Targeting the hedgehog transcription factors GLI1 and GLI2 restores sensitivity to vemurafenib-resistant human melanoma cells

BRAF inhibitor (BRAFi) therapy for melanoma patients harboring the V600E mutation is initially highly effective, but almost all patients relapse within a few months. Understanding the molecular mechanisms underpinning BRAFi-based therapy is therefore an important issue. Here we identified a previously unsuspected mechanism of BRAFi resistance driven by elevated Hedgehog (Hh) pathway activation that is observed in a cohort of melanoma patients after vemurafenib treatment. Specifically, we demonstrate that melanoma cell lines, with acquired in vitro-induced vemurafenib resistance, show increased levels of glioma-associated oncogene homolog 1 and 2 (GLI1/GLI2) compared with naive cells. We also observed these findings in clinical melanoma specimens. Moreover, the increased expression of the transcription factors GLI1/GLI2 was independent of canonical Hh signaling and was instead correlated with the noncanonical Hh pathway, involving TGF beta/SMAD (transforming growth factor-beta/Sma- and Mad-related family) signaling. Knockdown of GLI1 and GLI2 restored sensitivity to vemurafenib-resistant cells, an effect associated with both growth arrest and senescence. Treatment of vemurafenib-resistant cells with the GLI1/GLI2 inhibitor Gant61 led to decreased invasion of the melanoma cells in a three-dimensional skin reconstruct model and was associated with a decrease in metalloproteinase (MMP2/MMP9) expression and microphthalmia transcription factor upregulation. Gant61 monotherapy did not alter the drug sensitivity of naive cells, but could reverse the resistance of melanoma cells chronically treated with vemurafenib. We further noted that alternating dosing schedules of Gant61 and vemurafenib prevented the onset of BRAFi resistance, suggesting that this could be a potential therapeutic strategy for the prevention of therapeutic escape. Our results suggest that targeting the Hh pathway in BRAFi-resistant melanoma may represent a viable therapeutic strategy to restore vemurafenib sensitivity, reducing or even inhibiting the acquired chemoresistance in melanoma patients.Fapesp-grant number 2012/04194-1, 2013/05172-4, 2014/24400-0 and 2015/10821-7, CNPq-grant number 150447/2013-2 and 471512/2013-3 and PRODOC-grant no 3193-32/2010. Work in the lab of KS Smalley was supported by the National Institutes of Health grants R01 CA161107, R21 CA198550, and Skin SPORE grant P50 CA168536info:eu-repo/semantics/publishedVersio

Physicochemical characteristics of citrus jelly with non cariogenic and functional sweeteners

In this study the effect of sweeteners with low glycemic index and non-cariogenic characteristics (isomaltulose, oligofructose and tagatose) in jelly prepared with citrus juice has been evaluated considering a citrus jelly formulated with sucrose as reference. The soluble solids, moisture content, pH, water activity, antioxidant capacity, optical and mechanical properties of jelly made using different sweeteners was determined during storage. Besides, mesophilic aerobics and moulds and yeasts was also counted to determine their stability over time. Sensory evaluation of the citrus jelly has also been done. The results showed the antioxidant activity decreased during storage in all formulations. Tagatose increased lightness whereas coordinates a*, b* and chrome of all the jellies prepared using new sweeteners were lower than jellies with sucrose. However, citrus jelly with only oligofructose or tagatose or with the mixture of isomaltulose and tagatose were most closely resembled to the control jelly with respect to mechanical properties. Jelly prepared with the combination of isomaltulose and tagatose in equal proportions obtained the best score in the sensorial analysis.The authors would like to thank the Serigo-Andres family for donating the raw materials, and also the GVA projects GV/2013/029, GV/2014/012 as well as the Universitat Politecnica de Valencia (Spain) for the financial support given to this research study (UPV PAID-06-12 SP20120889).Rubio-Arraez, S.; Capella Hernández, JV.; Castelló Gómez, ML.; Ortolá Ortolá, MD. (2016). Physicochemical characteristics of citrus jelly with non cariogenic and functional sweeteners. 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