Genome-wide shRNA screening identifies genes involved in fulvestrant resistance in breast cancer
Autor(a) principal: | |
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Data de Publicação: | 2016 |
Tipo de documento: | Dissertação |
Idioma: | eng |
Título da fonte: | Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
Texto Completo: | http://hdl.handle.net/10362/19135 |
Resumo: | Breast cancer accounts as the most prevalent cancer and the leading cause of cancer death worldwide among women. Estrogen is one main factor responsible for tumour growth in breast cancer patients through stimulation of estrogen receptor (ER) signalling. ER positive (ER+) breast cancer patients are eligible for anti-estrogenic drugs. Fulvestrant (Faslodex®) represents a second-line therapy for the treatment of postmenopausal women with ER+ advanced breast cancer. Unfortunately, a significant number of ER+ patients will develop resistance to second-line fulvestrant treatment. It is therefore important to understand the molecular mechanisms of resistance and to identify biomarkers capable of predicting response to this treatment. The aim of this project is to establish a genome-wide shRNA functional screening to identify key proteins central in resistance mechanisms and potentially predictive biomarkers capable of identifying ER+ patients that are responsive or resistant to fulvestrant treatment. To do so, a MCF-7-based fulvestrant resistant breast cancer cell line was used. MCF-7/LCC1 and MCF-7/LCC9 were transduced with shRNA libraries covering over 15,000 mRNAs and treated with fulvestrant. This led to depletion and/or enrichment of shRNAs targeting genes evaluated by next generation sequencing (NGS). Deconvolution of NGS data from genomic DNA of LCC1 and LCC9 cells transduced by shRNA libraries led to identification of 206 genes that may have functional significance in fulvestrant resistance. Ingenuity Pathway Analysis of the candidate genes identified HSD17B10 and HSPE1 as key-molecules in networks related to cell proliferation and death. We have found that these genes are upregulated in different fulvestrant-resistant cell lines when compared to their fulvestrant-sensitive parental cell line at gene and protein expression levels using RT-qPCR and Western blotting. This expression is enhanced in fulvestrant presence, suggesting that these proteins may have critical importance in the resistance phenotype. Further studies on these proteins may elucidate on how to overcome fulvestrant resistance. |
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Genome-wide shRNA screening identifies genes involved in fulvestrant resistance in breast cancerER+ breast cancerFulvestrant resistanceGenome-wide shRNA screeningHSD17B10HSPE1Domínio/Área Científica::Engenharia e Tecnologia::Outras Engenharias e TecnologiasBreast cancer accounts as the most prevalent cancer and the leading cause of cancer death worldwide among women. Estrogen is one main factor responsible for tumour growth in breast cancer patients through stimulation of estrogen receptor (ER) signalling. ER positive (ER+) breast cancer patients are eligible for anti-estrogenic drugs. Fulvestrant (Faslodex®) represents a second-line therapy for the treatment of postmenopausal women with ER+ advanced breast cancer. Unfortunately, a significant number of ER+ patients will develop resistance to second-line fulvestrant treatment. It is therefore important to understand the molecular mechanisms of resistance and to identify biomarkers capable of predicting response to this treatment. The aim of this project is to establish a genome-wide shRNA functional screening to identify key proteins central in resistance mechanisms and potentially predictive biomarkers capable of identifying ER+ patients that are responsive or resistant to fulvestrant treatment. To do so, a MCF-7-based fulvestrant resistant breast cancer cell line was used. MCF-7/LCC1 and MCF-7/LCC9 were transduced with shRNA libraries covering over 15,000 mRNAs and treated with fulvestrant. This led to depletion and/or enrichment of shRNAs targeting genes evaluated by next generation sequencing (NGS). Deconvolution of NGS data from genomic DNA of LCC1 and LCC9 cells transduced by shRNA libraries led to identification of 206 genes that may have functional significance in fulvestrant resistance. Ingenuity Pathway Analysis of the candidate genes identified HSD17B10 and HSPE1 as key-molecules in networks related to cell proliferation and death. We have found that these genes are upregulated in different fulvestrant-resistant cell lines when compared to their fulvestrant-sensitive parental cell line at gene and protein expression levels using RT-qPCR and Western blotting. This expression is enhanced in fulvestrant presence, suggesting that these proteins may have critical importance in the resistance phenotype. Further studies on these proteins may elucidate on how to overcome fulvestrant resistance.Ditzel, HenrikElias, DanielRUNRamos, Pedro André Dias2016-10-18T09:14:26Z20162016-102016-01-01T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/masterThesisapplication/pdfhttp://hdl.handle.net/10362/19135enginfo:eu-repo/semantics/openAccessreponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãoinstacron:RCAAP2024-03-11T03:59:11Zoai:run.unl.pt:10362/19135Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireopendoar:71602024-03-20T03:25:18.490159Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãofalse |
dc.title.none.fl_str_mv |
Genome-wide shRNA screening identifies genes involved in fulvestrant resistance in breast cancer |
title |
Genome-wide shRNA screening identifies genes involved in fulvestrant resistance in breast cancer |
spellingShingle |
Genome-wide shRNA screening identifies genes involved in fulvestrant resistance in breast cancer Ramos, Pedro André Dias ER+ breast cancer Fulvestrant resistance Genome-wide shRNA screening HSD17B10 HSPE1 Domínio/Área Científica::Engenharia e Tecnologia::Outras Engenharias e Tecnologias |
title_short |
Genome-wide shRNA screening identifies genes involved in fulvestrant resistance in breast cancer |
title_full |
Genome-wide shRNA screening identifies genes involved in fulvestrant resistance in breast cancer |
title_fullStr |
Genome-wide shRNA screening identifies genes involved in fulvestrant resistance in breast cancer |
title_full_unstemmed |
Genome-wide shRNA screening identifies genes involved in fulvestrant resistance in breast cancer |
title_sort |
Genome-wide shRNA screening identifies genes involved in fulvestrant resistance in breast cancer |
author |
Ramos, Pedro André Dias |
author_facet |
Ramos, Pedro André Dias |
author_role |
author |
dc.contributor.none.fl_str_mv |
Ditzel, Henrik Elias, Daniel RUN |
dc.contributor.author.fl_str_mv |
Ramos, Pedro André Dias |
dc.subject.por.fl_str_mv |
ER+ breast cancer Fulvestrant resistance Genome-wide shRNA screening HSD17B10 HSPE1 Domínio/Área Científica::Engenharia e Tecnologia::Outras Engenharias e Tecnologias |
topic |
ER+ breast cancer Fulvestrant resistance Genome-wide shRNA screening HSD17B10 HSPE1 Domínio/Área Científica::Engenharia e Tecnologia::Outras Engenharias e Tecnologias |
description |
Breast cancer accounts as the most prevalent cancer and the leading cause of cancer death worldwide among women. Estrogen is one main factor responsible for tumour growth in breast cancer patients through stimulation of estrogen receptor (ER) signalling. ER positive (ER+) breast cancer patients are eligible for anti-estrogenic drugs. Fulvestrant (Faslodex®) represents a second-line therapy for the treatment of postmenopausal women with ER+ advanced breast cancer. Unfortunately, a significant number of ER+ patients will develop resistance to second-line fulvestrant treatment. It is therefore important to understand the molecular mechanisms of resistance and to identify biomarkers capable of predicting response to this treatment. The aim of this project is to establish a genome-wide shRNA functional screening to identify key proteins central in resistance mechanisms and potentially predictive biomarkers capable of identifying ER+ patients that are responsive or resistant to fulvestrant treatment. To do so, a MCF-7-based fulvestrant resistant breast cancer cell line was used. MCF-7/LCC1 and MCF-7/LCC9 were transduced with shRNA libraries covering over 15,000 mRNAs and treated with fulvestrant. This led to depletion and/or enrichment of shRNAs targeting genes evaluated by next generation sequencing (NGS). Deconvolution of NGS data from genomic DNA of LCC1 and LCC9 cells transduced by shRNA libraries led to identification of 206 genes that may have functional significance in fulvestrant resistance. Ingenuity Pathway Analysis of the candidate genes identified HSD17B10 and HSPE1 as key-molecules in networks related to cell proliferation and death. We have found that these genes are upregulated in different fulvestrant-resistant cell lines when compared to their fulvestrant-sensitive parental cell line at gene and protein expression levels using RT-qPCR and Western blotting. This expression is enhanced in fulvestrant presence, suggesting that these proteins may have critical importance in the resistance phenotype. Further studies on these proteins may elucidate on how to overcome fulvestrant resistance. |
publishDate |
2016 |
dc.date.none.fl_str_mv |
2016-10-18T09:14:26Z 2016 2016-10 2016-01-01T00:00:00Z |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/masterThesis |
format |
masterThesis |
status_str |
publishedVersion |
dc.identifier.uri.fl_str_mv |
http://hdl.handle.net/10362/19135 |
url |
http://hdl.handle.net/10362/19135 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.rights.driver.fl_str_mv |
info:eu-repo/semantics/openAccess |
eu_rights_str_mv |
openAccess |
dc.format.none.fl_str_mv |
application/pdf |
dc.source.none.fl_str_mv |
reponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação instacron:RCAAP |
instname_str |
Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação |
instacron_str |
RCAAP |
institution |
RCAAP |
reponame_str |
Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
collection |
Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
repository.name.fl_str_mv |
Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação |
repository.mail.fl_str_mv |
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1799137884643524608 |