In vivo evaluation of the role of Delta-like 4/Notch signaling in the development of intestinal tumors

Abstract

Colorectal cancer (CRC) is the third most common malignancy and the second leading cause of cancer-related death in the Western world. Dll4/Notch signaling has been shown to regulate tumor angiogenesis and cancer stem cell maintenance in CRC, but how it affects the intestinal precancerous lesions that lead to CRC initiation is not known. Therefore we evaluated the role of Dll4/Notch pathway during intestinal tumorigenesis. For that we used two well-established mouse models of CRC, the ApcMin/+ autochthonous transgenic model and the azoxymethane plus dextran sodium sulphate chemically induced model of chronic colitis associated-cancer (CAC). First we analyzed the protein expression pattern of Dll4 and other Notch pathway members in these settings relatively to that in the normal gut. Then we evaluated the effect of endothelial-specific or ubiquitous Dll4 deregulation and performed a therapeutic trial with the Dll4 inhibitor Dll4-Fc. This protein was administered alone, and in combination with the epidermal growth factor receptor (EGFR)-specific tyrosine kinase inhibitor erlotinib to assess if the anti-Dll4 therapy mediated vascular defects impaired the delivery of other anti-cancer drugs to the tumors. We observed that the Notch pathway is activated in the two studied models of CRC. The normal protein expression pattern of Notch pathway members in the gut is altered in chronic colitis and in ApcMin/+ and colitis-driven intestinal tumors. Dll4 is the most upregulated ligand in the intestinal adenomas in both models of CRC and is present in both tumor epithelium and stroma. Both Dll4 blockade (endothelial-specific and ubiquitously) and activation (endothelial-specific) have an inhibitory effect on intestinal tumor initiation and growth by promoting a noncompetent vasculature or decreasing the vessel density, respectively. Besides its angiogenic related effects, Dll4/Notch pathway promotes excessive inflammation in CAC, sustains the tumor stem cell pool and tumor proliferation synergistically with Wnt signaling, and inhibits differentiation mainly of the secretory cells. In addition, the effectiveness of erlotinib is not affected by Dll4-Fc, where these therapies additively inhibit intestinal tumorigenesis.

RESUMO - Avaliação in vivo da função da sinalização intercelular Dll4/Notch no desenvolvimento de tumores intestinais - O cancro colo-rectal (CCR) é o terceiro tipo de cancro mais comum e é uma das principais causas de morte no Mundo Ocidental. O CCR geralmente desenvolve-se esporadicamente, mas também pode ser hereditário como na síndrome polipose adenomatosa familiar (PAF). A PAF está associada à ativação da via de sinalização Wnt através de mutações no gene supressor tumoral Adenomatous Polyposis Coli (APC), que também ocorre frequentemente nos CCR esporádicos. O desenvolvimento do CCR também pode estar associado a inflamação crónica, nomeadamente à doença de Crohn (CD) e à colite ulcerativa (UC). Infelizmente o desenvolvimento de novas estratégias terapêuticas ou preventivas que tenham como alvo vias de sinalização críticas no desenvolvimento do CCR continua a ser extremamente necessário. Uma dessas estratégias tem como alvo a angiogénese tumoral. O primeiro agente anti-angiogénico a ser aprovado foi o Bevacizumab, que é usado em combinação com outros fármacos no tratamento do CCR metastático. No entanto, a utilização deste fármaco leva ao aparecimento de efeitos secundários e resistência tumoral e em tumores não metastáticos não se tem mostrado eficaz. Assim, continua a ser necessário desenvolver melhores estratégias terapêuticas anti-angiogénicas...