wortmannin and LY294002 and by rapamycin, a mammalian target of rapamycin (mTOR) inhibitor, 72%, 64%, and 65% (P<.05), respectively. High glucose also activated extracellular signal-regulated kinase 1/2 activity with peak stimulation at 5 minutes. In addition, PD98059, an inhibitor of mitogen-activated protein kinase kinase, attenuated HG-induced leucine incorporation. These data show for the first time that elevated glucose increases protein synthesis in cardiac myocytes. The increase appears to be mediated by activation of PI3K-PKB/Akt and/or PI3K-mTOR as well as extracellular signal-regulated kinase 1/2. These results provide new evidence for a direct effect of glucose independent of insulin on cardiac myocyte growth.

Overexpression of 12-lipoxygenase and cardiac fibroblast hypertrophy.

Leukocyte-type 12-lipoxygenase (12-LO) catalyzes the conversion of arachidonic acid (C20:4) to 12-hydroperoxyeicosatetraenoic acid, which in turn reduces to 12-hydroxyeicosatetraenoic acid (12-HETE) by glutathione peroxidase. Results of studies in vascular smooth muscle and in adrenal glomerulosa cells have supported the concept that 12-LO is an important mediator of angiotensin II (Ang II) action. Studies also indicate that 12-HETE is a potent growth-promoting factor and facilitates proliferation in Chinese hamster ovary (CHO) fibroblast cells overexpressing the Ang II AT1a receptor (CHO-AT1a cells). However, until recently, the role of 12-LO in cardiac cells had not been explored. Cardiac fibroblasts are a major source of matrix proteins, which can lead directly to extracellular matrix deposition and cardiac fibrosis. To elucidate the role of the 12-LO pathway in fibroblast cell growth, 12-LO cDNA was stably transfected into fetal rat cardiac fibroblasts. The cells overexpressing 12-LO showed an increase in cell protein content and enlargement in cell size with a slowing of cell division rate. Furthermore, the cells overexpressing 12-LO showed increases in fibronectin and collagen deposition compared with mock-transfected cells. These features are most consistent with cellular hypertrophy instead of proliferation. It is proposed that cardiac fibroblast cells overexpressing 12-LO retain the characteristics of fibroblasts, but with additional features of myocytes that have the function of showing cell hypertrophy. These results provide the basis for proposing the hypothesis that enhanced 12-LO expression or activity could play a role in pathogenic cardiac enlargement.