Lysine Deprivation Induces AKT-AADAT Signaling and Overcomes EGFR-TKIs Resistance in EGFR-Mutant Non-Small Cell Lung Cancer Cells.

Epidermal growth factor receptor (EGFR) mutations are the most common driver genes in non-small cell lung cancer (NSCLC), especially in the Asian population. Although EGFR-tyrosine kinase inhibitors (TKIs) are influential in the treatment of EGFR-mutant NSCLC patients, acquired resistance inevitably occurs. Therefore, there is an urgent need to develop strategies to overcome this resistance. In addition, cancer cells with particular mutations appear more vulnerable to deficiency related to the availability of specific amino acids. However, it is still unknown which amino acid is affected in the case of EGFR-mutant NSCLC. In the present study, we established a screening platform based on amino acid deprivation and found that EGFR-mutant NSCLC cells are sensitive to short-term lysine deprivation. Moreover, we found that expression of the gene for the lysine catabolism enzyme α-aminoadipate aminotransferase (AADAT) increased under lysine deprivation, revealing that AADAT can be regulated by EGFR-AKT signaling. Finally, we found that lysine reduction can not only enhance the cytostatic effect of single-agent osimertinib but also overcome the resistance of EGFR-TKIs in EGFR-mutant NSCLC cells. In summary, our findings suggest that the introduction of lysine stress might act as an advancement in EGFR-mutant NSCLC therapy and offer a strategy to overcome EGFR-TKI resistance.

Uncertain Associations of Major Bleeding and Concurrent Use of Antiplatelet Agents and Chinese Medications: A Nested Case-Crossover Study.

Despite the evidence that some commonly used Chinese medications (CMs) have antiplatelet/anticoagulant effects, many patients still used antiplatelets combined with CMs. We conducted a nested case-crossover study to examine the associations between the concomitant use of antiplatelets and CMs and major bleeding using population-based health database in Taiwan. Among the cohort of 79,463 outpatients prescribed antiplatelets (e.g., aspirin and clopidogrel) continuously, 1,209 patients hospitalized with new occurring bleeding in 2012 and 2013 were included. Those recruited patients served as their own controls to compare different times of exposure to prespecified CMs (e.g., Asian ginseng and dong quai) and antiplatelet agents. The periods of case, control 1, and control 2 were defined as 1-4 weeks, 6-9 weeks, and 13-16 weeks before hospitalization, respectively. Conditional logistic regression analyses found that concurrent use of antiplatelet drugs with any of the prespecified CMs in the case period might not significantly increase the risks of bleeding over that in the control periods (OR = 1.00, 95% CI 0.51 to 1.95 and OR = 1.13, 95% CI 0.65 to 1.97). The study showed no strong relationships between hospitalization for major bleeding events and concurrent use of antiplatelet drugs with the prespecified CMs.

Increased adiponectin associated with poor survival in hepatocellular carcinoma.

BACKGROUND: Alterations of adiponectin (APN), one of the adipokines, have been associated with human cancers. However, the clinical significance and impacts of APN on hepatocellular carcinoma (HCC) remain undetermined. METHODS: Using immunohistochemistry, expression patterns of APN were semiquantitatively scored and further statistically correlated with clinicopathological characteristics and patient survival. Furthermore, the bioeffects and underlying mechanisms of ectopic APN overexpression were determined in Hep3B and HepG2 cells by XTT, immunoblotting, flowcytometry, and invasion assays with or without chemical inhibitors and neutralization antibody. RESULTS: We found that cytoplasmic APN staining in 85 cancerous lesions was increased and associated with a poor survival rate (P = 0.007), even when using the Cox regression model (OR = 3.590; 95 % CI = 1.240-10.394; P = 0.018). Ectopic overexpression of APN in Hep3B and HepG2 cells increased proliferation and invasion as well as the levels of p-AKT (Ser473), p-STAT3 (Tyr705), and those downstream, i.e., cyclin D1 and ß-catenin. Similar results were also demonstrated in a stable APN-overexpressing clone, HepG2#136. APN neutralization antibody and LY294002 blocked the APN-mediated effects via inhibition of activated AKT. CONCLUSIONS: Our results suggest that increased APN may contribute to HCC at least in part through its activation of AKT signalling and may serve as a prognostic factor in HCC.

Effects of surface functionality of carbon nanomaterials on short-term cytotoxicity and embryonic development in zebrafish.

Nanomaterials have been widely used in the biomedical field as gene/drug carriers, magnetic resonance imaging (MRI) contrast reagents, photothermal therapy reagents, fluorescent cellular markers, etc. The origins and working mechanisms of cytotoxicities of nanomaterials, however, are not well understood. It is often stated in the literature that a nanomaterial is non-toxic and biocompatible. In this study, we show that the short term cytotoxicity of a nanomaterial is determined by the surface functionality, rather than the core nanomaterial. A so-called "non-toxic and biocompatible" nanomaterial, such as core/shell iron-filled carbon nanoparticles (Fe@CNPs) and nanodiamonds (NDs), can become cytotoxic when a cationic surface functionality, such as imidazolium (IM) and tertiary methyl ammonium ethyl methacrylate (TMAEA) moieties, was grafted onto the surface. To investigate the contributions of surface functionalities and the core nanomaterials on cytotoxicity, two "non-toxic and biocompatible" Fe@CNPs and NDs were surface-modified with different surface functionalities, including anionic COOH, zwitterionic PVP, neutral OH, cationic IM and TMAEA, and investigated for their cytotoxicities in both in vitro cancer cells (HeLa and U-87MG cells) and in vivo embryo development of zebrafish. Among these surface functionalities, cationic IM and TMAEA functionalities of both Fe@CNPs and NDs cause acute cytotoxicity to a similar extent in the in vitro cancer cell experiments, as well as affect severely the embryonic development and survival rates of zebrafish. Other surface functionalities do not show particularly strong cytotoxicities. To obtain information regarding the origins of cytotoxicities, the effects of surface functionalities were also examined on the lactate dehydrogenase (LDH) levels, cellular ROS generation, apoptosis, and changes in lysosomal membrane integrity, mitochondrial membrane potential, the intracellular pH (pHi), and cell cycles. Our results clearly point out that surface functionality, rather than the core nanomaterials, plays a critical role in dictating the short-term cytotoxicities.

Would It Matter to Expose Elderly Patients Who Took Digoxin to Chinese Medications?

OBJECTIVES: Elderly patients seem vulnerable to digoxin toxicity because of their diminished organ functions and tendency to encounter drug interactions. The aim of this research was to explore the extent of the concurrent use of digoxin with Chinese medications (CMs), its contributing factors, and the relevant consequences. METHODS: A retrospective population-based cohort study was conducted using Longitudinal Health Insurance databases in Taiwan. Those elderly patients being prescribed with digoxin in outpatient settings in 2006 were evaluated for the incidence, prevalence, and duration of concurrent use with concentrated CMs in 2006. After 1:1 random matching to select the corresponding digoxin-only elderly users, univariate and multivariate logistic regression analyses were performed to explore factors associated with concomitant incident digoxin-CM use and incident digoxin-specific CM use. The relevant clinical and economic outcomes for a 3-month follow-up period from the initial exposure of incident digoxin-CM use were compared. RESULTS: Of 185,076 elderly, 6,374 were prescribed with digoxin and 789 were CM-digoxin users in 2006. The prevalence and incidence of concomitant CM use among digoxin elderly users were 0.43% and 0.22%, respectively. Although the other factors were not statistically significantly associated with incident CM-digoxin use, patients with heart diseases and with benign prostate hypertrophy had an increased likelihood of incident CM-digoxin use of 115% and 102%, respectively. Almost all the concerned clinical and economic outcomes were not statistically significantly different between incident exposure or not, except for the use of potassium-sparing and nonsteroidal anti-inflammatory drugs. CONCLUSIONS: There was a relatively low incidence of digoxin-CM use among the elderly in Taiwan. Although no significant effects on clinical and economic outcomes occurred, it is necessary to monitor potential side effects of digoxin more aggressively for those vulnerable elderly using digoxin with CMs, especially for those who tended to expose to incident digoxin-CM use elderly patients.

A review of potential harmful interactions between anticoagulant/antiplatelet agents and Chinese herbal medicines.

BACKGROUND: The risks attributed to drug-herb interactions, even when known, are often ignored or underestimated, especially for those involving anti-clotting drugs and Chinese medicines. The aim of this study was to structurally search and evaluate the existing evidence-based data associated with potential drug interactions between anticoagulant/antiplatelet drugs and Chinese herbal medicines (CHMs) and evaluate the documented mechanisms, consequences, and/or severity of interactions. METHODOLOGY AND FINDINGS: Information related to anticoagulant/antiplatelet drug-CHM interactions was retrieved from eight interaction-based textbooks, four web resources and available primary biomedical literature. The primary literature searches were conducted in English and/or Chinese from January 2000 through December 2011 using the secondary databases (e.g., PubMed, Airiti Library, China Journal full-text database). The search terms included the corresponding medical subject headings and key words. Herbs or natural products not used as a single entity CHM or in Chinese Medicinal Prescriptions were excluded from further review. The corresponding mechanisms and severity ratings of interactions were retrieved using MicroMedex®, Lexicomp® and Natural Medicines Comprehensive Database®. Finally, we found 90 single entity CHMs contributed to 306 documented drug-CHM interactions. A total of 194 (63.4%) interactions were verified for its evidence describing possible mechanisms and severity. Of them, 155 interactions (79.9%) were attributable to pharmacodynamic interactions, and almost all were rated as moderate to severe interactions. The major consequences of these interactions were increased bleeding risks due to the additive anticoagulant or antiplatelet effects of the CHMs, specifically danshen, dong quai, ginger, ginkgo, licorice, and turmeric. CONCLUSIONS/SIGNIFICANCE: Conventional anticoagulants and antiplatelet drugs were documented to have harmful interactions with some commonly used single entity CHMs. For those patients who are taking conventional anti-clotting medications with CHMs for cardiovascular or cerebrovascular diseases, the potential risks of increased bleeding due to drug-CHM interactions should not be ignored.

Concurrent use of antiplatelets, anticoagulants, or digoxin with Chinese medications: a population-based cohort study.

PURPOSE: We examined the extent of concurrent use of antiplatelets, anticoagulants, or digoxin with Chinese medications (CMs) and identified its associated factors. METHODS: A retrospective cohort study was conducted using one million random samples from the Longitudinal Health Insurance Database 2005 in Taiwan. High-risk Western medications (HRWMs) focused on in this study were antiplatelets (aspirin, clopidogrel, dipyridamole, ticlopidine), anticoagulants (heparin, warfarin), and digoxin. Concurrent use was described as having an overlapping use period of HRWM with CMs any time in 2005. Baseline demographics, comorbidities, and health service utilizations between patients with and without concurrent HRWM-CM use were compared. Logistic regression analyses were performed to identify factors associated with incident concurrent use. RESULTS: Of the 70,698 eligible HRWM users, 13.2 % used CMs concurrently for an average duration of 26.7 ± 43 days. The incidence of concurrent HRWM-CM use, which excluded prior CM use within 6 months preceding the first CM use, was 6.3 %. Warfarin or ticlopidine users were more likely to be prescribed with CMs than were the other HRWM users. Factors associated with an increasing incidence of concurrent HRWM-CM use included female sex, age 45-54 years, middle monthly income, higher number of outpatient visits or distinct prescribed medications, and a previous diagnosis of heart diseases, stroke, or hypertension. In contrast, age ≥ 65 years and higher medical expenditure were associated with a lower incidence of concurrent use. CONCLUSIONS: In the Taiwanese population, approximately one in eight HRWM users were concomitantly prescribed CMs. Whether such concurrent use is associated with adverse clinical outcomes warrants further investigations.

MEK inhibitors reverse resistance in epidermal growth factor receptor mutation lung cancer cells with acquired resistance to gefitinib.

Lung adenocarcinoma cells harboring epidermal growth factor receptor (EGFR) mutations are sensitive to EGFR tyrosine kinase inhibitors (TKIs), including gefitinib. Acquired resistance to EGFR-TKIs develops after prolonged treatments. The study was prompt to explore effective strategies against resistance to EGFR-TKIs. We established gefitinib resistant PC-9 cells which harbor EGFR exon 19 deletion. Known mechanisms for intrinsic or acquired EGFR-TKI resistance, including KRAS mutation, HER2 mutation, EGFR T790M mutation and MET gene amplification, were studied, and we did not observe any known mechanisms for intrinsic or acquired resistance to EGFR-TKIs in the resistant cells. In the parental PC-9 cells, labeled as PC-9/wt, gefitinib completely inhibited EGF-induced phosphorylation of EGFR, AKT and ERK. Gefitinib inhibited EGFR phosphorylation, but was unable to block EGF-induced phosphorylation of ERK in resistant cells, labeled as PC-9/gef cells, including PC-9/gefB4, PC-9/gefE3, and PC-9/gefE7 subclones. We detected NRAS Q61K mutation in the PC-9/gef cells but not the PC-9/wt cells. MEK inhibitors, either AZD6244 or CI1040, inhibited ERK phosphorylation and sensitized gefitinib-induced cytotoxicity in PC-9/gef cells. Whereas MEK inhibitors or gefitinib alone did not activate caspases in PC-9/gef cells, combination of gefitinib and AZD6244 or CI1040 induced apoptosis. Our in vivo studies showed that gefitinib inhibited growth of PC-9/wt xenografts but not PC-9/gef xenografts. Furthermore, combination of a MEK inhibitor and gefitinib inhibited growth of both PC-9/wt xenografts and PC-9/gefB4 xenografts. To conclude, persistent activation of ERK pathway contributes to the acquired gefitinib-resistance. Combined treatment of gefitinib and MEK inhibitors may be therapeutically useful for acquired gefitinib-resistance lung adenocarcinoma cells harboring EGFR mutations.