Development of benchmark datasets for text mining and sentiment analysis to accelerate regulatory literature review.

In the field of regulatory science, reviewing literature is an essential and important step, which most of the time is conducted by manually reading hundreds of articles. Although this process is highly time-consuming and labor-intensive, most output of this process is not well transformed into machine-readable format. The limited availability of data has largely constrained the artificial intelligence (AI) system development to facilitate this literature reviewing in the regulatory process. In the past decade, AI has revolutionized the area of text mining as many deep learning approaches have been developed to search, annotate, and classify relevant documents. After the great advancement of AI algorithms, a lack of high-quality data instead of the algorithms has recently become the bottleneck of AI system development. Herein, we constructed two large benchmark datasets, Chlorine Efficacy dataset (CHE) and Chlorine Safety dataset (CHS), under a regulatory scenario that sought to assess the antiseptic efficacy and toxicity of chlorine. For each dataset, ∼10,000 scientific articles were initially collected, manually reviewed, and their relevance to the review task were labeled. To ensure high data quality, each paper was labeled by a consensus among multiple experienced reviewers. The overall relevance rate was 27.21% (2,663 of 9,788) for CHE and 7.50% (761 of 10,153) for CHS, respectively. Furthermore, the relevant articles were categorized into five subgroups based on the focus of their content. Next, we developed an attention-based classification language model using these two datasets. The proposed classification model yielded 0.857 and 0.908 of Area Under the Curve (AUC) for CHE and CHS dataset, respectively. This performance was significantly better than permutation test (p < 10E-9), demonstrating that the labeling processes were valid. To conclude, our datasets can be used as benchmark to develop AI systems, which can further facilitate the literature review process in regulatory science.

Overexpression of LINE-1 Retrotransposons in Autism Brain.

Long interspersed nuclear elements-1 (LINE-1 or L1) are mobile DNA sequences that are capable of duplication and insertion (retrotransposition) within the genome. Recently, retrotransposition of L1 was shown to occur within human brain leading to somatic mosaicism in hippocampus and cerebellum. Because unregulated L1 activity can promote genomic instability and mutagenesis, multiple mechanisms including epigenetic chromatin condensation have evolved to effectively repress L1 expression. Nonetheless, L1 expression has been shown to be increased in patients with Rett syndrome and schizophrenia. Based on this evidence and our reports of oxidative stress and epigenetic dysregulation in autism cerebellum, we sought to determine whether L1 expression was increased in autism brain. The results indicated that L1 expression was significantly elevated in the autism cerebellum but not in BA9, BA22, or BA24. The binding of repressive MeCP2 and histone H3K9me3 to L1 sequences was significantly lower in autism cerebellum suggesting that relaxation of epigenetic repression may have contributed to increased expression. Further, the increase in L1 expression was inversely correlated with glutathione redox status consistent with reports indicating that L1 expression is increased under pro-oxidant conditions. Finally, the expression of transcription factor FOXO3, sensor of oxidative stress, was significantly increased and positively associated with L1 expression and negatively associated with glutathione redox status. While these novel results are an important first step, future understanding of the contribution of elevated L1 expression to neuronal CNVs and genomic instability in autism will depend on emerging cell-specific genomic technologies, a challenge that warrants future investigation.

Effect of methapyrilene hydrochloride on hepatic intracellular iron metabolism in vivo and in vitro.

The liver, a central detoxification organ and main regulator of systemic iron homeostasis, is prone to damage by xenobiotics. In the present study, we investigated the effect of the hepatotoxicant and hepatocarcinogen methapyrilene hydrochloride on iron metabolism in rat liver in a repeat-dose in vivo toxicity study and in human HepaRG cells in vitro. Treatment of male Fischer 344 (F344) rats with methapyrilene at doses 40 and 80mg/kg body weight (bw)/day by gavage for 6 weeks resulted in changes in the expression of classic hepatotoxicity-related marker genes and iron homeostasis-related genes, especially a prominent, dose-dependent down-regulation of the transferrin (Tf) gene and an up-regulation of the ferritin, light chain (Ftl) gene. A decrease in the level of TF and an increase in the level of FTL also occurred in methapyrilene-treated differentiated HepaRG cells, indicating the existence of interspecies and in vitro-in vivo similarities in the disturbance of cellular iron homeostasis upon liver injury. In contrast, there was minimal overlap in the expression of liver toxicity-marker genes in the livers of rats and in HepaRG cells treated with methapyrilene. Importantly, the decrease of transferrin at mRNA and protein levels occurred after the treatment with a low dose of methapyrilene that exhibited minimal cytotoxicity. These results demonstrate the significance of the dysregulation of hepatic iron metabolism in the pathogenesis and mechanism of chemical-induced liver toxicity and suggest that these changes may be sensitive and useful indicators of potentially hepatotoxic chemicals.

Chemo brain or tumor brain - that is the question: the presence of extracranial tumors profoundly affects molecular processes in the prefrontal cortex of TumorGraft mice.

Cancer chemotherapy causes numerous persistent central nervous system complications. This condition is known as chemo brain. Cognitive impairments occur even before treatment, and hence are referred to as cancer associated cognitive changes, or tumor brain. There is much yet to be learned about the mechanisms of both chemo brain and tumor brain. The frequency and timing of chemo brain and tumor brain occurrence and persistence strongly suggest they may be epigenetic in nature and associated with altered gene expression. Here we used TumorGraftTM models wherein part of a patient's tumor is removed and grafted into immune-deficient mice and conducted global gene expression and DNA methylation analysis. We show that malignant non-central nervous system tumor growth causes profound molecular alterations in the brain. Mice harbouring triple negative or progesterone positive breast cancer TumorGrafts exhibited altered gene expression, decreased levels of DNA methylation, increased levels of DNA hydroxymethylation, and oxidative stress in the prefrontal cortex. Interestingly, chemotherapy did not have any additional synergistic effects on the analyzed processes. The molecular changes observed in this study are known signs of neurodegeneration and brain aging. This study provides an important roadmap for future large-scale analysis of the molecular and cellular mechanisms of tumor brain.

Editor's Highlight: Organ-Specific Epigenetic Changes Induced by the Nongenotoxic Liver Carcinogen Methapyrilene in Fischer 344 Rats.

Continuous lifetime exposure to certain natural and man-made chemicals is a major cause of cancers in humans; therefore, evaluating the carcinogenic risks of chemicals remains important. Currently, substantial progress has been made in identification of genotoxic carcinogens; in contrast, predicting the carcinogenic potential of nongenotoxic compounds is a challenge due to many different modes of action that may lead to tumorigenesis. In the present study, we investigated the effects of the nongenotoxic liver carcinogen methapyrilene and the nongenotoxic noncarcinogen usnic acid, at doses that do not exhibit organ cytotoxicity, on epigenomic alterations in the livers and kidneys of Fischer 344 (F344) rats. We demonstrate that a repeat-dose oral treatment of male F344 rats with methapyrilene for 6 weeks caused target organ-specific epigenetic alterations in the livers. In contrast, only very slight epigenetic changes were found in the livers of F344 rats treated with hepatotoxicant, but noncarcinogen, usnic acid. The methapyrilene-induced epigenetic changes consisted of changes in histone lysine acetylation and methylation, with the greatest increase occurring in global and gene-specific histone H3 lysine 9 (H3K9) deacetylation. Importantly, the results of the present study show an association between gene-specific histone H3K9 deacetylation and a reduced expression of critical cancer-related genes, including prospero homeobox 1 (Prox1), HNF1 homebox A (Hnf1a), and peroxisome proliferator activated receptor alpha (Ppara), which provides a mechanistic link between methapyrilene-induced epigenetic aberrations and liver carcinogenesis.

Sex-specific effects of cytotoxic chemotherapy agents cyclophosphamide and mitomycin C on gene expression, oxidative DNA damage, and epigenetic alterations in the prefrontal cortex and hippocampus - an aging connection.

Recent research shows that chemotherapy agents can be more toxic to healthy brain cells than to the target cancer cells. They cause a range of side effects, including memory loss and cognitive dysfunction that can persist long after the completion of treatment. This condition is known as chemo brain. The molecular and cellular mechanisms of chemo brain remain obscure. Here, we analyzed the effects of two cytotoxic chemotherapy drugs-cyclophosphamide (CPP) and mitomycin C (MMC) - on transcriptomic and epigenetic changes in the murine prefrontal cortex (PFC) and hippocampal regions. We for the first time showed that CPP and MMC treatments led to profound sex- and brain region-specific alterations in gene expression profiles. Gene expression changes were most prominent in the PFC tissues of female mice 3 weeks after MMC treatment, and the gene expression response was much greater for MCC than CPP exposure. MMC exposure resulted in oxidative DNA damage, evidenced by accumulation of 8-oxo-2'-deoxyguanosine (8-oxodG) and a decrease in the level of 8-oxodG repair protein OGG1 in the PFC of female animals 3 weeks after treatment. MMC treatment decreased global DNA methylation and increased DNA hydroxymethylation in the PFC tissues of female mice. The majority of the changes induced by chemotherapy in the PFC tissues of female mice resembled those that occur during the brain's aging processes. Therefore, our study suggests a link between chemotherapy-induced chemo brain and brain aging, and provides an important roadmap for future analysis.

Suppressing activity of tributyrin on hepatocarcinogenesis is associated with inhibiting the p53-CRM1 interaction and changing the cellular compartmentalization of p53 protein.

Hepatocellular carcinoma (HCC), an aggressive and the fastest growing life-threatening cancer worldwide, is often diagnosed at intermediate or advanced stages of the disease, which substantially limits therapeutic approaches for its successful treatment. This indicates that the prevention of hepatocarcinogenesis is probably the most promising approach to reduce both the HCC incidence and cancer-related mortality. In previous studies, we demonstrated a potent chemopreventive effect of tributyrin, a butyric acid prodrug, on experimental hepatocarcinogenesis. The cancer-inhibitory effect of tributyrin was linked to the suppression of sustained cell proliferation and induction of apoptotic cell death driven by an activation of the p53 apoptotic signaling pathway. The goal of the present study was to investigate the underlying molecular mechanisms linked to tributyrin-mediated p53 activation. Using in vivo and in vitro models of liver cancer, we demonstrate that an increase in the level of p53 protein in nuclei, a decrease in the level of cytoplasmic p53, and, consequently, an increase in the ratio of nuclear/cytoplasmic p53 in rat preneoplastic livers and in rat and human HCC cell lines caused by tributyrin or sodium butyrate treatments was associated with a marked increase in the level of nuclear chromosome region maintenance 1 (CRM1) protein. Mechanistically, the increase in the level of nuclear p53 protein was associated with a substantially reduced binding interaction between CRM1 and p53. The results demonstrate that the cancer-inhibitory activity of sodium butyrate and its derivatives on liver carcinogenesis may be attributed to retention of p53 and CRM1 proteins in the nucleus, an event that may trigger activation of p53-mediated apoptotic cell death in neoplastic cells.

MicroRNA-152-mediated dysregulation of hepatic transferrin receptor 1 in liver carcinogenesis.

Over-expression of transferrin receptor 1 (TFRC) is observed in hepatocellular carcinoma (HCC); however, there is a lack of conclusive information regarding the mechanisms of this dysregulation. In the present study, we demonstrated a significant increase in the levels of TFRC mRNA and protein in preneoplastic livers from relevant experimental models of human hepatocarcinogenesis and in human HCC cells. Additionally, using the TCGA database, we demonstrated an over-expression of TFRC in human HCC tissue samples and a markedly decreased level of microRNA-152 (miR-152) when compared to non-tumor liver tissue. The results indicated that the increase in levels of TFRC in human HCC cells and human HCC tissue samples may be attributed, in part, to a post-transcriptional mechanism mediated by a down-regulation of miR-152. This was evidenced by a strong inverse correlation between the level of TFRC and the expression of miR-152 in human HCC cells (r = -0.99, p = 4. 7 × 10-9), and was confirmed by in vitro experiments showing that transfection of human HCC cell lines with miR-152 effectively suppressed TFRC expression. This suggests that miR-152-specific targeting of TFRC may provide a selective anticancer therapeutic approach for the treatment of HCC.

Differentially expressed MicroRNAs provide mechanistic insight into fibrosis-associated liver carcinogenesis in mice.

Hepatocellular carcinoma (HCC) is one of the most prevalent human cancers, with a rising incidence worldwide. The molecular mechanisms associated with the development of HCC are complex and include multiple interconnected molecular alterations with mounting evidence indicating an important role of microRNAs (miRNAs) in the pathogenesis of HCC. In humans, the development of HCC is commonly associated with liver cirrhosis. To study fibrosis-associated liver carcinogenesis, we used a mouse model designed to emulate the development of HCC in cirrhotic liver. Specifically, we were interested in evaluating the role of miRNAs in the molecular pathogenesis of liver carcinogenesis in male B6C3F1/J mice treated with N-nitrosodiethylamine (DEN) or carbon tetrachloride (CCl4 ) alone or a combination of DEN and CCl4 and characterized by a differential tumor incidence that increased in the following order: DEN

Cerebellar oxidative DNA damage and altered DNA methylation in the BTBR T+tf/J mouse model of autism and similarities with human post mortem cerebellum.

The molecular pathogenesis of autism is complex and involves numerous genomic, epigenomic, proteomic, metabolic, and physiological alterations. Elucidating and understanding the molecular processes underlying the pathogenesis of autism is critical for effective clinical management and prevention of this disorder. The goal of this study is to investigate key molecular alterations postulated to play a role in autism and their role in the pathophysiology of autism. In this study we demonstrate that DNA isolated from the cerebellum of BTBR T+tf/J mice, a relevant mouse model of autism, and from human post-mortem cerebellum of individuals with autism, are both characterized by an increased levels of 8-oxo-7-hydrodeoxyguanosine (8-oxodG), 5-methylcytosine (5mC), and 5-hydroxymethylcytosine (5hmC). The increase in 8-oxodG and 5mC content was associated with a markedly reduced expression of the 8-oxoguanine DNA-glycosylase 1 (Ogg1) and increased expression of de novo DNA methyltransferases 3a and 3b (Dnmt3a and Dnmt3b). Interestingly, a rise in the level of 5hmC occurred without changes in the expression of ten-eleven translocation expression 1 (Tet1) and Tet2 genes, but significantly correlated with the presence of 8-oxodG in DNA. This finding and similar elevation in 8-oxodG in cerebellum of individuals with autism and in the BTBR T+tf/J mouse model warrant future large-scale studies to specifically address the role of OGG1 alterations in pathogenesis of autism.

Interstrain differences in the progression of nonalcoholic steatohepatitis to fibrosis in mice are associated with altered hepatic iron metabolism.

Nonalcoholic fatty liver disease (NAFLD) is a major health problem worldwide. Currently, there is a lack of conclusive information to clarify the molecular events and mechanisms responsible for the progression of NAFLD to fibrosis and cirrhosis and, more importantly, for differences in interindividual disease severity. The aim of this study was to investigate a role of interindividual differences in iron metabolism among inbred mouse strains in the pathogenesis and severity of fibrosis in a model of NAFLD. Feeding male A/J, 129S1/SvImJ and WSB/EiJ mice a choline- and folate-deficient diet caused NAFLD-associated liver injury and iron metabolism abnormalities, especially in WSB/EiJ mice. NAFLD-associated fibrogenesis was correlated with a marked strain- and injury-dependent increase in the expression of iron metabolism genes, especially transferrin receptor (Tfrc), ferritin heavy chain (Fth1), and solute carrier family 40 (iron-regulated transporter), member 1 (Slc40a1, Fpn1) and their related proteins, and pronounced down-regulation of the iron regulatory protein 1 (IRP1), with the magnitude being A/J<129S1/SvImJ

Iron metabolism disturbances in the MCF-7 human breast cancer cells with acquired resistance to doxorubicin and cisplatin.

The development of resistance of cancer cells to therapeutic agents is the major obstacle in the successful treatment of breast cancer and the main cause of breast cancer recurrence. The results of several studies have demonstrated an important role of altered cellular iron metabolism in the progression of breast cancer and suggested that iron metabolism may be involved in the acquisition of a cancer cell drug-resistant phenotype. In the present study, we show that human MCF-7 breast cancer cells with an acquired resistance to the chemotherapeutic drugs doxorubicin (MCF-7/DOX) and cisplatin (MCF-7/CDDP) exhibited substantial alterations in the intracellular iron content and levels of iron-regulatory proteins involved in the cellular uptake, storage and export of iron, especially in profoundly increased levels of ferritin light chain (FTL) protein. The increased levels of FTL in breast cancer indicate that FTL may be used as a diagnostic and prognostic marker for breast cancer. Additionally, we demonstrate that targeted downregulation of FTL protein by the microRNA miR-133a increases sensitivity of MCF-7/DOX and MCF-7/CDDP cells to doxorubicin and cisplatin. These results suggest that correction of iron metabolism abnormalities may substantially improve the efficiency of breast cancer treatment.

Modulation of intracellular iron metabolism by iron chelation affects chromatin remodeling proteins and corresponding epigenetic modifications in breast cancer cells and increases their sensitivity to chemotherapeutic agents.

Iron plays a vital role in the normal functioning of cells via the regulation of essential cellular metabolic reactions, including several DNA and histone-modifying proteins. The metabolic status of iron and the regulation of epigenetic mechanisms are well-balanced and tightly controlled in normal cells; however, in cancer cells these processes are profoundly disturbed. Cancer-related abnormalities in iron metabolism have been corrected through the use of iron-chelating agents, which cause an inhibition of DNA synthesis, G1-S phase arrest, an inhibition of epithelial-to-mesenchymal transition, and the activation of apoptosis. In the present study, we show that, in addition to these well-studied molecular mechanisms, the treatment of wild-type TP53 MCF-7 and mutant TP53 MDA-MB-231 human breast cancer cells with desferrioxamine (DFO), a model iron chelator, causes significant epigenetic alterations at the global and gene-specific levels. Specifically, DFO treatment decreased the protein levels of the histone H3 lysine 9 demethylase, Jumonji domain-containing protein 2A (JMJD2A), in the MCF-7 and MDA-MB-231 cells and down-regulated the levels of the histone H3 lysine 4 demethylase, lysine-specific demethylase 1 (LSD1), in the MDA-MB-231 cells. These changes were accompanied by alterations in corresponding metabolically sensitive histone marks. Additionally, we demonstrate that DFO treatment activates apoptotic programs in MCF-7 and MDA-MB-231 cancer cells and enhances their sensitivity to the chemotherapeutic agents, doxorubicin and cisplatin; however, the mechanisms underlying this activation differ. The induction of apoptosis in wild-type TP53 MCF-7 cells was p53-dependent, triggered mainly by the down-regulation of the JMJD2A histone demethylase, while in mutant TP53 MDA-MB-231 cells, the activation of the p53-independent apoptotic program was driven predominantly by the epigenetic up-regulation of p21.

Complex epigenetic regulation of engrailed-2 (EN-2) homeobox gene in the autism cerebellum.

The elucidation of epigenetic alterations in the autism brain has potential to provide new insights into the molecular mechanisms underlying abnormal gene expression in this disorder. Given strong evidence that engrailed-2 (EN-2) is a developmentally expressed gene relevant to cerebellar abnormalities and autism, the epigenetic evaluation of this candidate gene was undertaken in 26 case and control post-mortem cerebellar samples. Assessments included global DNA methylation, EN-2 promoter methylation, EN-2 gene expression and EN-2 protein levels. Chromatin immunoprecipitation was used to evaluate trimethylation status of histone H3 lysine 27 (H3K27) associated with gene downregulation and histone H3 lysine 4 (H3K4) associated with gene activation. The results revealed an unusual pattern of global and EN-2 promoter region DNA hypermethylation accompanied by significant increases in EN-2 gene expression and protein levels. Consistent with EN-2 overexpression, histone H3K27 trimethylation mark in the EN-2 promoter was significantly decreased in the autism samples relative to matched controls. Supporting a link between reduced histone H3K27 trimethylation and increased EN-2 gene expression, the mean level of histone H3K4 trimethylation was elevated in the autism cerebellar samples. Together, these results suggest that the normal EN-2 downregulation that signals Purkinje cell maturation during late prenatal and early-postnatal development may not have occurred in some individuals with autism and that the postnatal persistence of EN-2 overexpression may contribute to autism cerebellar abnormalities.

Chronic administration of 2-acetylaminofluorene alters the cellular iron metabolism in rat liver.

Dysregulated intracellular iron homeostasis has been found not only in rodent and human hepatocellular carcinomas but also in several preneoplastic pathological states associated with hepatocarcinogenesis; however, the precise underlying mechanisms of metabolic iron disturbances in preneoplastic liver and the role of these disturbances remain unexplored. In the present study, using an in vivo model of rat hepatocarcinogenesis induced by 2-acetylaminofluorene, we found extensive alterations in cellular iron metabolism at preneoplastic stages of liver carcinogenesis. These were characterized by a substantial decrease in the levels of cytoplasmic non-heme iron in foci of initiated hepatocytes and altered expression of the major genes responsible for the proper maintenance of intracellular iron homeostasis. Gene expression analysis revealed that the decreased intracellular levels of iron in preneoplastic foci might be attributed to increased iron export from the cells, driven by upregulation of ferroportin (Fpn1), the only known non-heme iron exporter. Likewise, increased Fpn1 gene expression was found in vitro in TRL1215 rat liver cells with an acquired malignant phenotype, suggesting that upregulation of Fpn1 might be a specific feature of neoplastically transformed cells. Other changes observed in vivo included the downregulation of hepcidin (Hamp) gene, a key regulator of Fpn1, and this was accompanied by decreased levels of CCAAT/enhancer binding proteins alpha and beta, especially at the Hamp promoter. In conclusion, our results demonstrate the significance of altered intracellular iron metabolism in the progression of liver carcinogenesis and suggest that correction of these alterations could possibly affect liver cancer development.

Role of ferritin alterations in human breast cancer cells.

Breast cancer is the most common malignancy in women. Successful treatment of breast cancer relies on a better understanding of the molecular mechanisms involved in breast cancer initiation and progression. Recent studies have suggested a crucial role of perturbations in ferritin levels and tightly associated with this, the deregulation of intracellular iron homeostasis; however, the underlying molecular mechanisms for the cancer-linked ferritin alterations remain largely unknown and often with conflicting conclusions. Therefore, this study was undertaken to define the role of ferritin in breast cancer. We determined that human breast cancer cells with an epithelial phenotype, such as MCF-7, MDA-MB-361, T-47D, HCC70 and cells, expressed low levels of ferritin light chain, ferritin heavy chain, transferrin, transferring receptor, and iron-regulatory proteins 1 and 2. In contrast, expression of these proteins was substantially elevated in breast cancer cells with an aggressive mesenchymal phenotype, such as Hs-578T, BT-549, and especially MDA-MB-231 cells. The up-regulation of ferritin light chain and ferritin heavy chain in MDA-MB-231 cells was accompanied by alterations in the subcellular distribution of these proteins as characterized by an increased level of nuclear ferritin and a lower level of the cellular labile iron pool as compared to MCF-7 cells. We established that ferritin heavy chain is a target of miRNA miR-200b, suggesting that its up-regulation in MDA-MB-231 cells may be triggered by the low expression of miR-200b. Ectopic up-regulation of miR-200b by transfection of MDA-MB-231 cells with miR-200b substantially decreased the level of ferritin heavy chain. More importantly, miR-200b-induced down-regulation of ferritin was associated with an increased sensitivity of the MDA-MB-231 cells to the chemotherapeutic agent doxorubicin. These results suggest that perturbations in ferritin levels are associated with the progression of breast cancer toward a more advanced malignant phenotype.

Comparative analysis of promoter methylation and gene expression endpoints between tumorous and non-tumorous tissues from HCV-positive patients with hepatocellular carcinoma.

Transcriptional silencing of tumor suppressor genes and other cancer-related genes induced by promoter CpG island hypermethylation is an important epigenetic mechanism of hepatocarcinogenesis. Previous studies have established methylation profiles of hepatocellular carcinomas (HCCs) and demonstrated that methylation of several candidate genes in resected tissues may be associated with time to recurrence. The goals of our study were to test whether specific promoter methylation and mRNA levels of candidate genes, as well as global changes in DNA methylation, can be linked with time to recurrence and clinicopathological variables in a homogenous study group of HCC patients. Forty-three tumorous and 45 non-tumorous liver tissue samples from the surgical margin were obtained from HCV-positive, HBV-negative HCC patients who underwent tumor resection surgery and who were monitored for tumor recurrence thereafter (median follow-up time: 16 months (range, 0-79 months)). Methylation-specific PCR was used to assess the promoter methylation status of P16(INK4a), SOCS-1, RASSF1A, APC, GSTP1, RIZ1, and MGMT genes, while the level of LINE-1 methylation was used as marker of global DNA methylation levels. Methylation frequencies in P16(INK4a), RASSF1A, APC, GSTP1, and RIZ1 genes were significantly greater in tumorous versus non-tumorous tissues. Methylation of RIZ1 in non-tumorous tissues was significantly associated with time to recurrence. Additionally, genomic DNA was significantly more hypomethylated in tumorous tissues, and this change was associated with shorter recurrence, but not with clinicopathological features. In conclusion, this study supports the role of aberrant methylation in the pathobiology of HCV-positive HCCs. The finding that RIZ1 methylation and increased levels of LINE-1 hypomethylation in non-tumorous tissues are associated with time to recurrence underscores the importance of assessing the epigenetic state of the liver remnant.

Alterations of microRNAs and their targets are associated with acquired resistance of MCF-7 breast cancer cells to cisplatin.

Cancer cells that develop resistance to chemotherapeutic agents are a major clinical obstacle in the successful treatment of breast cancer. Acquired cancer chemoresistance is a multifactorial phenomenon, involving various mechanisms and processes. Recent studies suggest that chemoresistance may be linked to drug-induced dysregulation of microRNA function. Furthermore, mounting evidence indicates the existence of similarities between drug-resistant and metastatic cancer cells in terms of resistance to apoptosis and enhanced invasiveness. We studied the role of miRNA alterations in the acquisition of cisplatin-resistant phenotype in MCF-7 human breast adenocarcinoma cells. We identified a total of 103 miRNAs that were overexpressed or underexpressed (46 upregulated and 57 downregulated) in MCF-7 cells resistant to cisplatin. These differentially expressed miRNAs are involved in the control of cell signaling, cell survival, DNA methylation and invasiveness. The most significantly dysregulated miRNAs were miR-146a, miR-10a, miR-221/222, miR-345, miR-200b and miR-200c. Furthermore, we demonstrated that miR-345 and miR-7 target the human multidrug resistance-associated protein 1. These results suggest that dysregulated miRNA expression may underlie the abnormal functioning of critical cellular processes associated with the cisplatin-resistant phenotype.

Role of DNA damage and alterations in cytosine DNA methylation in rat liver carcinogenesis induced by a methyl-deficient diet.

Currently, cancer is recognized as a disease provoked by both genetic and epigenetic events. However, the significance of early genetic and epigenetic alterations with respect to carcinogenic process in general and to liver carcinogenesis in particular remains unexplored. A lack of knowledge regarding how specific alterations during early preneoplasia may be mechanistically related to tumor formation creates a major gap in understanding the role of these genetic and epigenetic abnormalities in carcinogenesis. In the present study we investigated the contribution of DNA damage and epigenetic alterations to liver carcinogenesis induced by a methyl-deficient diet. Feeding Fisher 344 rats a methyl-deficient diet for 9 weeks resulted in DNA damage and aberrant DNA methylation. This was evidenced by an early up-regulation of the base excision DNA repair genes, accumulation of 8-oxodeoxyguanosine and 3'OH-end strand breaks in DNA, pronounced global loss of DNA methylation, and hypermethylation of CpG islands in the livers of methyl-deficient rats. These abnormalities were completely restored in the livers of rats exposed to methyl-deficiency for 9 weeks after removal of the methyl-deficient diet and re-feeding a methyl-sufficient diet. However, when rats were fed a methyl-deficient diet for 18 week and then given a methyl-sufficient diet, only DNA lesions were repaired. The methyl-sufficient diet failed to restore completely the altered DNA methylation status and prevent the progression of liver carcinogenesis. These results suggest that stable alterations in DNA methylation are a factor that promotes the progression of liver carcinogenesis. Additionally, the results indicate that epigenetic changes may be more reliable markers than DNA lesions of the carcinogenic process and carcinogen exposure.