143 research outputs found
Biomarkers for classification and risk assessment of pancreatic cystic neoplasms
Pancreatic cystic lesions (PCLs) are increasing incidental findings due to increased ageing of the population and widespread use of imaging. The main problem in clinical practice has to do with distinguishing the high-risk premalignant and malignant cysts that require surgical treatment from the benign or low-grade dysplastic cysts, which should not be over-treated and might not even require surveillance. The goal of the present work is to perform a comprehensive analysis of biomarkers and diagnostic approaches by Endoscopic Ultrasound with Fine-needle Aspiration (EUS-FNA), in a cohort of patients harboring mostly low-risk cysts under surveillance, which are far more frequent in clinical practice.
The PCF analysis performed in this cohort includes studies of genomics (DNA mutations), epigenomics (methylation analysis), metabolomics (glucose), and proteomics (CEA, chromogranin A, NSE), with putative biomarkers encompassing the diagnosis of mucinous and malignant cysts, that require surveillance and surgical resection, respectively.
We performed a first meta-analysis comparing current diagnostic methods - CEA and cytology - with KRAS mutations for the diagnosis of mucinous cysts. CEA was the best test for clinically significant cysts (AUC=0.69), cytology performed better in malignant cysts (AUC=0.78), surpassing KRAS mutations (AUC=0.53 and AUC=0.56, respectively). In a second meta-analysis we compared the accuracy of molecular analysis versus micro forceps biopsy (MFB) in the diagnosis of PCLs. The two approaches were identical for diagnosing benign cysts, but molecular analysis was superior for diagnosing both low and high-risk mucinous cysts.
In addition to these two meta-analyses, we performed a retrospective study evaluating the added value of KRAS and GNAS mutations in PCF of 52 frozen PCF samples. We conclude that, as compared with conventional tests, these had no added value in the differential diagnosis of PCLs. In another publication, we compared glucose level in PCF with CEA in 82 patients. For mucinous cyst diagnosis, a CEA >192 ng/ml showed an AUC of 0.84 while glucose <50 mg/dl revealed an AUC of 0.86. Besides its higher accuracy, PCF glucose evaluated “on site” with a glucometer is easy, immediate, and requires a minimal amount of PCF.
In the next study we sought to determine whether a second EUS-FNA changed the diagnosis or management of pancreatic cysts. We compared the outcome of 105 patients with a single EUS-FNA with that of 23 patients who had a second EUS-FNA. EUS-FNA may be recommended, as it changed management toward surgery in approximately 20% of the patients, particularly with diagnosis of cystic NETs. Following these results, we explored the role of EUS-FNA in small PCLs (<3 cm) in 115 patients with PCLs <3 cm who underwent EUS-FNA. 19/115 were submitted to surgery with 15 malignant or premalignant lesions and the remaining 4 were benign lesions. We conclude that EUS-FNA in lesions <3 cm may improve outcome and cost-effectiveness of surveillance programs, as it confirmed malignancy in 2 out of 5 resected lesions, while it also diagnosed benign cysts who could be released from these programs. In a pilot study with 16 patients, including 4 cystic NETs we aimed at assessing the value of Chromogranin A (CroA) and neuron-specific enolase (NSE) levels in PCF. CroA and NSE levels were higher in cystic NETs with an AUC of 0.94 for CroA and 1 for NSE. These are promising biomarkers to identify pancreatic cystic NETs.
Finally, we studied epigenetic changes in the diagnosis of malignant cysts. Methylation changes of GNAS locus were evaluated to understand whether they may contribute to malignant progression of PCLs. Fifty-two samples of PCF were studied. We observed that GNAS locus methylation changes were significantly associated with malignancy. This is the first study to identify methylation changes in the GNAS locus improving diagnosis of malignant PCLs.
We end this work proposing a revised diagnostic organogram of PCLs established by current guidelines, that incorporates the results obtained in this dissertation’s research.As lesões quísticas pancreáticas (PCL) têm incidência crescente devido ao envelhecimento da população e ao aumento da utilização dos métodos de imagem. Na prática clínica pretende-se distinguir os quistos mucinosos, de alto risco e malignos, que requerem tratamento cirúrgico, dos quistos benignos ou pré-malignos de baixo risco, que no máximo requerem vigilância. O objetivo do presente trabalho é analisar de forma abrangente, biomarcadores em líquido de quisto pancreático (PCF) obtido por Ecoendoscopia com punção (EUS-FNA), numa coorte de quistos predominantemente de baixo risco sob vigilância imagiológica, que são os mais comuns na prática clínica.
A análise de PCF nesta coorte inclui estudos de genómica (mutações no DNA), epigenómica (análise de metilação), metabolómica (glicose) e proteómica (CEA, cromogranina A, NSE), com avaliação de biomarcadores para diagnóstico de quistos mucinosos e quistos malignos, que beneficiam de vigilância e ressecção cirúrgica, respetivamente.
Numa primeira meta-análise comparámos a metodologia diagnóstica atual - CEA e citologia - com as mutações do KRAS para diagnóstico dos quistos mucinosos. O CEA foi o melhor teste em quistos clinicamente significativos (AUC=0.69), e a citologia em quistos malignos (AUC=0.78), superando as mutações do KRAS (AUC=0.53 e AUC=0.56, respetivamente). Numa segunda meta-análise comparámos a precisão diagnóstica da análise molecular versus biópsia com micropinça (MFB) no diagnóstico de PCL. As duas abordagens foram idênticas em quistos benignos, mas a análise molecular foi superior em quistos mucinosos tanto de baixo como de alto risco.
Além das duas meta-análises, realizámos um estudo retrospetivo para avaliar o valor das mutações do KRAS e do GNAS em 52 amostras de PCF congeladas. Concluímos que não têm valor adicional no diagnóstico diferencial das PCL, relativamente aos testes convencionais. Noutra publicação comparámos o nível de glicose em PCF com o CEA para diagnóstico de quistos mucinosos em 82 doentes. O CEA >192 ng/ml apresentou uma AUC de 0.84 e a glicose <50 mg/dl de 0.86. Além da maior precisão diagnóstica, a glicose avaliada in loco com um glicosímetro, é fácil, imediata e requer um volume mínimo de PCF.
No estudo seguinte, avaliámos se uma segunda EUS-FNA alterou o diagnóstico ou a decisão de quistos pancreáticos. Comparámos 105 doentes com uma única EUS-FNA com 23 doentes com uma segunda EUS-FNA. Esta pode ser recomendada, pois cerca de 20% dos doentes foram referenciados para cirurgia após repetição da EUS-FNA, incluindo dois com tumores neuroendócrinos (NET) quísticos. Seguidamente, explorámos o papel da EUS-FNA em pequenas PCL (<3 cm), num estudo com 115 PCL <3 cm. 19/115 foram operadas, correspondendo a 15 lesões malignas ou pré-malignas e 4 benignas. Concluímos que a EUS-FNA em quistos com <3 cm pode melhorar o diagnóstico e o custo-efetividade, pois confirmou malignidade em lesões ressecadas, e diagnosticou quistos benignos que podem ser libertados de vigilância. Num estudo piloto com 16 doentes, incluindo 4 com NET quísticos, avaliámos o valor diagnóstico da cromogranina A (CroA) e da enolase específica neuronal (NSE) em PCF. Os níveis de CroA e NSE foram mais elevados nos NET quísticos, com uma AUC de 0.94 para a CroA e 1 para a NSE. Estes revelaram-se biomarcadores promissores
Por fim, estudámos alterações epigenéticas no diagnóstico de quistos malignos. Analisámos a metilação do locus GNAS em PCF para perceber se se associa à progressão maligna de PCL. Estudámos 52 amostras e observámos que a alteração da metilação se associou significativamente a malignidade. Trata-se do primeiro trabalho a avaliar alterações de metilação no locus GNAS no diagnóstico de PCL.
Terminamos este trabalho com uma proposta de revisão do organograma de diagnóstico das PCL baseado nas guidelines atuais, que incorpora os resultados desta tese
Risk stratification in pancreatic cystic lesions: towards a precision medicine-based approach
The contribution of endoscopic ultrasound and biomarkers in the management of pancreatic adenocarcinoma and its precursor lesions.
Experimental acute pancreatitis models : history, current status, and role in translational research
Early detection of pancreatic cancer:the juicy details
Pancreatic cancer is a lethal disease, for which timely detection and early surgical resection provide the only chance of cure. In individuals with an increased risk of developing pancreatic cancer (high-risk individuals), including those with a hereditary predisposition or neoplastic pancreatic cyst, surveillance may lead to earlier detection and improved survival. However, it is debatable whether these benefits weigh up to its drawbacks, such as patient anxiety, harm by overtreatment and increased healthcare expenses.While malignant transformation is difficult to identify with current imaging modalities, (cancer) cells constantly shed various materials to surrounding tissues. We postulate that measuring these substances in blood and pancreatic juice may serve as indicators for high-grade dysplasia or early-stage cancer. The detection of a biomarker signature in blood and pancreatic juice may stratify high-risk individuals by cancer risk and bring forth a surveillance program with tailored intervals and modalities. The ultimate goal of such a marker panel is to detect pancreatic cancer at a curable stage, while minimizing surveillance-related harms.This thesis critically appraises current recommendations for pancreas surveillance (PART I) and evaluates the potential of biomarker analysis in blood and pancreatic juice (PART II).<br/
Prognostic Role of High-Grade Tumor Budding in Pancreatic Ductal Adenocarcinoma: A Systematic Review and Meta-Analysis with a Focus on Epithelial to Mesenchymal Transition
This study aims at clarifying the prognostic role of high-grade tumor budding (TB) in pancreatic ductal adenocarcinoma (PDAC) with the first systematic review and meta-analysis on this topic. Furthermore, we analyzed with a systematic review the relationship between TB and a recently suggested TB-associated mechanism: the epithelial to mesenchymal transition (EMT). Analyzing a total of 613 patients, 251 of them (40.9%) with high grade-TB, we found an increased risk of all-cause mortality (RR, 1.46; 95% CI, 1.13–1.88, p = 0.004; HR, 2.65; 95% CI, 1.79–3.91; p < 0.0001) and of recurrence (RR, 1.61; 95% CI, 1.05–2.47, p = 0.03) for PDAC patients with high-grade TB. Moreover, we found that EMT is a central process in determining the presence of TB in PDAC. Thanks to this meta-analysis, we demonstrate the potential clinical significance of high-grade TB for prognostic stratification of PDAC. TB also shows a clear association with the process of EMT. Based on the results of the present study, TB should be conveyed in pathology reports and taken into account by future oncologic staging systems
CHRONIC PANCREATITIS, PAIN, AND ANXIETY IN AN ALCOHOL AND HIGH FAT MOUSE MODEL
Homeodynamic space (HDS) shrinks as vulnerability increases with aging and repeated damage to the cells. HDS is lost in alcoholic pancreatitis patients due to overconsumption of alcohol, smoking, and high fat diets. Etiologically relevant animal models for study of chronic pancreatitis (CP) are needed. In order to begin filling this gap a central purpose of this dissertation research was to examine relationships between the alcohol and high fat diet (AHF) and pancreatitis with attention to hypersensitivity and anxiety-like behaviors. The AHF diet induced pancreatitis described here etiologically mimics human risk factors of AHF consumption for advancement to alcoholic CP.
In this study one group of mice was fed long term with a diet of high fat and alcohol for comparison with a group fed normal chow. Mice consumed a liquid diet containing 6% alcohol and a high fat supplement ad libitum over a period of five months. Each group was evaluated for heat and mechanical hypersensitivity, and histology indicative of CP.
The association of pancreatitis pathology with anxiety has been understudied. Anxiety, like pain, is useful as a transient state but when anxiety is prolonged it is termed a disorder. Anxiety is often comorbid with pain and depression. Therefore, it is important to determine anxiety in mice with CP histology.
This model was characterized for the interaction of pancreatitis histology, as well as persisting pain-, anxiety-, and fear-like behaviors. The AHF diet mice developed hypersensitivity, demonstrated anxiety-like behaviors, and showed concurrent histology consistent with CP. Nontransgenic mouse models where pancreatitis is induced only by a combination of ad libitum liquid food with added alcohol and lard supplementation do not currently exist, nor has an in-depth study of anxiety-like behaviors been conducted in this mouse model. This dissertation research addresses this knowledge gap
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