Comprehensive Molecular Characterization of Childhood Liver Cancer: Identification of Prognostic Biomarkers
Malignant tumors in children and adolescents are one of the leading causes of death from disease in this population despite being rare events. The main liver tumor in children is Hepatoblastoma (HB) representing two thirds of the total while pediatric Hepatocellular Carcinoma (pHCC) is rarer than HB...
| Autor: | |
|---|---|
| Tipo de recurso: | tesis doctoral |
| Estado: | Versión publicada |
| Fecha de publicación: | 2017 |
| País: | España |
| Institución: | CBUC, CESCA |
| Repositorio: | TDR. Tesis Doctorales en Red |
| OAI Identifier: | oai:www.tdx.cat:10803/664344 |
| Acceso en línea: | http://hdl.handle.net/10803/664344 |
| Access Level: | acceso abierto |
| Palabra clave: | Càncer en els infants Cáncer en niños Cancer in children Càncer de fetge Càncer de hígado Liver cancer Marcadors bioquímics Marcadores bioquímicos Biochemical markers Proteòmica Proteómica Proteomics Ciències de la Salut 616 |
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Comprehensive Molecular Characterization of Childhood Liver Cancer: Identification of Prognostic Biomarkers |
| title |
Comprehensive Molecular Characterization of Childhood Liver Cancer: Identification of Prognostic Biomarkers |
| spellingShingle |
Comprehensive Molecular Characterization of Childhood Liver Cancer: Identification of Prognostic Biomarkers Simon Coma, Marina Càncer en els infants Cáncer en niños Cancer in children Càncer de fetge Càncer de hígado Liver cancer Marcadors bioquímics Marcadores bioquímicos Biochemical markers Proteòmica Proteómica Proteomics Ciències de la Salut 616 |
| title_short |
Comprehensive Molecular Characterization of Childhood Liver Cancer: Identification of Prognostic Biomarkers |
| title_full |
Comprehensive Molecular Characterization of Childhood Liver Cancer: Identification of Prognostic Biomarkers |
| title_fullStr |
Comprehensive Molecular Characterization of Childhood Liver Cancer: Identification of Prognostic Biomarkers |
| title_full_unstemmed |
Comprehensive Molecular Characterization of Childhood Liver Cancer: Identification of Prognostic Biomarkers |
| title_sort |
Comprehensive Molecular Characterization of Childhood Liver Cancer: Identification of Prognostic Biomarkers |
| dc.creator.none.fl_str_mv |
Simon Coma, Marina |
| author |
Simon Coma, Marina |
| author_facet |
Simon Coma, Marina |
| author_role |
author |
| dc.contributor.none.fl_str_mv |
Armengol Niell, Carolina Bachs Valldeneu, Oriol Universitat de Barcelona. Facultat de Medicina |
| dc.subject.none.fl_str_mv |
Càncer en els infants Cáncer en niños Cancer in children Càncer de fetge Càncer de hígado Liver cancer Marcadors bioquímics Marcadores bioquímicos Biochemical markers Proteòmica Proteómica Proteomics Ciències de la Salut 616 |
| topic |
Càncer en els infants Cáncer en niños Cancer in children Càncer de fetge Càncer de hígado Liver cancer Marcadors bioquímics Marcadores bioquímicos Biochemical markers Proteòmica Proteómica Proteomics Ciències de la Salut 616 |
| description |
Malignant tumors in children and adolescents are one of the leading causes of death from disease in this population despite being rare events. The main liver tumor in children is Hepatoblastoma (HB) representing two thirds of the total while pediatric Hepatocellular Carcinoma (pHCC) is rarer than HB and is usually diagnosed in older patients. Patient survival at 5 years is higher than 75% for HB and less than 30% for pHCC. At molecular level, 2 different subclasses of HBs have been described based on a 16-gene signature, C1 and C2, being the latest more aggressive. Nowadays patient stratification is based only in clinical and pathological parameters. For this reason, the identification of prognostic markers easy to apply at the clinical practice is mandatory in order to better stratify patients and diminish the side effects of chemotherapy treatment, moving towards a more personalized medicine. The proteomic profile of 16HBs and 8 paired normal liver (NL) tissue was obtained by 2 different techniques, two-dimensional gel electrophoresis and label-free LC-MS. The differential expressed proteins were validated by western blot (WB) in the same patients and the final protein signature was validated by immunohistochemistry in additional 144 patients. Furthermore, RNA sequencing and copy number variation analysis were performed in 31 HB samples, 11 patient-derived xenografts (PDXs) and 5 pHCC. Results were validated by Sanger sequencing, droplet digital PCR or real time PCR and correlated with clinical features. Two hundred and thirty proteins were identified as deregulated in aggressive C2 tumors and 8 of them were selected and validated by WB considering also their expression in NL. After WB, 2 proteins were found significantly upregulated in C2 tumors while 1 was downregulated. A score was calculated for each protein and biomarkers 1 and 2 were considered altered when their staining was at least 2-fold higher than the NL while the biomarker 3 was considered as altered when no staining was observed. The 3-protein signature was defined by the number of altered biomarkers in each tumor and was highly correlated with patient survival and complementary to the current clinical stratification. RNA-sequencing data revealed that fusion proteins are rare events in HB as they were found in only 2% of patients. Mutational analysis allowed us to identify mutations in CTNNB1 (30%), NFE2L2 (7%) and EPHB4 (7%). Interestingly we identified a downregulation of the RNA editing which is correlated with patient outcome. The copy number variation analysis showed that gains are more frequent than losses in HB tumors and that PDXs maintain 78% of the aberrations, being a good model for the study of HB. In contrast, pHCC are characterized by more aberrations than HB and mainly losses. Thus, we stablished a molecular classification that includes 3 HB types: stable (no big CNVs), gains-enriched and losses-enriched classes. This classification is correlated with event-free survival, CTNNB1 mutations and the expression of stem cell markers. As a result of this thesis, we stablished a 3-protein signature that could be easily applied at the clinical practice and increased the molecular knowledge of childhood liver tumors. |
| publishDate |
2017 |
| dc.date.none.fl_str_mv |
2017 2018 2019 |
| dc.type.none.fl_str_mv |
info:eu-repo/semantics/doctoralThesis info:eu-repo/semantics/publishedVersion |
| format |
doctoralThesis |
| status_str |
publishedVersion |
| dc.identifier.none.fl_str_mv |
http://hdl.handle.net/10803/664344 |
| url |
http://hdl.handle.net/10803/664344 |
| dc.language.none.fl_str_mv |
Inglés |
| language_invalid_str_mv |
Inglés |
| dc.rights.none.fl_str_mv |
info:eu-repo/semantics/openAccess |
| eu_rights_str_mv |
openAccess |
| dc.format.none.fl_str_mv |
219 p. application/pdf application/pdf |
| dc.publisher.none.fl_str_mv |
Universitat de Barcelona |
| publisher.none.fl_str_mv |
Universitat de Barcelona |
| dc.source.none.fl_str_mv |
TDX (Tesis Doctorals en Xarxa) reponame:TDR. Tesis Doctorales en Red instname:CBUC, CESCA |
| instname_str |
CBUC, CESCA |
| reponame_str |
TDR. Tesis Doctorales en Red |
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TDR. Tesis Doctorales en Red |
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|
| repository.mail.fl_str_mv |
|
| _version_ |
1869402990425669632 |
| spelling |
Comprehensive Molecular Characterization of Childhood Liver Cancer: Identification of Prognostic BiomarkersSimon Coma, MarinaCàncer en els infantsCáncer en niñosCancer in childrenCàncer de fetgeCàncer de hígadoLiver cancerMarcadors bioquímicsMarcadores bioquímicosBiochemical markersProteòmicaProteómicaProteomicsCiències de la Salut616Malignant tumors in children and adolescents are one of the leading causes of death from disease in this population despite being rare events. The main liver tumor in children is Hepatoblastoma (HB) representing two thirds of the total while pediatric Hepatocellular Carcinoma (pHCC) is rarer than HB and is usually diagnosed in older patients. Patient survival at 5 years is higher than 75% for HB and less than 30% for pHCC. At molecular level, 2 different subclasses of HBs have been described based on a 16-gene signature, C1 and C2, being the latest more aggressive. Nowadays patient stratification is based only in clinical and pathological parameters. For this reason, the identification of prognostic markers easy to apply at the clinical practice is mandatory in order to better stratify patients and diminish the side effects of chemotherapy treatment, moving towards a more personalized medicine. The proteomic profile of 16HBs and 8 paired normal liver (NL) tissue was obtained by 2 different techniques, two-dimensional gel electrophoresis and label-free LC-MS. The differential expressed proteins were validated by western blot (WB) in the same patients and the final protein signature was validated by immunohistochemistry in additional 144 patients. Furthermore, RNA sequencing and copy number variation analysis were performed in 31 HB samples, 11 patient-derived xenografts (PDXs) and 5 pHCC. Results were validated by Sanger sequencing, droplet digital PCR or real time PCR and correlated with clinical features. Two hundred and thirty proteins were identified as deregulated in aggressive C2 tumors and 8 of them were selected and validated by WB considering also their expression in NL. After WB, 2 proteins were found significantly upregulated in C2 tumors while 1 was downregulated. A score was calculated for each protein and biomarkers 1 and 2 were considered altered when their staining was at least 2-fold higher than the NL while the biomarker 3 was considered as altered when no staining was observed. The 3-protein signature was defined by the number of altered biomarkers in each tumor and was highly correlated with patient survival and complementary to the current clinical stratification. RNA-sequencing data revealed that fusion proteins are rare events in HB as they were found in only 2% of patients. Mutational analysis allowed us to identify mutations in CTNNB1 (30%), NFE2L2 (7%) and EPHB4 (7%). Interestingly we identified a downregulation of the RNA editing which is correlated with patient outcome. The copy number variation analysis showed that gains are more frequent than losses in HB tumors and that PDXs maintain 78% of the aberrations, being a good model for the study of HB. In contrast, pHCC are characterized by more aberrations than HB and mainly losses. Thus, we stablished a molecular classification that includes 3 HB types: stable (no big CNVs), gains-enriched and losses-enriched classes. This classification is correlated with event-free survival, CTNNB1 mutations and the expression of stem cell markers. As a result of this thesis, we stablished a 3-protein signature that could be easily applied at the clinical practice and increased the molecular knowledge of childhood liver tumors.Els tumors malignes en nens i adolescents són una de les principals causes de mort per malaltia en aquesta població malgrat ser poc freqüents. El principal tumor de fetge en la infància és l’Hepatoblastoma (HB) mentre que el Carcinoma Hepatocel·lular pediàtric (pHCC) és menys freqüent i normalment es diagnostica en pacients més grans. La supervivència als 5 anys és superior al 75% per l’HB i menor del 30% pel pHCC. A nivell molecular, s’han descrit 2 subclasses d’HB en base a una signatura de 16 gens, C1 i C2, essent la segona més agressiva. Actualment l’estratificació dels pacients es basa només en paràmetres clínics i patològics. Per aquesta raó, la identificació de marcadors pronòstic que siguin fàcilment aplicables a la pràctica clínica és imprescindible per a una millor estratificació els pacients per tal de disminuir els efectes secundaris de la quimioteràpia, avançant cap a una medicina personalitzada. Es va estudiar el perfil proteòmic de 16 HBs i 8 teixits no tumorals (NL) mitjançant 2 tècniques, l’electroforesi bidimensional amb fluorescència i una tècnica sense marcatge LC-MS. Les proteïnes amb expressió diferencial es van validar per western blot (WB) en els mateixos pacients i la signatura final es va validar per immunohistoquímica en 144 pacients. A més, es va realitzar seqüenciació de RNA i un array genòmic en 31HBs, 11 xenògrafts derivats de pacients (PDXs) i 5 pHCC. Els resultats es van validar per seqüenciació Sanger, droplet digital PCR o PCR a temps real i correlacionar amb característiques clíniques. Es van identificar 230 proteïnes desregulades en els tumors agressius C2 i 8 d’aquestes es van seleccionar per ser validades per WB considerant també la seva expressió en NL. Els resultats del WB van confirmar que 2 de les proteïnes estaven sobreexpressades en els tumors C2 mentre que una d’elles estava infraexpressada. Es va calcular una puntuació per cada proteïna, de manera que els biomarcadors 1 i 2 es consideren desregulats si la seva expressió és superior a 2 vegades l’expressió del NL mentre que BM3 es considera alterat si no es detecta expressió. La firma de 3 proteïnes, que es va definir com el número de biomarcadors alterats, estava fortament correlacionada amb la supervivència i era complementària a l’actual estratificació clínica. Les dades de seqüenciació de RNA van revelar que les proteïnes de fusió són poc freqüents en HB i l’anàlisi de mutacions ens va permetre identificar mutacions en CTNNB1 (30%), NFE2L2 (7%) i EPHB4 (7%). A més, vam identificar una infra-regulació del mecanisme d’edició del RNA correlacionat amb el pronòstic. L’anàlisi genòmic va mostrar que els guanys cromosòmics són més freqüents que les pèrdues en l’HB i que els PDXs mantenen un 78% de les alteracions, representant un bon model per a l’estudi del HB. Per contra, el pHCC presenta més alteracions que l’HB i majoritàriament pèrdues. Finalment, vam establir una classificació molecular que inclou 3 classes d’HB: estable, enriquida en guanys i enriquida en pèrdues. Aquesta classificació està correlacionada amb la supervivència, mutacions de CTNNB1 i expressió de marcadors de cèl·lules progenitores. Amb aquesta tesi, hem establert una signatura de 3 proteïnes fàcilment aplicable a la pràctica clínica i augmentat el coneixement molecular dels tumors hepàtics pediàtrics.Universitat de BarcelonaArmengol Niell, CarolinaBachs Valldeneu, OriolUniversitat de Barcelona. Facultat de Medicina201820192017info:eu-repo/semantics/doctoralThesisinfo:eu-repo/semantics/publishedVersion219 p.application/pdfapplication/pdfhttp://hdl.handle.net/10803/664344TDX (Tesis Doctorals en Xarxa)reponame:TDR. Tesis Doctorales en Redinstname:CBUC, CESCAInglésADVERTIMENT. L'accés als continguts d'aquesta tesi doctoral i la seva utilització ha de respectar els drets de la persona autora. Pot ser utilitzada per a consulta o estudi personal, així com en activitats o materials d'investigació i docència en els termes establerts a l'art. 32 del Text Refós de la Llei de Propietat Intel·lectual (RDL 1/1996). Per altres utilitzacions es requereix l'autorització prèvia i expressa de la persona autora. En qualsevol cas, en la utilització dels seus continguts caldrà indicar de forma clara el nom i cognoms de la persona autora i el títol de la tesi doctoral. No s'autoritza la seva reproducció o altres formes d'explotació efectuades amb finalitats de lucre ni la seva comunicació pública des d'un lloc aliè al servei TDX. Tampoc s'autoritza la presentació del seu contingut en una finestra o marc aliè a TDX (framing). Aquesta reserva de drets afecta tant als continguts de la tesi com als seus resums i índexs.info:eu-repo/semantics/openAccessoai:www.tdx.cat:10803/6643442026-06-14T12:46:07Z |
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15,301603 |