Clinical and Functional Characterization of a Patient Carrying a Compound Heterozygous Pericentrin Mutation and a Heterozygous IGF1 Receptor Mutation

Intrauterine and postnatal longitudinal growth is controlled by a strong genetic component that regulates a complex network of endocrine factors integrating them with cellular proliferation, differentiation and apoptotic processes in target tissues, particularly the growth centers of the long bones. Here we report on a patient born small for gestational age (SGA) with severe, proportionate postnatal growth retardation, discreet signs of skeletal dysplasia, microcephaly and moyamoya disease. Initial genetic evaluation revealed a novel heterozygous IGF1R p.Leu1361Arg mutation affecting a highly conserved residue with the insulin-like growth factor type 1 receptor suggestive for a disturbance within the somatotropic axis. However, because the mutation did not co-segregate with the phenotype and functional characterization did not reveal an obvious impairment of the ligand depending major IGF1R signaling capabilities a second-site mutation was assumed. Mutational screening of components of the somatotropic axis, constituents of the IGF signaling system and factors involved in cellular proliferation, which are described or suggested to provoke syndromic dwarfism phenotypes, was performed. Two compound heterozygous PCNT mutations (p.[Arg585X];[Glu1774X]) were identified leading to the specification of the diagnosis to MOPD II. These investigations underline the need for careful assessment of all available information to derive a firm diagnosis from a sequence aberration

SPARC functions as an inhibitor of adipogenesis

Adipogenesis, a key step in the pathogenesis of obesity, involves extensive ECM remodeling, changes in cell-ECM interactions, and cytoskeletal rearrangement. Matricellular proteins regulate cell-cell and cell-ECM interactions. Evidence in vivo and in vitro indicates that the prototypic matricellular protein, SPARC, inhibits adipogenesis and promotes osteoblastogenesis. Herein we discuss mechanisms underlying the inhibitory effect of SPARC on adipogenesis. SPARC enhances the Wnt/β-catenin signaling pathway and regulates the expression and posttranslational modification of collagen. SPARC might drive preadipocytes away from the status of growth arrest and therefore prevent terminal differentiation. SPARC could also decrease WAT deposition through its negative effects on angiogenesis. Therefore, several stages of white adipose tissue accumulation are sensitive to the inhibitory effects of SPARC

Cancer cell-derived long pentraxin 3 (PTX3) promotes melanoma migration through a toll-like receptor 4 (TLR4)/NF-kappa B signaling pathway

Cutaneous melanoma is one of the most aggressive cancers characterized by a high plasticity, a propensity for metastasis, and drug resistance. Melanomas are composed of phenotypically diverse subpopulations of tumor cells with heterogeneous molecular profiles that reflect intrinsic invasive abilities. In an attempt to identify novel factors of the melanoma invasive cell state, we previously investigated the nature of the invasive secretome by using a comparative proteomic approach. Here, we have extended this analysis to show that PTX3, an acute phase inflammatory glycoprotein, is one such factor secreted by invasive melanoma to promote tumor cell invasiveness. Elevated PTX3 production was observed in the population of MITFlow invasive cells but not in the population of MITFhigh differentiated melanoma cells. Consistently, MITF knockdown increased PTX3 expression in MITFhigh proliferative and poorly invasive cells. High levels of PTX3 were found in tissues and blood of metastatic melanoma patients, and in BRAF inhibitor-resistant melanoma cells displaying a mesenchymal invasive MITFlow phenotype. Genetic silencing of PTX3 in invasive melanoma cells dramatically impaired migration and invasion in vitro and in experimental lung extravasation assay in xenografted mice. In contrast, addition of melanoma-derived or recombinant PTX3, or expression of PTX3 enhanced motility of low migratory cells. Mechanistically, autocrine production of PTX3 by melanoma cells triggered an IKIC/NF kappa B signaling pathway that promotes migration, invasion, and expression of the EMT factor TWIST1. Finally, we found that TLR4 and MYD88 knockdown inhibited PTX3-induced melanoma cell migration, suggesting that PTX3 functions through a TLR4-dependent pathway. Our work reveals that tumor-derived PTX3 contributes to melanoma cell invasion via targetable inflammation-related pathways. In addition to providing new insights into the biology of melanoma invasive behavior, this study underscores the notion that secreted PTX3 represents a potential biomarker and therapeutic target in a subpopulation of MITFlow invasive and/or refractory melanoma

Hypoxia and MITF control metastatic behaviour in mouse and human melanoma cells

International audienceMelanomas are very aggressive neoplasms with notorious resistance to therapeutics. It was recently proposed that the remarkable phenotypic plasticity of melanoma cells allows for the rapid development of both resistance to chemotherapeutic drugs and invasive properties. Indeed, the capacity of melanoma cells to form distant metastases is the main cause of mortality in melanoma patients. Therefore, the identification of the mechanism controlling melanoma phenotype is of paramount importance. In the present report, we show that deletion of microphthalmia-associated transcription factor (MITF), the master gene in melanocyte differentiation, is sufficient to increase the metastatic potential of mouse and human melanoma cells. MITF silencing also increases fibronectin and Snail, two mesenchymal markers that might explain the increased invasiveness in vitro and in vivo. Furthermore, ablation of this population by Forskolin-induced differentiation or MITF-forced expression significantly decreases tumour and metastasis formation, suggesting that eradication of low-MITF cells might improve melanoma treatment. Moreover, we demonstrate that a hypoxic microenvironment decreases MITF expression through an indirect, hypoxia-inducible factor 1 (HIF1)α-dependant transcriptional mechanism, and increases the tumourigenic and metastatic properties of melanoma cells. We identified Bhlhb2, a new factor in melanoma biology, as the mediator of hypoxia/HIF1α inhibitory effect on MITF expression. Our results reveal a hypoxia-HIF1α-BHLHB2-MITF cascade controlling the phenotypic plasticity in melanoma cells and favouring metastasis development. Targeting this pathway might be helpful in the design of new anti-melanoma therapies

Two distinct mutations of the RET receptor causing Hirschsprung's disease impair the binding of signalling effectors to a multifunctional docking site.

The RET gene codes for a transmembrane tyrosine kinase which is a subunit of a multimeric complex that acts as a receptor for four structurally related molecules: the glial cell line-derived neurotrophic factor (GDNF), neurturin, artemin and persephin. Germline mutations of RET cause a dominantly inherited dysgenesis of the enteric nervous system known as Hirschsprung's disease (HSCR; aganglionosis megacolon). The majority of HSCR mutations results either in a reduction of dosage of the RET protein or in the loss of RET function. Two novel distinct mutations of RET that led either to the deletion of codon 1059 (denoted Delta1059) or to the substitution of a Pro for Leu1061 have been identified in five HSCR families. In one large pedigree, two children born from asymptomatic consanguineous parents presented a severe form of HSCR and were found to carry the mutation at codon 1061 in the homozygous state. A tyrosine residue at position 1062 is an intracytoplasmic docking site that enables RET to recruit several signalling molecules, including the Shc adaptor protein. We now report that both HSCR mutations impair the fixation of Shc to RET and consequently prevent its phosphorylation. In addition, quantitative analysis in PC12 cells reveals that mutation Delta1059 inactivates the ability of RET to transduce a downstream signal whereas mutation L1061P only partially inhibits the signalling of RET. Finally, we provide evidence that these effects are partly mediated via the disruption of the RET/Shc interaction. Collectively, these results demonstrate that HSCR can be ascribed to mutations of RET which interfere with the binding of transduction effectors, such as Shc, and further provide a biochemical explanation for the phenotype of patients carrying a homozygous mutation at codon 1061. Finally, these data indicate that Y1062 is a multifunctional docking site that confers to RET the capacity to engage downstream signalling pathways which exert a crucial role during enteric neurogenesis

In vitro and in vivo anti-melanoma effects of ciglitazone

Activation of PPARgamma by synthetic ligands, thiazolidinediones, inhibits the proliferation of cancer cells. In this report, focusing our attention on ciglitazone, we show that ciglitazone inhibits melanoma growth by inducing apoptosis and cell-cycle arrest, whereas normal melanocytes are resistant to ciglitazone. In melanoma cells, ciglitazone-induced apoptosis is associated with caspase activations and a loss of mitochondrial membrane potential. Induction of cell-cycle arrest by ciglitazone is associated with changes in expression of key cell-cycle regulators such as p21, cyclin D1, and pRB hypophosphorylation. Cell-cycle arrest occurs at low ciglitazone concentrations and through a PPARgamma-dependent pathway, whereas the induction of apoptosis is caused by higher ciglitazone concentrations and independently of PPARgamma. These results allow an effective molecular dissociation between proapoptotic effects and growth inhibition evoked by ciglitazone in melanoma cells. Finally, we show that in vivo treatment of nude mice by ciglitazone dramatically inhibits human melanoma xenograft development. The data presented suggest that ciglitazone might be a better candidate for clinical trials in melanoma treatment than the thiazolidinediones currently used in the treatment of type 2 diabetes, such as rosiglitazone, which is devoid of a proapoptotic PPARgamma-independent function

Metformin Blocks Melanoma Invasion and Metastasis Development in AMPK/p53-Dependent Manner

International audienceMetformin was reported to inhibit the proliferation of many cancer cells, including melanoma cells. In this report, we investigated the effect of metformin on melanoma invasion and metastasis development. Using different in vitro approaches, we found that metformin inhibits cell invasion without affecting cell migration and independently of antiproliferation action. This inhibition is correlated with modulation of expression of proteins involved in epithelial-mesenchymal transition such as Slug, Snail, SPARC, fibronectin, and N-cadherin and with inhibition of MMP-2 and MMP-9 activation. Furthermore, our data indicate that this process is dependent on activation of AMPK and tumor suppressor protein p53. Finally, we showed that metformin inhibits melanoma metastasis development in mice using extravasation and metastasis models. The presented data reinforce the fact that metformin might be a good candidate for clinical trial in melanoma treatment