Cancer stem cells and the slow cycling phenotype. how to cut the gordian knot driving resistance to therapy in melanoma

Cancer stem cells (CSCs) have historically been defined as slow cycling elements that are able to differentiate into mature cells but without dedifferentiation in the opposite direction. Thanks to advances in genomic and non-genomic technologies, the CSC theory has more recently been reconsidered in a dynamic manner according to a “phenotype switching” plastic model. Transcriptional reprogramming rewires this plasticity and enables heterogeneous tumors to influence cancer progression and to adapt themselves to drug exposure by selecting a subpopulation of slow cycling cells, similar in nature to the originally defined CSCs. This model has been conceptualized for malignant melanoma tailored to explain resistance to target therapies. Here, we conducted a bioinformatics analysis of available data directed to the identification of the molecular pathways sustaining slow cycling melanoma stem cells. Using this approach, we identified a signature of 25 genes that were assigned to four major clusters, namely 1) kinases and metabolic changes, 2) melanoma-associated proteins, 3) Hippo pathway and 4) slow cycling/CSCs factors. Furthermore, we show how a protein−protein interaction network may be the main driver of these melanoma cell subpopulations. Finally, mining The Cancer Genome Atlas (TCGA) data we evaluated the expression levels of this signature in the four melanoma mutational subtypes. The concomitant alteration of these genes correlates with the worst overall survival (OS) for melanoma patients harboring BRAF-mutations. All together these results underscore the potentiality to target this signature to selectively kill CSCs and to achieve disease control in melanoma

The educational value of the rules in handball

The game rules of team sports and the technical and tactical methods applicable to them are indispensable for practicing competitive sports. The correct competition arose from the automatic mechanism of the sanction following the infringement of a game rule which is also aided by the relational dynamics of the individual members of the group who demand the application of the rule to continue playing. The competition can also be self-regulated by the two groups who compete for the victory without even the decisive action of the referee as always happens in training activities. This phenomenon is found only in those contexts where the rule is necessary and alone sufficient to ensure the orderly development of activities. The objective of the study is to identify the significant elements of the handball game rules, and the related technical and tactical behaviours, to identify an inventory of significant behaviours. The method is documentary archival research for the analysis of the game rules of the team sport in question, and a subsequent comparative method between grids of indicators, descriptors and weights that classify rule, technique and tactics. The expected results will focus on the appropriate presence of the significant elements and the commonality or discrepancy between team sports. The data is useful for measuring the quantity of significant behaviours in order to qualitatively elaborate the value of each of them with respect to the other current behaviours of quantitative performance and establish the connections

MicroRNAs in melanoma development and resistance to target therapy

microRNAs constitute a complex class of pleiotropic post-transcriptional regulators of gene expression involved in the control of several physiologic and pathologic processes. Their mechanism of action is primarily based on the imperfect matching of a seed region located at the 5' end of a 21-23 nt sequence with a partially complementary sequence located in the 3' untranslated region of target mRNAs. This leads to inhibition of mRNA translation and eventually to its degradation. Individual miRNAs are capable of binding to several mRNAs and several miRNAs are capable of influencing the function of the same mRNAs. In recent years networks of miRNAs are emerging as capable of controlling key signaling pathways responsible for the growth and propagation of cancer cells. Furthermore several examples have been provided which highlight the involvement of miRNAs in the development of resistance to targeted drug therapies. In this review we provide an updated overview of the role of miRNAs in the development of melanoma and the identification of the main downstream pathways controlled by these miRNAs. Furthermore we discuss a group of miRNAs capable to influence through their respective up- or down-modulation the development of resistance to BRAF and MEK inhibitors

ErbB3 Phosphorylation as Central Event in Adaptive Resistance to Targeted Therapy in Metastatic Melanoma. Early Detection in CTCs during Therapy and Insights into Regulation by Autocrine Neuregulin

In recent years the introduction of target therapies with BRAF and MEK inhibitors (MAPKi) and of immunotherapy with anti-CTLA-4 and anti-PD-1 monoclonal antibodies have dramatically improved survival of metastatic melanoma patients. Despite these changes drug resistance remains a major hurdle. Several mechanisms are at the basis of drug resistance. Particular attention has been devoted over the last years to unravel mechanisms at the basis of adaptive/non genetic resistance occurring in BRAF mutated melanomas upon treatment with to MAPKi. In this paper we focus on the involvement of activation of ErbB3 receptor following early exposure of melanoma cells to BRAF or MEK inhibitors, and the following induction of PI3K/AKT pathway. Although different mechanisms have been invoked in the past at the basis of this activation we show here with a combination of approaches that autocrine production of neuregulin by melanoma cells is a major factor responsible for ErbB3 phosphorylation and downstream AKT activation. Interestingly the kinetic of neuregulin production and of the ensuing ErbB3 phosphorylation is different in different melanoma cell lines which underscores the high degree of tumor heterogeneity. Moreover, heterogeneity is further highlighted by the evidence that in different cell lines neuregulin upregulation can occur at the transcriptional or at the post-transcritpional level. Finally we complement our study by showing with a liquid biopsy assay that circulating tumor cells (CTCs) from melanoma patients undergo upregulation of ErbB3 phosphorylation in vivo shortly after initiation of therapy

Combination of antibodies directed against different ErbB3 surface epitopes prevents the establishment of resistance to BRAF/MEK inhibitors in melanoma

Patients with metastatic melanoma bearing V600 mutations in BRAF oncogene clinically benefit from the treatment with BRAF inhibitors alone or in combination with MEK inhibitors. However, a limitation to such treatment is the occurrence of resistance. Tackling the adaptive changes helping cells survive from drug treatment may offer new therapeutic opportunities. Very recently the ErbB3 receptor has been shown to act as a central node promoting survival of BRAF mutated melanoma. In this paper we first demonstrate that ErbB3/AKT hyperphosphorylation occurs in BRAF mutated melanoma cell lines following exposure to BRAF and/or MEK inhibitors. This strongly correlates with increased transcriptional activation of its ligand neuregulin. Anti-ErbB3 antibodies impair the establishment of de novo cell resistance to BRAF inhibition in vitro. In order to more potently ablate ErbB3 activity we used a combination of two anti-ErbB3 antibodies directed against distinct epitopes of its extracellular domain. These two antibodies in combo with BRAF/MEK inhibitors potently inhibit in vitro cell growth and tumor regrowth after drug withdrawal in an in vivo xenograft model. Importantly, residual tumor masses from mice treated by the antibodies and BRAF/ERK inhibitors combo are characterized almost exclusively by large necrotic areas with limited residual areas of tumor growth. Taken together, our findings support the concept that triple therapy directed against BRAF/MEK/ErbB3 may be able to provide durable control of BRAF mutated metastatic melanoma

Activation of an early feedback survival loop involving phospho-ErbB3 is a general response of melanoma cells to RAF/MEK inhibition and is abrogated by anti-ErbB3 antibodies

BACKGROUND: Treatment of advanced melanoma has been improved with the advent of the BRAF inhibitors. However, a limitation to such treatment is the occurrence of resistance. Several mechanisms have been identified to be responsible for the development of resistance, either MEK-dependent or MEK-independent. In order to overcome resistance due to reactivation of MEK signaling, MEK inhibitors are being clinically developed with promising results. However, also in this case resistance inevitably occurs. It has been recently reported that ErbB3, a member of the EGFR receptor family, may be involved in the establishment of drug resistance. METHODS: Three melanoma cell lines were tested: LOX IMVI (BRAF V600E), MST-L (BRAF V600R) and WM266 (BRAF V600D). Phosphorylation of Receptor Tyrosine Kinases (RTKs) was assessed by an RTK array. Western blot analysis was performed on total protein extracts using anti-ErbB3, anti-AKT and anti-ERK 1/2 antibodies. The expression of neuregulin after vemurafenib treatment was assessed by Real Time PCR and Western blotting. The growth inhibitory effects of vemurafenib, GSK1120212b and/or anti-ErbB3 mAbs were evaluated by in vitro colony formation assays. RESULTS: In the present study we demonstrate that ErbB3 is the main RTK undergoing rapidly hyperphosphorylation upon either treatment with a BRAF inhibitor or with a MEK inhibitor in a panel of melanoma cell lines harboring a variety of V600BRAF mutations and that this results in a strong activation of phospho-AKT. Importantly, ErbB3 activation is fully abrogated by the simultaneous use of anti-ErbB3 monoclonal antibodies, which are also shown to potently synergize with BRAF inhibitors in the inactivation of both AKT and ERK pathways and in the inhibition of melanoma cell growth. We show that upregulation of phospho-ErbB3 is due to an autocrine loop involving increased transcription and production of neuregulin by melanoma cells. CONCLUSIONS: On the basis of these results, we propose that initial co-treatment with BRAF and/or MEK inhibitors and anti-ErbB3 antibodies should be pursued as a strategy to reduce the ErbB3-dependent feedback survival mechanism and enhance duration of clinical response

Dental twinning in the primary dentition: new archaeological cases from Italy

Dental twinning (or “double teeth”) is a rare developmental condition that implies the fusion of two or more adjacent teeth. Clinical literature reports individual cases and extensive population studies to clarify causation, distribution, heritability and differential diagnosis of the different types of dental twinning (i.e. fusion, gemination, and accretion) whereas, documentation for past populations is still scarce. Aims: the present study documents four new archaeological cases of dental twinning of deciduous teeth from four different Italian archeological sites and positions them within the framework of the known literature.Materials and methods: the observed cases include five deciduous teeth from four subadults from Sardinia (Monte Sirai, 7th-4th cent. BCE and Santa Filitica, 7th cent. CE), Campania (Velia, 1st-2nd cent. CE) and Latium (Villamagna, 13th-15th cent. CE). The identification, descriptions and differential diagnoses of the anomalies were performed with the use of morphological analyses and, in one case, radiographic means.Results: all cases fall within the category of double teeth; each involving a different set of processes (gemination and dental fusion), teeth (deciduous central incisors, lateral incisors and canines), locations (upper and lower) and occurrence (unilateral and bilateral). Conclusion: to this day, cases reported in literature of dental twinning in archaeological samples are sparse and limited to specific geographical areas. This study adds four more cases from Italy suggesting such anomalies should be recorded in dental analyses in order to, one day, obtain a more reliable modelling of the frequencies and distributions in past populations

Dental twinning in the primary dentition: new archaeological cases from Italy

Dental twinning (or “double teeth”) is a rare developmental condition that implies the fusion of two or more adjacent teeth. Clinical literature reports individual cases and extensive population studies to clarify causation, distribution, heritability and differential diagnosis of the different types of dental twinning (i.e. fusion, gemination, and accretion) whereas, documentation for past populations is still scarce. Aims: the present study documents four new archaeological cases of dental twinning of deciduous teeth from four different Italian archeological sites and positions them within the framework of the known literature.Materials and methods: the observed cases include five deciduous teeth from four subadults from Sardinia (Monte Sirai, 7th-4th cent. BCE and Santa Filitica, 7th cent. CE), Campania (Velia, 1st-2nd cent. CE) and Latium (Villamagna, 13th-15th cent. CE). The identification, descriptions and differential diagnoses of the anomalies were performed with the use of morphological analyses and, in one case, radiographic means.Results: all cases fall within the category of double teeth; each involving a different set of processes (gemination and dental fusion), teeth (deciduous central incisors, lateral incisors and canines), locations (upper and lower) and occurrence (unilateral and bilateral). Conclusion: to this day, cases reported in literature of dental twinning in archaeological samples are sparse and limited to specific geographical areas. This study adds four more cases from Italy suggesting such anomalies should be recorded in dental analyses in order to, one day, obtain a more reliable modelling of the frequencies and distributions in past populations

Inhibition of Stearoyl-CoA desaturase 1 reverts BRAF and MEK inhibition-induced selection of cancer stem cells in BRAF-mutated melanoma

Combination therapy with BRAF and MEK inhibitors significantly improves survival in BRAF mutated melanoma patients but is unable to prevent disease recurrence due to the emergence of drug resistance. Cancer stem cells (CSCs) have been involved in these long-term treatment failures. We previously reported in lung cancer that CSCs maintenance is due to altered lipid metabolism and dependent upon Stearoyl-CoA-desaturase (SCD1)-mediated upregulation of YAP and TAZ. On this ground, we investigated the role of SCD1 in melanoma CSCs