All-trans retinoic acid (ATRA) is the active metabolite of vitamin A and a promising agent in the prevention and treatment of breast cancer. In view of the development of ATRA-based therapeutic strategies aimed at the personalized treatment of mammary tumours, it has been recently demonstrated that approximately 70% of oestrogen-receptor-positive (ER+) breast cancer cell lines and primary tumours are sensitive to the anti-proliferative effects of ATRA. In contrast, only 10-20% of the HER2-positive and triple-negative counterparts respond to the retinoid. The present study is aimed at getting insights into the molecular mechanisms underlying the anti-tumour action of ATRA in the specific subsets of breast cancer identified. In addition, we intend to identify specific genes and gene-networks modulated by ATRA which may represent pharmacological targets for the design and development of rational combinations between the retinoid and unrelated therapeutic agents to be used in the personalized treatment of breast cancers. A final goal is the identification of potential bio-markers of the anti-tumour response to ATRA to be used in the clinics. To address all these points, we analysed data from high throughput technologies (RNA-sequencing and ChIP-sequencing) and Methylation arrays, to go insight the gene expression profiles inducted by the pharmacological treatment with retinoic acid. We performed deep-sequencing experiments on a panel of sixteen cell lines exposed to ATRA (1 µM) for 24 hours, recapitulating the heterogeneity of the breast cancer phenotype and characterized for their anti-proliferative response to ATRA. Moreover, we extracted public data available on immune-chromatin precipitation sequencing experiments and methylation arrays on breast cancer cell lines. The global gene-expression, chromatin occupation and methylation data were analyzed with a number of complementary bio-informatic tools which resulted in the identification of a few genes whose expression is up- or down-regulated specifically in ATRA-sensitive cell lines. Pathway and gene-network analyses indicate a strong enrichment in the up-regulation of genes involved in the pathways modulated by interferons. These last results are consistent with the idea that ATRA exerts a strong immuno-modulatory action in breast cancer cells and represents proof of principle for the evaluation of combinations between the retinoid and check-point inhibitors in the treatment of breast cancer.
L'acido retinoico tutto-trans (ATRA) è un metabolita attivo della vitamina A e un promettente agente nella prevenzione e nel trattamento del cancro al seno. Nell’ambito dello sviluppo di strategie terapeutiche basate su ATRA, volte al trattamento personalizzato dei tumori mammari, è stato recentemente dimostrato che circa il 70% delle linee cellulari di carcinoma mammario positive al recettore per l’estrogeno (ER+) sono sensibili all’effetto antiproliferativo di ATRA. Al contrario, solo il 10-20% delle controparte HER2-positiva e dei TNBC risponde al trattamento con il retinoide. Il presente studio è volto ad approfondire i meccanismi molecolari alla base dell'azione anti-tumorale di ATRA nei sottotipi di cancro al seno identificati. Inoltre, intendiamo identificare specifici geni e reti genetiche modulate da ATRA che possano rappresentare bersagli farmacologici per la progettazione e lo sviluppo di combinazioni terapeutiche di retinoidi e altri agenti terapeutici, da utilizzare nella trattamento personalizzato del carcinoma della mammella. Un obiettivo finale è l'identificazione di potenziali bio-markers della risposta anti-tumorale a ATRA da utilizzare in ambito clinico. Per fare ciò, sono stati analizzati i dati di sequenziamento di RNA e di sequenziamento di immuno-precipitazione di cromatina (Chip-sequencing) e arrays di metilazione, per identificare il profilo di espressione genica indotto dal trattamento farmacologico con acido retinoico. Abbiamo effettuato esperimenti di sequenziamento di RNA su un pannello di sedici linee cellulari trattate con ATRA (1 µ M) per 24 ore. Inoltre, sono stati processati e analizzati i dati depositati di Chip-sequencing a arrays di metilazione, su un ampio pannello di linee di cellulari del cancro del seno. L'espressione genica globale, l'occupazione della cromatina e i dati di metilazione sono stati analizzati con una serie di strumenti bio-informatici complementari, che hanno portato all'identificazione di alcuni geni la cui espressione è regolata in modo specifico dal trattamento con acido retinoico, e correlata con la risposta al trattamento farmaologico. Le vie molecolari e l'analisi delle interazioni geniche, indicano un forte arricchimento nella up-regolazione dei geni legati alle vie modulate da interferone. Questi ultimi risultati sono coerenti con l'idea che ATRA possa esercitare una forte azione immuno-modulatoria nelle cellule tumorali mammarie e rappresenta il razionale per la valutazione delle combinazioni tra i retinoidi e gli inibitori del check-point immunitario nel trattamento del carcinoma della mammella.
A multi-omics approach to study mechanisms of action of all-trans retinoic acid in breast cancer
VALLERGA, ARIANNA
2017/2018
Abstract
All-trans retinoic acid (ATRA) is the active metabolite of vitamin A and a promising agent in the prevention and treatment of breast cancer. In view of the development of ATRA-based therapeutic strategies aimed at the personalized treatment of mammary tumours, it has been recently demonstrated that approximately 70% of oestrogen-receptor-positive (ER+) breast cancer cell lines and primary tumours are sensitive to the anti-proliferative effects of ATRA. In contrast, only 10-20% of the HER2-positive and triple-negative counterparts respond to the retinoid. The present study is aimed at getting insights into the molecular mechanisms underlying the anti-tumour action of ATRA in the specific subsets of breast cancer identified. In addition, we intend to identify specific genes and gene-networks modulated by ATRA which may represent pharmacological targets for the design and development of rational combinations between the retinoid and unrelated therapeutic agents to be used in the personalized treatment of breast cancers. A final goal is the identification of potential bio-markers of the anti-tumour response to ATRA to be used in the clinics. To address all these points, we analysed data from high throughput technologies (RNA-sequencing and ChIP-sequencing) and Methylation arrays, to go insight the gene expression profiles inducted by the pharmacological treatment with retinoic acid. We performed deep-sequencing experiments on a panel of sixteen cell lines exposed to ATRA (1 µM) for 24 hours, recapitulating the heterogeneity of the breast cancer phenotype and characterized for their anti-proliferative response to ATRA. Moreover, we extracted public data available on immune-chromatin precipitation sequencing experiments and methylation arrays on breast cancer cell lines. The global gene-expression, chromatin occupation and methylation data were analyzed with a number of complementary bio-informatic tools which resulted in the identification of a few genes whose expression is up- or down-regulated specifically in ATRA-sensitive cell lines. Pathway and gene-network analyses indicate a strong enrichment in the up-regulation of genes involved in the pathways modulated by interferons. These last results are consistent with the idea that ATRA exerts a strong immuno-modulatory action in breast cancer cells and represents proof of principle for the evaluation of combinations between the retinoid and check-point inhibitors in the treatment of breast cancer.| File | Dimensione | Formato | |
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2018_12_Vallerga.pdf
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https://hdl.handle.net/10589/145219