Thesis (Ph. D.)--Case Western Reserve University, 2005. / [School of Medicine] Department of Epidemiology and Biostatistics. Includes bibliographical references. Available online via OhioLINK's ETD Center.

In this thesis Prognoslic factors of early recurrence of colorectal liver metastases after surgical therapy the autor characterizes the epidemidemiology, diagnostics and treatment of colorectal liver metastases (CLM) in relation to biological activity of tumour and the possibilities of determination. Contemporary the author introduces to problematics of tumour markers, which determine CLM and to clinical prognostic factors of CLM.

Hyperactive Wnt signaling is the seminal event in colorectal cancer (CRC) pathogenesis, where β-catenin serves as a key Wnt mediator enhancing CRC cell proliferation and survival. c-Cbl is a unique E3 ligase, which degrades both mutant and active (tumorigenic) β-catenin. c-Cbl phosphorylation at tyrosine 731 (Y731) regulates its binding and down regulation of β-catenin specifically in the presence of Wnt ligand (Wnt-on state). Since aberrant Wnt signaling activation is found in almost all cases of human CRC, it would be critical to understand the influence of c-Cbl phosphorylation on CRC cell survival. We hypothesized that c-Cbl phosphorylation regulates CRC cell survival in a Wnt dependent manner, a state that is mediated through mutations in β-catenin or adenomatosis polyposis coli (APC). Cbl phosphorylation was examined in a panel of Wnt-off cells with wild-type β-catenin and APC CRC cell line (RKO cell line) and Wnt-on cell lines with mutant APC (Wnt-on- DLD1, HCT15 cell line) or mutant β-catenin (HCT116) using phospho-specific antibodies to c-Cbl tyrosine residues at 700 (Y700), 731 and 774 (Y774) positions. Biological significance of specific phosphorylation sites was evaluated with phospho-inactive mutants of c-Cbl (Y700F, Y731F and Y774F) using both the MTT cell proliferation assay and the non-adherent colony formation assay. Potential meditators of c-Cbl were examined using immunoblotting. Here we show that c-Cbl was phosphorylated at all three major phosphorylation sites (Y700, Y731 and Y774) in both Wnt-off and Wnt-on CRC cell lines. However, the amount of phosphorylation was reduced in Wnt-on CRC cell lines (DLD1, HCT116 and HCT15) compared to Wnt-off (RKO) cell line. Wild-type c-Cbl significantly enhanced survival in RKO cell lines and reduced survivability in DLD1 cell lines. In contrast to the effect of wild-type c-Cbl, Y731F increased CRC cell survival and non-adherent colony forming units. Our preliminary data suggests that c-Cbl Y731 mutation regulates CRC survival through β-catenin. c-Cbl is heavily phosphorylated in CRC cell lines, where wild-type c-Cbl significantly inhibits cell survival in Wnt-on and enhances cell survival in Wnt-off CRC cell lines. Furthermore, our data indicates that Y731 influences CRC survival and colony formation only in Wnt-on cell lines. Though further validation is required, this dichotomy in the effect of c-Cbl phosphorylation on CRC survival being mediated by Wnt status can be further explored as a potentially novel therapeutic target in mutant CRC tumors, which represent more than 90% of CRC cases in humans.

BACKGROUND: The gold standard treatment for colorectal cancer liver metastases (CRCLM) is surgical resection. Unfortunately, the majority of patients with colorectal hepatic metastases are not candidates for resection. In recent years, several alternatives have emerged for patients whom are not resection candidates including modern systemic chemotherapy, targeted biologic treatments, regional therapies and local tumor ablation options. Microwave ablation (MWA) therapy is one such treatment alternative, based on thermal tissue ablation. This modality in concert with the most recent published literature on its use for patients with CRCLM will be reviewed in this paper. LITERATURE REVIEW FINDINGS: A structured review of the literature on ablative technologies was performed. In recent years, there has been an evolution from radiofrequency ablation (RFA) to microwave ablation therapy for the treatment of CRCLM. RFA has several limitations to its use and MWA theoretically avoids such limitations making it the currently preferable treatment option. There are limited publications comparing the use of RFA to MWA and limited publications on the use of microwave ablation for CRCLM. This paper will focus on the most recent data on MWA for CRCLM. This data can then be compared to the already published data on RFA. PROPOSED METHODS: Given the relative novel status for MWA as a treatment option for CRCLM, a potential disadvantage for its use is the perceived lack of knowledge across the medical professional spectrum. In an effort to expand the knowledge of MWA, the proposed outcomes for this study include creating a curriculum to be offered as a CME course focused for Primary Care Providers (PCPs) to provide a basis of clinical familiarity for its use. This effort will familiarize providers who may have patients diagnosed with CRCLM and also allow them to initiate the conversation about this therapy with their patients who may be candidates for this treatment. CONCLUSIONS: MWA therapy is a safe and effective treatment modality for CRCLM. Due to this new development in treating liver lesions originating from colorectal cancer, it’s imperative for providers to become familiar with these new technologies especially considering the high incidence of CRCLM. Therefore, a curriculum for PCPs will allow for a better understanding of this new technology and foster better provider-patient relationships.

The anticancer drug Camptothecin (CPT) specifically targets topoisomerase I (topoI). While this class of drug is used to treat various solid tumors, CPT is only effective in 13-30 percent of the patient population. Although the mechanism of the CPT resistance pathway is not fully understood, we found cancer cells degrade topoisomerase I (topoI) in response to CPT. The observed relationship between higher basal DNA-PKcs mediated phosphorylation of topoI, rapid degradation of topoI, and resistance to CPT suggested that DNA-PKcs is a critical regulator of the phosphorylation status of topoI and the rate of topoI degradation in response to CPT. The data shows CTDSP1 (a dual phosphatase) acts as a negative regulator of DNA-PKcs. Because CTDSP1 functionality plays a role in maintaining higher basal phosphorylation levels of topoI, CTDSP1 impairment contributes to greater CPT resistance. This renders the CPT chemotherapy treatment ineffective. We hypothesized inducing the catalytically inactive form of CTDPS1 via both knockdown and inhibition experiments would increase cancer cell resistance to CPT as a result of an overactive topoI degradation pathway. The function of CTDSP1 was impaired in the two colon cancer cell lines, HCT15 and HCT116, by silencing with siRNA and using Rabeprazole (a pharmacological agent known to inhibit CTDSP1). Inhibition of CTDSP1 phosphatase activity resulted in greater phosphorylation of DNA-PKcs and increased the rate of topoI degradation in the cells in response to CPT. Cellular resistance was also more notable in the sensitive HCT116 cell lines. HCT15 cells degrade topoI rapidly and are resistant to CPT. Thus, the effect of CPT is not as pronounced in this cell line. CTDSP1 knock down cells showed the greatest DNA-PKcs phosphorylation and topoI degradation when treated with CPT. In addition, the present study includes a clonogenic assay that shows a larger cell survival fraction in Rabeprazole treated HCT116 cells in comparison to controls after exposure to CPT. A higher survival fraction after CPT exposure is reflective of greater CPT resistance. This suggests that cell viability is enhanced during CPT chemotherapy treatment in CTDSP1 null colorectal cancer cells. Lastly, An EGFP read out experiment of topoI tagged to EGFP demonstrated CTDSP1 inhibition results in reduced topoI levels in colorectal cancer cells. The present study showed the potential link between lower topoI levels and greater resistance to CPT by showing that both effects are outcomes of silencing CTDSP1. / 2018-07-11T00:00:00Z

A Caderina-E està intimamente relacionada com a transiÃÃo epitelial-mesenquimal e com a progressÃo tumoral em muitos tipos de cÃncer, inclusive no cÃncer colorretal. O objetivo deste trabalho foi avaliar a imunoexpressÃo de Caderina-E no cÃncer colorretal primÃrio e nas respectivas metÃstases linfonodais, na mucosa colÃnica normal, e investigar possÃveis correlaÃÃes desta expressÃo com parÃmetros clÃnicopatolÃgicos. Setenta e sete casos de colectomias por carcinoma colorretal e dez casos de linfonodos metastÃticos, dos arquivos do Departamento de Patologia e Medicina Legal/Universidade Federal do CearÃ, foram utilizados. Realizou-se o Tissue Microarray e imunohistoquÃmica, com anticorpo monoclonal anti-Caderina-E. Foram avaliados os seguintes escores: 0 = ausÃncia de expressÃo; 1 = expressÃo citoplasmÃtica; 2 = expressÃo mista (citoplasmÃtica e membranar); 3 = expressÃo membranar pura. Foi utilizada tanto a classificaÃÃo proposta por Jawhari et al., agrupando os casos em expressÃo anormal (escores 0, 1 e 2) e expressÃo normal (escore 3), como os critÃrios propostos por Almeida et al., agrupando os casos como expressÃo nÃo-membranar (escores 0 e 1) e expressÃo membranar (escores 2 e 3). Os tumores primÃrios tiveram mais casos de expressÃo de Caderina-E anormal em comparaÃÃo com a mucosa normal (p < 0.0001). NÃo houve diferenÃa significante entre expressÃo de Caderina-E no tumor intestinal e em metÃstases linfonodais, embora nestas a expressÃo membranar tenha sido mais freqÃente do que no sÃtio primÃrio. Tumores de cÃlulas agrupadas apresentaram maior expressÃo de Caderina-E membranar do que os de cÃlulas isoladas, tanto utilizando a classificaÃÃo de Jawhari et al. (p = 0.0230), como os critÃrios propostos por Almeida et al. (p = 0.0043). Em conclusÃo, a expressÃo anormal de Caderina-E no tumor primÃrio, com persistÃncia freqÃente da imunomarcaÃÃo membranar associada à marcaÃÃo citoplasmÃtica (marcaÃÃo anormal heterogÃnea ou mista), reforÃa as evidÃncias de que esta alteraÃÃo no cÃncer à mais qualitativa do que propriamente quantitativa. O predomÃnio da expressÃo membranar no sÃtio primÃrio da neoplasia e na metÃstase, com ou sem expressÃo citoplasmÃtica associada, principalmente em tumores de cÃlulas agrupadas, sugere que a presenÃa da Caderina-E à essencial para a invasÃo local e progressÃo tumoral, em oposiÃÃo ao clÃssico paradigma de que a progressÃo tumoral se exacerba com a perda desta molÃcula de adesÃo. / E-cadherin is closely related to epitelial-mesenchymal transition and tumor progression in many cancers, including colorectal cancer. The aim of this study is to evaluate the expression of E-cadherin in primary colorectal cancer as well as in lymph node metastasis, establishing also a comparison with the expression of E-cadherin in normal colonic mucosa. We utilized 77 cases of colectomies for colorectal carcinoma and 10 cases of metastatic lymph nodes from the files of the Department of Pathology and Forensic Medicine/Federal University of Ceara. Tissue microarray and immunohistochemistry were performed with monoclonal anti-E-cadherin, evaluated using the following scores: 0 = no staining; 1 = cytoplasmic staining; 2 = mixed staining (cytoplasmic and membranous); 3 = membranous staining. It was used the classification proposed by Jawahri et al. which includes cases of abnormal expression (0, 1 and 2 scores) and cases of normal expression (3 score), and was also used the classification proposed by Almeida et al. which includes cases of non-membranous expression (0 and 1 scores) and membranous expression (2 and 3 scores). Primary tumors presented more cases of abnormal E-cadherin expression in comparison to normal colonic mucosa (p < 0.0001). There were no differences between E-cadherin expression in the primary tumor in comparison to lymph node metastasis. The grouped cell tumors showed increased expression of E-cadherin in comparison to isolated cell tumors, either using the classification proposed by Jawhari et al. (p = 0.0230) and the classification proposed by Almeida et al. (p = 0.0043). In conclusion, abnormal expression of E-cadherin in the primary tumor, with frequent membranar immunostaining associated with the cytoplasmic marking (abnormal heterogeneous or mixed staining), reinforces the evidence that E-cadherin expression change in cancer is more qualitative than quantitative. The predominance of membranar expression in primary tumor and lymph node metastasis, with or without associated cytoplasmatic expression, particularly in cell-grouped tumors, suggests that E-cadherin presence is essential for local invasion and tumor progression, as opposed to the classical paradigm that tumor progression is exacerbated by the loss of this adhesion molecule.

O cÃncer coloretal (CRC) à a 3 neoplasia mais prevalente no mundo. O irinotecano (IRI), fÃrmaco de primeira linha para os tratamentos do CRC e sua metÃstase hepÃtica, tem aumentado a sobrevivÃncia dos pacientes. Contudo, seus efeitos colaterais, incluindo a esteato-hepatite nÃo alcoÃlica (NASH), podem limitar o curso do tratamento. Os protocolos baseados em irinotecano foram associados com um aumento no risco de NASH de 3,4 vezes. A silimarina (SIL) tem mostrado ser capaz de prevenir doenÃas do fÃgado gorduroso no contexto clÃnico e em modelos de danos hepÃticos induzidos quimicamente. Assim, nosso objetivo foi estudar o efeito da SIL na NASH induzida pelo IRI, assim como o mecanismo envolvido. MÃtodos e Resultados: Camundongos Swiss fÃmeas (n=8-10), foram divididos em 6 grupos e injetados com salina (SAL 5ml/kg i.p.), IRI (50 mg/kg i.p.), SIL (150 mg/kg v.o.) ou IRI (50 mg/kg i.p.) + SIL (SIL 1,5, 15, 150 mg/kg v.o.) 3x/semana/7 semanas. Amostras de sangue foram coletadas na sÃtima semana para determinar a concentraÃÃo sÃrica das enzimas hepÃticas ALT e AST (U/L). Animais foram mortos para a coleta do fÃgado para avaliar do dano tecidual (escores de Kleiner), dosagem de lipÃdeos totais (mg/g de tecido), MDA (nmol/g tecido), NPSH (mg de NPSH/g tecido), IL-6 (pg/mg de tecido), IL-10 (pg/mg de proteÃna) e IL-1&#946; (pg/mg de tecido), imunomarcaÃÃo de Ãxido nÃtrico sintase induzida (iNOS), 3-Nitrotirosina (Ntyr), e Receptor Toll Like tipo 4 (TLR4), quantificaÃÃo do fator nuclear kappa B (NF&#954;B) e da &#945;-actina de mÃsculo liso (&#945;-SMA) e expressÃo do gene RSS. ANOVA/Teste de Newman-Keuls ou Kruskal Wallis/ Teste de Dunn foram utilizados para anÃlise estatÃstica. Foram consideradas diferentes amostras onde o nÃvel descritivo era inferior a 5%. O trabalho foi aprovado pelo comità de Ãtica em pesquisa animal sob o nÃmero de protocolo: 21/12. O IRI aumentou de forma significante as transaminases hepÃticas, o infiltrado neutrofÃlico, o acÃmo de lipÃdeos, o acÃmulo de MDA, a expressÃo de NTyr, a expressÃo de &#945;-SMA, a expressÃo de NF&#954;B, a expressÃo de IL-1&#946;, a expressÃo de IL-6, a expressÃo de TLR4 e quantificaÃÃo de DNA bacteriano, quando comparados ao grupo SAL. SIL (1,5 mg/kg) melhorou esses parÃmetros, exceto a infiltraÃÃo neutrofÃlica e a quantificaÃÃo do DNA bacteriano quando comparados ao o grupo IRI (P<0,05). Por outro lado, a dose media de SIL (15 mg/kg) foi efetiva apenas no Infiltrado NeutrofÃlico, na expressÃo de NTyr, na expressÃo de NF&#954;B e na expressÃo de IL-6. Adicionalmente, essa dose aumentou a expressÃo hepÃtica de IL-10, a quantificaÃÃo de DNA bacteriano e a expressÃo da &#945;-SMA quando comparados com o grupo IRI (p<0,05). Contudo, a expressÃo de iNOS nÃo foi afetada pelo prÃ-tratamento com SIL (P<0,05) e a maior dose foi ainda mais deletÃria. ConclusÃes: O prÃ-tratamento com SIL previne o dano hepÃtico causado pelo IRI provavelmente atravÃs da mudanÃa da resposta inflamatÃria mediada por receptores TLR4 e citocinas IL-1, IL-6, IL-10 e NF&#954;B. O dano observado no grupo de animais tratados com as maiores doses de SIL parece ser dependente da translocaÃÃo bacteriana do intestido que à associada a ativaÃÃo do TLR4. Adicionalmente, a silimarina contribui para hepatoproteÃÃo por inibir o estresse oxidativo e a nitrosilaÃÃo proteica, prevenindo a ativaÃÃo de mecanismos de fibrose hepÃtica

Orientadores: Wu Feng Chung, Huei Diana Lee / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-26T05:29:12Z (GMT). No. of bitstreams: 1 Espindola_Bianca_M.pdf: 2713191 bytes, checksum: 789ce5a7fa04c538dd3cc79742af1522 (MD5) Previous issue date: 2014 / Resumo: A manometria anorretal (MA) é um exame de especial interesse na área da Coloproctologia, pois, permite a avaliação de diversos parâmetros relacionados à fisiologia anorretal como a pressão máxima de contração voluntária (PMCV) e a capacidade de sustentação da pressão de contração voluntária (CS). Entretanto, apesar desses atributos serem amplamente utilizados, ainda persistem dúvidas em relação à melhor maneira de se avaliar a função de continência fecal, pois, diversos pacientes avaliados por meio desses parâmetros podem ser portadores de distúrbios funcionais relacionados ao ato de evacuação e apresentar resultados manométricos normais. Sob esse escopo, neste trabalho, analisou-se um novo parâmetro de avaliação da função anorretal, a área média resultante (AMR) sob as curvas pressão versus tempo do exame MA em comparação com os atributos biomecânicos PMCV e média da capacidade de sustentação (MCS). Foram incluídos no estudo 64 exames de MA, sendo 12 pertencentes a pacientes em condição de continência fecal (Controle), quatro representativos de indivíduos portadores de IF grau I (IF GI), 20 provenientes de pessoas com característica de IF grau II (IF GII) e 28 representaram o conjunto de sujeitos com IF grau III (IF GIII). Para o delineamento dos gráficos de manometria e cálculo da PMCV, MCS e AMR de cada exame, foi utilizado um aplicativo computacional desenvolvido por meio das linguagens de programação R e Java. As médias da PMCV (mmHg) encontradas nos grupos Controle, IF GII e IF GIII foram 247,58 mmHg, 142,40 mmHg e 153,36 mmHg, respectivamente (p = 0,0001), enquanto na MCS (segundos), 37,44 segundos, 35,97 segundos e 37,31 segundos, respectivamente (p = 0,1155) e, por final, na AMR (mmHg x segundos) 3934,32 mmHg x segundo, 2031,49 mmHg x segundo e 1855,60 mmHg x segundo, respectivamente (p = 0,0001). Nas comparações entre pares, a PMCV mostrou-se adequada para diferenciar pacientes continentes de incontinentes GII e de GIII (p < 0,001), no entanto, não ocorreu diferença estatística significativa após a comparação feita entre o grupo IF GII e o IF GIII (p > 0,05). De modo semelhante, a AMR foi eficiente para distinguir o Grupo Controle do IF GII e do GIII, todavia, não se mostrou adequada para diferenciar os dois últimos grupos entre si. A MCS não se mostrou adequada para diferenciar pacientes continentes de incontinentes fecal grau II e grau III. Por meio desses resultados, a PMCV e a AMR podem auxiliar no diagnóstico de pacientes com incontinência fecal. Todavia, novos estudos precisam ser realizados com a finalidade de aumentar a compreensão da fisiopatogenia dessa condição patológica / Abstract: The anorectal manometry (AM) is an exam of special interest in the area of Coloproctology, because it permits the evaluation of several parameters related to the anorectal¿s physiology, such as the maximum voluntary contraction pressure (MVCP) and the support¿s capacity of voluntary squeeze pressure (SCVSP). However, despite the fact that these parameters are widely used, questions related to the best way to evaluate the fecal continence function still remain. This is due to the fact that several of the evaluated patients possibly have functional disorders related with the defecation act and yet have normal manometrics¿ results. Under this scope, in this work a new anorectal function evaluation¿s parameter has been analysed: the average resulting area (ARA) obtained from anorectal manometry exam¿s curves "Pressure versus Time" compared with the biomechanics attributes MVCP and average support¿s capacity (ASC). During the research 64 manometric exams were included, of which 12 of patients with fecal continence (Control), 4 representing individuals with FI degree I (FI DI), 20 with characteristics of FI degree II (FI DII) and 28 representing a group of patients with FI degree III (FI DIII). For the manometric graphics¿ delineation and for calculating the MVCP, SCVSP and ARA of each exam, a computational application developed using Java and R languages was used. The average values of MVCP (in mmHg) found on the groups Control, FI DII and FI DIII were respectively 247.58 mmHg, 142.40 mmHg and 153.36 mmHg (p = 0.0001). The SCVSP results (in seconds) were respectively 37.44 seconds, 35.97 seconds and 37.31 seconds (p = 0.1155). Finally, the ARA results (in mmHg x seconds) were correspondingly 3934.32 mmHg x second, 2031.49 mmHg x second and 1855.60 mmHg x second (p = 0.0001). In the comparison between pairs, the MVCP parameter has demonstrated to be appropriate to distinguish continent from incontinent patients DII and DIII (p > 0.001). However, there was no significant statistic difference when the comparison was made between the group FI DII and FI DIII (p > 0.05). In a similar way, the ARA parameter was efficient to differentiate the Control group from the groups FI DII and FI DIII, although it was not adequate to distinguish the last two ones between themselves. There was no evidence that SCVSP was adequate to differentiate continent patients from fecal incontinent patients degree II and degree III. According to these results, the MVCP and the ARA parameters have shown to be able to assist the patient¿s diagnosis with fecal incontinence. However, new researches need to be developed to better understand the pathogenesis of different degrees of this illness / Mestrado / Fisiopatologia Cirúrgica / Mestra em Ciências

Colorectal cancer is a common disease, representing the third and second most common cause of cancer death in the United States in women and men, respectively. [Ahnen et al.: Mayo Clin Proc 2014;89:216-224; Siegel et al.: CA Cancer J Clin 2016;66:7]. It is estimated that 20% of patients have distant metastatic disease at time of diagnosis [Ahnen et al.: Mayo Clin Proc 2014;89:216-224; Siegel et al.: CA Cancer J Clin 2016;66:7]. The most common metastatic sites include regional lymph nodes, liver, lungs, and peritoneum via lymphatic/hematogenous dissemination as well as contiguous and transperitoneal routes [Ahnen et al.: Mayo Clin Proc 2014;89:216-224; Siegel et al.: CA Cancer J Clin 2016;66:7]. Upon review of the literature, we found that metastatic colon cancer to the scrotum is rare. The following case report proved to be a unique example of this type of metastasis. This rare regional metastasis is theorized to have resulted from a colo-urethro-scrotal fistula that precipitated from the patient's prior traumatic event. (C) 2017 The Author(s) Published by S. Karger AG, Basel

Class of 2017 Abstract / Objectives: To determine if the addition of oral alvimopan, liposomal bupivacaine and intravenous acetaminophen as part of a comprehensive enhanced recovery after surgery (ERAS) program decreases length of stay, recovery time and narcotic/acetaminophen use without affecting colorectal surgery patient outcomes. Methods: Patients were compared before and after the implementation of alvimopan, liposomal bupivacaine and intravenous acetaminophen with an ERAS program. The primary outcome was hospital length of stay (measured in hours). Secondary outcomes included change in time to first meal, bowel sounds, and bowel movement (measured in hours), pain scores (visual analog scale 0-10), opioid use (measured in morphine equivalent milligrams), and acetaminophen use (measured in mg). Results: Thirty-seven individuals were included in the pre implementation population and fifty one patients were included in the post implementation population. The mean length of stay decreased from 124.3 hours to 100.2 hours (P equals 0.13) with the addition of the ERAS program with the three medications. The 24 hour morphine equivalent intervals for seventy-two hours following surgery decreased from 125.8 mg (day 1), 81.9 mg (day 2) and 44.5 mg (day 3) to 44.3 mg (day 1), 22.8 mg (day 2) and 13.2 mg (day 3) (P less than 0.005 for each one). Conclusions: The addition of alvimopan, liposomal bupivacaine and intravenous acetaminophen as part of a comprehensive ERAS program decreased length of stay but not significantly. However, the addition of these three medications with the ERAS program changes was associated with a statistically significant decrease in opioid use.