Tesis: "Carcinoma colorettale (Colorectal cancer)"

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Mescoli, Claudia. "FATTORI PROGNOSTICI DEL CARCINOMA COLO-RETTALE: IL FENOTIPO E IL RUOLO DELLE CELLULE TUMORALI ISOLATE STUDIO PROSPETTICO". Doctoral thesis, Università degli studi di Padova, 2011. http://hdl.handle.net/11577/3427520.

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AIM.To date, lymph node (LN) involvement is the most important prognostic factor in patients undergoing radical surgery for CRC, and pN+ve status identifies patients who require adjuvant chemotherapy. About 20-30% of patients with pathological-negative LNs (pN0), however, develop recurrent disease. Thus, there is a subgroup of pN0-CRC patients harbouring occult metastatic disease that is undetected by the current pathological and clinical evaluations. The need for more accurate prognostic factors has addressed the aim of this study to look for new morphologic reliable markers. PATIENTS AND METHODS. The study included: - Group A: 1606 consecutive pN0-CRC patients surgically treated at the University of Padova between 2002 and 2008; - Group B: 944 pN0-CRC patients (obtained from group A); - Group C: 361 consecutive pN0-CRC patients (obtained from group B, ruling out rectal carcinomas undergone neoadjuvant radio-chemotherapy) in order to immunohistochemically assess the prevalence of ITC and the clinical outcome of pN0-ITC+ve-CRC patients. All the 1606 surgical resections were sampled and diagnosed at the Pathology Department according to a standardized precedure in order to recorder the clinico-pathological features. From 361 pN0 cases, ITC detection has been performed on serial sections (at different levels: 80-100 μm) of the 5920 originally dissected (formalin fixed paraffin embedded) lymph nodes. Each section has been stained with appropriate antibodies to reveal the presence of tumoral cells undetected at the routine observation. The clinico-pathological features has been compared between each other and the clinical outcome. RESULTS. (a) The importance of histopathological variables (Infiltration Depth, Lymphonodal Status, Growth Pattern, Vascular Invasion, Rectal localization, Tumoral de- differentiation Grade ) as prognostic factors has been confirmed; (b) Independent paramenters influencing relapse in negative nodes are : Tumoral de differentiation Grade, Histotype, Growth Pattern and peri and intra neoplastic inflammation; (c) Multivariate Analisys shows that ITC presence in nodes of CRC pN0 patients is the only variable significantly influencing prognosis. CONCLUSIONS. The present study confirms the importance of those histopathological parameters which are not primary involved in patient stratification risk of relapsing disease, in particular of ITC. Our results elucidate the role of ITC as prognostic factors clinically significant and they suggest the possible utilization of ITC as selection criteria in pN0 patients for adiuvant therapy.
SCOPO. Il carcinoma del colon-retto (CCR) è la seconda neoplasia maligna causa di morte nel mondo occidentale. La presenza di metastasi nei linfonodi regionali è il fattore prognostico più importante e, distinguendo lo stadio III (pN1/2) dagli stadi I e II (pN0) (in assenza di metastasi extranodali) identifica i pazienti da sottoporre a terapia adiuvante. Tuttavia, il 20-30% dei pazienti senza metastasi linfonodali (pN0) sviluppa malattia recidiva. Esiste, pertanto, tra gli stadi precoci, un sottogruppo di soggetti “portatori” di metastasi occulte che non sono identificabili con le comuni tecniche di valutazione. La necessità di nuovi e più accurati fattori prognostici ha indirizzato lo scopo di questo lavoro alla ricerca di parametri morfologici clinicamente rilevanti per lo sviluppo di nuove strategie terapeutiche. PAZIENTI E METODI. Lo studio include: - Gruppo A: 1606 pazienti consecutivi con CCR sottoposti a resezione curativa; - Gruppo B: 944 pazienti pN0 (estratti dal gruppo A); - Gruppo C: 361 pazienti consecutivi pN0 (estratti dal gruppo B, escludendo i cancri rettali sottoposti a terapia neo-adiuvante) nei cui linfonodi è stata indagata la presenza/prevalenza di Cellule Tumorali Isolate. 1606 resezioni consecutive colo-rettali sono state campionate e refertate secondo un protocollo standardizzato per raccoglierne le caratteristiche clinico-patologiche. Da 361 resezioni colo-rettali, campionate e refertate come sopra, sono stati analizzati con metodica immunoistochimica (anticorpo anti-citocheratina MNF116) 5920 linfonodi, per ognuno dei quali sono state esaminate due sezioni a distanza di 80-100 μm. Le caratteristiche clinico-patologiche sono state comparate tra loro e con i risultati dell’ outcome secondo i parametri “ricorrenza di malattia” e “decesso per malattia”. RISULTATI. (a) L’importanza di variabili isto-patologiche (Profondità d’infiltrazione, Status Linfonodale, Pattern di crescita, presenza di Invasione Vascolare, insorgenza in Sede Rettale, Grado di de-differenziazione tumorale) come fattori prognostici è stata consolidata; (b) Parametri indipendenti che influenzano la recidiva in assenza di metastasi linfonodali sono : Grado di de-differenziazione tumorale , Istotipo, Pattern di crescita e Reazione flogistica intra e peritumorale; (c) Analisi in multivariata dimostrano che la presenza di ITC nei linfonodi di pazienti con CCR pN0 è l’unica variabile che influenza significativamente la prognosi. CONCLUSIONI. Il presente studio conferma l’importanza di parametri isto-patologici “secondari” nella stratificazione del rischio di ricorrenza di malattia, in particolare delle Cellule Tumorali Isolate. I risultati ottenuti chiariscono il ruolo delle ITC come indicatori prognostici clinicamente rilevanti e ne suggeriscono il possibile utilizzo come criterio di selezione di pazienti pN0 da sottoporre a terapia adiuvante.

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Bertazza, Loris. "Analisi delle Cellule Tumorali Circolanti nel carcinoma gastrico e nelle metastasi epatiche da cancro del colon-retto: ruolo di Survivin e CD133 come fattori prognostici". Doctoral thesis, Università degli studi di Padova, 2011. http://hdl.handle.net/11577/3427460.

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Background At present the only prognostic system routinely employed for the management of gastric cancer patients is the TNM staging classification, that identifies broad risk categories with a significant prognostic variability within each stage, which makes TNM a suboptimal predictive tool on the single patient basis. Only 10-20% of patients with liver metastases from colorectal cancer is resectable with radical intent and 60-70% will develop a relapse despite apparently curative surgery. Both these classes of patients therefore would benefit from additional treatments after surgery such as adjuvant chemotherapy. We need new prognostic factors that can identify patients at high risk to be submitted to treatment. Aim of the study To study circulating tumor cells by gene profiling the peripheral blood in order to identify prognostic biomarkers that add independent prognostic power to the conventional staging systems. This might allow for a better stratification of patients’ risk and thus a better therapeutic management of gastric cancer and metastatic colorectal cancer patients, especially in the adjuvant setting. Patients, materials and methods 70 patients, affected with gastric adenocarcinoma at different TNM stages of disease who underwent radical surgery and 50 patients undergoing liver resection for metastases from colorectal cancer (stage IV) were enrolled in the study. Immediately before surgery, a sample of peripheral blood was withdrawn from each patient. For each sample, RNA was extracted and utilized for quantitative real time PCR evaluation of the expression of the following genes: CK19, CEA, VEGF, and Survivin for gastric cancer patients; CK19, CK20, CEA, VEGF, EGFR, CD133 and Survivin for colorectal liver metastases patients. Univariate and multivariate survival analysis was performed to investigate on the prognostic role of these biomarkers. Results After stepwise variable selection, Cox multivariate analysis of survival showed a significant association between overall survival and both TNM stage ad Survivin gene expression levels in peripheral blood of gastric cancer patients, while multivariate analysis confirmed the statistically significant association between both the radical resection and the transcriptional levels of CD133 and overall survival in colorectal liver metastases patients. In addition, Survivin transcriptional levels were higher in patients with gastric cancer as compared to the calibrator reference (obtained from the peripheral blood of healthy donors) in 98.6% of cases; analogously, CK19 was upregulated in 97.1% of cases. These findings support the hypothesis that the peripheral blood gene profile might be utilized also as a diagnostic marker in gastric cancer. Concluding remarks The positive findings of this pilot study are the basis for larger prospective studies in larger groups of gastric cancer and colorectal liver metastases patients, aimed to validate the prognostic power of Survivin and CD33 expression in peripheral blood of these two groups, respectively. Moreover it would be interesting to further explore also the potential diagnostic value of the peripheral blood gene profile in gastric cancer.
Presupposti dello studio Attualmente l'unico sistema prognostico utilizzato in clinica per i pazienti con cancro gastrico è la stadiazione TNM, che crea classi di rischio con prognosi significativamente diversa, ma con un’alta variabilità del rischio all’interno delle singole classi, risultando così uno strumento prognostico non ottimale a livello di singolo paziente. Solo il 10-20% dei pazienti con metastasi epatiche da carcinoma del colon-retto (CRC) risulta resecabile con intento radicale e di questi il 60-70% svilupperà una recidiva nonostante l’intervento potenzialmente curativo. Entrambe queste classi di pazienti necessitano di trattamenti aggiuntivi alla chirurgia come la chemioterapia adiuvante. Sono quindi necessari fattori prognostici nuovi, che permettano di individuare i pazienti ad alto rischio da indirizzare alla terapia. Scopo dello studio Studiare le cellule tumorali circolanti, attraverso il profilo di espressione genica nel sangue periferico, per individuare fattori prognostici indipendenti, in modo da rendere migliore la stratificazione del rischio e di conseguenza la cura dei pazienti con adenocarcinoma gastrico e con metastasi epatiche da carcinoma del colon-retto, con particolare riguardo alla selezione dei pazienti da trattare con terapia adiuvante. Pazienti, materiali e metodi Nello studio sono stati inclusi 70 pazienti con adenocarcinoma gastrico in diverso stadio TNM sottoposti a gastrectomia con intento radicale e 50 pazienti con metastasi epatiche da CRC sottoposti a chirurgia. Prima dell’intervento chirurgico, a ogni paziente è stato eseguito un prelievo di sangue venoso periferico, se ne è estratto l’RNA totale ed il corrispondente cDNA è stato utilizzato per l’analisi di espressione genica mediante PCR quantitativa. Per i pazienti con carcinoma gastrico sono stati valutati i geni CK19, CEA, VEGF, Survivin; per i pazienti con metastasi epatiche da CRC sono stati valutati i geni CK19, CK20, CEA, VEGF, EGFR, CD133 e Survivin. Per valutare il ruolo prognostico di ogni marcatore sono state effettuate le analisi di sopravvivenza uni- e multivariata. Risultati All’analisi multivariata secondo Cox della sopravvivenza globale, dopo selezione stepwise, sono risultati fattori prognostici indipendenti per i pazienti con cancro gastrico la stadiazione TNM e l’espressione del gene codificante per Survivin, mentre per i pazienti con CRC metastatico sono risultati fattori prognostici indipendenti la radicalità dell’intervento e l’espressione di CD133 nel sangue periferico. Inoltre Survivin era maggiormente espressa nei pazienti con carcinoma gastrico rispetto al calibratore (ottenuto dal sangue di donatori sani) nel 98.6% dei casi; analogamente CK19 era maggiormente espressa nel 97.1% dei casi. Questi dati supportano la possibilità dell’utilizzo dell’espressione genica nel sangue periferico anche come marcatore diagnostico del carcinoma gastrico. Conclusioni I risultati positivi di queste analisi costituiscono la base per la conduzione di più ampi studi prospettici nelle due patologie considerate, al fine di poter validare il valore prognostico dell’espressione di Survivin e CD133 nel sangue periferico dei pazienti rispettivamente con carcinoma gastrico e con CRC metastatico. Sarebbe inoltre di sicuro interesse confermare il significato diagnostico del profilo genico del sangue periferico nel cancro gastrico.

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La, Morella Carla Maria Alessia. "La fibrinogenemia nel K colorettale (prospettive come fattore prognostico)". Doctoral thesis, Università di Catania, 2012. http://hdl.handle.net/10761/949.

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Scopo della presente tesi è quello di valutare, in base ai dati della letteratura, se l iperfibrinogenemia costituisca un elemento clinicamente rilevante ai fini predittivi della recidiva del carcinoma colorettale. Viene inoltre presentata la casistica relativa ai casi di CRC operati presso la Clinica Chirurgica II (Direttore Prof. Salvatore Berretta) del P.O. Policlinico dal 2001 al 2010.

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Schutte, Berend. "Cancer cell ploidy and proliferation in colorectal carcinoma". Maastricht : Maastricht : Rijksuniversiteit Limburg ; University Library, Maastricht University [Host], 1987. http://arno.unimaas.nl/show.cgi?fid=5360.

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Gruppo, Mario. "Subclinical myopathy and colorectal cancer: identification and role of new muscle damage and regeneration biomarkers". Doctoral thesis, Università degli studi di Padova, 2015. http://hdl.handle.net/11577/3424623.

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Background Skeletal muscle is the major reservoir of body proteins and it can be affected in conditions associated to altered protein turnover and metabolism such as cancer. Although severe wasting is seen primarily in patients with advanced malignancy, some of them present degree of wasting at the onset of disease. Autophagy has been recently described to play a relevant role in muscle wasting. Materials and Methods We performed morphometric studies and immunohistochemical analyses on intraoperative rectus abdominis muscle biopsies from 50 consecutive weight stable colorectal patients and 25 weight-stable patients operated for non-inflammatory benign diseases with no clinical signs of myopathies. Biochemical and molecular analyses have been performed in order to evaluate protein profile, the presence of autophagy induction and their correlation with clinical outcome. Results In cancer patients, we observed a subclinical myopathy characterized by an abnormal distribution of myonuclei relocated from the periphery inside the myofiber. The percentage of myofibers with abnormally located myonuclei was significantly higher in patients compared to controls. Analyses on serum samples showed that, in the absence of systemic inflammation, in the prevalence of cancer patients the levels of albumin and prealbumin were below the normal range and the mean value was significantly lower compared to that detected in controls. Molecular analyses showed an accumulation of p62, a typical marker of autophagy induction, significantly higher in cancer patients compared to controls. We found an inverse correlation between the number of abnormally nucleated myofibers and the presence of lymph node metastasis. Cancer relapse was correlated with low serum levels of prealbumin and high levels of p62 in myofibers of cancer patients. Conclusions Colorectal cancer patients have a subclinical myopathy characterized by myofibers with internally located myonuclei. In the absence of inflammation, cancer patients show low levels of prealbumin and albumin as markers of altered protein turnover and persistent high levels of p62 in myofibers as expression of autophagy induction with an impairment in physiological autophagic flux. Up to now our data indicate that skeletal muscle fibers show nuclear abnormalities that seems to be associated to a better prognosis, while the presence of an altered protein turnover at an early stage of disease, with an impairment in the physiological autophagic flux, that could be predictive of cancer relapse and onset of cancer cachexia.
Introduzione Il muscolo scheletrico rappresenta la principale riserva proteica del corpo e può essere compromesso in varie affezioni metaboliche e di alterato turnover proteico, quale il cancro. Benchè una severa perdita di massa sia generalmente presente in quadri neoplastici avanzati, in alcuni casi può essere già evidente in una fase di malattia iniziale. L’autofagia è stata recentemente descritta come uno dei possibili fattori responsabili del processo catabolico. Materiali e Metodi 50 pazienti sottoposti ad intervento chirurgico per neoplasia colorettale e 25 pazienti operati per patologia benigna non infiammatoria, in assenza di segni clinici di miopatia, sono stati sottoposti a biopsia muscolare su cui sono state eseguite analisi di carattere morfometrico ed istochimico. Sono state, inoltre, eseguite analisi biochimiche e molecolari al fine di valutare l’assetto proteico e lo stato di attivazione del processo autofagico e la loro correlazione con l’outcome clinico dei pazienti. Risultati Nei pazienti neoplastici abbiamo riscontrato la presenza di una miopatia subclinica, caratterizzata dalla presenza di fibre muscolari con un’anomala localizzazione del nucleo cellulare al centro della fibra, significativamente maggiore rispetto ai controlli. L’analisi dell’assetto proteico ha dimostrato valori sierici di albumina e prealbumina significativamente più bassi nei pazienti oncologici, mentre l’analisi molecolare ha documentato elevati livelli di p62 nelle fibre muscolari dei pazienti affetti da carcinoma colorettale, rispetto ai controlli. La valutazione dell’outcome clinico ha dimostrato una correlazione inversa tra la percentuale di miofibre anomale e l’insorgenza di metastasi linfonodali, mentre bassi livelli sierici di prealbumina ed alti livelli di p62 nelle fibre muscolari sono risultati correlati con un aumentato rischio di ripresa di malattia Conclusioni I pazienti affetti da carcinoma colorettale presentano una miopatia subclinica già all’insorgenza della malattia, caratterizzata dalla presenza di fibre con alterata posizione del nucleo nella cellula. In assenza di infiammazione sistemica e tissutale, i pazienti oncologici presentano bassi livelli sierici di albumina e prealbumina, come espressione di un alterato turnover proteico, nonché elevati livelli di p62 nelle fibre muscolari, a dimostrazione dell’attivazione del processo autofagico che risulta tuttavia compromesso. Tali dati suggeriscono, pertanto, un verosimile ruolo protettivo per le anomalie nucleari descritte, mentre un alterato turnover proteico ed una compromissione del normale flusso autofagico, in concomitanza dell’insorgenza della neoplasia, costituiscono un potenziale fattore predittivo negativo in termini di ripresa di malattia ed evoluzione verso uno stato cachettico.

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Barrow, Emma. "Cancer risk and biomarker analysis in mismatch repair deficient colorectal carcinoma". Thesis, University of Manchester, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.516737.

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Lynch Syndrome is caused by mutations in the DNA mismatch repair (MMR) genes; MLH1, MSH2, MSH6 and PMS2. An accurate estimation of cancer risk for MMR mutation carriers is essential for counselling and screening. United Kingdom (UK) specific cancer risks for MMR mutation carriers and cumulative colorectal cancer risks by decade have not been described to date. Using data from 121 Lynch Syndrome families, the cumulative lifetime risks of Lynch Syndrome spectrum cancers in mutation carriers were calculated, correcting for ascertainment bias. This data provides reassurance that current UK screening guidelines are appropriate. For colorectal cancer, tables of risk by decade were compiled for carriers of the different gene mutations. This novel data represents a useful counselling tool. Despite an increased cancer risk, individuals with Lynch Syndrome lack clear phenotypic characteristics. Mutation carrier identification is desirable as screening reduces mortality. After counselling and assessment of family history against clinical criteria, molecular diagnosis of tumour tissue is performed. Immunohistochemistry (IHe) of the MMR proteins has great potential as a method of identifying the most likely mutation and guiding mutation analyse. However IHC of the MMR proteins is not yet well established and problems have been reported with staining and slide interpretation. IHC slide assessment methods in the reported literature have been qualitative. This thesis describes the development and validation of a robust, semiquantitative scoring technique to be used in the assessment of IHC stained tumour sections from patients with possible Lynch Syndrome. Colorectal tumour sections from 51 MMR mutation carriers were stained with 3,3' Diaminobenzidine (DAB) using antibodies against the MMR proteins. Slide assessment was semiquantified using a 0-12 scale, and was found to be highly sensitive and specific for the identification of mutation carriers. It was found however, that protein expression may occur in the context of known pathogenic mutations, a potential pitfall in the screening process. Two quantitative techniques of slide assessment that could potentially obviate the need for human operator slide analysis were explored. Semiconductor quantum dots (QOs) are quantifiable fluorescent labels that can be used for multiplex staining. Quantitative QO IHC was compared to quantitative DAB IHC. Both are novel methodologies. Tumour sections from 36 mutation carriers were stained using each method, and multispectral analysis of the slides was performed. Quantitative analysis of the DAB stained slides was less sensitive than semi-quantitative analysis in this study, but was sufficiently sensitive to be used as a pre-screen in Lynch Syndrome. However despite the potential advantages of quantitative QD staining, it was less sensitive and specific than quantitative DAB IHC. The development of automated DAB IHC staining and quantitative multispectral slide analysis may enable future high throughput IHC for MMR mutation carrier identification. Around 15% of sporadic colorectal cancers develop along the MSI pathway due epigenetic MLH1 inactivation. The evidence suggests that patients with such tumours do not benefit from 5-Fluorouracil based chemotherapy. The cost effectiveness of identifying patients with such tumours was explored. Potential chemotherapeutic savings are substantial, supporting the introduction of this biomarker into clinical practice. The improved diagnostic accuracy of semi-quantitative IHC slide assessment and the potential for automation of quantitative IHC slide assessment would facilitate the introduction of this service.

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Fernando, Winnie Collet. "Molecular Pathogenesis of Serrated Carcinoma of the Colon". Thesis, Griffith University, 2014. http://hdl.handle.net/10072/367873.

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Colorectal cancer (CRC) progresses from precursor lesions or polyps via two main pathways – the traditional and serrated pathway. The traditional pathway is the conventional type, while the serrated neoplastic pathway has been recently identified and is not well characterised. The aim of this study was to evaluate molecular factors associated with the serrated pathway and study its progression from a serrated polyp to colorectal cancer using patient samples and mouse models. The first part of the study was to evaluate CIMP status of all the serrated polyp subtypes and its association with functionally important genes such as MLH1, p16 and IGFBP7 in patient samples. The mouse models were then utilized to assess the contribution of BRAF V600E mutation along with the cell cycle regulators and tumour suppressor genes, p16Ink4a and p19Arf in the initiation and progression of CRC. The CIMP study was a prospective study in which the real-time based MethyLight assay was employed to evaluate CIMP and methylation status of p16, IGFBP7 and MLH1 in 154 serrated polyps and 63 adenomas. Samples were subjected to bisulfite modification and methylation levels were assessed using the Weisenberger et al panel of methylation markers (IGF2, SOCS1, NEUROG1, RUNX3 and CACNA1G) with ALU as the reference gene.
Thesis (PhD Doctorate)
Doctor of Philosophy (PhD)
School of Biomolecular and Physical Sciences
Science, Environment, Engineering and Technology
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Jankowski, Janusz Antoni Zygmunt. "Constitutive gene expression during colorectal tumorigenesis : in vivo and in vitro". Thesis, Imperial College London, 1996. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.321753.

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Melville, D. M. "The cancer risk in ulcerative colitis : the accuracy of markers of premalignant change and their value in clinical practice". Thesis, University of Oxford, 1988. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.235940.

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Majorana, Alessandra. "Alterazioni specifiche del trascrittoma dei microrna e struttura globale del network in carcinoma colorettale dopo trattamento con Cetuximab". Thesis, American Association for Cancer Research, 2011. http://hdl.handle.net/10761/129.

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Il carcinoma del colon-retto (CRC) e' una delle neoplasie piu' frequenti nel mondo occidentale e rappresenta un problema socio-sanitario di notevole rilievo. E' noto che non tutti i pazienti affetti da CRC rispondono in modo positivo alla terapia con i piu' comuni agenti antitumorali (es. Cetuximab): vi sono infatti dei fattori genetici predittivi della risposta, tra i quali sicuramente uno dei piu' noti e' lo stato mutazionale di KRAS. Tuttavia, questo approccio diagnostico molecolare e' comunque invasivo, poiche' presuppone un prelievo bioptico. Per questa ragione, diversi gruppi di ricercatori stanno cercando di individuare biomarcatori che, oltre ad essere specifici, siano identificabili con procedure non invasive. Indubbiamente, tra i possibili marcatori, i microRNA (miRNA) rivestono un ruolo particolarmente rilevante. I miRNA sono piccole molecole di RNA non codificante che svolgono un ruolo critico nella regolazione dell ' espressione genica in tutti i processi cellulari. Ad oggi, sono stati pubblicati diversi dati che suggeriscono come il profilo di espressione dei miRNA vari in modo specifico nei diversi tipi di cellule neoplastiche, in correlazione con il fenotipo del tumore e con la sua evoluzione. L' oggetto di questa tesi e' stato lo studio della relazione tra risposta terapeutica e modifiche del trascrittoma dei miRNA nel tumore colorettale (CRC), che ad oggi rimane sconosciuta. Per raggiungere questo obiettivo abbiamo effettuato il profiling dell'espressione di 667 miRNA in due linee cellulari umane CRC, una sensibile e l'altra resistente al Cetuximab (Caco-2 e HCT-116, rispettivamente) mediante RT-PCR con sonde TaqMan. Le Caco-2 e le HCT-116 esprimono diversi set di miRNA dopo il trattamento: in particolare, mentre nelle Caco-2 sono differenzialmente espressi 21 miRNA (DE miRNA), nelle HCT-116 i DE miRNA sono 22 (t-test, p <0.01). Testando l'espressione dei DE miRNA in campioni paraffinati di pazienti affetti da CRC, abbiamo scoperto che il miR-146b-3p e il miR-486-5p sono piu' abbondanti nei campioni con il gene KRAS mutato rispetto a quelli wild-type (test di Wilcoxon, p <0.05). Dall' analisi dei cluster e delle famiglie geniche dei DE miRNA e' emerso che miRNA localizzati in stretta prossimita' genomica o appartenenti alla stessa famiglia mantengono spesso lo stesso trend di espressione in seguito al trattamento. Secondo dati di letteratura, il 67% dei DE miRNA e' coinvolto nel cancro, incluso il CRC, mentre 19 targets dei DE miRNA sono stati precedentemente associati alla pathway del Cetuximab e del CRC, come ad esempio KRAS (targets dei DE miRNA let-7b e let-7e), PTEN e PIK3R1 (entrambi targets del miR-486-5p). Abbiamo identificato 25 fattori di trascrizione che putativamente controllerebbero questi DE miRNA; 11 di questi sono gia' stati individuati per essere coinvolti nella patogenesi del CRC, come ad esempio MYC, che controlla positivamente l ' espressione del miR-17* (un marcatore del CRC i cui livelli sono abbondanti in biopsie e plasma di pazienti). Sulla base di questi dati, suggeriamo che la sottoespressione di let-7b e let-7e e la sovraespressione del miR-17* potrebbero far considerare questi miRNA come marcatori molecolari candidati per la resistenza al Cetuximab. L'analisi funzionale globale delle network dei targets dei DE miRNA ha mostrato una significativa sovra-rappresentazione di processi biologici correlati al cancro, all' angiogenesi e alla risposta immunitaria, e di moduli centrati sui nodi critici coinvolti nell' internalizzazione di EGFR e sua degradazione ubiquitina-mediata. L'identificazione di miRNA la cui espressione e' legata all' efficacia della terapia, dovrebbe quindi consentire di predire la risposta dei pazienti al trattamento e potrebbe condurre ad una migliore comprensione dei meccanismi molecolari della risposta farmacologica. Il lavoro esposto in questa tesi e' stato pubblicato nel 2010 su Molecular Cancer Therapeutics (E-pub 29 Settembre).
The relationship between therapeutic response and modifications of miRNA transcriptome in Colorectal Cancer (CRC) remains unknown. We investigated this issue by profiling the expression of 667 miRNAs in two human CRC cell lines, one sensitive and the other resistant to cetuximab(Caco-2 and HCT-116, respectively) through TaqMan RT-PCR. Caco-2 and HCT-116 expressed different sets of miRNAs after treatment: specifically, 21 and 22 miRNAs were differentially expressed (DE) in Caco-2 or HCT-116, respectively (t-test, p<0.01). By testing the expression of DE miRNAs in CRC patients, we found that miR-146b-3p and miR-486-5p are more abundant in KRAS mutated samples respect to wild-type ones (Wilcoxon test, p<0.05). 67% of DE miRNAs were involved in cancer, including CRC, while 19 miRNA targets had been previously reported to be involved in the cetuximab pathway and CRC. We identified 25 TFs putatively controlling these miRNAs, 11 of which already reported to be involved in CRC. Based on these data, we suggest that the down regulation of let-7b and let-7e (targeting KRAS) and the up regulation of miR-17* (a CRC marker) could be considered as candidate molecular markers of cetuximab resistance. Global network functional analysis, based on miRNA targets, showed a significant overrepresentation of cancer-related biological processes and networks centered on critical nodes involved in EGFR internalization and ubiquitin-mediated degradation. The identification of miRNAs, whose expression is linked to the efficacy of therapy, should allow to predict the response of patients to treatment and possibly lead to a better understanding of the molecular mechanisms of drug response.

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Nordman, Ina IC Clinical School St Vincent's Hospital Faculty of Medicine UNSW. "Surrogate endpoints of survival in metastatic carcinoma". Publisher:University of New South Wales. Clinical School - St Vincent's Hospital, 2008. http://handle.unsw.edu.au/1959.4/42791.

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In most randomised controlled trials (RCTs), a large number of patients need to be followed over many years, for the clinical benefit of the drug to be accurately quantified (1). Using an early proxy, or a surrogate endpoint, in place of the direct endpoint of overall survival (OS) could theoretically shorten the duration of RCTs and minimise the exposure of patients to ineffective or toxic treatments (2, 3). This thesis examined the relationship between surrogate endpoints and OS in metastatic colorectal cancer (CRC), advanced non-small cell lung cancer (NSCLC) and metastatic breast cancer (MBC). A review of the literature identified 144 RCTs in metastatic CRC, 189 in advanced NSCLC and 133 in MBC. The publications were generally of poor quality with incomplete reporting on many key variables, making comparisons between studies difficult. The introduction of the CONSORT statement was associated with improvements in the quality of reporting. For CRC (337 arms), NSCLC (429 arms) and MBC (290 arms) there were strong relationships between OS and progression free survival (PFS), time to progression (TTP), disease control rate (DCR), response rate (RR) and partial response (PR). Correlation was also demonstrated between OS and complete response (CR) in CRC and duration of response (DOR) in MBC. However, while strong relationships were found, the proportion of variance explained by the models was small. Prediction bands constructed to determine the surrogate threshold effect size indicated that large improvements in the surrogate endpoints were needed to predict overall survival gains. PFS and TTP showed the most promise as surrogates. The gain in PFS and TTP required to predict a significant gain in overall survival was between 1.2 and 7.0 months and 1.8 and 7.7 months respectively, depending on trial size and tumour type. DCR was a better potential predictor of OS than RR. The results of this study could be used to design future clinical trials with particular reference to the selection of surrogate endpoint and trial size.

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12

Vargas, Ashley Joy. "Assessing the Role of Dietary Polyamines on the Continuum of Colorectal Carcinoma". Diss., The University of Arizona, 2013. http://hdl.handle.net/10150/293416.

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Putrescine, spermidine and spermine are the polyamines biosynthesized by human cells via ornithine decarboxylase (ODC) and are also sourced from the diet. Polyamines are required for malignant and normal cell growth and development. Pharmacological suppression of polyamine biosynthesis, by difluoromethylornithine, and inflammation, via sulindac, has demonstrated ~70% efficacy in preventing premalignant colorectal adenomas (CRA) in a clinical trial; however, high polyamine intakes mitigated this preventative action. Further, dietary polyamines increase the dysplasia of CRA in initiated animal models of colorectal cancer (CRC) and are hypothesized to function as tumor promoters. Human research on dietary polyamines was limited until the development of a dietary database in 2007 but, continues to be limited by the lack of a biomarker of exposure. Chapter 1 of this dissertation tests the hypothesis that dietary polyamines increase risk of CRA in polyp-formers (n = 1164) and found evidence to support this hypothesis. However, only women, younger participants and certain genotypes experienced more risk of CRA with high polyamine exposure. Chapter II tests the hypothesis that dietary polyamines increase the risk for CRC in an average risk cohort of post-menopausal women (n = 87,620) and did not find evidence to support this hypothesis in the whole population. Rather, dietary polyamines were non-significantly protective against CRC and significantly protective when paired with aspirin use and against CRC-specific death. There was some evidence to support an increase in risk of CRC in younger participants with high polyamine exposure. Overall, the first two chapters suggest that dietary polyamines protect the colorectum in normal risk individuals but promote carcinogenesis in high risk individuals. Chapter III tests the hypothesis that dietary polyamine intake correlates with urinary polyamine output in a group of overweight/obese, older men (n = 36) and Chapter IV tests the hypothesis that intake of highly ripe sweet cherries will increase urinary polyamine output in a subgroup of 10 men from Chapter III. The findings from these chapters suggest there may be a positive correlation, but that a better measure of dietary polyamine intake is needed to determine if urinary polyamines are biomarkers of exposure to polyamines.

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13

Hartley, John Edward. "An evaluation of the use of laparoscopic techniques for the resection of colorectal carcinoma". Thesis, University of Hull, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.322452.

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14

Kerr, Ian Balfour. "A study of metastasis in colorectal carcinoma using DNA recombinant and molecular hybridisation techniques". Thesis, University of Edinburgh, 1989. http://hdl.handle.net/1842/19010.

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15

Haagensen, Emma Joanne. "Pre-clinical evaluation of P13K and MEK inhibitor combinations in colorectal cancer tumour models". Thesis, University of Newcastle Upon Tyne, 2012. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.633014.

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16

Law, Kevin L. "Detection of colorectal carcinoma-associated antigens using specific antibodies against human milk oligosaccharides". Thesis, Virginia Polytechnic Institute and State University, 1986. http://hdl.handle.net/10919/54876.

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Rabbit antibodies against human milk sialyltetrasaccharide b Galßl—3[NeuAca2—6]GlcNAcßl—3Galßl—4Glc) and sialyltetrasaccharide a (NeuAca2-3Ga|ßl·3GlcNAcß1-3Galßl-4Glc) were used to detect their homologous haptens as gangliosides in the human colorectal carcinoma cell line SW'lll6. Sialyltetrasaccharide b-ceramide was detected in the monosialylganglioside fraction from human meconium and a total ganglioside fraction from SWlll6 cells on thin layer chromatograms by radioimmune staining using anti-sialyltetrasaccharide . b. Sialyltetrasaccharide b-ceramide was not detected in a total Iipid extract from normal intestinal mucosa, thus suggesting that it may represent another tumor·associated antigen. Two novel disialylgangliosides recently reported in human colonic adenocarcinoma -- disialylIactotetraosylceramide (NeuAc¤2·3Ga|ß1-3[NeuAca2·6]GlcNAcßl-3 Galßl-4Glcl-lCer) and disialyl Lea (NeuAc¤2-3Ga|B1—3[NeuAc¤2-6 (Fucal·4)]GIcNAcßl-3Ga|ßl-4Glcl·lCer) -· both contain the sialyltetrasaccharide a and b structures, either of which may represent the biosynthetic precursor of these disialylgangliosides. The anti·sialy|tetrasaccharide a antibody specifically recognizes its reduced homologous hapten and was used in a radioimmmune binding assay to detect sialyltetrasaccharide a as a reduced and tritiated, ganglioside·derived sialyloligosaccharide from SWlll6 cells. Sialyltetrasaccharide a-ceramide was recently detected in human embryonal carcinoma cells, and it is the biosynthetic precursor of the sialyl Lea antigen, a tumor-associated ganglioside in SWH16 cells. This report confirms the existence of sialyltetrasaccharide a-ceramide in SW11l6 cells.
Master of Science

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17

Du, Toit Joe-Lin. "The modulation of various signal transduction pathways in colorectal carcinoma cells by docosahexaenoic acid". Thesis, Stellenbosch : University of Stellenbosch, 2006. http://hdl.handle.net/10019.1/17350.

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Thesis (MSc)--University of Stellenbosch, 2006.
ENGLISH ABSTRACT: Introduction: The ability of different polyunsaturated fatty acids (PUFAs), especially n-3 PUFAs, to prevent the development of cancer has been under intense investigation the past three decades. Numerous studies have shown that these fatty acids can kill cancer cells in vitro as well as in vivo whilst normal cells remain unaffected. Unfortunately, the cellular and molecular mechanisms responsible for this phenomenon are still poorly understood. This study investigated the signalling pathways modulated by docosahexaenoic acid (DHA) in an adenocarcinoma cell line, in order to shed some light on these unknown mechanisms. Materials & Methods: NCM460 (normal colon epithelial) and CaCo2 (colon adenocarcinoma) cells were cultured and treated with low doses of palmitic acid (PMA), oleic acid (OA), arachidonic acid (AA), and DHA. The effects of these fatty acids on the proliferation of the cells were measured with the MTT assay. The composition of membrane phospholipids of CaCo2 cells was determined after 48h supplementation with different fatty acids by gas chromatography. Also, CaCo2 cells were treated with DHA (10 μM) only and proteins were harvested at fixed time points ranging from 2 minutes to 48 hours. The protein inhibitors wortmannin (PI3 kinase inhibitor), PD 98059 (MEK inhibitor) and SB 203580 (p38 inhibitor) and also RNA interference (RNAi) of the p38 MAPK protein were used to investigate cross-talk between signalling pathways. ERK, p38 MAP kinase, Akt, and p53 were then analysed by Western blotting using phospho-specific and total antibodies. The cleavage of the apoptotic proteins, caspase-3 and PARP were also analysed. Results and discussion: MTT assays revealed that none of the fatty acids were toxic to normal cells. In addition, DHA was shown to be most effective to kill CaCo2 cells whilst protecting NCM460 cells and a subsequent dose response experiment revealed that lower concentrations are most suitable for this purpose. DHA was also shown to be readily incorporated into phospholipids, along with AA. This is associated with increased membrane fluidity, which could affect the localisation, and downstream effects, of various signalling proteins within the membrane. Western blot analysis revealed a rapid increase in activity in most proteins under investigation, especially ERK and Akt (Ser473). Long-term DHA supplementation suppressed the full activation of Akt. This down regulation of survival signalling could lead to cell death in CaCo2 cells. In addition, it was shown that after 48h, DHA induced the cleavage of caspase-3 and PARP, which is indicative of apoptosis. RNAi experiments suggested a possible role for p38 MAPK in the phosphorylation of p53 at Ser15, a site which is associated with DNA damage. Conclusion: DHA exerts its effects by means of cellular signal transduction pathways, particularly by suppression of the important survival-related kinase, Akt. This could have implications for future therapeutic interventions in cancer patients, as fatty acids are safe to use and do not interfere with the functionality of normal tissue.
AFRIKAANSE OPSOMMING: Inleiding: Die vermoë van verskillende poli-onversadigde vetsure (POVSe), veral n-3 POVSe, om die ontstaan van kanker te voorkom, is intens nagevors die afgelope drie dekades. Menigte studies het aangevoer dat hierdie vetsure kankerselle in vitro asook in vivo kan doodmaak, terwyl normale selle nie daardeur beïnvloed word nie. Ongelukkig word die sellulêre and molekulêre meganismes onderliggend tot hierdie verskynsel nie goed begryp nie. Hierdie studie het verskeie seintransduksie-paaie wat deur dokosaheksaenoësuur (DHS) in ‘n adenokarsinoom sellyn gemoduleer word, ondersoek. Materiale & Metodes: NCM460 (normale kolonepiteel) en CaCo2 (kolon adenokarsinoom) selle is onderhou in ‘n selkultuur-laboratorium en behandel met lae dosisse palmitiensuur (PMS), oleïensuur (OS), aragidoonsuur (AS), en DHS. Die invloed van hierdie vetsure op die proliferasie van die selle is d.m.v. die MTT toets bepaal. The samestelling van membraan-fosfolipiede van CaCo2 selle is na 48h behandeling met die verskillende vetsure bepaal deur middel van gaschromatografie. Die CaCo2 selle is ook met DHA (10 μM) alleenlik behandel en teen vaste tydpunte wat wissel van 2 minute tot 48h, waarna proteïene geëkstraeer is. Die proteïen-inhibitore wortmannin (PI3 kinase inhibitor), PD 98059 (MEK inhibitor), en SB 203580 (p38 inhibitor) asook RNAinterferensie (RNAi) teen die p38 MAPK proteïen is ingespan om oorvleueling tussen seintransduksie–weë te ondersoek. ERK, p38 MAPK, Akt, en p53 is geanaliseer deur middel van die Western–klad metode met fosfo–spesifieke en totale antiliggame. Die kliewing van die apoptotiese proteïene caspase-3 en PARP is ook bepaal. Resultate en bespreking: MTT toetse het ontul dat geen vetsure toksies was vir die normale selle nie. Daar is ook gevind dat DHS die mees effektiewe vetsuur was om CaCo2 selle te dood, terwyl NCM460 selle beskerm word. Gevolglik het ‘n dosis-respons eksperiment getoon dat laer konsentrasies die beste geskik is vir hierdie doel. Daar is ook gevind dat DHA maklik in fosfolipiede geïnkorporeer word, tesame met AS. Dit word geassosieer met verhoogde membraan-vloeibaarheid, wat die ligging, en ook stroom-af werking, van verskeie seintransduksie proteïene in die membraan, kan beïnvloed. Westernklad analises het ‘n vinnige verhoging in die aktiwiteite van die meeste proteïene onder die soeklig, getoon, veral ERK en Akt (Ser473). Langdurige DHS behandeling het die maksimale aktiwiteit van Akt onderdruk. Hierdie afname van oorlewing-gerigte seine kan lei tot seldood in CaCo2 selle. Daar is boonop geving dat DHS die kliewing van caspase-3 en PARP geïnduseer het na 48, wat dui op apoptose. Uit die RNAi eksperiment kon daar ook ‘n moontlike rol vir p38 MAPK in die fosforilering van p53 by Ser15, wat geassosieer word met DNS-skade, getoon word. Gevolgtrekking: DHS beoefen sy effekte deur middel van seintransduksie paaie, veral deur die oorlewing-geassosieerde kinase, Akt, te onderdruk. Dit kan implikasies hê vir toekomende terapeutiese ingrypings in kankerpasiënte, aangesien vetsure veilig is om te gebruik en nie skadelik is vir normale weefsel nie.

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18

Enayat, Shabnam. "Investigating The Anticarcinogenic Role Of Salix Aegyptiaca L. In Colorectal Carcinoma". Master's thesis, METU, 2009. http://etd.lib.metu.edu.tr/upload/3/12610372/index.pdf.

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In this study, extracts from bark, leaves and catkins of Salix aegyptiaca L. were investigated for their antioxidant content by 2,2-diphenyl-2-picrylhydrazyl hydrate (DPPH) free radical quenching assay, total phenolic and total flavonoid assays. The highest antioxidant activity (19 ug/ml IC50 for inhibition of DPPH radical activity), total phenolic content (212 mg gallic acid equivalents/g of dried extract) and total flavonoid (479 mg catechin equivalents/g of dried extract) was observed in the ethanolic extract of bark. High performance liquid chromatography (HPLC) analyses revealed the presence of gallic acid, caffeic acid, vanillin and p-coumaric acid, myricetin, catechin, epigallocatechin gallate, rutin, quercetin as well as salicin. In addition, the anti-proliferative effects of the ethanolic extracts on colorectal cancer cell lines (HCT-116 and HT-29) were examined by an MTT cell viability assay while their apoptotic effects were assayed by acridine orange staining and caspase 3 activity. The results indicate that the ethanolic extract of bark of S. aegyptiaca can strongly inhibit cell proliferation and induces apoptosis in a dose dependent manner on both cell lines. We propose that extracts from this plant may be utilized as a source of health promoting antioxidants. Our data provide a perspective for more detailed study of biochemical pathways associated with the cancer preventive effects of active components of the extracts from S. aegyptiaca.

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19

Roberts, Kirsty Anne. "Analysis of 14-3-3σ methylation and associated changes in gene expression and function in colorectal carcinoma". Thesis, University of Edinburgh, 2010. http://hdl.handle.net/1842/4821.

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The aims of the work presented in this thesis were: to investigate the role of methylation of 14-3-3σ (a key regulator of p53-mediated G2/M arrest and of translational control during mitosis) in colorectal cancer using colorectal cancer cell lines and fresh colorectal tumours; to investigate any relationship between 14-3-3σ methylation status and gene expression; to determine whether aberrant methylation is associated with cell cycle defects and other factors known to contribute to colorectal carcinogenesis. PCR bisulphite sequencing showed that 78% (7/9) of colorectal cancer cell lines were unmethylated in the 14-3-3σ upstream promoter region (UPR). The unmethylated cell lines expressed high levels of 14-3-3σ, while methylated cell lines expressed negligible levels of 14-3-3σ protein or mRNA. Methylated colorectal cancer cell lines were treated with 5-aza-2’-deoxycytidine and demethylation was confirmed by MSP analysis. However, demethylation did not induce 14-3-3σ re-expression in the methylated cell lines, suggesting that CpG methylation may not be the only mechanism of transcriptional control. In contrast to colorectal cancer cell lines, 90% (89/99) of fresh colorectal tumours were methylated at CpG dinucleotides within the 14-3-3σ UPR. Bisulphite sequencing analysis of individual clones from 14-3-3σ methylated tumours (n =3) demonstrated that the clones displayed methylated CpG levels of approximately 41%. In agreement with previous PCR bisulphite sequencing analysis, there were a low percentage of methylated CpG dinucleotides (~ 15%) in clones from the 14-3-3σ unmethylated tumours (n =3). Unmethylated tumours expressed significantly higher levels of 14-3-3σ in comparison to methylated tumours (p =0.03), indicating that 14-3-3σ methylation may be associated with expression. PCR bisulphite sequencing analysis of matched normal mucosa tissues indicated that the 14-3-3σ UPR was methylated in all samples. Preliminary studies therefore suggest that there is tumour-specific loss of 14-3-3σ methylation in colorectal tumours within the 14-3-3σ UPR and CpG island. There were no apparent clinico-pathological correlations with 14-3-3σ methylation status. Whilst 14-3-3σ methylation was associated with expression in fresh colorectal tumours, there was no significant difference in expression levels between unmethylated colorectal tumours and matched methylated normal tissue. Bisulphite sequencing analysis of individual clones from normal tissues (from patients free of cancer) revealed that the 14-3-3σ UPR and CpG island was methylated at the majority of CpG sites analysed in colonic tissue (422/495, ~85.2%) and approximately half (795/1557, 51.1%) of CpG sites in skin samples (n =3). Furthermore, higher levels of 14-3-3σ protein were observed in skin tissue samples compared to normal colonic tissue, suggesting that 14-3-3σ CpG island methylation may be associated with tissue-specific expression. Experiments to assess the relationship between 14-3-3σ methylation and general methylation defects, suggest that methylation differences in 14-3-3σ were not simply a consequence of more general methylation phenomena well described in colorectal cancer. Nearest Neighbor analysis showed no evidence of generalised hypomethylation. Furthermore, MethyLight analysis of the CpG Island Methylator Phenotype (CIMP) showed no relationship between 14-3-3σ methylation status and CIMP; since, 1/5 (20%) tumours methylated at 14-3-3σ UPR and 1/5 (20%) tumours unmethylated at 14-3-3σ UPR were CIMP positive. In vitro functional assays showed that overexpression of 14-3-3σ in SW480 cells (14-3-3σ methylated) delayed the apoptotic response to UV-C, compared to control SW480 cells. This suggests that 14-3-3σ may protect colorectal cancer cells from apoptosis. MTT assays showed that overexpression of 14-3-3σ in SW480 cells resulted in a trend of increasing proliferation with a significant increase on day 4, compared to controls SW480 cells (p <0.01). Furthermore, FACS-sorted SW480 cells overexpressing 14-3-3σ, showed a significant shift to S-phase from G1 compared to control SW480 cells (p <0.01). Western blot analysis and immunohistochemistry revealed no relationship between p53 status and methylated 14-3-3σ in fresh tumours, while there was no relationship between published p53 status for colorectal cancer cell lines and 14-3-3σ methylation status defined experimentally. I have presented data which shows that methylation status of 14-3-3σ varies between colorectal cancer tissue, colorectal cancer cell lines and normal colonic tissue. Overexpression of 14-3-3σ appears to contribute to colorectal cancer carcinogenesis, raising the hypothesis that 14-3-3σ expression and function may at least in part be dependent on CpG methylation.

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20

Bedin, Chiara. "Utilizzo di Nanoporous Silica Chip nello studio del profilo peptidico plasmatico: applicazione nello sviluppo e progressione del cancro colorettale". Doctoral thesis, Università degli studi di Padova, 2014. http://hdl.handle.net/11577/3423727.

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Background: To date single tumour molecule identification approach for the protein biomarkers discovery is unabled to unequivocally recognize the cancer, because current tumor biomarkers are also expressed in normal cells. The protein profiling identify a specific pattern of hundred of protein in numerous specimens. Low molecular weight (LMW) proteins/peptides, the fraction less abundant of the biological fluids, seem to contain disease-specific information and correlate to the tissue pathological status. The detection by MS-analysis of low abundant fraction and LMW peptides remains a critical challenge. Aim: A Nanoporous Silica Chip (NSC) was used to select and purify LMW plasma peptides in samples of colorectal cancer (CRC) patients to study the peptide profiling in association to development and progression of tumour. Materials and Methods: NSC is a patented prototype of Prof. M Ferrari Labs (Dept. of Nanomedicine of The Methodist Hospital Research Institute, Houston TX). It is a disc of 10 cm of diameter with a superficial nanoporous silica thin film. A standardized, fast and simple protocol was validated to perform the selection of plasma LMW peptides. 34 health subjects, 27 patients with pre-cancer lesion (adenoma ), 33 patients with early stage of CRC (stage I-II) and 34 patients with late stage of CRC (stage III-IV) was enrolled for this study. MS-analysis by MALDI-TOF instrument was performed on fractionated LMW plasma samples. Data were calibrated, aligned and normalized, and then they were undergone to accurate univariate and multivariate statistical analysis to highlight and identify difference of plasma peptide profile intensity comparing the 4 study group. Results: Good classification of control group was obtained regard patient group, but poor discrimination was observed between adenoma, early stage CRC and late stage CRC. Consequently, 29 ionic species was differentially expressed in study groups. MALDI-TOF/TOF analysis was identified the aminoacid sequence of several ionic species. All of them were fragment peptide of plasma protein arising to inflammatory response and system of complement. Moreover, peptide fragments of C3f and C4-A/B could be generated from precursor peptide by endopreotease and exoprotease cleavage. A peptide fragment was originated from propeptide of ITIH4 (Inter-?-trypsin inhibitor heavy chain H4). ITIH4 was secreted to liver and it belongs to phase acute proteins involved in inflammatory responses. It may also play a role in liver development and regeneration. Some studies were identified some ITIH4 peptide fragments involved in several pathological status as ischemic stroke, breast, ovarian and prostate cancer. Conclusions: A fast and simple method was set to perform study with NSC. NSC is a new tool with wide potential application. A deep study of obtained peptide proteolytic pattern was necessary suggesting a colorectal cancer specific proteases and exoprotease activity.
Background: Per la ricerca di biomarcatori proteici, l’approccio basato sulla identificazione di un singolo marcatore hanno, finora, dimostrato l’incapacità di individuare inequivocabilmente il cancro, in parte perché gli attuali biomarcatori tumorali sono anche espressi nelle cellule normali. Il profilo proteico si basa, invece, sulla rilevazione di pattern identificativo di centinaia di proteine in un alto numero di campioni. Il contenuto informativo più alto sembra risiedere nelle proteine/peptidi a basso peso molecolare (LMW), la frazione meno abbondante nei fluidi biologici, che sembrano rispecchiame meglio gli stati fisiopatologici dei tessuti. Per l’analsi in spettrometrometria di massa e’ necessario selezionare e arrichire questa frazione del plasma. Scopo: Si è focalizzato sull'utilizzo di un dispositivo nanoporoso (Nanoporous Silica Chip, NSC) per il recupero della frazione a basso peso molecolare da plasma, in campioni di pazienti con cancro colorettale (CRC) a vari stadi di progressione tumorale e sullo studio del relativo profilo peptidico mediante tecnica MALDI-TOF. Materiale e metodi: NSC è un prototipo creato e brevettato dal Laboratorio del Prof. M. Ferrari (Dip. di Nanomedicina presso The Methodist Hospital Research Institute, Houston, TX, USA) costituito da un supporto in silicone, di circa 10cm, rivestito da un sottile strato di silice con una struttura a nano-pori. È stato necessario lo sviluppo di un protocollo semplice e veloce di frazionamento dei peptidi plasmatici. Il protocollo standardizzato è stato applicato per il frazionamento di campioni di plasma di 34 soggetti sani (controlli), 27 con lesione pre-cancerosa (adenoma) e 33 con CRC a stadio precoce (stadio I-II) e 34 con CRC a stadio tardivo (stadio III-IV). La frazione ottenuta è stata analizzata con spettrometria di massa MALDI-TOF e i dati calibrati, allineati e normalizzati sono stati sottoposti ad attenta e accurata analisi statistica univariata e multivariata con lo scopo di identificare differenze nel profilo peptidico plasmatico nei diversi gruppi di soggetti. Risultati: Si è ottenuta una buona classificazione dei controlli rispetto ai pazienti, ma una scarsa distinzione tra i gruppi di soggetti con adenoma, CRC con stadio precoce e tardivo. Da tale analisi, si sono individuate alcune specie ioniche rappresentate con diversa intensità nei vari gruppi, che sono state sottoposte a identificazione della sequenza amminoacidica mediante MALDI-TOF/TOF. Sono risultate essere tutte frammenti peptidici di proteine plasmatiche appartenenti alla risposta infiammatoria e al sistema del complemento. In particolare i frammenti peptidici del C3f e C4-A/B sembrano originare dal proprio precursore a seguito di tagli enzimatici a carico di endoproteasi ed esoproteasi. La presenza di alcuni di questi frammenti è risultata essere variabile nei gruppi in esame. Inoltre si è identificato un interessante frammento peptidico che deriva dal propeptide di ITIH4 (Inter-?-trypsin inhibitor heavy chain H4), una proteina di fase acuta secreta dal fegato e coinvolta nello sviluppo e rigerazione epatica. Una ricerca bibliografica ha identificato che frammenti di ITIH4, possono essere possibili biomarcatori per vari stati patologici come infarto e cancro alla mammella e prostata. Conclusioni: La metodica ottimizzata è semplice e veloce e NSC è un dispositivo con ampie potenzialità di utilizzo. È necessario approfondire lo studio sul particolare pattern proteolitico di peptidi ottenuti, che suggerisce il coinvolgimento di un'attività esoproteasica ben distinta e la presenza di proteasi specifiche del tumore del colon-retto.

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21

Pizzini, Silvia/P S. "A bioinformatic approach to the study of gene, microRNA expression and alternative splicing regulation in colorectal cancer progression and liver metastasis". Doctoral thesis, Università degli studi di Padova, 2012. http://hdl.handle.net/11577/3422457.

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Expression profiles are increasingly used in cancer research, and a growing number of microarray data is available in public databases such as Gene Expression Omnibus (GEO) and ArrayExpress. Therefore, it is possible to compare studies with similar research aims through a meta-analysis approach. Colorectal cancer (CRC) is a very common disease and represents a fundamental biological model of tumorigenesis. Many experiments comparing the expression profiles of normal colonic mucosa and colorectal cancer (CRC) samples have been conducted and results are available. Using A-MADMAN, a web application that allows the retrieval, annotation, organization and analysis of public available gene expression datasets, we downloaded from the GEO and collected the largest-collected normal colon, adenoma and primary colorectal tumor, with and without metastasis, gene expression dataset which includes 1,795 experiments pertaining to 27 distinct series, obtained from two different generations of Affymetrix, HG-U133A and HG-U133 plus2.0. After we performed the quality control of the downloaded data and have designed three different workflows for signal reconstruction, we conducted preliminary analysis of differential expression. In this first project we focused on right and left colon differences. In particularly, we analyzed normal, adenoma and tumor expression profiles focusing on changes among topologically different samples, under the hypothesis that the use of site-specific normal tissue as reference for the study of right and left tumor samples may facilitate to shed light on specific cell functions, pathways and regulatory circuits altered in each tumor type, and help deepening our understanding of this complex disease. From preliminary data, we concluded that the workflow based on the virtual chips production and on a chip-specific background correction is the most robust. Higher numbers of differentially expressed genes were identified when comparing, respectively, tumor and adenoma versus normal tissue (647 and 683 genes). These tissue samples were then pairwise compared by topology to identify those genes that are differentially expressed, when the matched by topology set of normal samples is considered. We found 72 genes colon left-specific and 1183 colon-right specific in the comparison from adenoma with normal tissues, and 46 genes and 69 genes, respectively, in colon left- and colon right- specific in the comparison from CRC and adenoma tissues. We focused also to the functional information associated to gene expression differences between tumor and normal tissue samples, compared considering only right or left samples, to identify specific functions and pathways involved with cancer present, such as DNA repair, adhesion and cell interaction. In the second project we reconstructed regulatory networks of colon cancer by integrating gene expression, alternative splicing and microRNA (miRNAs) expression data. We carried out a genome-wide integrative analysis of miRNA and genes and exon expression profiles in tissues of 55 colon cancer patients, comprehensive of normal colon mucosa, primary tumor and liver metastasis. GeneChip Human Exon 1.0 ST (Affymetrix) and Affymetrix GeneChip miRNA arrays have been used to obtain high quality gene, exon and miRNA expression quantification respectively. We analyzed both differential gene expression and alternative splicing applying AltAnalyze software to exon-level analysis, identifying 33,740 genes involved in alternative splicing events. We considered a custom set of expression signals of 449,810 probesets, using MIDAS and FIRMA statistics, and we combined the results with MMBGX software output, to identify a more restricted, but perhaps more robust set of candidate alternative splicing events, possibly relevant for colon cancer biology. When comparing liver metastasis with normal colon tissue whereas 182 genes were identified; comparable numbers of genes were identified when comparing metastasis versus colorectal tumor and colorectal tumor versus normal tissue (51 and 10 respectively). We validated two genes with alternative splicing events, VCL and CALD1. We then identified respectively 62, 63 and 11 differentially expressed miRNAs in tumors and metastases compared to normal tissue, and comparing metastasis with primary tumors. To assess the reproducibility and robustness of the miRNA signature identified, we measured by qRT-PCR the expression of 5 miRNA in all samples (hsa miR-150, hsa miR-10b, has miR-146a, miR-210 and has miR-122). For each considered contrast, KEGG pathways enriched in genes resulting supported target of differentially expressed miRNAs were identified. The integrated analysis of miRNA and genes expression profiles with target predictions was performed with the purpose of reconstruct post-transcriptional regulatory networks governing tumor and metastases development. Considering expression profiles in the same set of samples of 305 miRNAs and 12,748 genes with variable expression profile, we reconstructed post-transcriptional regulatory networks involving modulated miRNAs. In particular, considering the network associated to the tumor versus normal contrast, we experimentally validated miR-145 – c-Myc and miR-182 – ENTPD5 relationships. This latter is new and may have a relevant pathogenetic role. Looking at our results, we can say that the regulatory signatures that affect tumor progression are complex and difficult to interpret. They involve interactions involving different modulated miRNAs that control the expression of multiple genes belonging to the same pathway in different ways, and alternative transcripts of genes that are differentially expressed in normal tissues, tumors and metastases.
Nella ricerca oncologica sono sempre più utilizzati i profili di espressione genica e un numero crescente di dati di microarray è disponibile in database pubblici come NCBI GEO e ArrayExpress. Diventa quindi possibile il confronto di studi con obiettivi di ricerca simili attraverso approcci di “meta-analisi”. Il cancro colorettale (CRC) rappresenta un fondamentale modello biologico di tumorigenesi, oltre ad essere una patologia molto diffusa. Sono a disposizione nei database pubblici molti esperimenti sul CRC, che confrontano mucosa normale con mucosa tumorale del colon attraverso metodologia microarray. Noi abbiamo utilizzato A-MADMAN, un’applicazione web open source di supporto per la meta-analisi di dati grezzi d’espressione genica ottenuti con microarray, per scaricare dal database dell’NCBI Gene Expression Omnibus (GEO), 27 collezioni di esperimenti per un totale di 1045 campioni ottenuti da mucosa normale, adenoma e CRC con e senza metastasi di colon-retto su cui erano state eseguite analisi di espressione genica con tecnologia gene chip Affymetrix. Dopo aver effettuato un controllo di qualità dei dati scaricati e aver disegnato 3 diversi flussi di lavoro per la ricostruzione del segnale d’espressione, sono state condotte delle analisi preliminari di espressione differenziale. In questo primo progetto ci siamo focalizzati sulle differenze molecolari sito–specifiche. Abbiamo quindi analizzato campioni di mucosa sana, di adenoma e di CRC suddivisi per localizzazione in colon destro e colon sinistro. Prendendo come riferimento il tessuto normale destro e sinistro, abbiamo ipotizzato che è possibile discriminare pattern di espressione genica tumorale sede specifica. Dal disegno dei tre flussi di lavoro abbiamo dedotto che il flusso di lavoro basato sulla generazione di chip virtuali e su una correzione del background chip-specifica è il più affidabile. Abbiamo identificato 647 geni differenzialmente espressi confrontando tessuto tumorale con tessuto normale e 683 geni nel confronto adenoma con tessuto normale, poi abbiamo individuato 72 geni nel confronto adenoma-tessuto normale specifici del colon sinistro e 1183 geni nello stesso confronto, colon destro specifici, mentre per quanto riguarda il confronto CRC e adenoma, 46 geni sono colon sinistro specifici e 69 sono colon destro specifici. Abbiamo inoltre trovato termini funzionali e pathway diversi sovra-rappresentati in colon destro rispetto a colon sinistro e rispetto ai corrispondenti normali. Nel secondo progetto, abbiamo ricostruito una rete regolatoria riguardante il CRC e la metastasi epatica da CRC integrando dati di originali di espressione genica, di splicing alternativo e di espressione di microRNA (miRNA). Lo studio si basa sulla raccolta di biopsie di tumore primario al colon, mucosa adiacente normale e metastasi al fegato di 55 pazienti, che sono state analizzate utilizzando piattaforme Affymetrix per l'analisi di espressione genica/esonica (GeneChip Human Exon 1,0 ST), e di microRNA (GeneChip® miRNA Array ). Analizzando l’ espressione differenziale a livello genico ed esonico mediante il software AltAnalyze identificando 33.740 geni coinvolti in probabili eventi di splicing alternativo. Integrando i dati risultati dalle statistiche di questo programma con i risultati ottenuti con un software basato su un approccio Bayesiano, MMBGX, abbiamo identificato una lista più ristretta ma anche più robusta di candidati eventi di splicing possibilmente rilevanti per la formazione e la progressione del CRC. Confrontando metastasi epatiche con tessuto normale del colon sono stati identificati 182 geni, mentre un numero inferiore di geni sono stati identificati nel confronto contro le metastasi del tumore colorettale e del tumore del colon-retto rispetto al tessuto normale (51 e 10 rispettivamente). A partire da questi risultati, abbiamo scoperto il coinvolgimento di trascritti alternativi di due geni, VCL e CALD1, nella progressione tumorale. In parallelo, sono stati identificati microRNA modulati in seguito allo sviluppo del tumore e della metastasi, trovandone rispettivamente 62, 63 e 11 differenzialmente espressi nel tumore rispetto al normale, nella metastasi rispetto al normale e nella metastasi rispetto al tumore. Abbiamo quindi confermato la robustezza dei risultati validando cinque miRNA presenti nella lista dei differenzialmente espressi (hsa miR-150, hsa miR-10b, has miR-146a, miR-210 and has miR-122) mediante RT-PCR. Per ogni contrasto considerato, sono stati identificati i KEGG pathway modulate e quindi sotto putativamente controllate dai miRNA. Grazie all’analisi integrata di profili di espressione di miRNA e dei loro geni target anti-correlati, sono state definite le principali reti di regolazione post-trascrizionale coinvolte nella cancerogenesi. Particolarmente rilevante è la rete che coinvolge il sottoinsieme dei miRNA differenzialmente espressi nel tumore rispetto al normale. Tra le interazioni inferite, abbiamo convalidato sperimentalmente le relazioni miR-145 - c-Myc e miR-182 - ENTPD5. Quest’ultima rappresenta una relazione nuova, il cui ruolo patogenetico può essere rilevante. Dai nostri risultati possiamo concludere che le vie di regolazione che interessano la progressione tumorale sono complesse e difficili da interpretare, che implicano interazioni che coinvolgono miRNA diversamente modulati che agiscono in diversi modi sull’espressione di più geni appartenenti ad una stessa pathway e da trascritti alternativi di geni che vengono espressi in modo differenziale nei tessuti sani, nei tumori e nelle metastasi.

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Aldecoa, Ansórregui Iban. "Caracterización de la estadificación molecular en carcinoma de colon. Correlación clínico-histológica". Doctoral thesis, Universitat de Barcelona, 2016. http://hdl.handle.net/10803/401651.

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ANTECEDENTES: El tratamiento del carcinoma colorectal (CCR) en estadios I-II es quirúrgico, aunque un 25% de pacientes pueden recidivar. El estadio ganglionar es un factor pronóstico independiente y determina el tratamiento quimioterápico. El estadiaje ganglionar histológico (pN) se realiza con tinción de hematoxilina-eosina (HE), método poco sensible para detectar pacientes con riesgo de recidiva. Las técnicas moleculares detectan células tumorales en el 25-50% de los ganglios linfáticos (GLs) de CCR que se han diagnosticado como negativos por HE, las cuales se asocian en el CCR precoz a mayor riesgo de recidiva y peor supervivencia. Sin embargo, el diagnóstico molecular es complejo y costoso, impidiendo su uso en la práctica clínica. OBJETIVOS, ESTUDIOS REALIZADOS Y RESULTADOS OBTENIDOS: El primer estudio que compone la tesis doctoral es multicéntrico y prospectivo. Tiene como objetivo determinar la relación entre la carga tumoral molecular en GLs y los factores de riesgo convencionales en pacientes con cáncer de colon en estadios I-II. Se obtuvieron 1940 GLs de 149 pacientes con cáncer de colon con estadio histológico pN0. Se cuantificó la cantidad de ARNm mensajero (ARNm) de citoqueratina 19 (CK19) en los GLs mediante la técnica Reverse Transcription Loop-Mediated Isothermal Amplification (RT-LAMP) denominada One-Step Nucleic Acid Amplification. Se definió la carga tumoral total (CTT) de cada paciente como la suma de todas las copias de ARNm de CK19/μL de cada GL positivo por colectomía. Se obtuvo una mediana de 15 GLs por caso (RIC 12;20). La positividad molecular se correlacionó con la presencia de áreas de alto grado (p <0,01), histología mucinosa/anillo de sello (p = 0,017), sexo masculino (p = 0,02), número GLs aislados (p = 0,012) y el peso total de GLs por caso (p < 0,01). La CTT se relacionó con el estadio pT (p = 0,01) y tamaño tumoral (p <0,01) en los tumores de bajo grado. El estudio de regresión logística multivariante mostró una correlación independiente de positividad molecular con el género, grado tumoral y número de GLs en fresco [AUC = 0,71 (IC del 95% = 0,62 a 0,79)]. El objetivo del segundo estudio es evaluar el impacto del tatuaje endoscópico prequirúrgico en la detección molecular de carga tumoral en GLs de neoplasias de colon en estadios iniciales. Métodos: Se trata de un estudio de cohorte prospectivo basado en una población de cribado de CCR en un hospital universitario terciario. Se evaluaron los GLs de colectomías con y sin tatuaje endoscópico preoperatorio mediante dos métodos, HE y RT-LAMP. Se comparó la cantidad de la carga tumoral y los valores de GLs obtenidos entre especímenes tatuados y no tatuados. Resultados: Se evaluaron mediante HE y RT-LAMP 936 GLs obtenidos de 71 colectomías que contenían carcinomas precoces y adenomas endoscópicamente irresecables (8 pT0, 17 pTis, 27 pT1, 19 pT2); 47 de 71 casos (66,2%) estaban tatuados. La positividad molecular en GLs se correlacionó con la presencia de tatuaje ganglionar [p <0,001; odds ratio 3.1 (95% IC 1.7 a 5.5)]. Se obtuvo un número significativamente mayor de GLs en especímenes tatuados en comparación con los no tatuados (mediana 17 GLs vs. 14,5 GLs; p = 0,019). CONCLUSIONES: Los resultados de los estudios de la presente tesis doctoral muestran que la detección de ARNm de CK19 en ganglios linfáticos se correlaciona con factores de alto riesgo clásicos en pacientes con cáncer de colon en estadio I-II. La CTT es una medida cuantitativa y objetiva que puede contribuir a una mejor estadificación de pacientes con cáncer de colon precoz. Asimismo, el tatuaje endoscópico permite la detección de los ganglios linfáticos más proclives a albergar células tumorales y aumenta el número de ganglios linfáticos aislados.
Stage I–II colorectal cancer patients are surgically treated although up to 25 % will recur from disease. Molecular tumour detection in LN of early-stage patients is associated with an increased risk of disease recurrence and poor survival. The first study is a prospective multicentre study aimed to determine the relationship between LN molecular tumour burden and conventional high-risk factors in stage I–II colon cancer patients. A total of 1940 LN from 149 pN0 colon cancer patients were analysed for the amount of tumour CK19 mRNA with the quantitative OSNA molecular assay. Patient’s total tumour load (TTL) resulted from the sum of all CK19 mRNA tumour copies/μL of each positive LN from the colectomy specimen. A median of 15 LN were procured per case. Molecular positivity correlated with high-grade, mucinous/signet ring type, male gender, number of collected LN and total LN weight per case (p≤0.02). The TTL was related to pT stage and tumour size in low-grade tumours (p≤0.01). Multivariate logistic regression showed independent correlation of molecular positivity with gender, tumour grade and number of fresh LN (AUC=0.71). The second study aimed to evaluate the impact of presurgical endoscopic tattooing in early colon neoplasms (i.e. amount of tumour burden in LNs and LN yields). A prospective cohort study from a CRC screening-based population was performed. LNs from colectomies with and without preoperative endoscopic tattooing were assessed by HE and OSNA. HE and OSNA analyses of 936 LNs were performed from 71 colectomies containing early carcinomas and endoscopically unresectable adenomas; 47/71 cases were tattooed. Molecular positivity correlated with the presence of tattoo in LN (p<0.001; OR 3.1). A significantly higher number of LNs were obtained in tattooed specimens (median 17 LN vs. 14.5 LN; p=0.019). In conclusion, lymph node CK19 mRNA detection correlates with classical high-risk factors in stage I–II colon cancer patients. TTL is a quantitative and objective measure that may help to better stage early colon cancer patients. Additionally, endoscopic tattooing enables the analysis of those LNs most prone to harbour tumour cells and improves the number of LN harvested.

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Libanje, Fotine. "Etude de la contribution des voies de signalisation dépendantes des RhoGTPases à l'invasion collective des carcinomes colorectaux". Thesis, Université Paris-Saclay (ComUE), 2017. http://www.theses.fr/2017SACLS503/document.

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La progression métastatique des cancers est responsable de 90% des décès liés à la maladie. Cette cascade est initiée par l’invasion des cellules cancéreuses du stroma péritumoral, et conduit à leur dissémination dans l’organisme.Mon travail de thèse a eu pour but d’identifier les mécanismes moléculaires et cellulaires régulant l’invasion des cancers colorectaux (CRC), qui est le 2ème cancer le plus répandu dans le monde. Grâce à une analyse réalisée sur des échantillons humains de tumeurs primaires, nous avons révélés que les cellules de CRC utilisent un mode d’invasion collective dans lequel elles gardent une architecture glandulaire spécifique des épithelia. Afin d’étudier les voies de signalisation régulant cette invasion, nous avons utilisé des modèles organotypiques récapitulant l’architecture des glandes de CRC (cystes de Caco-2 et tumoroïdes de xénogreffes derivés de patients) dans des tests d’invasion utilisant du collagen-I. Du fait de son rôle central dans la régulation de la motilité cellulaire, la voie des RhoGTPases était un bon candidat à la régulation de l’invasion collective des CRC. Dans un screen utilisant des siRNA ciblant tous les effecteurs connus des RhoGTPases, seule la déplétion des protéines kinases ROCK a déclenché l’invasion collective dans notre système expérimental. Nous avons démontré que l’inhibition de ROCK2 et non de ROCK1 était suffisante pour induire la formation de cellules leader, permettant la polarisation leader/follower requise pour l’invasion collective. Nos résultats montrent que l’inhibition de ROCK2 déclenche l’invasion collective par l’inhibition de MyosinII combinée à l’activation du facteur d’échange nucléotide guanine (GEF), FARP2, et de RAC1. Notre étude permet donc d’identifier FARP2 comme un nouvel effecteur de ROCK2 et le positionne comme un nouveau médiateur du crosstalk entre RhoA et RAC1 dans la régulation de l’invasion collective des CRC. En conclusion, nous avons décrit une nouvelle voie de signalisation dépendante de ROCK2 contrôlant l’invasion collective de glandes de CRC. De façon intéressante, notre étude révèle un rôle anti-invasive de ROCK2 contredisant son rôle pro-invasive décrit dans l’invasion de cellules individuelles. Cela suggère que ROCK2 assure des rôles distincts en fonction du mode d’invasion adopté par les cellules cancéreuses et remet en question le bénéfice thérapeutique de l’inhibition de ROCK proposé pour bloquer l’invasion des cellules cancéreuses
Metastatic progression of cancer is responsible for 90% of the disease related death. It is a multi-step process which is initiated by invasion of the peritumoral stroma by cancer cells and which leads to the dissemination of cancer cells in the organism.My PhD work aimed at identifying the molecular and cellular process driving colorectal carcinoma (CRC) invasion, which is the 2nd most frequent cancer worldwide. Our analysis of live and human primary cancer specimen revealed that CRC cells used a collective mode of invasion to disseminate, in which cells retain an epithelium specific -glandular architecture. To investigate the signaling pathways regulating this mode of invasion, we used 3D organotypic models recapitulating the features of CRC glands (Caco-2 cysts and Patient derived Xenografts (PDX) tumoroids) in collagen-I based organotypic invasion assays and in microscopy-based analyses. Because of its central role in the regulation of cell motility, we postulated that RhoGTPases signaling pathways could control the collective of CRC. In a siRNA based- screen targeting all the known effectors of RhoGTPases we found that only ROCK kinases downregulation induced collective invasion in our experimental settings. We demonstrated that ROCK2 but not ROCK1 inhibition was sufficient to promote Leader cell formation, which induced the leader/follower polarization necessary for collective invasion. Our results revealed that ROCK2 inhibition triggered collective invasion through the concomitant inhibition of MyosinII and activation of the guanine nucleotide exchange factor (GEF) FARP2 and the RhoGTPase RAC1. We therefore identify FARP2 as a new effector of ROCK2 and a mediator of the RhoA-RAC1 crosstalk in the regulation of collective invasion. In conclusion our study proposes a new ROCK dependent-signaling pathway in the regulation of collective invasion of highly polarized CRC glands models. Importantly, we found ROCK2 to be an anti-invasive protein which is in contradiction with its described pro-invasive role in single cell invasion. This suggests distinct roles of ROCK which may depend on the mode of invasion adopted by the cells and questions the benefice of proposed ROCK inhibition strategies to block cancer cell invasion

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Wong, Chi-wai y 黃志偉. "High resolution mapping of loss of heterozygosity and chromosomal aberrations using oligonucleotide single nucleotide polymorphismgenotyping arrays in colorectal adenoma to carcinoma progression". Thesis, The University of Hong Kong (Pokfulam, Hong Kong), 2006. http://hub.hku.hk/bib/B3871923X.

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Zhang, Yuan. "Circadian clocks and cancer : The implication of BMAL1 (brain and muscle Arnt-like protein-1) in colorectal and breast carcinoma development and treatment". Thesis, Université Paris-Saclay (ComUE), 2019. http://www.theses.fr/2019SACLS422.

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BMAL1, une protéine centrale de l'horloge circadienne.L’inactivation de BMAL1 (BMAL1-KO) entraîne une perte complète de la rythmicité dans les horloges central et périphérique. Le travail de ma thèse se concentre sur le rôle du gène BMAL1 dans la développement et le traitement des cancers du sein et du côlon.1. Pharmacodynamique in vitro de l’Everolimus en fonction du temps d’administration malgré une horloge circadienne défectueuse ((Zhang et al., 2018) (Zhang, Levi and Chang, 2018)L’everolimus (EV) est un inhibiteur de la mTOR chez les mammifères et il est utilisé pour traiter le cancer du sein positif aux oestrogènes (ER+). Nous avons focalisé nos recherches sur la chronopharmacologie de l’Everolimus administré sur des cellules MCF-7 (ER+). Les MCF-7 présentent une oscillation circadienne de l’activité de mTOR sans mise en évidence d’une oscillation des gènes d’horloge. L’oscillation d’activité de mTOR induirait une oscillation de synthèse et/ou de phosphorylation de protéines importantes dans la progression de la phase G1, notamment la Cycline D1 et RB phosphorylée. Ces variations rythmiques des MCF-7 synchronisées expliquent la chrono-efficacité de l’Everolimus selon des temps différents d’administration.Ce travail a révélé que même dans un système de cellules cancéreuses dont l’horloge était perturbée, l'intégration d'autres rythmes cellulaires dans la chronothérapie pouvait augmenter l'efficacité du médicament. Ce principe peut être appliqué à des traitements du cancer pour optimiser la chronothérapie du cancer.2. Le Knockdown BMAL1 a déclenché différents destins de cellules du carcinome du côlon (CRC) en modifiant l'équilibre délicat entre les voies AKT / mTOR et P21 / P53 (Article soumis)Premièrement, nos résultats ont révélé que le knockdown BMAL1 par le shRNA (BMAL1-KD) avait déclenché une activation plus évidente de l’AKT / mTOR dans deux lignées cellulaires primaires (HCT116 et SW480) que une lignée métastatique de CRC, SW620. De plus, bien que les deux lignées cellulaires primaires de CRC aient présenté une augmentation significative de l'activité de l'AKT/mTOR, elles avaient des statuts différents de P53 (WT ou mutant). Dans ce contexte, les cellules SW480 BMAL1-KD avec P53 mutant présentaient une sénescence accrue, mais les cellules HCT116 BMAL1-KD avec P53 WT présentaient d’abord une apoptose transitoire, puis un taux de prolifération plus élevé.Ainsi, nos travaux ont révélé le rôle crucial de BMAL1 pour équilibrer un régulateur central du métabolisme AKT / mTOR et une voie de réponse au stress P53 / P21 dans des lignées cellulaires de CRC, ce qui met en évidence l’importance de BMAL1 dans le développement de CRC et la progression du vieillissement.3. BMAL1 renforce les propriétés épithéliales et diminue la chimiorésistance des cellules du CRC (article en préparation)La transition épithélo-mésenchymateuse (EMT) est un événement critique dans l'invasion et la métastase des carcinomes, y compris le CRC.Dans ce travail, nous avons étudié comment BMAL1 knockdown (Bmal1-KD) altère l’équilibre délicat entre les propriétés épithéliales et mésenchymateuse de trois lignées cellulaires de CRC (HCT116, SW480 et SW620).Après BMAL1-KD, la diminution de l’expression Twist, un facteur de transcription favorisé l’EMT et des marqueurs mésenchymateux (N-Cadhérine, Vimentine) étaient associées à une expression accrue des marqueurs épithéliaux (E-cadhérine, CK20 et EpCAM). De manière constante, l'augmentation de l'expression de l’E-cadhérine après BMAL1-KD était accompagnée d'une co-localisation membranaire accrue de la β-caténine avec l'E-cadhérine, ainsi que d'une diminution de la localisation nucléaire de la β-caténine, suggérant une diminution de l'activation de la voie Wnt. De plus, les cellules BMAL1-KD ont montré une diminution des capacités de migration et de la résistance aux médicaments.Au total, ces données soulignent l’importance de BMAL1 dans l’EMT des cellules de CRC
BMAL1 is a core circadian clock protein, forming a heterodimer with CLOCK to initiate the transcription of circadian and output genes. Among canonical clock genes, only BMAL1 knockout results in complete loss of rhythmicity in both the SCN and peripheral tissues. My thesis work focuses on exploring the important role of BMAL1 in human breast and colon cancer progression and treatment. My work is divided into three main parts:1. Dosing time dependent in vitro pharmacodynamics of Everolimus despite a defective circadian clock (Zhang et al., 2018)(Zhang, Levi and Chang, 2018) Everolimus (EV) is an inhibitor of mammalian target of Rapamycin (mTOR) and is used to treat estrogen positive (ER+) breast cancer. Here, we investigated whether EV efficacy varied according to administration timing by using the ER+ breast cancer cell line MCF-7 as a model system. Serum shock synchronization induced a circadian oscillation in mTOR activity in MCF-7 cells, which rhythmically regulated the synthesis or phosphorylation of key G1 progression proteins, such as Cyclin D1 and phosphorylated RB, ultimately resulting in different G0/G1 blockage efficiency according to different EV administration timing. Thus, the different delivery schedule of EV presented different efficacy in G0/G1 phase blockage in serum shocked MCF-7 cells.This investigation revealed that, even in a breast cancer cell system with disrupted circadian organization, modulating drug administration according to other protein rhythms could still increase drug efficacy. This principle may be applied to many other cancer systems and treatment types to optimize cancer chronotherapy.2. Knockdown BMAL1 triggered different colon carcinoma cells fates by altering the delicate equilibrium between AKT/mTOR and P21/P53 pathways (Article in preparation)We tried to evaluate in vitro how knockdown BMAL1 (BMAL1-KD) by shRNA influences human colorectal cancer cell (CRC) behavior.The results revealed that BMAL1-KD triggered different CRC cell fates based on distinct p53 status in different cell lines. First, after BMAL1 knockdown, two primary CRC cell lines (HCT116 and SW480) presented a more evident AKT/mTOR activation than the metastatic colon carcinoma cell line, SW620. Furthermore, although both primary CRC cell lines presented a significant increase of AKT/mTOR activity, they had different P53 status (WT or mutant) and activation pattern. Under these context, SW480 BMAL1-KD cells exhibited increased senescence but HCT116 BMAL1-KD cells showed firstly a transient apoptosis and then higher proliferation rate.Thus, our work uncovered the crucial role of BMAL1 to balance a central metabolism regulator AKT/mTOR and a stress response pathway P53/P21 in CRC cell lines, which highlighted the importance of BMAL1 in CRC development and aging progression.3. BMAL1 knockdown leans epithelial–mesenchymal balance toward epithelial properties and decreased the chemoresistance of colon carcinoma cell (Article in preparation)Epithelial-mesenchymal transition (EMT) is a critical early event in the invasion and metastasis of carcinoma, including colorectal cancer (CRC). In this work, we studied how BMAL1-KD alters the delicate equilibrium between epithelial and mesenchymal properties of three colon carcinoma cell lines (HCT116, SW480 and SW620).The results showed the molecular alterations after BMAL1-KD promote mesenchymal-to-epithelial transition-like changes mostly appeared in two primary CRC cell lines (HCT116 and SW480) compared to the metastatic cell line SW620. Subsequently, BMAL1-KD HCT116 and SW480 cells harbored a decreased migration, invasiveness and drug resistance capacities relative to their scramble counterpart cells. All these data suggested the importance of BMAL1 on EMT inducing in colon carcinoma cells

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Carluccio, S. "GENERATION OF TUMOR-SPECIFIC CYTOTOXIC T-LYMPHOCYTES FROM PEROPHERAL BLOOD OF COLORECTAL CANCER PATIENTS FOR ADOPTIVE T-CELL TRANSFER". Doctoral thesis, Università degli Studi di Milano, 2014. http://hdl.handle.net/2434/231155.

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Colorectal cancer (CRC) is the third most common cancer worldwide and the fourth most common cause of death in the developed Western countries. Adoptive T-cell transfer (ACT) refers to an immunotherapeutic approach in which anti-tumor T lymphocytes, usually the tumor infiltrating lymphocytes (TIL), are identified, grown ex vivo and then re-infused into the cancer patient. ACT of EBV-specific T-cell lines and T Cytotoxic Lymphocytes (CTLs) for the therapy of EBV-induced lymphomas is the best demonstration of clinically efficacious ACT, but there are many evidences also for leukemia and multiple myeloma. As regards to the solid tumors, ACT using autologous TIL, grown ex-vivo and then re-infused into the cancer patient, has emerged as an effective treatment for metastatic melanoma and renal cell carcinoma (RCC), that are the most immunogenic tumors in humans. Randomized clinical trials are ongoing for gastric cancer, hepatocellular carcinoma and lung cancer. These approaches mainly use the TIL and the definition of tumor associated antigen (TAA), tumor specific antigen (TSA) or cancer testis antigen (CTA), that are generally correlated with tumor progression and immunogenicity in various types of cancer. However these antigens are often found to be poorly expressed in CRC, and few is known about their relationship with this type of neoplasia. In addition, although a clear association between TIL and clinical outcome of CRC has been documented, active and adoptive immunotherapy do not play yet an important role in the treatment of advanced CRC. In order to develop an ACT protocol for CRC treatment, we designed an experimental approach that does not require neither the definition of molecular defined tumor antigens, nor the availability of TIL. Our strategy was based on the in vitro stimulation of patient’s CD8+-enriched T-cells from peripheral blood mononuclear cells (PBMCs) with dendritic cells (DCs), pulsed with apoptotic tumor cells as a source of tumor antigens, in order to generate autologous CTLs with strong anti-tumor activity. In this study, 78 CRC patients were enrolled. Tumor biopsies were obtained at surgery, together with 100 ml of heparinized peripheral blood (PB). Tumors were mechanically dissociated to a single-cell suspension and cultured to obtain tumor cell line from each patient. DCs were generated from previously separated PBMCs, using a magnetic positive selection of CD14+ monocytes, cultured in presence of recombinant human Interleukin-4 (rh IL-4) and recombinant human Granulocyte-Macrophage Colony-Stimulating Factor (rh GM-CSF). Anti-tumor CTLs were elicited in co/micro-culture using DCs as antigen-presenting cells, autologous apoptotic tumor cells as source of antigens and T CD8+ lymphocytes enriched effectors, with weekly stimulation. CTLs Interferon-γ (IFN-γ) secretion was assessed by ELISpot assay to evaluate their activation in response to autologous tumor. Tumor cell lines were obtained from 20 out of 78 patients (25,6%), because gut intestinal flora had adversly affected the establishment of primary tumor cell line and a loss of expansion of tumor cells was observed. DCs were generated from 26 patients, but only 6 patients had the corresponding tumor cell line, indispensable for the co-culture setting up. This was the reason why co/micro-cultures were set up only for 6 patients. ELISpot assay was performed at the end of co/micro-culture stimulations to evaluate effectors IFN-γ secretion. ELISpot results showed that strong and significant IFN-γ secretion was detected at the third, fourth and fifth stimulations for one patient and at the second for another patient, whereas for three patients a weak secretion was detected during the second and third stimulations. Although our immunological study must be performed on an increased number of CRC patients, and the CTLs expansion, together with CTLs lytic ability against autologous tumor cells, must be still performed, our results suggested that the generation of tumor-specific CTLs could be useful for supporting an ACT approach in CRC.

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Thoresen, Lene. "Nutrition Care in Cancer Patients : Nutrition assessment: diagnostic criteria and theassociation to survival and health-related quality oflife in patients with advanced colorectal carcinoma". Doctoral thesis, Norges teknisk-naturvitenskapelige universitet, Institutt for kreftforskning og molekylær medisin, 2012. http://urn.kb.se/resolve?urn=urn:nbn:no:ntnu:diva-16470.

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God ernæringspraksis for kreftpasienter. Ernæringsutredning; diagnostiske kriterier og sammenheng med overlevelse og helserelatert livskvalitet hos pasienter med avansert tykk- og endetarmskreft. Avhandlingen bygger på fire studier. Artikkel 1. Denne studien undersøker meninger til 2759 sykepleiere, 1753 sykehusleger og 359 kliniske ernæringsfysiologer (kef) fra Skandinaviske sykehus om bruk av kef’enes fagkompetanse i sykehus. Sykepleiere og leger som ser kef to eller flere ganger per uke i motsetning til de som ser kef sjeldnere enn to ganger per uke prioriterte klinisk ernæring høyere i avdeling, hadde oftere internundervisning om ernæring, fant det enklere å identifisere underernærte pasienter og pasienter som trengte ernæringsstøtte. Studien viser at sykepleiere og leger som ser kef oftere enn to ganger per uke har større fokus på ernæring. Artikkel 2. I denne studien ble ernæringsstatus til 46 kreftpasienter som ble innlagt i en palliativ enhet undersøkt med hjelp av objektive kriterier og skjemaet ”Subjective Global Assessment” (SGA). I følge de objektive kriteriene; vekttap, BMI, hudfoldtykkelse, armmuskelomkrets, S-Albumin og S-Pre-albumin var 28 pasienter underernært. Med SGA var 30 pasienter vurdert som underernært. SGA hadde en sensitivitet på 96% for å påvise underernæring. Underernærte pasienter hadde flere spiserelaterte symptomer og spiste mindre matporsjoner. Vi fant at to tredjedeler av pasientene var underernært og at SGA var valid som metode for å undersøke ernæringsstatus blant kreftpasienter med avansert sykdom. Artikkel 3. Her er ulike metoder for å måle nedgang i ernæringsstatus hos 77 pasienter med avansert tykk- og endetarmskreft undersøkt. Videre ble metodenes evne til å predikere pasientenes overlevelse studert. 28 pasienter hadde sarkopeni, 32 hadde ernæringsrisiko, 26 var underernært og 16 hadde kakeksi (CCSG) mens 41 hadde kakeksi (EPCRC). De ulike metodene overlappet hverandre ufullstendig. Studien viste at en stor andel av pasientene hadde dårlig ernæringsstatus. Det å ha kakeksi (CCSG) eller å være underernært predikerte kortere overlevelse. Artikkel 4. I denne studien ble ernæringsstatus og livskvalitet undersøkt hos 50 nyhenviste pasienter til Kreftavdelingen for vurdering av kjemoterapi for avansert tykk- og endetarmskreft. Pasientene hadde lavere livskvalitet sammenliknet med normalbefolkningen. De som var underernærte eller hadde kakeksi (EPCRC-SGA) hadde både statistisk og klinisk signifikant dårligere livskvalitet enn de øvrige pasientene. Etter 3 måneder økte 13 pasienter vekt og de forbedret flere livskvalitetsparametre. Syv pasienter tapte vekt og de forverret livskvaliteten signifikant, mens de pasientene som var vektstabile hadde uendret livskvalitet. Tolking. Den høye andelen av underernæring blant pasientene indikerer at mer bør gjøres for å forebygge underernæring tidligere i pasientforløpet. Underernæring og kakeksi kunne ikke holdes i fra hverandre med de metodene som ble undersøkt. Vekttap som kriterium er for uspesifikt til å diagnostisere kakeksi. Det er behov for å utvikle metoder for å påvise nedbrytning av muskulatur som kjennetegner kakeksi. En undersøkelse av ernæringsstatus gjennom et pasientforløp vil kunne avdekke tidspunkter for når intervensjoner bør settes inn mot underernæring.

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28

Pez, Floriane. "Implication de la lysyl oxydase dans la réponse hypoxique et dans la progression tumorale des cellules de carcinome colorectal humain". Thesis, Lyon 1, 2010. http://www.theses.fr/2010LYO10141.

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Au sein d’une tumeur des régions hypoxiques se forment ce qui conduit à l’activation du facteur de transcription HIF1. HIF1, composé des deux protéines HIF1a et HIF1b, permet l’adaptation des cellules à des faibles concentrations d’oxygène en activant la transcription de gène cible. Un de ces gènes est celui codant pour la lysyl oxydase (LOX). Cette enzyme structure la matrice extracellulaire et est impliquée dans la tumorigénèse. Pour comprendre les liens entre LOX et HIF1a, nous avons modulé leurs expressions dans des lignées humaines de carcinome du côlon. En condition hypoxique, HIF1a contrôle l’expression de LOX et réciproquement, LOX régule la synthèse protéique d’HIF1a via l’activation de la voie de signalisation PI3K/AKT. Nous avons donc mis en évidence l’existence d’une boucle de régulation positive entre LOX et HIF1 en conditions hypoxique. Sachant que ces deux protéines sont des acteurs majeurs de la progression tumorale, nous avons cherché à comprendre le rôle de cette régulation mutuelle dans ce processus. Nos résultats démontrent que l’activité enzymatique de LOX promeut la croissance tumorale in vitro et in vivo et que son action est potentialisée par la présence de son partenaire HIF1a. De plus, LOX et HIF1a agissent en synergie afin d’augmenter la potentiel métastatiques des cellules tumorale de côlon in vitro. Ainsi, ce travail de thèse a permis de mettre en évidence l’existence d’une boucle de régulation entre HIF1a et LOX qui est critique dans la progression tumorale et semble également être impliquée dans le processus métastatiques
The microenvironment of solid tumors is exposed to hypoxic conditions which lead to the activation of Hypoxia‐Inducible Factor 1 (HIF1). HIF1, composed by a heterodimer of HIF1a and HIF1b protein, is a key transcription factor involved in cellular adaptation to changes in oxygen level, inducing the expression of several transcriptional targets such as Lysyl Oxidase (LOX). LOX is an amine oxidase that catalyzes crosslinking of fibrillar collagens and elastin in the extracellular matrix. Furthermore, LOX is implied in tumor progression. To clarify, the link between LOX and HIF1a, their expression were modulated in human colorectal carcinoma cell lines. We pointed out that besides HIF1‐dependant regulation of LOX, LOX can also act on the HIF1 pathway under hypoxic conditions. Indeed, LOX enzymatic activity upregulates HIF1a protein synthesis, and this action is mediated by the PI3K/AKT pathway. Thus, these results emphasize the existence of a regulation loop between HIF‐1a and LOX, which represent two main actors of tumoral progression. Thus, we wanted to determine the implication of this amplification loop in tumor progression. Our results show that LOX enzymatic activity increase tumor growth in vitro and in vivo, and this role is partially dependant of its partner HIF1a. Furthermore, we established that that LOX and HIF1a act in synergy to foster metastatic potential in colorectal carcinoma cell lines. Taken together, our results demonstrate a regulation loop between LOX and HIF1a with is critical for tumor progression and metastasis formation

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Hildebrandt, Bert, I. Heide, Christian Thiede, S. Nagel, Annette Dieing, S. Jonas, Peter Neuhaus, Christoph Rochlitz, Hanno Riess y Andreas Neubauer. "Lack of Point Mutations in Exons 11–23 of the Retinoblastoma Susceptibility Gene RB-1 in Liver Metastases of Colorectal Carcinoma". Saechsische Landesbibliothek- Staats- und Universitaetsbibliothek Dresden, 2014. http://nbn-resolving.de/urn:nbn:de:bsz:14-qucosa-133565.

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Diouf, Momar. "Valeur pronostique de la qualité de vie en cancérologie". Thesis, Besançon, 2014. http://www.theses.fr/2014BESA3018/document.

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L'objectif principal de cette thèse est d'étudier la valeur pronostique de la qualité de vie (QdV) en cancérologie etd'explorer son utilité tant pour la pratique clinique que pour la planification des essais.Pour ce faire, nous évaluerons la significativité statistique et clinique de rapport d'un modèle de prédiction composé defacteurs cliniques et de la Qdv par rapport à un modèle de prédiction composé uniquement de facteurs cliniques.La validation statistique et clinique de cette plus-value de la Qdv en oncologie, permettrait de construire des index plusprécis en termes de pronostic vital, et mieux adapter les traitements. En fonction de sa valeur pronostic, la Qdv pourraitservir de critère de stratification dans les essais randomisés en oncologie.Pour les différentes localisations et le stade concernés, la recherche des facteurs pronostics sera réalisée selon lesrecommandations publiées dans la littérature. La procédure suivante sera utilisée :> Sélection des facteurs pronostics par un modèle univarié.> Construction d'un modèle multivarié par sélection « forward », « backward » ou « stepwise » en respectant aumieux la « règle de 10 événements par variable ».> Si existence de scores pronostics étudier l'apport de la Qdv à ces scores.> Vérifications à posteriori des hypothèses du modèle.> Mesure de la concordance entre les valeurs prédites et les valeurs observées à l'aided'indicateurs comme le C-index d'Harell, la statistique de Schemper, le NR1 (Net Reclassification Improvement) et l'IDI (Integrated Discrimation Improvement) de d'Agostino. Analyse de sensibilité par rapport aux données manquantes dans les différentes dimensions de la qualité de vie.Validation interne et/ou externe du modèle. Recherche de valeurs seuil pour les différents scores de qualité de vie dans le but d'une meilleure applicationclinique.Trois localisations seront étudiées : Carcinome hépatocellulaire en situation palliative. Cancer du pancréas métastatique. Cancer colorectal métastatique
The primary objective of this thesis is to evaluate the added prognostic value of health-related quality of life (QoL) inoncology and to explore ils utility for routine clinical practice as well as for design of clinical trials.In a methodological point of view, we will fïrst compare statistical and clinical performances of a model based onclinico-biological variables and a model based on clinico-biological variables and QoL scores.After validation of ils prognostic value, optimal cut-off points for QoL scores will be explored and revised prognosticclassifications including QoL measures will be built. QoL could then be used to guide treatment assignment and asinclusion/exclusion criteria or as stratification criteria in randomized clinical trials.For the different types of cancer, the validation of the prognostic value of QoL for advanced cancer patients will beperformed according to standard recommendations. The following steps will be performed:> Selection of prognostic factors based on univariable Cox models.> Multivariable Cox models using stepwise procedure. The number of variables entering the multivariable modelwill be selected in such a way that the thumb rule (10 events per variable) will be respected.> If a prognostic System exists, compare its performance alone with ils performance after addition of QoL factorsas well as other clinico-biological factors not included in the prognostic System.> Verify model hypotheses.> Assess model performance using HarreH"s C-index, Schempers V statistic, The NRI (Net ReclassificationImprovement) and the IDI (Integrated Discrimination Improvement). Perform sensitivity analysis after imputation of missing QoL data. Internai validation ofnew models. Find cut-off values for QoL scales to facilitate their use in daily practice.Three type of cancer will be studied: Advanced hepatocellular carcinoma. Metastatic pancreatic adenocarcinoma. Metastatic colorectal cancer

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31

Hildebrandt, Bert, I. Heide, Christian Thiede, S. Nagel, Annette Dieing, S. Jonas, Peter Neuhaus, Christoph Rochlitz, Hanno Riess y Andreas Neubauer. "Lack of Point Mutations in Exons 11–23 of the Retinoblastoma Susceptibility Gene RB-1 in Liver Metastases of Colorectal Carcinoma". Karger, 2000. https://tud.qucosa.de/id/qucosa%3A27516.

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Chu, Céline. "Etude de l’effet sur la P‐glycoprotéine (ABCB1) de deux médicaments dirigés contre le récepteur de facteur de croissance épithélial (EGFR), le cétuximab et le lapatinib et conséquence sur la pharmacocinétique et l’efficacité anti‐tumorale de médicaments substrats de ABCB1". Thesis, Paris 11, 2013. http://www.theses.fr/2013PA114806/document.

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La P-glycoprotéine (P-gp) est une protéine transmembranaire de la famille des ATP binding cassette transporteurs. Elle est impliquée dans l’efflux du milieu intracellulaire vers le milieu extracellulaire d’une grande variété de médicaments anticancéreux. Elle peut être responsable de la diminution de la biodisponibilité orale et de la concentration intra-tumorale des médicaments qui en sont substrats. Elle peut notamment être surexprimée par les cellules cancéreuses des adénocarcinomes du colon naïfs de tout traitement, suggérant une résistance naturelle de cette tumeur et également après une chimiothérapie. Notre premier travail in vivo a documenté le caractère substrat de la P-gp de l’evérolimus, inhibiteur de mTOR indiqué dans divers cancers (rein, tumeurs neuroendocrines d’oringine pancréatique et sein), jusqu’à maintenant uniquement étudié dans des modèles in vitro. Une augmentation significative de l’AUC de l’evérolimus administré par voie orale est observée chez des souris mdr1a-/b- comparées à des souris mdr1a+/1b+. Une amélioration significative de la biodisponibilité orale de l’evérolimus est aussi notée chez des souris prétraitées par le lapatinib (Tyverb®), inhibiteur des tyrosines kinases (EGFR et HER2) indiqué dans le cancer du sein, par rapport aux souris ayant reçu l’evérolimus seul. Ce résultat est accompagné d’une inhibition de l’expression de la P-gp intestinale par le lapatinib mesurée par la technique de Western Blot. Enfin, une étude préclinique menée chez des souris porteuses d’une xénogreffe colorectale mutée KRAS montre une activité anti-tumorale certaine des deux médicaments utilisés seuls et en schéma séquentiel. Notre seconde étude a montré pour la première fois que le cétuximab (Erbitux®), anticorps anti-EGFR, inhibe la fonctionnalité de la P-gp dans deux lignées cellulaires surexprimant la P-gp (les cellules IGROV-1 et les HEK P-gp) indépendamment de leur statut EGFR et entraîne chez des souris porteuses d’une xénogreffe colorectale une augmentation significative de la biodisponibilité orale et de la concentration intra-tumorale du SN-38, métabolite actif de l’irinotécan (Campto®) administré par voie orale. Le cétuximab étant prescrit en association avec l’irinotécan chez des patients atteints d’un cancer colorectal métastasé, initialement réfractaire à l’irinotécan, ces résultats pourraient en partie expliquer la réversion de la résistance à l’irinotécan par le cétuximab par une inhibition de l’efflux de la P-gp. Grâce à l’étude de deux associations de médicaments «lapatinib-evérolimus» et «cétuximab-irinotécan», nous avons démontré l’intérêt de l’étude de l’inhibition de la P-gp avec les traitements les plus récents, notamment son rôle dans l’amélioration de la biodisponibilité orale de chimiothérapies utilisées par voie orale
P-glycoprotein (P-gp) is a membrane transporter and belongs to the ATP-binding cassette (ABC) transporter super family. P-gp decreases oral bioavailability of substrate drugs and can cause multidrug resistance in tumor cells by decreasing intracellular drug levels. P-gp is overexpressed in colorectal carcinoma naturally resistant to chemotherapy. The aim of our first study was to document the in vivo transport of everolimus (Afinitor®), a mTOR inhibitor, by P-gp. A signiﬁcant increase of everolimus oral bioavaibility was observed in mdr1a-/1b- mice compared to the wild type. In addition, a signiﬁcant increase of everolimus oral bioavaibility was showed in mice that received a lapatinib pre-treatment (a dual EGFR/HER2 tyrosine kinase inhibitor) compared to mice that received everolimus alone. These results were accompanied by a signiﬁcant decrease of P-gp expression in duodenum segment in lapatinib pre-treated group as compared to control group. Finally, each drug given alone or in association showed a major antitumor activity in a xenograft model of human colorectal carcinoma with KRAS mutation. Our second study showed for the first time that cetuximab (Erbitux®), a monoclonal antibody directed towards EGFR, inhibits P-gp functionality in two cell lines overexpressing P-gp (IGROV-1 and HEK P-gp cells) independently of EGFR status and leads to significant increases of oral bioavailability and intratumoral concentration of SN-38, the active metabolite of irinotecan (Campto®) in mice bearing colorectal carcinoma xenograft. Cetuximab is used in combination with irinotecan in patients with metastatic colorectal cancer, initially refractory to irinotecan, our results may partly explain the reversion of resistance to irinotecan by inhibiting P-gp efflux by cetuximab. In conclusion, our results showed the interest to study the effect of recent anticancerous drugs on P-gp, including their ability to improve oral bioavailability of oral chemotherapy used

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33

Ask, Alexandra. "REPLICATING THE TUMOUR MICROENVIRONMENT:CHEMOSENSITIVITY TESTING IN FIBROBLAST COCULTURES". Thesis, Uppsala universitet, Institutionen för kvinnors och barns hälsa, 2017. http://urn.kb.se/resolve?urn=urn:nbn:se:uu:diva-327344.

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Kucukdemir, Mumine. "Investigation Of The Effect Of Sodium Butyrate Induced Differentiation On Inflammatory Pathways In Colon Cancer Cells". Master's thesis, METU, 2012. http://etd.lib.metu.edu.tr/upload/12614490/index.pdf.

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Sodium butyrate (NaBt) is a four-carbon short chain fatty acid, produced naturally in colon as the end product of the bacterial anaerobic metabolism on dietary fibers. It was previously shown that NaBt can induce differentiation and may inhibit proliferation. The objective of this study was to investigate the effect of NaBt-induced differentation on inflammatory pathways in HT29 colon cancer cells. For this purpose, first, cells were treated with varying concentrations of NaBt from 1-5 mM and amount required to induce differentiation was determined as 3 mM. To understand the effect of NaBt on inflammation, the NF-kappaB pathway (p50 and p65) was investigated. Immunofluorescent staining showed increased nuclear translocation of p50 subunit with no remarkable change in subcellular localization of p65
moreover a synergistic effect was observed when cells were co-treated with NaBt and an NF-kappaB repressor, Bay 11-7085
implying the formation of repressive p50 homodimers in the nucleus. Our preliminary chromatin immunoprecipitation results showed that p65 recruitment v to the promoters of ICAM-1 was reduced, whereas p50 recruitment was increased. However, analysis of NF-kappaB target genes showed that cells treated with 3 mM NaBt have higher expression of the cytokines IL1-&beta
and TNF-&alpha
, adhesion molecules ICAM-1 and VCAM-1 but not COX-2. These results suggest that NaBt-induced differentiation could cause the emergence of an inflammatory signal in HT29 cells as an anti-tumor mechanism, independent from the NFkappaB activity. This work will be important in understanding the role of SCFAs in the colon microenvironment and may provide alternative therapeutic options in colorectal cancer.

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Fohlen, Audrey. "Optimisation des stratégies thérapeutiques en radiologie interventionnelle : application en embolisation et en oncologie Transhepatic forceps biopsy combined with biliary drainage in obstructive jaundice: safety and accuracy ransarterial chemoembolization (TACE) in the management of hepatocellular carcinoma: Results of a French national survey on current practices". Thesis, Normandie, 2019. http://www.theses.fr/2019NORMC424.

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La radiologie interventionnelle (RI) présente des avantages indéniables. Grâce aux techniques d’embolisation, les patients présentant des hémorragies pouvant engager le pronostic vital, peuvent être traités. Les agents liquides collants peuvent être utilisés or, leur technique d’utilisation reste très empirique. Les complications des embolisations à la colle peuvent être gravissimes. Leur maniabilité peut être appréhendée. Dans la 1ère partie de ce travail nous avons étudié in vitro les caractéristiques de plusieurs colles et notamment les dernières mises sur le marché (Glubran 2 et Purefill, par rapport à l’Histoacryl), en fonction de leur préparation (température, ratio du mélange avec le lipiodol). Nous avons également étudié les temps caractéristiques de ces 3 colles. La 2ème partie de ce travail a porté sur la RI appliquée à l’oncologie. Elle est scindée en plusieurs travaux ayant comme objectif commun l’optimisation des techniques de RI en hépato-biliaire. Une étude clinique a porté sur l’intérêt et la sécurité des biopsies endobiliaires. Une étude a consisté en la réalisation d’une enquête de pratiques en chimio-embolisation artérielle (TACE) du carcinome hépato-cellulaire en France. Deux études ont porté sur la recherche in vitro du ou des meilleurs agents cytotoxiques sur des lignées cellulaires (mélanomes de l’uvée murines µ2 et µ2F et humaines mel 270 et OMM 1.3 et de cancer colo-rectal HT29, SW 48, SW 480, SW 620, HCT116, CaCo2) en condition de temps d’exposition court (30 minutes) pour imiter les conditions de TACE. L’objectif final était de pouvoir trouver le ou les agents cytotoxiques les plus efficaces pour optimiser les traitements par TACE des métastases hépatiques de ces 2 origines. Pour les mélanomes de l’uvée, il est apparu que le paclitaxel était significativement le meilleur agent parmi ceux testés. Pour les cellules de CRC, l’idarubicine était le meilleur agent cytotoxique, suivi par la gemcitabine Un dernier travail a été réalisé pour appréhender in vitro la partie embolisation de la TACE. L’embolisation a été assimilée à une hypoxie. Nous avons ainsi pu évaluer l’effet de l’hypoxie à 2 niveaux (1 % et 0.2 %) sur la viabilité cellulaire, après que celles-ci aient été mises en contact (30 minutes) avec de la chimiothérapie. Une hypoxie sévère entraine une chimiorésistance dans ces conditions d’évaluation. Ces travaux doivent amener à des études sur modèle animal ainsi qu’à la mise en place d’essais cliniques
Interventional radiology (IR) has undeniable advantages. With embolization techniques, patients with potentially life-threatening haemorrhage can be treated. Glue can be used but, their technique of use remains very empirical. The complications of glue embolization can be very serious. Their maneuverability can be apprehended. In the first part of this work we studied in vitro the characteristics of several glues and in particular the latest releases on the market (Glubran 2 and Purefill, compared to Histoacryl), according to their preparation (temperature, mixing ratio with lipiodol). We also studied the characteristic times of these 3 glues. The second part of this work focused on the RI applied to oncology. It is divided into several works with the common objective of optimizing the RI techniques in hepatobiliary. A clinical study focused on the interest and safety of endobiliary biopsies. A study consisted in the realization of a survey of practices in arterial chemo-embolization (TACE) of hepatocellular carcinoma in France. Two studies focused on in vitro research of the best cytotoxic agent (s) on cell lines (uveal melanoma murine u2 and μ2F and human mel 270 and OMM 1.3 and colorectal cancer HT29, SW 48, SW 480, SW 620, HCT116, CaCo2) under short exposure time (30 min) to mimic TACE conditions. For uveal melanomas, it was found that paclitaxel was significantly the best agent among those tested. For CRC cells, idarubicin was the best cytotoxic agent, followed by gemcitabine. The ultimate goal was to find the most effective cytotoxic agent (s) to optimize the TACE treatment of liver metastases from these 2 origins. A final work was done to apprehend in vitro the embolization part of the TACE. Embolization has been equated with hypoxia. We were able to evaluate the effect of 2-level hypoxia (1% and 0.2%) on cell viability after they were exposed (30 minutes) with chemotherapy. Severe hypoxia leads to chemoresistance under these conditions of evaluation. This work should lead to studies on animal models as well as the establishment of clinical trials

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Cotte, Alexia. "Implication du métabolisme des phospholipides dans la progression et la résistance des cancers digestifs". Thesis, Bourgogne Franche-Comté, 2017. http://www.theses.fr/2017UBFCI019.

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Le métabolisme des lipides joue un rôle prépondérant dans le cancer. Ce métabolisme a pour effet, particulièrement grâce à la production de phospholipides (PLs), de supporter le niveau accru de prolifération mais aussi de réguler finement des mécanismes intra-cellulaires et extra-cellulaires qui promeuvent le maintien et la progression des cellules cancéreuses. Parmi tous ces acteurs, les gouttelettes lipidiques (GLs), connues pour leur fonction de réservoir, commencent à dévoiler leurs côtés sombres. Notre premier projet nous a permis de mettre en avant l’accumulation de GLs par des cellules de cancer colorectal (CCR) chimiorésistantes. La formation de GLs est régie par l’expression de l’enzyme lysophosphatidylcholine acyltransférase 2 (LPCAT2), permettant la production de phosphatidylcholine. Elle a pour effet de protéger le réticulum endoplasmique (RE) de l’induction d’un stress prévenant l’activation d’une mort cellulaire immunogène. Ces modulations lipidiques peuvent également se retrouver dans le plasma, où elles font l’objet de l’identification de biomarqueurs. Dans ce contexte, nous avons montré dans un second projet, que certains PLs pouvaient diagnostiquer la présence d’un carcinome hépatocellulaire (CHC) sur un foie cirrhotique. Ces deux aspects soulignent l’importance du métabolisme des PLs dans les cancers digestifs
Among all altered cancer metabolic pathways, lipid metabolism has a preponderant role in cancer development. This metabolism, especially through the production of phospholipids, supports high level of proliferation and carefully regulates intra-cellular and extra-cellular mechanisms promoting maintenance and progression of cancer cells. Among all metabolic players, lipid droplets (LD), known for their storage function, begin to reveal dark sides. Our first project led us to highlight LD involvement in the chemoresistance of colorectal cancer (CRC) cells. This resistance carries out thanks to LD accumulation during chemotherapy treatment. Their accumulation is regulated by the expression of lysophosphatidylcholine acyltransferase 2 (LPCAT2), leading to the production of phosphatidylcholine. It causes the protection of the endoplasmic reticulum (ER) stress induction preventing the activation of immunogenic cell death. These lipid modulations can also be found in plasma where they can be identified as biomarkers. In this context, we have shown that some phospholipids could prognosticate hepatocellular carcinoma (HCC) upon cirrhotic liver. These two aspects highlight the significance of phospholipid metabolism in digestive cancers

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37

NARLOCH, ROBERT. "Characterization of a novel isoform of Bruton's Tyrosine Kinase (BTK) involved in resistance to drug-induced apoptosis of carcinoma cells". Doctoral thesis, Università degli Studi di Milano-Bicocca, 2010. http://hdl.handle.net/10281/10304.

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Cancer is a multistep process in which mutation of 4 to 6 genes has to occur for the cell to become fully oncogenic. Very often, genes affected by such mutations encode for molecules belonging to the pathways allowing DNA repair or controlling the apoptotic process. Many anticancer drugs act by inducing DNA damage, which in turn triggers apoptosis. Inability of tumour cells to undergo chemotherapy-induced apoptosis is a main mechanism of drug resistance. Therefore, in order to find rational targets to tailor therapy is necessary to identify novel genes involved in modulating apoptosis induced by chemotherapeutics. Recently in our lab a loss-of-function RNAi-library-based phenotypic screen has been performed which identified 49 novel genes whose silencing reverts 5-fluoraouracil (FU) resistance in a model colon cancer cell line (HCT116p5KO). The aim of the project was to study the role of one of the targets identified in the abovementioned screen, the Bruton’s tyrosine kinase (BTK), so far assumed to be expressed only in hematopoietic lineages. Our findings suggest that BTK could be an interesting candidate to be targeted in the therapy of drug-resistant colon cancers and a marker of drug-resistance.

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38

Petitprez, Florent. "Integrated analysis and clinical impact of immune and stromal microenvironments in solid tumors Quantitative analyses of the tumor microenvironment composition and orientation in the era of precision medicine Transcriptomic analysis of the tumor microenvironment to guide prognosis and immunotherapies Tumor microenvironment quantification tool draws a comprehensive map of the tumor microenvironment of non-hematologic human cancers The mMCP-counter method to estimate abundance of tissue-infiltrating immune and stromal cell populations using gene expression in murine samples Immune sub-classes in sarcoma predict survival and immunotherapy response Intra-tumoral tertiary lymphoid structures are associated with a low risk of hepatocellular carcinoma early recurrence Association of IL-36γ with tertiary lymphoid structures and inflammatory immune infiltrates in human colorectal cancer Immune-based identification of cancer patients at high risk of progression Tumor-infiltrating and peripheral blood T-cell immunophenotypes predict early relapse in localized clear cell renal cell carcinoma PD-L1 expression and CD8+ T-cell infiltrate are associated with clinical progression in patients with node-positive prostate cancer Intratumoral classical complement pathway activation promotes cancer progression". Thesis, Sorbonne Paris Cité, 2018. http://www.theses.fr/2018USPCB104.

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Les tumeurs sont composées de cellules malignes et d'une grande variété de cellules non-tumorales, en particulier des cellules immunitaires qui forment le micro-environnement tumoral (MET). Il a été démontré que la composition du MET était associée au devenir clinique des patients, en termes de survie et de réponses thérapeutiques. Avec le développement récent des immunothérapies qui ciblent des éléments spécifiques du MET, l'immunité anti-tumorale a soulevé un intérêt majeur. Plusieurs méthodologies ont été mises au point afin d'étudier la composition du MET, avec une précision toujours plus grande. En particulier, des méthodes comme MCP-counter permettent d'exploiter les données transcriptomiques de la tumeur entière afin de quantifier les différentes populations qui composent le MET. Le volet méthodologique de ce travail de thèse a ainsi consisté à proposer une amélioration de MCP-counter, en particulier pour l'analyse de données RNA-Seq. Une adaptation de la méthode pour des données issues de modèles murins (mMCP-counter) est également proposée. MCP-counter permet d'analyser rapidement le MET de larges séries de tumeurs. Un second volet de cette thèse consiste en l'application de cette méthode pour établir une classification immunitaire des sarcomes des tissus mous, un type de cancer rare, hétérogène et agressif. Cette classification immunitaire a permis de mettre en évidence des groupes de tumeurs faiblement ou fortement infiltrés, ainsi qu'un groupe marqué par une forte vascularisation. De manière intéressante, la classification immunitaire permet de prédire la réponse des patients aux immunothérapies. Ce travail a aussi démontré un rôle important des structures lymphoïdes tertiaires (SLT). Les SLT sont des structures de type noeud lymphatique composées de lymphocytes B et T qui se forment dans la tumeur ou à proximité de celle-ci. Au sein des SLT, une réponse immunitaire anti-tumorale peut se former et maturer. L'intérêt porté aux SLT est de plus en plus important pour de nombreux types de cancers. Dans la plupart des types de cancer, une forte infiltration de la tumeur par des lymphocytes T, en particulier CD8+, est associée à une meilleure survie des patients. Cependant, le carcinome rénal à cellules claires et le cancer de la prostate sont des exceptions à cette règle. En effet, dans ces deux cancers urologiques, la présence dans la tumeur de lymphocytes T est associée à une survie plus courte des patients, ainsi qu'à une rechute et une progression plus précoce. Ces exceptions sont détaillées dans une troisième partie de cette thèse, par une description minutieuse du MET, ainsi que par l'analyse de l'implication du système du complément. Dans leur ensemble, les résultats présentés dans cette thèse démontrent qu'en combinant différentes méthodes d'analyse, in silico, in situ et in vivo, il est possible d'obtenir une vision extrêmement complète du MET. La connaissance des types cellulaires présents dans la tumeur ainsi que leur orientation fonctionnelle permet de guider le soin apporté aux patients et d'améliorer leur devenir clinique. La description complète du MET ouvre la voie à une médecine personnalisée pour les patients atteints de cancer
Tumors are composed not only of malignant cells but also contain a vast variety of non-malignant cells, notably immune cells forming the tumor microenvironment (TME). The composition of the TME was shown to be associated with clinical outcome for cancer patients, in terms of survival and therapeutic responses. With the relatively recent development of immunotherapies targeting specific elements of the TME, tumor immunology has risen a strong interest and holds a strong therapeutic potential. Several methodologies have been developed to study the composition of the TME with an increased precision. Notably, some methods such as MCP-counter enable the use of the tumor bulk transcriptome to quantify cell populations composing the TME. The methodological aspect of this PhD project consisted in setting up an enhanced version of MCP-counter that can be readily applied to RNA-Seq data, as well as propose an adaptation of the method for mouse models. Using MCP-counter, the TME of large series of tumors can be easily analyzed. The application part of this PhD work consisted of applying MCP-counter to establish an immune-based classification of soft-tissue sarcoma, a rare, aggressive and heterogeneous cancer type. The immune classification notably allowed to identify immune low and high groups, and a group characterized by a strong vasculature. Interestingly, the classification was notably found to be predictive of the patients' response to immunotherapies. It also highlighted an important role of tertiary lymphoid structures (TLS). TLS are lymph-node-like structures composed of T and B cells that form within the tumor or in close proximity. They are a site of formation and maturation of antitumoral immune responses. TLS are raising a growing interest in many malignancies. In most cancer types, a strong infiltration by T cells, in particular CD8+ T cells, is associated with a favorable clinical outcome. However, clear-cell renal cell carcinoma and prostate cancer are exceptions to this general rule. Indeed, in these urological cancers, an increased infiltration by T cells is associated with a decreased patient survival and with earlier relapse and disease progression. In a third part of this thesis, these exceptions are investigated with more details by scrutinizing the TME, and questioning the implication of the complement system. Overall, this thesis presents how the combination of several analysis methods, in silico, in situ and in vivo, can help achieve an extremely precise description of the TME. Knowing accurately what cell populations and what their functional orientation can help guide patients care and improve clinical outcome. Complete description of the TME opens the way towards personalized medicine for cancer patients

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39

NOBILI, STEFANIA. "Analisi dell'espressione di geni coinvolti nella resistenza al 5-fluorouracile nel carcinoma colorettale". Doctoral thesis, 2002. http://hdl.handle.net/2158/779872.

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5-FU-based chemotherapy (CT)is widely used in CC treatment. However, many patients (pts) still fail on these regimens due to natural or acquired tumor resistance mainly caused by increased expression of the target enzyme, thymidylate synthase (TS). In addition, a decreased activity of the folypolyglutamate synthetase (FPGS) enzyme has been associated with 5-FU resistance presumable due to the resulting depletion of folate polyglutamates, destabilizing the inhibitory TS-fluorodeoxyurilydate-folate coenzyme ternary complex. In tumor samples from 31CC pts, we measured TS and FPGS expression to establish their predictive role in prognosis for 5-FU based CT, by analizing TS and FGPS mRNA levels by RT-PCR and TS protein by immunohistochemistry. The median TS/beta-actin ratio was 41.36x10-3 (range 2,49x10-3-294,54x10-3). The TS immunostaining intensity was absent or weak in 54,8% and medium or strong in 45,2% samples. A statistically significant correlation was observed between TS mRNA and protein levels (p=0,0018). Overall median survival (OS) was significantly longer for pts with TS mRNA levels lower than the median value compared with those with higher levels (p=0.0103). Also the OS of pts with completely resected tumors (n=26) was longer for those with low TS expression compared with those with high TS expression (p=0.0092), as was disease-free survuval (DFS) (P=0.0923). Analysis of TS protein expression gave a similar correlation, although significnat only for OS of pts with completely resected tumors (p==.0041). DFS and OS data in the same pts demonstrated a significant difference between CT treated pts and surgical controls in pts with high Ts mRNA expression (p=0.0012 and 0.0006, respectively) but not in those with low TS expression. In pts undergoing surgery alone, high TS levels were unfavourable prognostic factor. Our data confirm the prognostic role of TS expression, as determined by both methods, for 5-FU based adjuvant CT. No correlation has been found between FPGS gene expression and usrvival of CC pts. The large scale evaluation of these and other molecular determinants of sensitivity may help tailor CT for CC pts and thus obtain improved efficacy.

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40

PERRONE, GABRIELE. "Identificazione di MTHFR 1298A>C come marcatore predittivo di sopravvivenza in due coorti di pazienti con carcinoma colorettale (stadio II e III) trattati con chemioterapia adiuvante a base di fluoropirimidine con o senza oxaliplatino". Doctoral thesis, 2014. http://hdl.handle.net/2158/854506.

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Adjuvant treatment based on fluoropyrimidines alone or in association with oxaliplatin improves both disease free and overall survival in stage II/III colorectal cancer. However, a certain percentage of patients do not take advantage of the treatment. The identification of predictive genetic biomarkers may be useful in the selection of patients for chemotherapy treatment by identifying responsive patients and avoiding treatment in non-responsive patients . The aim of this study was to compare the impact of a set of fluoropyrimidines-related polymorphisms on 5-year disease free survival in two groups of colorectal cancer patients treated with adjuvant fluoropyrimidines with or without oxaliplatin. A set of 23 polymorphisms in 10 fluoropyrimidines-related genes in two cohorts of stage II/III fluoropyrimidines-treated colorectal cancer patients, including a total of 262 cases were analysed. A concordant effect for MTHFR-1298A>C (rs1801131) polymorphism was found. Carriers of MTHFR-1298CC genotype had a worse disease free survival in both groups (HR=3.48, 95%CI 1.01-11.96 in fluoropyrimidines alone; HR=3.13, 95%CI 1.23-7.97 in fluoropyrimidines + oxaliplatin). In the pooled group of patients MTHFR-1298CC carriers had also a worse overall survival (HR=2.01, 95%CI 0.85-4.75, adjusted P-value=0.035). We computed a clinical score related to disease free survival including MTHFR-1298A>C, disease stage, sex, and tumor location, where MTHFR-1298A>C is the most detrimental factor. In conclusion MTHFR-1298A>C is a prognostic factor for disease free survival and overall survival in stage II/III colorectal cancer patients treated with a fluoropyrimidines-based treatment and could be used as an additional criteria for the choice of the proper adjuvant regimen.

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41

Federica, Ricci. "Gut microbiota and immune response in human model of colon inflammatory: Potential and Celiac disease, Polyps and Cancer". Doctoral thesis, 2020. http://hdl.handle.net/2158/1189487.

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Nell’insorgenza e nella progressione del carcinoma colorettale (CRC) e della malattia celiaca (MC), il sistema immunitario svolge un ruolo fondamentale: mentre in ambito neoplastico è coinvolto nel riconoscimento delle cellule trasformate e nella loro eliminazione, nella celiachia è il sistema immunitario stesso ad innescare il danno. Oltre al sistema immunitario, molti fattori sembrano influenzare l’insorgenza di tali patologie, e tra questi, sempre più studi indicano l’importanza del microbiota intestinale. Studi condotti su topi “germ free”, dimostrano come il microbiota intestinale sia in grado di modulare la risposta immune, ed essendo quest’ultima coinvolta nelle neoplasie e nelle malattie autoimmuni, è facile ipotizzare che una disbiosi intestinale possa contribuire alla patogenesi delle malattie in esame. Data dunque l’importanza della componente immunitaria nelle due patologie e l’ipotesi del coinvolgimento del microbiota intestinale, l’obiettivo di questo lavoro di tesi è stato quello di caratterizzare la risposta immune presente a livello tissutale e periferico nei pazienti con CRC e con MC e di analizzare la componente microbica presente nella sede intestinale dei pazienti. Individuare il tipo di risposta immune presente negli individui patologici ed i “cluster” microbici potenzialmente associati alle patologie, potrebbe indirizzare verso nuovi sistemi di prevenzione basati sull’immunoterapia oppure sull'impiego di trattamenti innovativi a base di probiotici (batteri vivi che potrebbero esercitare effetti positivi per la salute), prebiotici (ingredienti alimentari, non digeribili, che stimolano la crescita/attività dei batteri) e simbiontici. Lo studio ha inoltre previsto di valutare la risposta immune ed il microbiota intestinale in pazienti affetti da poliposi intestinale e da malattia celiaca potenziale visto che le poliposi intestinali, se non trattate, molto spesso posso evolvere in tumore e che i pazienti con malattia celiaca potenziale presentano la predisposizione genetica e gli autoanticorpi specifici della MC senza avere un danno intestinale. Riscontrare, nei pazienti di controllo, elementi predittivi l’insorgenza della patologia permetterebbe di intervenire in maniera preventiva al fine di evitare l’evolversi della patologia tumorale e della celiachia.

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42

Lipská, Ludmila. "Colorectal carcinoma and markers of biological activity". Doctoral thesis, 2006. http://www.nusl.cz/ntk/nusl-268921.

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43

Yang, Tzu-Wei y 楊子緯. "Explore the Dysbiosis of Gut Microbiota along the Conventional Adenoma-Carcinoma Colorectal Cancer Pathway". Thesis, 2019. http://ndltd.ncl.edu.tw/handle/3gsuz7.

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博士
國立交通大學
生物科技學系
107
CRC is one of the leading causes of cancer-related deaths worldwide. The adenoma-carcinoma sequence pathway accounts for more than 70 % of sporadic CRCs which developed from small adenomas. The change of microbial composition of intestinal microbiota, so-called "dysbiosis" was found to be is one of the multifactorial factors contributing to the pathogenesis of CRC recently. We conducted an observational study in the correlation between human gut microbiota and colorectal neoplasms, including colon polyp and CRC. Using next-generation sequencing targeting bacterial 16S rRNA, we analyzed 283 stool and 471 remnant cecal yellow fluid (CYF). We categorized all the subjects into normal, colon polyps, and CRCs according to the colonoscopic and pathologic results. In the stool samples, we identified 3 enterotypes with Bacteroids, Prevotella, and Escherichia-dominated using the principal component analysis (PCoA). In addition to previously reported, we found a significantly increased abundance of Morganella in all CRC stool samples (Kruskal–Wallis test, p<0.05). We also identified significantly decreased abundances of Oscillospira and Haemophilus in the transitional process from advanced adenoma, carcinoma in situ (stage 0 CRC) to the early stage of CRC, respectively (all p<0.001). Finally, we found a significantly lower abundance of co-abundance group (CAG) cluster 6 in the CRC group by hierarchical clustering and Spearman’s correlation (p<0.05). In the CYF samples, we found the microbial composition was different from stool samples. Nine genera were statistically significant abundant in the CRC (all p < 0.001). Seven genera were almost depleted in the early and late stage of CRC. In conclusion, CRC gut microbiota dysbiosis was found in both stool and CYF samples. The dysbiosis of gut microbiota had already occurred in the upper gastrointestinal tract and proximal colon. Enterotypes and CAGs analysis of fecal samples could be a potential clinical classifier for colon neoplasms which requires further study in a large cohort.

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Prince-Wright, Lawrence. "C-CBL phosphorylation status influences colorectal cancer cell survival in a Wnt-dependent manner". Thesis, 2015. https://hdl.handle.net/2144/16145.

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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.

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Shukla, Madhura Shirish. "Role of PERK in Anchorage-Independent Growth of Colorectal Carcinoma and Cell Migration In-Vitro". Thesis, 2020. http://hdl.handle.net/1805/24082.

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Indiana University-Purdue University Indianapolis (IUPUI)
The unfolded protein response (UPR) is important for cell adaptation to accumulation of unfolded proteins in the endoplasmic reticulum (ER). A central UPR sensor of ER stress is PKR- like ER Kinase (PERK), which phosphorylates eIF2 to reduce global translation and help mitigate ER stress. While this is a survival mechanism that serves to save the cell from catastrophic events during ER stress, PERK can also be activated in cancer cells due to genetic changes and exposure to stresses inherent in the tumor micro-environment. Published reports have indicated that PERK is activated in cancer cells in response to hypoxia, nutrient deprivation, matrix detachment, and increased protein load by oncogene activation to facilitate cell survival. The UPR features PERK and another ER stress sensory protein, IRE1α, which also regulates the dynamic assembly of the actin cytoskeleton; loss of either PERK or IRE1α functions decrease cell migration activity. We hypothesized that PERK is required for anchorage-independent survival of the cancer cell line HCT116 and that PERK is essential for cell migration. Consistent with these premises, inhibition of PERK using pharmacological inhibitors GSK2656157 and LY-4 in suspended cells showed reduced growth. Furthermore, PERK-deficient cells showed reduced migration in transwell migration assays as compared to their wild type counterpart. These results suggest that PERK facilitates anchorage-independent growth of cancer cells and cell migration.

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46

Tweel, Kristin. "Competing Influences Of The Tumor Microenvironment On CD26 And The Cancer Phenotype Of Colorectal Carcinoma Cells". Thesis, 2011. http://hdl.handle.net/10222/21904.

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In Canada, colorectal cancer is the second leading cause of cancer death for both men and women. There are many different factors that contribute to the progression and spread of the disease. However, increasing evidence now suggests that the tumor microenvironment plays a paramount role in these processes. CD26 is a multifunctional, cell-surface glycoprotein that has intrinsic enzyme activity, binds adenosine deaminase and interacts with the extracellular matrix. Through its various functions it serves to constrain cancer progression. For example, it is known to cleave CXCL12, the ligand for CXCR4. The CXCL12:CXCR4 axis is normally involved in cancer metastasis by promoting cancer cell migration, invasion and proliferation. Down-regulation of CD26 is observed in certain cancers - this has been shown in vitro to occur in response to certain soluble mediators. The first part of this study looked at the effects of glucose and its metabolic product lactate on CD26 expression in colorectal carcinoma cells. Our study showed that CD26 expression is lower in cancer cells that are grown in low-glucose, high-lactate conditions, which replicates the situation within a tumor. The second part of this study examined the effect of adenosine, a purine nucleoside, on colorectal carcinoma cells and supportive stromal cells - cancer-associated HS675.T fibroblasts (CAFs) and Met-5a mesothelial cells. Adenosine increased the proliferation of CAFs and increased CXCL12 mRNA in both stromal cell lines. It also increased MMP-13 mRNA in stromal cells as well as colorectal cancer cells, suggesting that adenosine may promote progression and metastasis through various mechanisms. The last section focused on the ability of cellular products and 3-dimensional tissue topology to coordinate and affect the behaviour of the different cell populations. Here we show that secretory products from colorectal cancer cells promote CAF proliferation but inhibit mesothelial cell proliferation, and are also able to modulate MMP-13 expression. Finally, certain responses are enhanced in multicellular spheroids. In conclusion, the tumor microenvironment represents a major consideration in the treatment of solid tumors. Our data suggest that various soluble mediators, such as adenosine, may have therapeutic implications in cancer treatment and might represent novel targets for future research.

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47

"Laparoscopic assisted resection of recto-sigmoid carcinoma: is it justified?" Thesis, 2005. http://library.cuhk.edu.hk/record=b6074005.

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Colorectal cancer is one of the commonest malignancies worldwide. Its prevention, diagnosis and treatments have attracted multidisciplinary attention. Surgery is the mainstay of treatment for colorectal cancer. It was estimated that up to 85% of colorectal cancer were amenable to surgical treatment, whether curative or palliative. Not surprisingly laparoscopic resection of colorectal cancer was reported soon after cholecystectomy. However, with the appearance of early port site recurrence, most authorities were concerned about the adequacy of tumour clearance and the long-term survival after laparoscopic resection.
In this thesis, comparative and randomized studies were conducted to answer the above questions. It was concluded that, as compared to conventional open surgery, laparoscopic assisted resection of recto-sigmoid carcinoma was less painful and allowed earlier post-operative recovery. Tissue trauma, as reflected by systemic cytokines response, was less after laparoscopic assisted resection. Some cellular components of immune system were also less suppressed. Most importantly, laparoscopic resection did not jeopardize the survival and disease control of patients. The justification of adopting laparoscopic technique would depend on the societal value of its effectiveness in improving the short-term post-operative outcomes.
Laparoscopic technology and its application may be the biggest advancement in nearly all surgical specialties in the last decade. Since the introduction of laparoscopic cholecystectomy, enthusiastic surgeons have attempted laparoscopic approach in almost every type of operations, and many of the techniques have gained public acceptance within a very short time. However, most of these developments were not based on good scientific evidence from comparative study. While laparoscopic cholecystectomy was shown to cause less pain and allow patients to recover earlier after operation, these benefits may or may not be conferred to other procedures and diseases.
Therefore, to justify the use of laparoscopic assisted colorectal resection for carcinoma, two criteria must be satisfied. Firstly the long term survival and the disease free interval of patients should not be adversely affected, as these are the most important endpoints in the success of tumour surgery. Secondly, the proposed benefits of minimally invasive surgery must be demonstrated, otherwise it is not worthwhile to adopt a new technique.
Leung Ka Lau.
"July 2005."
Source: Dissertation Abstracts International, Volume: 67-01, Section: B, page: 0174.
Thesis (M.D.)--Chinese University of Hong Kong, 2005.
Includes bibliographical references (p. 122-155).
Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web.
Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web.
School code: 1307.

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48

Chou, Yu-Ting y 邱于庭. "The molecular mechanisms of ribose-5-phosphate isomerase A in the formation of colorectal cancer and hepatocellular carcinoma". Thesis, 2018. http://ndltd.ncl.edu.tw/handle/s459u5.

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49

Ferreira, Bárbara Mendes. "Radioembolização de metástases hepáticas do carcinoma colorretal". Master's thesis, 2021. http://hdl.handle.net/10400.6/11316.

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Introdução: O papel da radioembolização (RE) em metástases hepáticas do carcinoma colorretal (mhCCR) permanece indefinido. Esta investigação pretende avaliar os resultados e possíveis fatores de prognóstico da RE nestes doentes. Metodologia: Uma análise retrospetiva foi realizada a todos os doentes com mhCCR que foram intervencionados com uma RE no Instituto Português de Oncologia do Porto, desde janeiro de 2011 a março de 2020. A sobrevida a um ano foi determinada pelo método de Kaplan-Meier e os possíveis fatores de prognóstico foram avaliados usando os testes logRank, Qui-quadrado, de Fisher, Mann-Whitney e teste-t para amostras independentes. Resultados: Trinta pacientes foram avaliados. A idade média foi de 61,5 anos e a maioria dos doentes eram do sexo masculino (63,3%). A dor abdominal foi a complicação mais frequente (40%). O sucesso da RE foi observado em 50% dos casos. Um estádio inferior ou igual a três, níveis inferiores a 20 ng/mL de CEA ao diagnóstico, um tempo livre de metastização mais longo e a ausência de invasão vascular ou linfática ao diagnóstico estão significativamente associados ao sucesso da RE (valores P de <0,001, 0,035, 0,036, 0,028 e 0,020, respetivamente). A sobrevida a um ano de pacientes com ou sem sucesso na RE foi de 9,4 (IC 95%, 1,8-17,1) e 8,9 meses (IC 95%, 7,5-10,2), respetivamente. Conclusão: Vários fatores aumentam a probabilidade de obter uma RE com sucesso. Foi considerada uma terapêutica bem-tolerada, com a maioria das complicações facilmente geridas. Ainda assim, são necessários mais estudos, com amostras maiores, para avaliar a validade da RE.
Introduction: The role of radioembolization (RE) in liver dominant metastatic colorectal cancer (lmCRC) is unclear. This research aims to assess the prognostic factors and outcomes of RE in these patients. Methodology: A retrospective analysis of all patients with lmCRC who underwent RE in Instituto Português de Oncologia do Porto, from January 2011 to March 2020, was performed. The one-year survival was evaluated with the Kaplan-Meier method and potential prognostic factors were analyzed using the log-rank test, Mann-Whitney test, chisquare test, Fisher’s test, and t-test for independent samples. Results: Thirty patients were analyzed. The median age was 61,5 years and most patients were male (63,3%). There was a low complication rate. Successful RE was observed in 50% of the cases. Lower cancer stage, CEA levels at diagnosis lower than 20ng/mL, more than one year between diagnosis of CRC and the emergence of liver metastases, absence of vascular or lymphatic invasion at the moment of diagnosis were significantly associated with a successful RE (P values of <0,001, 0,035, 0,036, 0,028 and 0,020, respectively). The one-year survival of patients with and without successful RE was 9,4 months (CI 95%, 1,8- 17,1) and 8,9 months (CI 95%, 7,5-10,2), respectively. Conclusion: Several factors increase the likelihood of achieving a successful RE. The RE was considered a well-tolerated procedure, with easily managed complications and a low complication rate. However, more studies with larger cohorts are needed to validate this procedure.

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Correia, Maria João Reis. "Efeito da exposição ambiental no desenvolvimento e prevenção do carcinoma colo-retal". Master's thesis, 2016. http://hdl.handle.net/10284/5388.

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Muitos fatores, como a dieta, a ingestão de álcool, o uso de tabaco, a obesidade, entre outros, têm sido estudados na etiologia do carcinoma colo-retal (CCR). Entre eles, os fatores alimentares têm um papel fundamental, sendo a incidência de CCR maior em países onde a ingestão de carne vermelha é maior. (Durko and Malecka-Panas, 2014). Os programas de rastreio de CCR apenas são possíveis em países economicamente desenvolvidos. Contudo, a atenção no futuro deve ser dada às áreas geográficas onde existe maior envelhecimento populacional e estilos de vida preocupantes (Tárraga López et al, 2014). Abordar os diversos aspetos do CCR, nomeadamente os pontos mais polémicos relacionados com fatores ambientais que afetam esta patologia, e abordar os principais fatores de prevenção do CCR, são os principais objetivos desta dissertação. Esta revisão bibliográfica teve como base as linhas de orientação acerca dos efeitos ambientais no desenvolvimento e prevenção do CCR, os quais são de uma forma geral abordados anualmente. Tem como objetivo pesquisar e sintetizar as evidências disponíveis na literatura científica, relacionadas com os fatores de risco e com os fatores preventivos para o cancro colo-retal, assim como selecionar as recomendações educativas em saúde, específica para a promoção e prevenção do CCR, aliada a uma alimentação saudável.
Many factors such as diet, alcohol intake, tobacco use, obesity, among others, have been studied, which are seen as the most important factors in the etiology of CCR. Among them, dietary factors play a key role, as the high incidence of CCR occurs in countries where red meat intake is higher (Durko and Malecka-Panas, 2014). CCR screening programs are only possible in economically developed countries. However, attention in the future should be given to geographical areas where there is higher population aging and worrying lifestyles (Tárraga López et al, 2014). Address the various aspects of the CCR, including the most controversial points related to environmental factors that affect this condition and address the main CCR prevention factors are the main objectives of this thesis. This literature review will inform the guidelines for the environmental effects in the development and prevention of CCR, which are, in general, covered annually. Aims to search and synthesize the available evidence in the scientific literature related to the risk factors and preventive factors for colorectal cancer, as well as categorizing the aspects related to the CCR and select educational recommendations on health for the promotion and prevention of CCR, combined with a healthy diet.

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Tesis sobre el tema "Carcinoma colorettale (Colorectal cancer)"

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