Lucile Grzesiak

L’athérosclérose est l’une des premières causes des maladies cardiovasculaires. Le développement de complications de plaque telles que la calcification et l’angiogénèse intra-plaque influence grandement la stabilité de la plaque et donc le pronostic vital du patient. Certains facteurs de risque sont connus pour expliquer le développement de ces complications comme le diabète ainsi que la maladie rénale chronique. Cependant, tous les mécanismes impliqués ne sont à ce jour pas élucidés. Il est donc essentiel de comprendre quels sont les mécanismes moléculaires mis en jeu dans la progression de ces lésions. Dans une étude préliminaire et dans la littérature, il a été montré que la LRG1 (leucine-rich α-2 glycoprotéine 1) s’accumulait dans le plasma de patients atteints de diabète de type II mais également dans un contexte de maladie rénale chronique. En premier lieu, nous avons démontré une élévation plasmatique de la LRG1 dans un modèle d’athérosclérose sévère chez la souris (souris ApoE KO nourries avec un régime riche en gras). Nous avons observé la présence de la LRG1 au sein même des plaques d’athérome compliquées et qu’elle se localisait préférentiellement au niveau des régions calcifiées à la fois dans le modèle murin mais aussi dans des plaques carotidiennes humaines. Nous avons montré in vitro que l’expression de la LRG1 était augmentée dans des cellules endothéliales à la suite d’une stimulation pro-inflammatoire avec du TNF-α. De manière intéressante, il a été décrit que la LRG1 pouvait se fixer à un récepteur accessoire de la famille du TGF-β. Sachant que cette voie est fortement impliquée dans la modulation de la calcification dans les cellules musculaires lisses vasculaires (CMLV), nous avons donc étudié le rôle de la LRG1 dans ces cellules. Nous avons identifié que la LRG1 était capable d’induire la calcification des CMLV in vitro en augmentant le contenu calcique et en modulant l’expression de gènes impliqués dans la transition ostéochondrogénique. Pour conclure, nos résultats montrent pour la première fois le rôle de la LRG1 dans la calcification vasculaire ce qui suggère un rôle de cette protéine dans le développement de complications de plaque d’athérome.

The colorectal epithelium is rapidly renewing, with remarkable capacity to regenerate following injury. In colorectal cancer (CRC), this regenerative capacity can be co-opted to drive epithelial plasticity. Although oncogenic MAPK signalling in CRC is common, with frequent mutations of both KRAS (40–50%) and BRAF (10%) ¹ , inhibition of this pathway typically drives resistance clinically. Here, given the development of KRAS inhibitors and licensing of BRAF inhibitor combinations ^2–4 , we have interrogated key mechanisms of resistance to these agents in advanced preclinical CRC models. We show that oncogenic MAPK signalling induces epithelial-state changes in vivo, driving adoption of a regenerative/revival stem-like population, whereas inhibition leads to rapid transcriptional remodelling of both Kras -mutant and Braf- mutant tumours, favouring a WNT-associated, canonical stem phenotype. This drives acute therapeutic resistance in Kras -driven and delayed resistance in Braf -driven models. Where plasticity is restrained, such as in early metastatic disease, or through targeting ligand-dependent WNT pathway Rnf43 mutations, marked therapeutic responses are observed. This explains the super response to BRAF + EGFR-targeted therapies previously observed in a BRAF–RNF43 co-mutant patient population, highlighting the criticality of cellular plasticity in therapeutic response. Together, our data provide clear insight into the mechanisms underpinning resistance to MAPK-targeted therapies in CRC. Moreover, strategies that aim to corral stem cell fate, restrict epithelial plasticity or intervene when tumours lack heterogeneity may improve therapeutic efficacy of these agents.

Atherosclerosis is the primary cause of cardiovascular disease. The development of plaque complications, such as calcification and neo-angiogenesis, strongly impacts plaque stability and is a good predictor of mortality in patients with atherosclerosis. Despite well-known risk factors of plaque complications, such as diabetes mellitus and chronic kidney disease, the mechanisms involved are not fully understood. We and others have identified that the concentration of circulating leucine-rich α-2 glycoprotein 1 (LRG1) was increased in diabetic and chronic kidney disease patients. Using apolipoprotein E knockout mice (ApoE−/−) (fed with Western diet) that developed advanced atherosclerosis and using human carotid endarterectomy, we showed that LRG1 accumulated into an atherosclerotic plaque, preferentially in calcified areas. We then investigated the possible origin of LRG1 and its functions on vascular cells and found that LRG1 expression was specifically enhanced in endothelial cells via inflammatory mediators and not in vascular smooth muscle cells (VSMC). Moreover, we identified that LRG1 was able to induce calcification and SMAD1/5-signaling pathways in VSMC. In conclusion, our results identified for the first time that LRG1 is a direct contributor to vascular calcification and suggest a role of this molecule in the development of plaque complications in patients with atherosclerosis.

Vascular calcification is an important risk factor for cardiovascular (CV) mortality in patients with chronic kidney disease (CKD). It is also a complex process involving osteochondrogenic differentiation of vascular smooth muscle cells (VSMCs) and abnormal deposition of minerals in the vascular wall. In an observational, multicenter European study, including 112 patients with CKD from Spain and 171 patients on dialysis from France, we used serum proteome analysis and further validation by ELISA to identify calprotectin, a circulating damage-associated molecular pattern protein, as being independently associated with CV outcome and mortality. This was confirmed in an additional cohort of 170 patients with CKD from Sweden, where increased serum calprotectin concentrations correlated with increased vascular calcification. In primary human VSMCs and mouse aortic rings, calprotectin exacerbated calcification. Treatment with paquinimod, a calprotectin inhibitor, as well as pharmacological inhibition of the receptor for advanced glycation end products and Toll-like receptor 4 inhibited the procalcifying effect of calprotectin. Paquinimod also ameliorated calcification induced by the sera of uremic patients in primary human VSMCs. Treatment with paquinimod prevented vascular calcification in mice with chronic renal failure induced by subtotal nephrectomy and in aged apolipoprotein E–deficient mice as well. These observations identified calprotectin as a key contributor of vascular calcification, and increased circulating calprotectin was strongly and independently associated with calcification, CV outcome, and mortality in patients with CKD. Inhibition of calprotectin might therefore be a promising strategy to prevent vascular calcification in patients with CKD.