Waterborne Tebuconazole Exposure Induces Male-Biased Sex Differentiation in Zebrafish (Danio rerio) Larvae via Aromatase Inhibition.

Tebuconazole is a widely used fungicide for various crops that targets sterol 14-α-demethylase (CYP51) in fungi. However, attention has shifted to aromatase (CYP19) due to limited research indicating its reproductive impact on aquatic organisms. Herein, zebrafish were exposed to 0.5 mg/L tebuconazole at different developmental stages. The proportion of males increased significantly after long-term exposure during the sex differentiation phase (0-60, 5-60, and 19-60 days postfertilization (dpf)). Testosterone levels increased and 17ß-estradiol and cyp19a1a expression levels decreased during the 5-60 dpf exposure, while the sex ratio was equally distributed on coexposure with 50 ng/L 17ß-estradiol. Chemically activated luciferase gene expression bioassays determined that the male-biased sex differentiation was not caused by tebuconazole directly binding to sex hormone receptors. Protein expression and phosphorylation levels were specifically altered in the vascular endothelial growth factor signaling pathway despite excluding the possibility of tebuconazole directly interacting with kinases. Aromatase was selected for potential target analysis. Molecular docking and aromatase activity assays demonstrated the interactions between tebuconazole and aromatase, highlighting that tebuconazole poses a threat to fish populations by inducing a gender imbalance.

Selenium-binding protein 1 inhibits malignant progression and induces apoptosis via distinct mechanisms in non-small cell lung cancer.

BACKGROUND: Selenium is an essential trace element in the human body. In epidemiological and clinical studies, Se supplementation significantly reduced the incidence of lung cancer in individuals with low baseline Se levels. The significant action of selenium is based on the selenium-containing protein as a mediator. Of note, the previous studies reported that the expression of selenium-binding protein 1 (SELENBP1) was obviously decreased in many human cancer tissues including non-small cell lung cancer (NSCLC). However, its roles in the origin and development of NSCLC are still unclear. METHODS: The expression of SELENBP1 was measured by qRT-PCR, Western blotting and IHC in our collected clinical NSCLC tissues and cell lines. Next, the CCK-8, colony formation, wound-haeling, Millicell, Transwell, FCM assay, and in vivo xenograft model were performed to explore the function of SELENBP1 in NSCLC. The molecular mechanisms of SELENBP1 were investigated by Western blotting or IF assay. RESULTS: We further identified that the expression of SELENBP1 was significantly decreased in NSCLC tissues in TCGA database and 45 out of 59 collected clinical NSCLC tissues compared with adjacent nontumor tissues, as well as in four NSCLC cell lines compared with normal lung cells. Particularly, we unexpectedly discovered that SELENBP1 was obviously expressed in alveolar type 2 (AT-II) cells for the first time. Then, a series of in vitro experiments uncovered that overexpression of SELENBP1 inhibited the proliferation, migration, and invasion of NSCLC cells, and induced cell apoptosis. Moreover, overexpression of SELENBP1 also inhibited growth and induced apoptosis of NSCLC cells in vivo. Mechanistically, we demonstrated that overexpression of SELENBP1 inhibited the malignant characteristics of NSCLC cells in part via inactivating the PI3K/AKT/mTOR signal pathway. Meanwhile, we found that overexpression of SELENBP1 inducing the apoptosis of NSCLC cells was associated with the activation of caspase-3 signaling pathway under nonhigh level of oxidative stress, but overexpression of SELENBP1 facilitating the cell apoptosis might be related to its combining with GPX1 and colocalizing in the nucleus under high level of oxidative stress. CONCLUSIONS: Our findings highlighted that SELENBP1 was an important tumor suppressor during the origin and development of NSCLC. It may help to discover novel biomarkers or drug therapy targets for NSCLC.

Association of Different Obesity Phenotypes with Sarcopenia in Han Chinese Middle-Aged and Elderly with Type 2 Diabetes Individuals.

Purpose: To investigate the relationship between different obesity phenotypes and sarcopenia in hospitalized Chinese patients with type 2 diabetes mellitus (T2DM). Methods: This cross-sectional study included 385 men. Anthropometric measurements including applied the determination method of Dual-energy X-ray absorptiometry (DXA) determination of limb skeletal muscle mass index (ASMI) and blood samples were analyzed. The people were divided into four groups according to body mass index (BMI) (≥24kg/m2) and waist circumference (WC) (female ≥85cm, male ≥90cm). Group A (BMI and WC were normal), Group B (BMI was normal and high WC), Group C (high BMI and WC were normal), and Group D (BMI and WC were abnormal). Results: The prevalence rates of sarcopenia and abdominal obesity were 32.2% and 74.0%, respectively. The detection rate of lower ASMI decreased gradually from Group A to Group D(74.6% vs 68.3% vs 54.5% vs 51.6%, χ 2 =14.243, P=0.003). Logistic analysis showed that the risk of lower ASMI were decreased by 62.4% (95% CI: 0.149-0.950, P = 0.039) in Group C and 68.8% (95% CI: 0.165-0.593, P = 0.000) in Group D compared with Group A, respectively. The risk of lower ASMI were increased 4.153-fold (95% CI: 2.623-6.576, P = 0.000) in male. Male (OR = 4.065, 95% CI: 2.246-7.356, P = 0.000) and WC (OR = 1.053, 95% CI: 1.004-1.104, P = 0.033) were risk factors for lower ASMI, but the risk of lower ASMI was decreased by 32% (95% CI: 0.5744-0.804, P = 0.000) by elevated BMI in the overweight and obese group (Group C and Group D). Conclusion: The prevalence of sarcopenia and abdominal obesity was elevated in han Chinese middle-aged and elderly patients with T2DM. Being overweight or obesity as defined by BMI protect against sarcopenia, while abdominal obesity increases the risk of sarcopenia.

ETS-1/c-Met drives resistance to sorafenib in hepatocellular carcinoma.

BACKGROUND: The purpose of this study was to clarify the molecular regulatory mechanism of c-Met up-regulated expression and elucidate the molecular mechanisms by which c-Met overexpression and activation drive progression and sorafenib resistance in hepatocellular carcinoma (HCC). METHODS: The resistance index was calculated. Bioinformatic techniques were applied to predict the transcription factors that bind and their binding sites on the c-Met promoter. Chromatin immunoprecipitation assays were implemented to verify the prediction results. To determine the regulatory mechanisms and effects of c-Met on sorafenib resistance in HCC, c-Met expression and activation were down-regulated by siRNA and inhibitor in in vivo and vitro experiments, while a parental cell line (Huh-7) was transfected with the adenovirus that upregulated c-Met expression. RESULTS: c-Met expression was increased in HCC sorafenib-resistant cells. Functional findings suggested that c-Met overexpression and activation drive HCC tumor progression and sorafenib resistance by promoting cell proliferation, migration, and stopping apoptosis. Molecular mechanism findings demonstrated that the MEK/ERK signaling pathway activated the expression and activity of ETS-1 mediated by p-ERK, which led to its binding to the c-Met gene promoter and upregulation of c-Met transcriptional expression. The activation of the HGF/c-Met pathway drives sorafenib resistance in HCC cells by activating the Ras/Raf/ERK and PI3K/Akt signaling pathways, which regulate biologic processes, including cell proliferation, migration and anti-apoptosis. CONCLUSION: c-Met overexpression and activation is an essential mechanism of sorafenib resistance in HCC. Combination therapy of sorafenib plus c-Met inhibitor overcame the resistance of sorafenib-targeted therapy for HCC.

IGF-1R down regulates the sensitivity of hepatocellular carcinoma to sorafenib through the PI3K / akt and RAS / raf / ERK signaling pathways.

BACKGROUND: Insulin-like growth factor-1 receptor (IGF-1R) promotes cell proliferation and migration and inhibitsapoptosis, all of which can contribute to the development of cancers. METHOD: This study investigated the effect and mechanism of IGF-1R in mediating the desensitization of hepatocellular carcinoma (HCC) to sorafenib. RESULTS: IGF-1R, highly expressed in the HCC cell lines SK-Hep1 and HepG2, promotes cell proliferation, migration, and anti-apoptosis through PI3K / Akt and RAS / Raf / ERK signaling pathways, resulting in HCC resistance to sorafenib. Knockdown of IGF-1R by RNA interference decreased proliferation and cell migration and upregulation of sorafenib-induced apoptosis of HCC cells. In vivo studies demonstrated that IGF-1R knockdown inhibited the growth of SK-Hep1 xenografts. CONCLUSION: These data are evidence that IGF-1R participates in regulating the survival and cell growth of HCC through the PI3K / Akt and RAS / Raf / ERK signaling pathways. Intervention in the expression of IGF-1R may increase the inhibitory effect of sorafenib on HCC.

Nanoparticles Loaded with GSK1059615 Combined with Sorafenib Inhibited Programmed Cell Death 1 Ligand 1 Expression by Negatively Regulating the PI3K/Akt/NF-κB Pathway, Thereby Reversing the Drug Resistance of Hepatocellular Carcinoma to Sorafenib.

Activation of the cellular signaling pathways can induce sorafenib-resistant hepatocellular carcinoma (HCCR). In this work, the PI3K/mTOR inhibitor GSK1059615 inhibited the proliferation and invasion of HCCR cells. PLGA-PEG-mal diblock copolymer was used to load GSK1059615 and sorafenib, and the vector was further modified with GPC3 antibody (hGC33) to obtain hGC33-modified GSK1059615 and sorafenib-loaded nanoparticles (Ab-G/S-NP). Ab-G/S-NP regulated the activation of cellular signaling pathways in HCCR cells by inhibiting the expression and activation of NF-κB and downregulating the level of programmed cell death 1 ligand 1(PD-L1) to reverse drug resistance of HCCR cells to sorafenib. These findings deserve further study in the combined treatment of HCCR cells with GSK1059615 in vivo to develop a more effective treatment of sorafenib-resistant cancers.

Prolonged exposure of azocyclotin induced inter- and transgenerational endocrine disruption on Danio rerio linked to transcriptomic and DNA methylomic alterations.

The transgenerational effect assessment linked to epigenetic analysis of environmental pollutants on eco (toxico)logical relevant species is regarded as a potential future risk-assessment tool. As an organotin acaricide widely used in China, azocyclotin can lead to endocrine disrupting effect on directly exposed environmental organisms, but whether it has transgenerational negative impact remains unknown. In order to illustrate this issue, in the present study, zebrafish, an aquatic model animal, was exposed to azocyclotin at less than µg/L level in a time span of embryonic stage to adult stage. Subsequently, the developmental and reproductive endocrine disrupting effects of azocyclotin on exposed F0 and unexposed offspring (F1 and F2) were evaluated. Result indicated that parentally exposed to 0.36 µg/L azocyclotin induced embryonic toxicity to unexposed offspring, and significantly (p < 0.05) reduced body weight (by 8.5%-13.9%), whole body length (by 4.8%-14.3%), hepatosomatic index (by 15.6%-24.3%), gonadosomatic index (by 5.3%-17.1%), egg production (by 19.5%-25.4%), estradiol content (47.0%-65.0%) and proportion of mature germ cells (by 29.3%-41.0% and 39.2%-47.7% for late oocytes and spermatozoa, respectively) in adults of F0 and offspring. Additionally, azocyclotin decreased the contents of 5-methycytosine in gonads of unexposed offspring (by 9.9%-38.6%, p < 0.05), led to genome-wide gene up-regulated expression bias and genomic DNA hypomethylation tendency in unexposed offspring. Moreover, based on the level of differentially methylated cytosine in promoter regions/gene body regions, it was found totally 5331/11,170 (in F1) and 3808/7507 (in F2) differentially expressed genes were closely related with differentially methylated genes (r > 0.6). The present study provided a primary evidence that prolonged exposure to low dose azocyclotin induced inter- and transgenerational endocrine disrupting effects on zebrafish probably linked to transcriptomic and DNA methylomic alterations.

The effects of coal dust exposure on DNA damage and repair of human bronchial epithelial cells.

To explore the effects of coal dust exposure on DNA damage and repair, human bronchial epithelial BEAS-2B cells were exposed to coal dust and the cellular response was investigated. It was found that γ-H2AX foci of DNA damage appeared, γ-H2AX protein level increased, and the rate of cell apoptosis was significantly elevated when BEAS-2B cells were exposed to coal dust for a short time. Phagocytized coal dust particles, swollen mitochondria, and reduced mitochondrial membrane potential were simultaneously identified. Moreover, Caspase-9, Caspase-3, and DFF45 proteins of the mitochondrial apoptotic pathway were activated. After the cells were exposed to coal dust chronically, phosphorylation levels of DNA repair kinases (ATM/ATR, DNA-PKcs) and downstream regulatory protein AKT were significantly upregulated. γ-H2AX foci and tail DNA of the cells following treatment with cisplatin were also reduced, and the colony formation rate was improved. It was concluded that coal dust could induce DNA damage, cause mitochondrial depolarization, and activate mitochondrial apoptosis pathways in BEAS-2B cells. Additionally, activated DNA repair kinases (ATM/ATR and DNA-PKcs) and their regulatory protein AKT increased DNA repair and proliferation of BEAS-2B cells chronically exposed to coal dust.

Estrogen receptor: A potential linker of estrogenic and dopaminergic pathways in zebrafish larvae following deltamethrin exposure.

Deltamethrin (DM), a type II pyrethroid insecticide, is widely used to control agricultural pests. However, its excessive use exerts a detrimental effect on the ecological environment and human health, indicating the need to study its potential risks in detail. In the present study, zebrafish embryos were exposed to varying concentrations of DM (0.1, 1, 10, and 25 µg/L) for 96 h to assess the alterations in the transcript levels of proteins of the estrogenic and dopaminergic pathways. In addition, its effect on zebrafish locomotor activity was studied. The mRNA expression of cyp19a1b, erα, erß2, fshr, gnrh2, gnrhr3, vtg3, dat, and dr1 significantly changed after exposing the embryos to DM. Deltamethrin at 10 and 25 µg/L significantly reduced the average swimming speed of zebrafish larvae. In addition, embryos injected with zebrafish estrogen receptor α (erα) and ß (erß) morpholinos and co-exposed to 25 µg/L DM for 96 h showed reduced expression of vtg3 mRNA compared to embryos exposed to 25 µg/L DM only. The locomotor activity of erα and erß knockdown zebrafish following DM exposure was increased significantly when compared with that of larvae exposed to 25 µg/L DM only. Our results demonstrated that DM altered the locomotor activity of zebrafish larvae and the transcript levels of the components of estrogenic and dopaminergic pathways; erα and erß knockdown weakened these effects.

Curcumin reverses hepatic epithelial mesenchymal transition induced by trichloroethylene by inhibiting IL-6R/STAT3.

OBJECTIVE: Epithelial mesenchymal transition (EMT) and inflammation have been identified as carcinogenic agents. This study aims to investigate whether inhibition of trichloroethylene (TCE) associated hepatocellular carcinoma (HCC) by curcumin is associated with inflammation and EMT. METHODS: In the current study, TCE sub-chronic cell model was induced in vitro, and the effects of TCE on cell proliferation, migration, invasion, and expression of functional proteins were verified by Western blot, MTT, clone formation, wound healing, Transwell. The detoxification effect of curcumin on TCE was explored by a mouse tumor-bearing experiment. RESULTS: TCE induces hepatocyte migration, colony formation, and EMT in vitro. In vivo studies have shown that curcumin significantly reduces the mortality of mice and control the occurrence and size of liver tumors by inhibiting the IL-6/STAT3 signaling pathway. In vitro, curcumin inhibits the proliferation of HepG2 cells as determined by MTT assay. In addition, curcumin significantly inhibited the protein expression of IL-6R, STAT3, snail, survivin, and cyclin D1 in THLE-2 and HepG2 cells induced by IL-6. CONCLUSION: Curcumin has anti-inflammatory and anti-proliferative effects, and inhibits the development of HCC induced by TCE by reversing IL-6/STAT3 mediated EMT.

Development of chemiluminescence method based on serum type I collagen hydroxyl terminal peptide ß special sequence (ß-CTX).

The objective of this work is to develop and verify the analytical performance of a chemiluminescence immunoassay for the specific sequence ß-carboxy-terminal cross-linking telopeptide of type I collagen (ß-CTX) in human serum. Two specific monoclonal antibodies (mAb-8A03 and mAb-3D12) with high affinity for ß-CTX were selected, and, under optimized conditions, a chemiluminescence immunoassay method (CLIA) for ß-CTX was established. The CLIA of ß-CTX detected ß-CTX in a wide range of 2.0-6000 ng/L. The recovery rate in serum is 95-105%, the specificity is high, and the cross-reaction rate with common easily interfering substances is low (not more than 0.01%). The CLIA correlates well with Roche electrochemiluminescence immunoassay (ECLIA), with a correlation coefficient of 0.9551, which fully meets the requirements of clinical analysis. The developed ß-CTX CLIA kit has high sensitivity and good stability. It has the same performance as the commercial Roche ECLIA kit and can be applied clinically.

BEZ235 Increases the Sensitivity of Hepatocellular Carcinoma to Sorafenib by Inhibiting PI3K/AKT/mTOR and Inducing Autophagy.

Acquired resistance of hepatocellular carcinoma (HCC) to sorafenib (SFB) is the main reason for the failure of SFB treatment of the cancer. Abnormal activation of the PI3K/AKT/mTOR pathway is important in the acquired resistance of SFB. Therefore, we investigated whether BEZ235 (BEZ) could reverse acquired sorafenib resistance by targeting the PI3K/mTOR pathway. A sorafenib-resistant HCC cell line Huh7R was established. MTT assay, clone formation assay, flow cytometry, and immunofluorescence were used to analyze the effects of BEZ235 alone or combined with sorafenib on cell proliferation, cell cycle, apoptosis, and autophagy of Huh7 and Huh7R cells. The antitumor effect was evaluated in animal models of Huh7R xenografts in vivo. Western blot was used to detect protein levels of the PI3K/AKT/mTOR pathway and related effector molecules. In vitro results showed that the Huh7R had a stronger proliferation ability and antiapoptosis effect than did Huh7, and sorafenib had no inhibitory effect on Huh7R. SFB + BEZ inhibited the activation of the PI3K/AKT/mTOR pathway caused by sorafenib. Moreover, SFB + BEZ inhibited the proliferation and cloning ability, blocked the cell cycle in the G0/G1 phase, and promoted apoptosis in the two cell lines. The autophagy level in Huh7R cells was higher than in Huh7 cells, and BEZ or SFB + BEZ further promoted autophagy in the two cell lines. In vivo, SFB + BEZ inhibited tumor growth by inducing apoptosis and autophagy. We concluded that BEZ235 enhanced the sensitivity of sorafenib through suppressing the PI3K/AKT/mTOR pathway and inducing autophagy. These observations may provide the experimental basis for sorafenib combined with BEZ235 in trial treatment of HCC.

LY­294002 enhances the chemosensitivity of liver cancer to oxaliplatin by blocking the PI3K/AKT/HIF­1α pathway.

Liver cancer remains one of the leading causes of cancer deaths worldwide. The therapeutic effect of oxaliplatin on liver cancer is often limited by acquired resistance of the cancer cells. Abnormal activation of the PI3K/AKT pathway plays an important role in the acquired resistance of oxaliplatin. The present study investigated the effects of the PI3K inhibitor LY­294002 and AKT inhibitor MK2206 on the chemosensitivity of oxaliplatin­resistant liver cancer cells and the molecular mechanism involved. An oxaliplatin­resistant liver cancer cell line HepG2R was developed. MTT assay, clone formation experiments, flow cytometry and Annexin V­FITC/PI staining were used to determine the proliferation, cycle and apoptosis of HepG2R cells when oxaliplatin was combined with LY­294002 or MK2206 treatment. The effects of LY­294002 and MK­2206 on the abnormal activation of PI3K/AKT pathway and hypoxia inducible factor (HIF)­1α protein level in HepG2R cells were detected using western blotting. The results indicated that the PI3K/AKT pathway is stably activated in HepG2R cells. Compared with the AKT inhibitor MK2206, the PI3K inhibitor LY­294002 more effectively downregulated the phosphorylation levels of p85, p110α, p110ß, p110γ and AKT in the PI3K/AKT pathway in HepG2R cells, and more effectively inhibited the proliferation of the cells. LY­294002 enhanced the chemotherapy sensitivity of HepG2R cells to oxaliplatin by inducing G0/G1 phase arrest and increasing the proportion of apoptotic cells. In addition, LY­294002 reduced the level of HIF­1α, which is highly expressed in HepG2R cells. It was concluded that LY­294002 enhanced the chemosensitivity of liver cancer cells to oxaliplatin by inhibiting the PI3K/AKT signaling pathway, which may be related to the inhibition of HIF­1α expression. These findings may have clinical significance for the treatment of oxaliplatin­resistant liver cancer.

LY3214996 relieves acquired resistance to sorafenib in hepatocellular carcinoma cells.

Background: Sorafenib, an oral multi-kinase inhibitor of rapidly accelerated fibrosarcoma; vascular endothelial growth factor receptor-2/3, platelet-derived growth factor receptor, c-Kit, and Flt-3 signaling, is approved for treatment of advanced hepatocellular carcinoma (HCC). However, the benefit of sorafenib is often diminished because of acquired resistance through the reactivation of ERK signaling in sorafenib-resistant HCC cells. In this work, we investigated whether adding LY3214996, a selective ERK1/2 inhibitor, to sorafenib would increase the anti-tumor effectiveness of sorafenib to HCC cells. Methods: The Huh7 cell line was used as a cell model for treatment with sorafenib, LY3214996, and their combination. Phosphorylation of the key kinases in the Ras/Raf/MAPK and PI3K/Akt pathways, protein expression of the cell cycle, and apoptosis migration were assessed with western blot. MTT and colony-formation assays were used to evaluate cell proliferation. Wound-healing assay was used to assess cell migration. Cell cycle and apoptosis analyses were conducted with flow cytometry. Results: LY3214996 decreased phosphorylation of the Ras/Raf/MAPK and PI3K/Akt pathways, including p-c-Raf, p-P90RSK, p-S6K and p-eIF4EBP1 activated by sorafenib, despite increased p-ERK1/2 levels. LY3214996 increased the anti-proliferation, anti-migration, cell-cycle progression, and pro-apoptotic effects of sorafenib on Huh7R cells. Conclusions: Reactivation of ERK1/2 appears to be a molecular mechanism of acquired resistance of HCC to sorafenib. LY3214996 combined with sorafenib enhanced the anti-tumor effects of sorafenib in HCC. These findings form a theoretical basis for trial of LY3214996 combined with sorafenib as second-line treatment of sorafenib-resistant in advanced HCC.

Targeted Delivery of Glypican 3 (GPC3) Antibody-Modified MicroRNA (miR let-7b-5p) Polymer Nanoparticles to Sorafenib-Resistant Hepatsocellular Carcinoma Cells.

The miR let-7b-5p (a kind of microRNAs) has many pathophysiological regulation effects, including human hepatocellular carcinoma (HCC) pathogenesis. This study investigated whether nanoparticle-mediated miR let-7b-5p could jointly enhance the therapeutic effect of sorafenib on HCC by inhibiting the proliferation of HCC cells, inducing apoptosis, and reversing drug resistance. We evaluated the level of miR let-7b-5p in sorafenib-resistant HepG2 cells (HepG2R) and HepG2 HCC cells by qRT-PCR and analyzed the biological effects of hepatocellular carcinoma treated with sorafenib with miR let-7b-5p, and further studied the toxicity of nanoparticles (Ab-miR-NPs) that deliver miR let-7b-5p mimics and target GPC3 on the surface of hepatocellular carcinoma cells. Results showed that, in HepG2 cells, the expression level of miR let-7b-5p was significantly higher than that in HepG2R cells. Targeted nanoparticle Ab-miR-NPs mediated the delivery of miR let-7b-5p to the HCC cytoplasm and released miRNA after being broken down, down-regulating the expression of IGF1R and inhibiting AKT/mTOR and Ras/Raf signal transmission. Ab-miR-NPs not only enhanced the proliferation of sorafenib in cultured HepG2R cells and induced cell apoptosis efficiency, but they also improved the anti-tumor activity in the mouse models. These results indicated that GPC3 antibody-modified PLGA-PLL (polylactic acid-glycolic acetic copolymer grafted hyper-branched polylysine) loaded miR let-7b-5p polymer nanoparticles combined with sorafenib may be a new treatment strategy for HCC resistant to sorafenib.

hGC33-Modified and Sorafenib-Loaded Nanoparticles have a Synergistic Anti-Hepatoma Effect by Inhibiting Wnt Signaling Pathway.

Delivery of tumor-specific inhibitors is a challenge in cancer treatment. Antibody-modified nanoparticles can deliver their loaded drugs to tumor cells that overexpress specific tumor-associated antigens. Here, we constructed sorafenib-loaded polyethylene glycol-b-PLGA polymer nanoparticles modified with antibody hGC33 to glypican-3 (GPC3 +), a membrane protein overexpressed in hepatocellular carcinoma. We found that hGC33-modified NPs (hGC33-SFB-NP) targeted GPC3+ hepatocellular carcinoma (HCC) cells by specifically binding to GPC3 on the surface of HCC cells, inhibited Wnt-induced signal transduction, and inhibited HCC cells in G0/1 by down-regulating cyclin D1 expression, thus attenuating HCC cell migration by inhibiting epithelial-mesenchymal transition. hGC33-SFB-NP inhibited the migration, cycle progression, and proliferation of HCC cells by inhibiting the Ras/Raf/MAPK pathway and the Wnt pathway in tandem with GPC3 molecules, respectively. hGC33-SFB-NP inhibited the growth of liver cancer in vivo and improved the survival rate of tumor-bearing mice. We conclude that hGC33 increases the targeting of SFB-NP to HCC cells. hGC33-SFB-NP synergistically inhibits the progression of HCC by blocking the Wnt pathway and the Ras/Raf/MAPK pathway.

Anti-GPC3 Antibody-Conjugated BEZ235 Loaded Polymeric Nanoparticles (Ab-BEZ235-NP) Enhances Radiosensitivity in Hepatocellular Carcinoma Cells by Inhibition of DNA Double-Strand Break Repair.

AIM: To assess AB-BEZ235-NP potential as a radio-sensitizer in hepatocellular carcinoma models. METHOD: By comparing hepatocellular carcinoma cell with simple radiation or combined AB-BEZ235-NP therapy, the HCC apoptosis and self-repair level have significant differences in mortality rates and cell migration abilities. RESULTS: Cell proliferation and DNA damage increased by pretreatment with AB-BEZ235-NP after irradiation; further studies on the repair pathway indicated that AB-BEZ235-NP inhibited the important pathway of DSB repair. Our results further show that AB-BEZ235-NP significantly inhibits the phosphorylation of the canonical protein, γ-H2AX, in the NHEJ DSB repair pathway and Serine Protein Kinase (SPK) ATM, and TP53-Binding Protein one. More importantly, AB-BEZ235-NP increased the mount of mean γ-H2AX Foci in irradiated cells, indicating that AB-BEZ235-NP can selectively inhibit DSB repair in HCC cells. Therefore, these results clearly eludicate that treatment with AB-BEZ235-NP is a potential promising therapy which can increase the radiosensitivity to HCC.

miR-410 induces both epithelial-mesenchymal transition and radioresistance through activation of the PI3K/mTOR pathway in non-small cell lung cancer.

Radiotherapy remains one of the major treatments for non-small cell lung cancer (NSCLC) patients; whereas intrinsic or acquired radioresistance limits its efficacy. Nevertheless, most studies so far have only focused on acquired resistance. The exact mechanisms of intrinsic radioresistance in NSCLC are still unclear. A few studies have suggested that epithelial-mesenchymal transition (EMT) is associated with radioresistance in NSCLC. However, little is known about whether the abnormal expression of specific microRNAs induces both EMT and radioresistance. We previously found that miR-410 has multiple roles as an oncomiRNA in NSCLC. In this study, we revealed that miR-410 overexpression promoted EMT and radioresistance, accompanied by enhanced DNA damage repair both in vitro and in vivo. Conversely, knockdown of miR-410 showed the opposite effects. We further demonstrated that PTEN was a direct target of miR-410 by using bioinformatic tools and dual-luciferase reporter assays, and the miR-410-induced EMT and radioresistance were reversed by PI3K, Akt, and mTOR inhibitors or by restoring the expression of PTEN in NSCLC cells. In addition, we preliminarily found that the expression of miR-410 was positively correlated with EMT and negatively associated with the expression of PTEN in NSCLC specimens. In summary, these results demonstrated that miR-410 is an important regulator on enhancing both NSCLC EMT and radioresistance by targeting the PTEN/PI3K/mTOR axis. The findings suggest that miR-410-induced EMT might significantly contribute to the enhanced radioresistance. Therefore, miR-410 may serve as a potential biomarker or therapeutic target for NSCLC radiotherapy.

Gefitinib sensitization of cisplatin-resistant wild-type EGFR non-small cell lung cancer cells.

PURPOSE: The usual first-line strategy of wild-type EGFR (wtEGFR) non-small cell lung cancer (NSCLC) remains cisplatin-based chemotherapy. However, cisplatin often loses effectiveness because most tumors acquire drug resistance over time. As EGFR is the most important pro-survival/proliferation signal receptor in NSCLC cells, we aimed at investigating whether cisplatin resistance is related to EGFR activation and further evaluating the combined effects of cisplatin/gefitinib (EGFR-tyrosine kinase inhibitor, EGFR-TKI) on cisplatin-resistant wtEGFR NSCLC cells. MATERIALS AND METHODS: EGFR activation was analysed in parental and cisplatin-resistant wtEGFR NSCLC cell lines (H358 and H358R, A549 and A549R). Cellular proliferation and apoptosis of H358R/A549R cells treated with cisplatin or gefitinib, alone or in combination were investigated, and the related effector protein was detected by western blot analysis. Anti-tumor effect of two drugs combined was evaluated in animal models of H358R xenografts in vivo. RESULTS: EGFR was significantly phosphorylated in cisplatin-resistant wtEGFR NSCLC cells H358R and A549R than their parental cells. In H358R and A549R cells, anti-proliferative ability of gefitinib was further improved, and gefitinib combined with cisplatin enhanced inhibition of cellular survive/proliferation, and promotion of apoptosis in vitro. The combined effects were also associated with the inhibition of EGFR downstream effector proteins. Similarly, in vivo, gefitinib and cisplatin in combination significantly inhibited tumor growth of H358R xenografts. CONCLUSION: Abnormal activation of EGFR may induce wtEGFR NSCLC cell resistance to cisplatin. The combined effects of cisplatin/gefitinib suggest that gefitinib, as a combination therapy for patients with cisplatin-resistant wtEGFR NSCLC should be considered.