Hepatocellular carcinoma (HCC) is the third leading cause of cancer mortality worldwide. Current treatments are subpar, with late stage diagnosis and poor prognosis contributing to limited treatment options. The evolutionarily conserved, ubiquitously expressed transcription factor LSF is overexpressed in HCC, and its expression is positively correlated with disease severity. Certain small molecules, known as Factor Quinolinone Inhibitors (FQIs), specifically inhibit LSF DNA-binding activity, inhibit HCC cell proliferation in vitro and prevent tumor growth in an endogenous mouse liver cancer model without apparent toxicity. The targeting of transcription factors by small molecule inhibitors has been historically difficult (Dunker and Uversky, 2010), warranting further molecular investigation into the requirement for LSF in HCC to confirm that the anti-tumor effects of FQIs are the consequence of LSF inhibition. This body of work investigates a dual approach for inhibiting LSF function in order to determine the molecular consequences for HCC cells. To identify the specific point of the cell cycle where LSF is required for HCC proliferation, synchronous HCC cells were treated with FQI or with short interfering RNA to reduce levels of LSF. The results indicate that LSF is required for proper mitotic progression in HCC cells. Specifically, these data show a reduction of key mitotic regulators Aurora Kinase B and Cdc20, at the level of mRNA and protein expression. Time-lapse microscopy also demonstrated an increase in the time for progression through mitosis, with a prometaphase/metaphase delay. Immunofluorescence analysis revealed a prometaphase delay plus aberrant cell division and generation of multi-nucleated cells. These findings were consistent with both FQI1 treatment and RNA interference. Additionally, shorter incubation with FQI1 surprisingly revealed a distinct, non-transcriptional regulation of mitosis in HCC cells, suggesting that mitotic regulation by LSF is multi-faceted. As a targeted therapy for use in the clinic, the in vivo toxicity of FQIs is critical to investigate. Whole blood provides populations of rapidly dividing normal cells that can test susceptibility to anti-mitotic compounds. When mice were treated with FQI1, the blood analysis showed no toxicity. Taken together, these findings indicate that LSF is a mitotic regulator in HCC, further supporting the therapeutic promise of molecular therapies targeting LSF. / 2019-03-04T00:00:00Z

SND1 MEDIATED DOWNREGULATION OF PTPN23 IN HEPATOCELLULAR CARCINOMA By Nidhi Jariwala, MS A thesis submitted in partial fulfillment of the requirements for the degree of Master of Science at Virginia Commonwealth University, 2014. ADVISOR: Dr. Devanand Sarkar Associate Professor, Department of Human and Molecular Genetics Blick Scholar Associate Scientific Director, Cancer Therapeutics VCU Institute of Molecular Medicine Massey Cancer Center ABSTRACT Staphyloccocal nuclease domain containing protein 1 (SND1) is identified as an oncogene in multiple cancers, including hepatocellular carcinoma (HCC). SND1 regulates gene expression at transcriptional as well as post-transcriptional level and mediates molecular pathways that culminate into carcinogenesis. SND1 is a component of RNA-induced silencing complex (RISC) and functions as a nuclease for RNAi-mediated mRNA degradation. On the other hand SND1 also binds to specific mRNAs, increasing their stability and hence expression. The aim of the present study is to identify mRNAs to which SND1 binds and modulates them either by degradation or increasing stability which might facilitate promotion of HCC by SND1. We performed RNA immunoprecipitation followed by RNA sequencing (RIP-Seq) using anti-SND1 antibody and human HCC cell line QGY-7703. More than 350 mRNAs were identified to be interacting with SND1, of which Protein tyrosine phosphatase non-receptor 23 (PTPN23) was of particular interest, since PTPN23 has been identified to be a tumor suppressor and its role in HCC has not been studied. We document that SND1 can bind to PTPN23 mRNA and induce its degradation. There is an inverse correlation between SND1 and PTPN23 levels in human HCC cell lines and PTPN23 level is downregulated in HCC. Our study thus identifies a novel mechanism by which SND1 promotes hepatocarcinogenesis and identifies PTPN23 as a potential tumor suppressor in HCC. Further studies need to be performed to explore the relationship of these two molecules in in vivo models and to develop PTPN23 overexpression as a potential therapeutic approach for HCC.

Background: Hepatocellular carcinoma (HCC) is the leading cause of cancer in Taiwan, which consumes high medical expense among the total healthcare expenditure and becomes the significant burden to the finance of National Health Insurance (NHI). However, population-based statistics about the cost with the survival rate is rare. Objective: Through analyzing the data from the Bureau of National Health Insurance and hospital clinical files, we estimated the medical expenditure for treating HCC patients and the factors associated with treatment outcome. Methods: The National Health Insurance data from 1996 to 2002 with ICD-9 code 155 were used, which include age, gender, visiting time, and medical cost for inpatient or outpatient visits. The population-based data were analyzed to estimate 1-5 year survival probability and medical cost of HCC patients. In hospital-based data, 189 patients from 1991 to 2004 were recorded by clinical chart and 62 of them diagnosed during 2002 and 2004 were further selected to match the claim data. These informations were used for computing survival or recurrence probability, related factors, and medical cost. Results: For all incident cases in 1997, the average 5-year cumulated cost was NT$ 219,398, while the cumulated cost for those patients survived more than five years was NT$ 491,288. The survival probability was 30.8% for more than one year, 20.9% for more than two years, 15.2% for more than three years, 11.3% for more than four years, and 9.7% for more than five years in 1997 respectively. Female and those age >45-65 years old seemed to have better survival outcomes than male and those age ¡Ø45 years old or >65 years old , the averaged medical cost per treated case surviving more than five years were T$2,457,214 for male and NT$ 1,987,874 for female. Hospital clinical data indicated that TNM stages, therapy choice, liver cirrhosis, and ascites are risk factors for surviving. Although pre-TACE treatment has higher expenditure, its five-year survival probability is better than other treatments in this research. Conclusions: This paper presents medical cost with survival probability and its associated factors for HCC patients and further estimates the medical cost per life saved for treating HCC patients. Our findings can offer the policy-maker, provider, and patient to evaluate the intervention or prevention program in the future.

By Western blotting and immunohistochemical analyses, we have demonstrated that Pin1 was overexpressed in 71.4% of hepatocellular carcinoma (HCC) and its levels correlated with the clinical survival rate. This conclusion was supported by the results from examining Pin1 protein in HCC cancer cell lines. RT-PCR was performed to examine the Pin1 transcription level in tumor part and was compared with that in non-tumor part. Our results indicated that pin1 overexpression was due to the upregulation of Pin1 transcription. Interestingly, most of the cases with upregulation of Pin1 have been shown to correlate with £]-catenin and Cyclin D1 accumulation in HCC specimens. These results were consistent with the previous studies that Pin1 caused £]-catenin and Cyclin D1 elevation in breast cancer. The concordance between hepatitis virus chronic infection and Pin1 overexpression of HCC patients was also analysis. Taken together, these data indicated that Pin1 overexpression leading to £]-catenin and Cyclin D1 accumulation might play a critical role in hepatocellular carcinogenesis and tumor progression. Pin1 levels therefore can be used as a prognostic marker for HCC, and our results suggested that Pin1 is a potential target for therapeutic intervention in hepatocellular carcinoma.

The objective was to study apoptotic effects and underlying molecular mechanisms of goniothalamin (GTN) in hepatocellular carcinoma (HCC)-derived cells. The GTN that isolated from Goniothalamus amuyon, was found to possess profound cytotoxic activities against human SK-Hep1 and Hep-3B cells in vitro. The cytotoxicity of GTN/cell viability was measured by MTT assay at 570-nm absorbance and the IC50 at 72 h were determined as 7.5, 17 mM in SK-Hep1 and Hep-3B respectively. The GTN induced cell death and accumulation of reactive oxygen species in HCC-derived cells. One reactive oxygen species inhibitor, N-actylcysteine, further restored cell viabilities post-GTN treatments. Formation of £^-H2AX foci suggested that GTN-induced DNA damages were double-strand breaks. The GTN arrested cell cycle at G0/G1 by regulation of CCND, CCNE1, RB1 and E2F1 proteins in SK-Hep1, and at G2/M by regulation of CCNB1 in Hep-3B cells. The GTN-induced apoptosis in HCC-derived cells were evidenced by phosphatidylserine externalization and involved both extrinsic and intrinsic pathways in SK-Hep1 cells, but only extrinsic pathway in Hep-3B cells. In SK-Hep1 cells, GTN induced apoptosis and cell cycle arrest through TP53-mediated pathway in contrast to that of TP53/FAS mutated Hep-3B cells. Importantly, GTN was able to induce apoptosis in both TP53 wild type and TP53/FAS mutated HCC-derived cells. On the other hand, GTN was able to induce TP53 and p21WAF1/Cip1 up-regulation in SK-Hep1 and Hep-3B cells and p27Kip1 up-regulation in Hep-3B cells. These results demonstrated that GTN induced apoptosis in HCC cells through distinct signaling pathways.

Information and communication technologies (ICTs) for chronic care are increasingly being researched in Human-Computer Interaction. One of the current health management areas where ICTs have been employed is in supporting communications between patients and physicians. This is particularly relevant for patients suffering from chronic diseases since there is evidence that better communication leads to better health outcomes. Researchers are investigating different chronic diseases to design and test technology interventions to promote better chronic disease care. However, few have investigated pediatric asthma as a case study for designing communication technologies. The World Health Organization (WHO) estimated in 2008 that 300 million people suffer from asthma, and that asthma is the most common chronic illness among children. Asthma interferes with breathing by preventing airflow into the lungs. It is difficult to determine the actual cause of asthma and to predict who will have asthma. These unique challenges provide opportunities to investigate pediatric asthma management. To address these challenges, I have conducted a series of studies with pediatric asthma patients, families, and healthcare providers to better understand their needs, challenges and strategies regarding the use of technologies. I have conducted interviews, a focus group, and a technology probe study to create and refine initial technology designs for children with asthma and their caregivers. Based on the Health Belief Model (HBM), patient-provider communications, and my findings in the prior studies, I designed a mobile and web service to increase asthma knowledge and symptom/management awareness in the child for better health outcomes, and to affect the perceived quality of interaction with healthcare providers. I show the results of my two field deployment studies with total 65 patients to learn how the system affected their practices and health outcomes. My contributions come from an increased understanding in three areas: physician-patient communication via a mobile and web services; ubiquitous communication technology designs to improve current pediatric asthma practices; and controlled evaluation of a ubiquitous communication technology, SMS, in the field.

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Agradecemos a compreensão. on 2016-09-02T19:13:42Z (GMT) / Submitted by MATHEUS ALVAREZ null (matheus@ibb.unesp.br) on 2016-09-08T12:51:41Z No. of bitstreams: 1 Matheus Tese Final - Apend e FC.pdf: 4914163 bytes, checksum: b130a3757b044bcbf1520f2e0437ba1a (MD5) / Approved for entry into archive by Juliano Benedito Ferreira (julianoferreira@reitoria.unesp.br) on 2016-09-08T16:26:08Z (GMT) No. of bitstreams: 1 alvarez_m_dr_bot.pdf: 4914163 bytes, checksum: b130a3757b044bcbf1520f2e0437ba1a (MD5) / Made available in DSpace on 2016-09-08T16:26:08Z (GMT). No. of bitstreams: 1 alvarez_m_dr_bot.pdf: 4914163 bytes, checksum: b130a3757b044bcbf1520f2e0437ba1a (MD5) Previous issue date: 2016-08-01 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / O carcinoma hepatocelular (CHC) é um tumor primário do fígado e envolve diferentes modalidades de tratamento de acordo com seu estadiamento. Depois de terapias locais, como a quiomioembolização transarterial (do inglês, Transarterial Chemoembolization), a avaliação do tumor se baseia nos critérios mRECIST (do inglês, Modified Response Evaluation Criteria In Solid Tumors), que envolvem a medição do diâmetro máximo da lesão viável. Esta pesquisa teve como objetivo o desenvolvimento de um algoritmo para a quantificação objetiva da eficiência da quimioembolização na redução de volume e diâmetro máximo do CHC. Foram utilizados 64 exames retrospectivos de Tomografia Computadorizada (TC) de 32 pacientes. Nesta pesquisa foram comparadas, em pacientes que receberam tratamento por quimioembolização, as medidas do diâmetro maior do CHC viável, estimadas por especialista da área de radiologia e pelo algoritmo desenvolvido neste estudo. O Volume Tumoral Total (VTT) dos CHC antes e após o processo de quimioembolização também foram estimados utilizando um algoritmo, desenvolvido com base em transformadas discretas de wavelet (DWT). Um fantoma virtual simulando as características do fígado normal e contrastado foi desenvolvido para o planejamento, desenvolvimento e validação do algoritmo de detecção. A comparação do diâmetro máximo do tumor, estimado por um especialista e pelo algoritmo revelou diferenças na ordem de 0,25 cm para os tumores de grandes dimensões (diâmetro > 5 centímetros), enquanto uma diferença menor que 0,1 cm foi encontrada para os tumores de pequenos diâmetros. Reduções de CHC na ordem de 51.5±70.0% e 62,8±75,6% após a quimioembolização foram encontradas no método do diâmetro e volume estimado, respectivamente, enquanto 57,8±64.1% e 70,6±36,4% foram encontrados pela segmentação do CHC para o maior diâmetro e volume, respectivamente. A maior diferença pode ser explicada pelo fato do algoritmo descontar áreas de necrose tumoral, visto que isso é difícil em alguns casos de CHC onde a necrose aparece difusa no tumor. A correlação de Pearson entre resposta e sobrevida analisada foi de 0,46 para volume segmentado e 0,17 para o diâmetro. Dessa forma, os resultados apresentados nesta pesquisa vêm colaborar com os métodos subjetivos de medição dos tumores apresentando maior confiabilidade na avaliação das áreas de realce do CHC. EssaO carcinoma hepatocelular (CHC) é um tumor primário do fígado e envolve diferentes modalidades de tratamento de acordo com seu estadiamento. Depois de terapias locais, como a quiomioembolização transarterial (do inglês, Transarterial Chemoembolization), a avaliação do tumor se baseia nos critérios mRECIST (do inglês, Modified Response Evaluation Criteria In Solid Tumors), que envolvem a medição do diâmetro máximo da lesão viável. Esta pesquisa teve como objetivo o desenvolvimento de um algoritmo para a quantificação objetiva da eficiência da quimioembolização na redução de volume e diâmetro máximo do CHC. Foram utilizados 64 exames retrospectivos de Tomografia Computadorizada (TC) de 32 pacientes. Nesta pesquisa foram comparadas, em pacientes que receberam tratamento por quimioembolização, as medidas do diâmetro maior do CHC viável, estimadas por especialista da área de radiologia e pelo algoritmo desenvolvido neste estudo. O Volume Tumoral Total (VTT) dos CHC antes e após o processo de quimioembolização também foram estimados utilizando um algoritmo, desenvolvido com base em transformadas discretas de wavelet (DWT). Um fantoma virtual simulando as características do fígado normal e contrastado foi desenvolvido para o planejamento, desenvolvimento e validação do algoritmo de detecção. A comparação do diâmetro máximo do tumor, estimado por um especialista e pelo algoritmo revelou diferenças na ordem de 0,25 cm para os tumores de grandes dimensões (diâmetro > 5 centímetros), enquanto uma diferença menor que 0,1 cm foi encontrada para os tumores de pequenos diâmetros. Reduções de CHC na ordem de 51.5±70.0% e 62,8±75,6% após a quimioembolização foram encontradas no método do diâmetro e volume estimado, respectivamente, enquanto 57,8±64.1% e 70,6±36,4% foram encontrados pela segmentação do CHC para o maior diâmetro e volume, respectivamente. A maior diferença pode ser explicada pelo fato do algoritmo descontar áreas de necrose tumoral, visto que isso é difícil em alguns casos de CHC onde a necrose aparece difusa no tumor. A correlação de Pearson entre resposta e sobrevida analisada foi de 0,46 para volume segmentado e 0,17 para o diâmetro. Dessa forma, os resultados apresentados nesta pesquisa vêm colaborar com os métodos subjetivos de medição dos tumores apresentando maior confiabilidade na avaliação das áreas de realce do CHC. Essa pesquisa traz contribuições originais para a comunidade cientifica com os algoritimos para quantificação objetiva de tumores no figado, a serem utilizados com confiabilidade de modo a otimizar a relação risco-beneficio para o paciente e custo-beneficio para a instituição. / Hepatocellular carcinoma (HCC) is a primary liver tumor and involves different types of treatment according to tumor staging. After local therapies such as transarterial chemoembolization (TACE), the evaluation of the tumor is based on mRECIST criteria (Modified Response Evaluation Criteria In Solid Tumors), which involves the measurement of the maximum diameter of the viable lesion. This research aimed to develop an algorithm for objective quantification of the efficiency of chemoembolization in reducing volume and maximum diameter CHC. 64 retrospective examination of computed tomography (CT) of 32 patients were used. Measures of patients who were treated by chemoembolization were compared using the measures of larger diameter viable CHC, estimated by specialist radiology and the algorithm developed in this study. The Total Tumor Volume (TTV) of CHC before and after chemoembolization process were also estimated using an algorithm developed based on Discrete Wavelet Transform (DWT). A virtual phantom simulating the characteristics of normal liver and contrasted was developed for planning, development and validation of detection algorithm. The comparison of the maximum tumor diameter, estimated by an expert and the algorithm revealed differences in the order of 0.25 cm for large tumors (diameter> 5 cm), while less than 0.1 cm difference was found for tumors of small diameters. CHC reductions of 51.5% ± 70.0 and 62.8 ± 75.6% were found after chemoembolization method in diameter and estimated volume, respectively, and 57.8% ± 64.1 and 70.6 ± 36.4% They were found by targeting the CHC for the larger diameter and volume, respectively. The biggest difference can be explained by the fact that the algorithm does not count areas of tumor necrosis, whereas it is difficult for the radiologist in some cases where CHC necrosis appears diffuse in the tumor. The Pearson correlation between response and analyzed survival was 0.46 and 0.17 for segmented volume and diameter. Thus, the results presented in this research come collaborate with subjective measurement of tumors showing higher reliability methods in the evaluation of the areas of enhancement of HCC. This research brings unique contributions to the scientific community with the algorithms for objective quantification of tumors in the liver, to be used in order to optimize the risk-benefit ratio for the patient and cost-effective for the institution.

Orientador: Diana Rodrigues de Pina / Resumo: O carcinoma hepatocelular (CHC) é um tumor primário do fígado e envolve diferentes modalidades de tratamento de acordo com seu estadiamento. Depois de terapias locais, como a quiomioembolização transarterial (do inglês, Transarterial Chemoembolization), a avaliação do tumor se baseia nos critérios mRECIST (do inglês, Modified Response Evaluation Criteria In Solid Tumors), que envolvem a medição do diâmetro máximo da lesão viável. Esta pesquisa teve como objetivo o desenvolvimento de um algoritmo para a quantificação objetiva da eficiência da quimioembolização na redução de volume e diâmetro máximo do CHC. Foram utilizados 64 exames retrospectivos de Tomografia Computadorizada (TC) de 32 pacientes. Nesta pesquisa foram comparadas, em pacientes que receberam tratamento por quimioembolização, as medidas do diâmetro maior do CHC viável, estimadas por especialista da área de radiologia e pelo algoritmo desenvolvido neste estudo. O Volume Tumoral Total (VTT) dos CHC antes e após o processo de quimioembolização também foram estimados utilizando um algoritmo, desenvolvido com base em transformadas discretas de wavelet (DWT). Um fantoma virtual simulando as características do fígado normal e contrastado foi desenvolvido para o planejamento, desenvolvimento e validação do algoritmo de detecção. A comparação do diâmetro máximo do tumor, estimado por um especialista e pelo algoritmo revelou diferenças na ordem de 0,25 cm para os tumores de grandes dimensões (diâmetro > 5... (Resumo completo, clicar acesso eletrônico abaixo) / Doutor

Intra-arterial or percutaneous locoregional therapies (LRT) are often employed to maintain potential liver transplant (LT) recipients with hepatocellular carcinoma (HCC) within T2/Milan criteria. Predictors of survival when LRT is used as destination therapy in those who are either ineligible or unwilling for LT remain poorly defined. We evaluated predictors of 3-year survival with destination LRT in a population of cirrhotic patients diagnosed with HCC, presenting within T2 criteria, and either ineligible or unwilling for LT. The cohort surviving 3 years had a significantly lower model for end-stage liver disease (MELD) score at HCC diagnosis (9.7 vs. 11.4, P= 0.037) and MELD following initial locoregional therapy (10.7 vs. 13.3, P= 0.008) compared to those not surviving three years despite similar demographic, tumor, and treatment variables. LRT as destination therapy results in modest intermediate term survival, with liver function at presentation and immediately following initiation of LRT predicting intermediate survival with this approach.

Hydroxycinnamoyl conjugates (HCC)s are an extremely diverse class of natural products that serve a wide variety of key functions in plant physiology, for example during wood formation, and in defence. They have diverse biological properties and act as antioxidants, antimicrobials, and antivirals. The biochemical basis of HCC diversity, however, has not yet been fully elucidated. Plants in the Populus genus are known to produce a particularly diverse range of HCCs and they constitute up to 5% of the leaf dry mass in some Populus species. HCCs can be formed by hydroxycinnamoyl transferases (HCTs) and distinct HCT isoforms in Populus may have distinct biological functions related to the synthesis of specific classes of HCCs. These can be identified on the basis of their evolutionary history and I show that many of the biochemically characterised HCTs belong to the BAHD superfamily of acyltransferases. My phylogenetic reconstruction of the BAHD superfamily has also defined a subclass containing most of the already-characterised HCTs, including nine potential HCT candidates in Populus. Caffeoyl-shikimate is a central precursor in the formation of lignin, a biopolymer (along with cellulose) that imparts mechanical stability to wood. Based on the transcript abundance of two candidate genes PtHCTA1 (Potri.001G042900)) and PtHCTA2 (Potri.003G183900) were hypothesised to be responsible for caffeoyl-shikimate formation in secondary xylem (i.e., wood). As part of this project, RNAi whole-plant knock-downs were generated for the xylem-associated PtHCTA1/2. The PtHCTA1/2 RNAi knock-downs have stunted growth, reminiscent of other mutants with impaired lignin biosynthesis. Based on thioacidolysis GC-MS, I found that the mutants produced a lignin with enriched hydroxyphenyl (H) subunits, which were derived from precursors upstream of the HCT-catalysed reaction and normally do not occur in Populus lignin. Interestingly, in one of the RNAi lines, the lignin phenotype was uncoupled from the developmental dwarfing phenotype. This is of high interest from a bioethanol perspective, since wood rich in H-lignin is more easily fermented than wood that is rich in guaiacyl (G) and syringyl (S) lignin. Another candidate gene (Potri.018G109900, HCT-E2) was linked to the formation of caffeoyl-spermidine in male catkins (which function in pollen coat formation), and one candidate gene (Potri.018G104700, HCT-C2) was associated with the formation of bioactive, soluble HCCs in leaves and roots. Since RNAi-mediated down-regulation proved ineffective, CRISPR-based gene knock-out methodology was developed and utilised for the Populus hairy root system. Targeted knock-out mutants for the leaf-associated HCT-C2 were generated. HCC identity was determined by metabolite purification and subsequent MS/MS/MS from leaf extracts, and the metabolite concentrations were determined by LC-MS. A decrease in chlorogenic acid concentration was apparent in CRISPR hairy-root knockouts of HCT-C2 indicating that HCTC2 is involved in HCC biosynthesis and can directly produce chlorogenic acid. Candidates for the HCTs involved in lignin biosynthesis, soluble ester biosynthesis, and pollen coat formation were identified and plant genetics confirmed the role of the lignin and soluble ester HCT candidates. / Graduate