ß-Lapachone promotes the recruitment and polarization of tumor-associated neutrophils (TANs) toward an antitumor (N1) phenotype in NQO1-positive cancers.

NAD(P)H:quinone oxidoreductase 1 (NQO1) is overexpressed in most solid cancers, emerging as a promising target for tumor-selective killing. ß-Lapachone (ß-Lap), an NQO1 bioactivatable drug, exhibits significant antitumor effects on NQO1-positive cancer cells by inducing immunogenic cell death (ICD) and enhancing tumor immunogenicity. However, the interaction between ß-Lap-mediated antitumor immune responses and neutrophils, novel antigen-presenting cells (APCs), remains unknown. This study demonstrates that ß-Lap selectively kills NQO1-positive murine tumor cells by significantly increasing intracellular ROS formation and inducing DNA double strand breaks (DSBs), resulting in DNA damage. Treatment with ß-Lap efficiently eradicates immunocompetent murine tumors and significantly increases the infiltration of tumor-associated neutrophils (TANs) into the tumor microenvironment (TME), which plays a crucial role in the drug's therapeutic efficacy. Further, the presence of ß-Lap-induced antigen medium leads bone marrow-derived neutrophils (BMNs) to directly kill murine tumor cells, aiding in dendritic cells (DCs) recruitment and significantly enhancing CD8+ T cell proliferation. ß-Lap treatment also drives the polarization of TANs toward an antitumor N1 phenotype, characterized by elevated IFN-ß expression and reduced TGF-ß cytokine expression, along with increased CD95 and CD54 surface markers. ß-Lap treatment also induces N1 TAN-mediated T cell cross-priming. The HMGB1/TLR4/MyD88 signaling cascade influences neutrophil infiltration into ß-Lap-treated tumors. Blocking this cascade or depleting neutrophil infiltration abolishes the antigen-specific T cell response induced by ß-Lap treatment. Overall, this study provides comprehensive insights into the role of tumor-infiltrating neutrophils in the ß-Lap-induced antitumor activity against NQO1-positive murine tumors.

Augmented Concentration of Isopentyl-Deoxynyboquinone in Tumors Selectively Kills NAD(P)H Quinone Oxidoreductase 1-Positive Cancer Cells through Programmed Necrotic and Apoptotic Mechanisms.

Lung and breast cancers rank as two of the most common and lethal tumors, accounting for a substantial number of cancer-related deaths worldwide. While the past two decades have witnessed promising progress in tumor therapy, developing targeted tumor therapies continues to pose a significant challenge. NAD(P)H quinone oxidoreductase 1 (NQO1), a two-electron reductase, has been reported as a promising therapeutic target across various solid tumors. ß-Lapachone (ß-Lap) and deoxynyboquinone (DNQ) are two NQO1 bioactivatable drugs that have demonstrated potent antitumor effects. However, their curative efficacy has been constrained by adverse effects and moderate lethality. To enhance the curative potential of NQO1 bioactivatable drugs, we developed a novel DNQ derivative termed isopentyl-deoxynyboquinone (IP-DNQ). Our study revealed that IP-DNQ treatment significantly increased reactive oxygen species generation, leading to double-strand break (DSB) formation, PARP1 hyperactivation, and catastrophic energy loss. Notably, we discovered that this novel drug induced both apoptosis and programmed necrosis events, which makes it entirely distinct from other NQO1 bioactivatable drugs. Furthermore, IP-DNQ monotherapy demonstrated significant antitumor efficacy and extended mice survival in A549 orthotopic xenograft models. Lastly, we identified that in mice IP-DNQ levels were significantly elevated in the plasma and tumor compared with IB-DNQ levels. This study provides novel preclinical evidence supporting IP-DNQ efficacy in NQO1+ NSCLC and breast cancer cells.

IP-DNQ induces mitochondrial dysfunction and G2/M phase cell cycle arrest to selectively kill NQO1-positive pancreatic cancer cells.

Pancreatic cancer is among the top five leading causes of cancer-related deaths worldwide, with low survival rates. Current therapies for pancreatic cancer lack tumor specificity, resulting in harmful effects on normal tissues. Therefore, developing tumor-specific agents for the treatment of pancreatic cancer is critical. NAD(P)H:quinone oxidoreductase 1 (NQO1), highly expressed in pancreatic cancers but not in normal tissues, makes NQO1 bioactivatable drugs a potential therapy for selectively killing NQO1-positive cancer cells. Our previous studies have revealed that novel NQO1 bioactivatable drug deoxynyboquinone (DNQ) is ten-fold more potent than the prototypic NQO1 bioactivatable drug ß-lapachone in killing of NQO1-positive cancer cells. However, DNQ treatment results in high-grade methemoglobinemia, a significant side effect that limits clinical development. Here, we report for the first time on a DNQ derivative, isopentyl-deoxynboquinone (IP-DNQ), which selectively kills pancreatic ductal adenocarcinoma cells in an NQO1-dependent manner with equal potency to the parent DNQ. IP-DNQ evokes massive ROS production and oxidative DNA lesions that results in PARP1 hyperactivation, mitochondrial catastrophe and G2/M-phase arrest, leading to apoptotic and necrotic programmed cell death. Importantly, IP-DNQ treatment causes mild methemoglobinemia in vivo, with a three-fold improvement in the maximum tolerated dose compared to DNQ, while significantly suppresses tumor growth and extends the lifespan of mice in subcutaneous and orthotopic pancreatic cancer xenograft models. Our study demonstrates that IP-DNQ is a promising therapy for NQO1-positive pancreatic cancers and may enhance the efficacy of other anticancer drugs. IP-DNQ represents a novel approach to treating pancreatic cancer with the potential to improve patient outcomes.

Pathological complete response rate and clinical outcome after neoadjuvant therapy of HER2-low breast cancer: A National Cancer Database Analysis.

BACKGROUND: The interest in breast cancer with low HER2 expression as a distinct subtype is increasing. We aimed to explore the differences between HER2-low and HER2-zero breast cancer in their prognosis and rate of pathological complete response (pCR) after neoadjuvant therapy. METHODS: The National Cancer Database (NCDB) was used to select patients with breast cancer who received neoadjuvant therapy from 2004 to 2017. Logistic regression model was constructed for analysis of pCR. Cox proportional hazards regression model and Kaplan-Meier method were used for survival analysis. RESULTS: A total of 41500 breast cancer patients were included, among which 14814 (35.7%) had HER2-zero tumors and 26686 (64.3%) had HER2-low. HER2-low tumors were more commonly HR-positive in comparison with HER2-zero (66.3% versus 47.1%, P < 0.001). A lower rate of pCR was observed in HER2-low tumors than in HER2-zero tumors after neoadjuvant therapy in the total cohort (OR = 0.90; 95% CI [0.86-0.95]; P < 0.001) and in the subset of HR-positive (OR = 0.87; 95% CI [0.81-0.94]; P < 0.001). Patients with HER2-low tumors had a significantly superior survival than those with HER2-zero tumors (HR = 0.90; 95% CI [0.86-0.94]; P < 0.001), regardless of the HR status. Additionally, a marginal survival difference was also observed between HER2 IHC1+ and HER2 IHC2+/ISH-negative (HR = 0.91; 95% CI [0.85-0.97]; P = 0.003) cohorts. CONCLUSION: HER2-low tumors are a clinically relevant breast cancer subtype that is distinct from HER2-zero tumors. These findings may provide clues to appropriate therapeutic strategies for this subtype in the future.

KP372-1-Induced AKT Hyperactivation Blocks DNA Repair to Synergize With PARP Inhibitor Rucaparib via Inhibiting FOXO3a/GADD45α Pathway.

Poly (ADP-ribose) polymerase (PARP) inhibitors (PARPi) have exhibited great promise in the treatment of tumors with homologous recombination (HR) deficiency, however, PARPi resistance, which ultimately recovers DNA repair and cell progress, has become an enormous clinical challenge. Recently, KP372-1 was identified as a novel potential anticancer agent that targeted the redox enzyme, NAD(P)H:quinone oxidoreductase 1 (NQO1), to induce extensive reactive oxygen species (ROS) generation that amplified DNA damage, leading to cancer cell death. To overcome PARPi resistance and expand its therapeutic utility, we investigated whether a combination therapy of a sublethal dose of KP372-1 with a nontoxic dose of PARPi rucaparib would synergize and enhance lethality in NQO1 over-expressing cancers. We reported that the combination treatment of KP372-1 and rucaparib induced a transient and dramatic AKT hyperactivation that inhibited DNA repair by regulating FOXO3a/GADD45α pathway, which enhanced PARPi lethality and overcame PARPi resistance. We further found that PARP inhibition blocked KP372-1-induced PARP1 hyperactivation to reverse NAD+/ATP loss that promoted Ca2+-dependent autophagy and apoptosis. Moreover, pretreatment of cells with BAPTA-AM, a cytosolic Ca2+ chelator, dramatically rescued KP372-1- or combination treatment-induced lethality and significantly suppressed PAR formation and γH2AX activation. Finally, we demonstrated that this combination therapy enhanced accumulation of both agents in mouse tumor tissues and synergistically suppressed tumor growth in orthotopic pancreatic and non-small-cell lung cancer xenograft models. Together, our study provides novel preclinical evidence for new combination therapy in NQO1+ solid tumors that may broaden the clinical utility of PARPi.

ß-Lapachone Selectively Kills Hepatocellular Carcinoma Cells by Targeting NQO1 to Induce Extensive DNA Damage and PARP1 Hyperactivation.

Hepatocellular carcinoma (HCC) is the second leading cause of cancer-related death globally. Currently there is a lack of tumor-selective and efficacious therapies for hepatocellular carcinoma. ß-Lapachone (ARQ761 in clinical form) selectively kill NADPH: quinone oxidoreductase 1 (NQO1)-overexpressing cancer cells. However, the effect of ß-Lapachone on HCC is virtually unknown. In this study, we found that relatively high NQO1 and low catalase levels were observed in both clinical specimens collected from HCC patients and HCC tumors from the TCGA database. ß-Lapachone treatment induced NQO1-selective killing of HCC cells and caused ROS formation and PARP1 hyperactivation, resulting in a significant decrease in NAD+ and ATP levels and a dramatic increase in double-strand break (DSB) lesions over time in vitro. Administration of ß-Lapachone significantly inhibited tumor growth and prolonged survival in a mouse xenograft model in vivo. Our data suggest that NQO1 is an ideal potential biomarker, and relatively high NQO1:CAT ratios in HCC tumors but low ratios in normal tissues offer an optimal therapeutic window to use ß-Lapachone. This study provides novel preclinical evidence for ß-Lapachone as a new promising chemotherapeutic agent for use in NQO1-positive HCC patients.

PCNA inhibition enhances the cytotoxicity of ß-lapachone in NQO1-Positive cancer cells by augmentation of oxidative stress-induced DNA damage.

ß-Lapachone is a classic quinone-containing antitumor NQO1-bioactivatable drug that directly kills NQO1-overexpressing cancer cells. However, the clinical applications of ß-lapachone are primarily limited by its high toxicity and modest lethality. To overcome this side effect and expand the therapeutic utility of ß-lapachone, we demonstrate the effects of a novel combination therapy including ß-lapachone and the proliferating cell nuclear antigen (PCNA) inhibitor T2 amino alcohol (T2AA) on various NQO1+ cancer cells. PCNA has DNA clamp processivity activity mediated by encircling double-stranded DNA to recruit proteins involved in DNA replication and DNA repair. In this study, we found that compared to monotherapy, a nontoxic dose of the T2AA synergized with a sublethal dose of ß-lapachone in an NQO1-dependent manner and that combination therapy prevented DNA repair, increased double-strand break (DSB) formation and promoted programmed necrosis and G1 phase cell cycle arrest. We further determined that combination therapy enhanced antitumor efficacy and prolonged survival in Lewis lung carcinoma (LLC) xenografts model. Our findings show novel evidence for a new therapeutic approach that combines of ß-lapachone treatment with PCNA inhibition that is highly effective in treating NQO1+ solid tumor cells.

A novel colorimetric immunoassay based on enzyme-regulated instant generation of Turnbull's blue for the sensitive determination of ochratoxin A.

The aim of this study was to develop a novel colorimetric sensing method based on enzyme-regulated instant generation of Turnbull's blue, serving as a chromogenic agent, for a sensitive immunoassay for the determination of ochratoxin A (OTA). Unlike the traditional enzyme-linked immunosorbent assay (ELISA), the chromogenic reaction reported herein relies on the immediate formation of Turnbull's blue. K3[Fe(CN)6] rapidly forms a coordinate bond with iron(ii), yielding a blue product. Meanwhile, glucose oxidase (GOx) catalyzes glucose hydrolysis to produce hydrogen peroxide (H2O2), which was used to inhibit the formation of Turnbull's blue by oxidizing iron(ii) to iron(iii). Thus, Turnbull's blue was generated in an enzyme-regulated manner. Accordingly, a competitive-type colorimetric enzyme immunoassay was established using a GOx based nanolabel. Under optimal conditions, the absorbance increased upon increasing the target OTA concentration in the range of 0.01-10 ng mL-1 with a detection limit of 8.3 pg mL-1 estimated at the 3Sblank level. The assay accuracy was validated by analyzing spiked wine samples. The present results potentially provide novel insights into the development of Turnbull's blue-based biological detection methods and colorimetric immunoassay strategies.

Modulation of HIF-2α PAS-B domain contributes to physiological responses.

Hypoxia-inducible factors (HIFs) are transcription factors in the basic helix-loop-helix PER-ARNT-SIM (bHLH-PAS) protein family that contain internal hydrophobic cavities within their PAS-A and PAS-B domains. Among HIFs, the HIF-2α PAS-B domain contains a relatively large cavity exploited for the development of specific artificial ligands such as PT2399. Administration of PT2399 could suppress HIF-2α target gene expression without affecting HIF-1 activity in mice under hypoxia conditions. A single mutation (S305M) within the HIF-2α PAS-B domain suppressed HIF-2α activity while conferring resistance to PT2399 in vivo, indicating the vital role of PAS-B domain in HIF-2α hypoxia response. In contrast, the mutant mice did not phenocopy PT2399 intervention in wild-type mice under metabolic stress. Under a high-fat diet (HFD), the mutant mice exert enhanced adipogenesis and obtain larger adipose mass and body weight gain compared to wild type. However, administration of PT2399 along with HFD feeding sufficiently suppressed HFD-induced body weight and adipose mass increase through suppression of adipogenesis and lipogenesis. The accompanying decreased lipid accumulation in the liver and improved glucose tolerance in wild-type mice were not observed in the mutant mice indicating negative regulation of HIF-2α on obesity and a complex role for the PAS-B domain in metabolic regulation. Notably, short-term administration of PT2399 to obese mice decreased adipose mass and improved metabolic condition. These results indicate a regulatory role for HIF-2α in obesity progression and suggest a therapeutic opportunity for PT2399 in obesity and associated metabolic disorders.

Bcl-2 promotes metastasis through the epithelial-to-mesenchymal transition in the BCap37 medullary breast cancer cell line.

Metastatic breast cancer is one of the major types of cancer in women. However, despite being the focus of considerable research efforts, its molecular mechanism remains to be fully elucidated. The B-cell lymphoma/leukemia gene-2 (Bcl-2) protein is well known for its role in inhibiting programmed cell death/apoptosis. However, little is known concerning its function in cell invasion and migration. In the present study, cell migration and invasion assays revealed that anti-apoptotic Bcl-2 protein induced migration and invasion without affecting cell proliferation in the BCap37 breast cancer cell line. In addition, it was found that the overexpression of Bcl-2 in BCap37 cells increased metastasis to the lung in a mouse model. Using western blotting and RT q-PCR analysis, it was demonstrated that the overexpression of Bcl-2 inhibited the expression of E-cadherin, an epithelial marker, whereas it increased the levels of mesenchymal markers N-cadherin and vimentin. Therefore, the results suggested that Bcl-2 may induce cellular metastasis in breast cancer via the epithelial-to-mesenchymal transition.

Promotional effect of microRNA-194 on breast cancer cells via targeting F-box/WD repeat-containing protein 7.

Breast cancer is the most common type of malignant cancer in females. An increasing number of studies have revealed that microRNAs (miR), which belong to a class of small non-coding RNAs, serve an important role in a number of human cancer subtypes. In the present study, the role of miR-194 in breast cancer cells and its underlying mechanisms were investigated. The results demonstrated that the serum levels of miR-194 were significantly higher in patients of the poorly differentiated and well-differentiated groups, compared with in healthy adults. Additionally, the serum level of miR-194 was significantly higher in the poorly differentiated group compared with in the well-differentiated group. In order to further investigate the role of miR-194 in breast cancer cells, the present study transfected two breast cancer cell lines, MCF-7 and MDA-MB-231, with an empty vector (control), miR-194 (overexpression), antagomiR-194 (inhibitor, functional knock down) or antagomiR-194 and miR-194. An MTT assay was performed in order to detect the proliferation of breast cancer cells in the various groups. The results revealed that the overexpression of miR-194 significantly accelerated cell proliferation, whereas the inhibition of miR-194 significantly decelerated the proliferation of MCF-7 and MDA-MB-231 cells. Furthermore, the expression levels of cyclin D and cyclin E were significantly upregulated in miR-194 overexpressing cells, and the expression levels of cyclin D and cyclin E were significantly downregulated in miR-194 inhibited cells, as compared with in control cells. No significant change was observed in the level of proliferation of cells co-transfected with miR-194 and antagomiR-194, compared with in the control cells. According to the hypothesis suggesting possible target genes of miR-194, the present study proposed that F-box/WD repeat-containing protein 7 (Fbxw-7) may be a direct target of miR-194, which was confirmed by a luciferase reporter assay. The present study suggested that miR-194 expression promoted the proliferation of breast cancer cells by targeting Fbxw-7, and may serve as a biomarker and a novel target for breast cancer therapy.

Effects of HGF gene polymorphisms and protein expression on transhepatic arterial chemotherapeutic embolism efficacy and prognosis in patients with primary liver cancer.

OBJECTIVE: To investigate the correlations of two hepatocyte growth factor (HGF) gene polymorphisms (rs5745652 and rs2074725) and their protein expression levels with the efficacy of transhepatic arterial chemotherapeutic embolism (TACE) and prognosis in patients with primary liver cancer (PLC). METHODS: From March 2011 to June 2012, 109 PLC patients (the case group) who chose TACE as primary treatment and 80 healthy people (the control group) who had undergone physical examination in The First Affiliated Hospital, Zhejiang University were selected during the same period. Gene polymorphisms of HGF rs5745652 and HGF rs2074725 were detected. Serum HGF level, treating efficacy, survival quality, and 3-year survival rate for PLC patients who received TACE were observed. RESULTS: There were significant differences in genotype and allele frequencies of HGF rs5745652 and HGF rs2074725, between the case and control groups (all P<0.05). Compared with CT+TT genotype of HGF rs5745652, patients carrying CC genotype had lower serum HGF levels, higher efficacy, better survival quality, and prolonged 3-year survival rate (all P<0.05). In rs2074725, patients carrying CA+AA genotype had lower serum HGF levels, higher efficacy, better survival quality, and prolonged 3-year survival rate compared with patients carrying rs2074725 CC genotype (all P<0.05). Gene polymorphisms of HGF rs5745652 and HGF rs2074725, tumor size, and Barcelona Clinic Liver Cancer stage were independent prognostic factors for PLC (P<0.05). CONCLUSION: Our results indicated that HGF gene polymorphisms affect TACE efficacy and survival quality of PLC patients. Patients with HGF CC genotype of rs5745652 and CA+AA genotype of rs2074725 had decreased HGF level, better curative effect, high survival quality, and a good prognosis after TACE treatment.

MIR196A2 rs11614913 C > T polymorphism correlates with an increased risk of hepatopulmonary syndrome in liver cirrhosis: a case-control study in China.

OBJECTIVE: This case-control study is designed to explore the relationship between microRNA-196a2 (MIR196A2) rs11614913 C > T polymorphism and the risk of hepatopulmonary syndrome (HPS) in liver cirrhosis. METHODS: From January 2013 to January 2015, 163 liver cirrhosis patients with HPS (case group), 264 liver cirrhosis patients without HPS (control group), and 195 healthy people (normal group) were selected. A DNA extraction kit was used to extract plasma DNA from peripheral blood. Polymerase chain reaction-restriction fragment length polymorphism was used to detect the allele and genotype frequencies of MIR196A2 C > T polymorphism. Real-time quantitative polymerase chain reaction was adopted to detect the relative expression of MIR196A. RESULTS: The frequencies of C allele in the case group were higher than those in the control and normal groups (all P < 0.05), whereas no significant difference was found between the control and normal groups, which indicated that MIR196A2 C > T polymorphism was closely associated with an increased risk of HPS in patients with liver cirrhosis. Compared with the normal group, the relative expression of MIR196A in the case group was significantly increased (P < 0.05), but there was no significant difference in the control group (P > 0.05). In the case group, compared with patients carrying the TT genotype, the relative expression of MIR196A of patients carrying the C allele (CT + CC) evidently increased (P < 0.05). CONCLUSIONS: The MIR196A2 rs11614913 C > T polymorphism may contribute to an increased risk of HPS in liver cirrhosis patients.

Large intergenic non-coding RNA-ROR reverses gemcitabine-induced autophagy and apoptosis in breast cancer cells.

The purpose of this study was to elucidate the potential role of long intergenic non-protein coding RNA, regulator of reprogramming (linc-ROR) in gemcitabine (Gem)-induced autophagy and apoptosis in breast cancer cells. MDA-MB-231 cells were treated with short hairpin RNA (shRNA) to knockdown Linc-ROR expression in the presence of Gem. Gem treatment alone decreased cell survival and increased both apoptosis and autophagy. Gem treatment also increased the expression of LC3-II, Beclin 1, NOTCH1 and Bcl-2, but decreased expression of p62 and p53. Untreated MDA-MB-231 cell lines strongly expressed linc-ROR, but linc-ROR knockdown decreased cell viability and expression of p62 and p53 while increasing apoptosis. Linc-ROR knockdown also increased LC3-II/ß-actin, Beclin 1, NOTCH1, and Bcl-2 expression, as well as the number of autophagic vesicles in MDA-MB-231 cells. Linc-ROR negatively regulated miR-34a expression by inhibiting histone H3 acetylation in the miR-34a promoter. We conclude that linc-ROR suppresses Gem-induced autophagy and apoptosis in breast cancer cells by silencing miR-34a expression.

Breast metastasis from signet ring cell carcinoma of the urinary bladder: A case report.

Carcinoma of the bladder metastatic to the breast is only sporadically reported in the literature. To the best of our knowledge, the present report is the first described case of signet ring cell carcinoma of the urinary bladder metastasizing to the breast. The patient was a 43-year-old woman who underwent transurethral partial cystectomy for signet ring cell carcinoma of the urinary bladder and adjuvant chemotherapy with cisplatin and gemcitabine. At 7 months postcystectomy, the patient presented with a solitary nodule in the right breast. Following transdermic core needle puncture biopsy of the lesion and histological examination, the tumor was found to be composed of signet ring cells, which were similar to the cells in the original cystectomy specimen. The patient underwent mastectomy without further chemotherapy and has remained free from metastasis to other organs during 1 year follow-up.

Expression of FOXO6 is Associated With Oxidative Stress Level and Predicts the Prognosis in Hepatocellular Cancer: A Comparative Study.

The aim of this study was to explore the association of Forkhead box O6 (FOXO6) expression with oxidative stress level and prognosis of hepatocellular cancer (HCC).The case group included tissues of HCC from 128 patients who were hospitalized in Division of Hepatobiliary and Pancreatic Surgery, Department of Surgery of First Affiliated Hospital, School of Medicine, Zhejiang University. The control group included normal liver tissues from 74 patients. RT-PCR and Western blot were used to test expressions of FOXO6, heme oxygenase (HO)-1, glutathione peroxidase (GPx), superoxide dismutase (SOD), and catalase (CAT). Dihydroethidium (DHE) was dyed to observe reactive oxygen species (ROS) level. Immunohistochemistry was used to test FOXO6 expression. FOXO6 was silenced in HepG2 cells to detect cell proliferation and apoptosis. The expressions of ROS, HO-1, GPx, SOD, CAT, p27, and cyclin D1 were also detected to further explore the possible mechanism.The expressions of FOXO6, HO-1, GPx, SOD, and CAT in HCC tissue was significantly higher than those in normal and adjacent HCC tissues (P <0.05). The tumor size, TNM stage, Alpha-fetoprotein (AFP) level, the presence or absence of hepatitis B surface antigen (HbsAg), and differentiation degree were related to FOXO6 expression level (all P <0.05). COX analysis showed that high FOXO6 expression, male, positive HBsAg, advanced TNM staging, high expression of AFP, and low degree of differentiation were all risk factors for prognosis in HCC (P <0.05). Compared with the blank group (C group, without transfection) and the negative control (NC) group, the mRNA expressions of ROS, FOXO6, HO-1, SOD, GPx, and CAT were decreased (P <0.05). si-RNA group had significantly decreased proliferation speed during 24 to 72 hours (P <0.05), whereas si-FOXO6 group had remarkably increased G0/G1 staged cells and decreased S-staged cells (P <0.05). The si-FOXO6 group showed notably increased apoptosis rate (P <0.05) and p27 expressions as well as decreased cyclin D1 expressions (P <0.05).FOXO6 was highly expressed in HCC tissue and was related to oxidative stress levels. Furthermore, FOXO6 expression can be used as a biomarker for deterioration and prognosis of liver cancer, which may provide a novel treatment target for HCC therapy.

Linc-ROR induces epithelial-mesenchymal transition and contributes to drug resistance and invasion of breast cancer cells.

We aimed to investigate the role of large intergenic noncoding RNA regulator of reprogramming (linc-ROR) in the chemotherapy resistance of human breast cancer (BC) cells and its mechanism. A total of 142 patients diagnosed with BC in the First Affiliated Hospital, Zhejiang University between January 2012 and January 2014 were enrolled in our study. The BC tissues and the adjacent normal tissues (5 cm away from tumor tissue) of the enrolled patients were selected, and human BC cell lines (MCF10A, SK-BR-3, MCF-7, Bcap-37, MDA-MB-231, and T47D) were also selected. Quantitative real-time polymerase chain reaction (qRT-PCR), Western blot, 3-(4, 5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt (MTS) assay, and Transwell were applied in our study. Expression level of linc-ROR messenger RNA (mRNA) in BC tissues was clearly higher than that in adjacent normal tissues, and significant difference was found between expression level of linc-ROR mRNA and lymph node metastasis (all P < 0.05). Linc-ROR was highly expressed in others BC cell lines compared with that in immortalized mammary epithelial cells (MECs) MCF10A (both P < 0.05), while MDA-MB231 cell presented the higher expression (P < 0.001). Under different concentrations of 5-FU and paclitaxel in MDA-MB231 cell, E-cadherin mRNA and protein expressions increased gradually with the increase of concentrations, and Vimentin and N-cadherin mRNA and protein expressions decreased gradually with the decrease of concentrations (all P < 0.05). Compared with shCtrl group, MDA-MB231 cell in shROR group presented higher sensibility of 5-FU and paclitaxel with increased E-cadherin expression, decreased Vimentin and N-cadherin expression and invasion ability (all P < 0.05). Compared with vector cell, overexpressed linc-ROR cell presented decreased sensibility of 5-FU and paclitaxel with decreased E-cadherin expression, increased Vimentin, N-cadherin expression, and invasion ability (all P < 0.05). Our study demonstrated that linc-ROR is an important marker for multidrug resistance of BC, and its up-regulation is important for chemotherapy tolerance and invasion of BC.

Breast cancer stem cells phenotype and plasma cell-predominant breast cancer independently indicate poor survival.

PURPOSE: Cancer stem cell-tumor microenvironment ecosystem is proposed to drive tumor heterogeneity. Tumor-infiltrating lymphocytes (TILs) in breast cancer ecosystem were demonstrated to indicate better prognosis and benefit from chemotherapy. This study sought to detect the association between breast cancer stem cells and TILs. METHODS: 92 patients with breast cancer were enrolled. Matched cancerous and paracancerous tissues were assembled in a tissue microarray and immunohistochemistry was employed to test expression of breast cancer stem cells (BCSCs) markers. TILs counts were estimated with global hematoxylin-eosin staining. The association between TILs and BCSCs phenotypes was analysed by multivariate analysis. RESULTS: Although it was unable to find direct significant association between BCSCs phenotypes and TILs, the BCSCs phenotype with CD44(+)CD24(-)ALDH1A1(+)EpCAM(+)CD49f(+) was proved to be associated with worse DFS and OS (P=0.037 and 0.001). This result was confirmed by cox proportional-hazards regression model (for DFS and OS respectively, HR=2.438 and 3.383, P=0.019 [95%CI 1.418-3.457] and 0.025 [95%CI 1.162-9.843]). Additionally, in results of TILs, plasma cell-predominant breast cancer (PPBC) was unexpectedly found to indicate worse OS and HR was 2.686 (P=0.038 [95%CI 1.582-3.789]). CONCLUSIONS: The BCSCs phenotype and PPBC may be helpful stratified factors in future clinical trials. The underlying mechanism needs further research.

RPS23RG1 reduces Aß oligomer-induced synaptic and cognitive deficits.

Alzheimer's disease (AD) is the most common form of dementia in the elderly. It is generally believed that ß-amyloidogenesis, tau-hyperphosphorylation, and synaptic loss underlie cognitive decline in AD. Rps23rg1, a functional retroposed mouse gene, has been shown to reduce Alzheimer's ß-amyloid (Aß) production and tau phosphorylation. In this study, we have identified its human homolog, and demonstrated that RPS23RG1 regulates synaptic plasticity, thus counteracting Aß oligomer (oAß)-induced cognitive deficits in mice. The level of RPS23RG1 mRNA is significantly lower in the brains of AD compared to non-AD patients, suggesting its potential role in the pathogenesis of the disease. Similar to its mouse counterpart, human RPS23RG1 interacts with adenylate cyclase, activating PKA/CREB, and inhibiting GSK-3. Furthermore, we show that human RPS23RG1 promotes synaptic plasticity and offsets oAß-induced synaptic loss in a PKA-dependent manner in cultured primary neurons. Overexpression of Rps23rg1 in transgenic mice consistently prevented oAß-induced PKA inactivation, synaptic deficits, suppression of long-term potentiation, and cognitive impairment as compared to wild type littermates. Our study demonstrates that RPS23RG1 may reduce the occurrence of key elements of AD pathology and enhance synaptic functions to counteract oAß-induced synaptic and cognitive deficits in AD.

Hsp90 chaperone inhibitor 17-AAG attenuates Aß-induced synaptic toxicity and memory impairment.

The excessive accumulation of soluble amyloid peptides (Aß) plays a crucial role in the pathogenesis of Alzheimer's disease (AD), particularly in synaptic dysfunction. The role of the two major chaperone proteins, Hsp70 and Hsp90, in clearing misfolded protein aggregates has been established. Despite their abundant presence in synapses, the role of these chaperones in synapses remains elusive. Here, we report that Hsp90 inhibition by 17-AAG elicited not only a heat shock-like response but also upregulated presynaptic and postsynaptic proteins, such as synapsin I, synaptophysin, and PSD95 in neurons. 17-AAG treatment enhanced high-frequency stimulation-evoked LTP and protected neurons from synaptic damage induced by soluble Aß. In AD transgenic mice, the daily administration of 17-AAG over 7 d resulted in a marked increase in PSD95 expression in hippocampi. 17-AAG treatments in wild-type C57BL/6 mice challenged by soluble Aß significantly improved contextual fear memory. Further, we demonstrate that 17-AAG activated synaptic protein expression via transcriptional mechanisms through the heat shock transcription factor HSF1. Together, our findings identify a novel function of Hsp90 inhibition in regulating synaptic plasticity, in addition to the known neuroprotective effects of the chaperones against Aß and tau toxicity, thus further supporting the potential of Hsp90 inhibitors in treating neurodegenerative diseases.