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Failure of normal intestinal homeostasis under the influence of tumour cell-derived paracrine signals

Abstract : Epithelial homeostasis and tumorigenesis are two intertwined concepts. Indeed, the formation of a tumour and its progression to aggressive stages are the consequence of a loss of control of the spatial and mechanical interactions of epithelial cells with their environment. Such perturbed tissue homeostasis can have two origins: an intrinsic one, often due to genetic mutations, causing mutant cells to lose the ability to correctly interpret environment signals, and an extrinsic or non-cell autonomous cause, as the environment surrounding mutant cells can no longer provides coherent information to correctly orchestrate tissue homeostasis.Previous results in the lab indicated that some tumour cells transcriptionally resemble normal stem cells. I was intrigued by this observation and decided to study the molecular basis of intratumoral heterogeneity, keeping in mind that normal-like stem cells could be present within tumours. My PhD was focused on examining interactions between normal and tumour epithelial cells, using the stroma-free model of organotypic cultures. During these studies, I discovered and characterised a hitherto unknown mechanism of cellular communication between tumour epithelial cells and genetically wild type epithelial cells in the context of colorectal cancer.Taking advantage of the intestinal organoid model system, allowing in vitro study of epithelial cells organising in a defined micro-environmental context free of stromal cells, I identified a rapid "transforming" effect of tumour cells on genetically wild type intestinal stem cells. I then demonstrated that this fast and reversible transformation was mediated by a secreted protein, and evaluated by SILAC mass spectrometry which proteins were specifically secreted by tumour but not normal organoids. This high-throughput quantitative analysis allowed us to identify a factor that was necessary for the observed transformation: thrombospondin-1 (Thbs1). Indeed, inhibition of Thbs1 by neutralizing antibodies or by genetic knock-out was sufficient to abolish the transforming capacity of tumour organoids. Transformation of wild type organoids by tumour organoids is manifested by a morphological change resulting in loss of cell polarisation and formation of hollow cysts, but also by a loss of compartmentalisation of proliferative cells, normally restricted to the crypt regions of organoids. In order to understand how Thbs1 induced such a change, I then analysed the transcriptome of transformed organoids by RNA sequencing and showed a specific activation of the Hippo signalling pathway in response to tumour-derived conditioned medium. This study shows how tumour cells can induce genetically wild type cells to switch to a tumour-like behaviour, using signal such as Hippo pathway activation normally employed during regeneration, making non-mutant cells dangerously adapted to survive and proliferate in a tumoral context.This work provides original mechanistic insights into the processes of early tumour remodelling and epithelial cell communication independently of stromal cells. The molecular mechanisms we have unveiled support the hypothesis that wild type stem cells can co-exist with mutant cells in tumours and contribute to tumour growth and clonal expansion, thanks to paracrine factors (like Thbs1) secreted by surrounding tumour cells, which allow them to thrive in the tumour environment
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Guillaume Jacquemin. Failure of normal intestinal homeostasis under the influence of tumour cell-derived paracrine signals. Cellular Biology. Université Paris sciences et lettres, 2019. English. ⟨NNT : 2019PSLET017⟩. ⟨tel-02873486⟩

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