Post-translational deregulation of YAP1 is genetically controlled in rat liver cancer and determines the fate and stem-like behavior of the human disease

Previous studies showed that YAP1 is over-expressed in hepatocellular carcinoma (HCC). Here we observed higher expression of Yap1/Ctgf axis in dysplastic nodules and HCC chemically-induced in F344 rats, genetically susceptible to hepatocarcinogenesis, than in lesions induced in resistant BN rats. In BN rats, highest increase in Yap1-tyr357, p73 phosphorylation and Caspase 3 cleavage occurred. In human HCCs with poorer prognosis ( 3 years survival; HCCB). In the latter, higher levels of phosphorylated YAP1-ser127, YAP1-tyr357 and p73, YAP1 ubiquitination, and Caspase 3 cleavage occurred. Expression of stemness markers NANOG, OCT-3/4, and CD133 were highest in HCCP and correlated with YAP1 and YAP1-TEAD levels. In HepG2, Huh7, and Hep3B cells, forced YAP1 over-expression led to stem cell markers expression and increased cell viability, whereas inhibition of YAP1 expression by specific siRNA, or transfection of mutant YAP1 which does not bind to TEAD, induced opposite alterations. These changes were associated, in Huh7 cells transfected with YAP1 or YAP1 siRNA, with stimulation or inhibition of cell migration and invasivity, respectively. Furthermore, transcriptome analysis showed that YAP1 transfection in Huh7 cells induces over-expression of genes involved in tumor stemness. In conclusion, Yap1 post-translational modifications favoring its ubiquitination and apoptosis characterize HCC with better prognosis, whereas conditions favoring the formation of YAP1-TEAD complexes are associated with aggressiveness and acquisition of stemness features by HCC cells

Trilayer dissolving polymeric microneedle array loading Rose Bengal transfersomes as a novel adjuvant in early-stage cutaneous melanoma management

Melanoma remains a global concern, but current therapies present critical limitations pointing out the urgent need for novel strategies. Among these, the cutaneous delivery of drugs selectively damaging cancer cells is highly attractive. Rose Bengal (RB) is a dye exhibiting selective cytotoxicity towards melanoma, but the high water solubility and low permeability hinder its therapeutic potential. We previously developed RB-loaded transfersomes (RBTF) to mediate the RB dermal delivery; however, a platform efficiently delivering RBTF in the deepest strata is essential for a successful therapeutic activity. In this regard, dissolving microneedles release the encapsulated cargo up to the dermis, painlessly piercing the outmost skin layers. Therefore, herein we developed and characterised a trilayer dissolving microneedle array (RBTF-TDMNs) loading RBTF to maximise RBTF intradermal delivery in melanoma management. RBTF-TDMNs were proven strong enough to pierce excised porcine skin and rapidly dissolve and deposit RBTF intradermally while maintaining their physicochemical properties. Also, 3D visualisation of the system itself and while penetrating the skin was performed by multi-photon microscopy. Finally, a dermatokinetic study showed that RBTF-TDMNs offered unique delivery efficiency advantages compared to RBTF dispersion and free drug-loaded TDMNs. The proposed RBTF-TDMNs represent a valuable potential adjuvant tool for the topical management of melanoma

Inhomogeneous Diastereomeric Composition of Mongersen Antisense Phosphorothioate Oligonucleotide Preparations and Related Pharmacological Activity Impairment

Mongersen is a 21-mer antisense oligonucleotide designed to downregulate Mothers against decapentaplegic homolog 7 (SMAD7) expression to treat Crohn's disease. Mongersen was manufactured in numerous batches at different scales during several years of clinical development, which all appeared identical, using common physicochemical analytical techniques, while only phosphorous-31 nuclear magnetic resonance (P-31-NMR) in solution showed marked differences. Close-up analysis of 27 mongersen batches revealed marked differences in SMAD7 downregulation in a cell-based assay. Principal component analysis of P-31-NMR profiles showed strong correlation with SMAD7 downregulation and, therefore, with pharmacological efficacy in vitro. Mongersen contains 20 phosphorothioate (PS) linkages, whose chirality (Rp/Sp) was not controlled during manufacturing. A different diastereomeric composition throughout batches would lead to superimposable analytical data, but to distinct P-31-NMR profiles, as indeed we found. We tentatively suggest that this may be the origin of different biological activity. As similar manifolds are expected for other PS-based oligonucleotides, the protocol described here provides a general method to identify PS chirality issues and a chemometric tool to score each preparation for this elusive feature

The Palaeocene Cerro Munro tonalite intrusion (Chubut Province, Argentina): A plutonic remnant of explosive volcanism?

The Cerro Munro sub-volcanic intrusion is emplaced in the back-arc (400 km from the trench) as small sub-circular tonalite-granodiorite plutons with abundant radial porphyritic dikes. U-Pb zircon SHRIMP data give an age of crystallization of 57 Ma ± 1.4 Ma. It is located to the east of the North Patagonian Batholith (NPB) that shows a protracted and episodic magmatic history from Cretaceous to Miocene time. The NPB Palaeogene episode is characterized by the lack of magmatic activity at the arc axis, as small plutonic emplacements move to the fore-arc and back-arc. This Palaeogene tectono-magmatic episode is ruled by the detachment of the Aluk plate during the Aluk-Farallon-SAM triple junction, active at that time along northern Patagonia active margin, changing the Cretaceous ?NPB orogenic? setting to a Palaeogene ?Munro transitional? tectono-magmatic setting. Near the contacts, the tonalite contains abundant enclaves of igneous appearance and variable size from several cm to dm, described as autoliths. The study of autoliths and host tonalite reveals interesting results on the processes of fractionation in a thermally zoned magma chamber. Autoliths, and in a large extent the host tonalite, represent disguised cumulates from which a hydrous silicic liquid was extracted. Barometry calculations from mineral chemistry in both autoliths and tonalites record a shallow pressure of emplacement of 0.5 kbar. Rhyolite-dacite flows and ignimbrites, surrounding the northern contact of the Cerro Munro tonalite, may represent the exsolved liquid from the plutonic cumulates. The study by cathodoluminiscence and electron backscattered diffraction techniques from a rhyolite-hosted quartz supports this protracted history of the Cerro Munro magma chamber.Facultad de Ciencias Naturales y MuseoCentro de Investigaciones GeológicasConsejo Nacional de Investigaciones Científicas y Técnica

The Palaeocene Cerro Munro tonalite intrusion (Chubut Province, Argentina): A plutonic remnant of explosive volcanism?

The Cerro Munro sub-volcanic intrusion is emplaced in the back-arc (400 km from the trench) as small sub-circular tonalite-granodiorite plutons with abundant radial porphyritic dikes. U-Pb zircon SHRIMP data give an age of crystallization of 57 Ma ± 1.4 Ma. It is located to the east of the North Patagonian Batholith (NPB) that shows a protracted and episodic magmatic history from Cretaceous to Miocene time. The NPB Palaeogene episode is characterized by the lack of magmatic activity at the arc axis, as small plutonic emplacements move to the fore-arc and back-arc. This Palaeogene tectono-magmatic episode is ruled by the detachment of the Aluk plate during the Aluk-Farallon-SAM triple junction, active at that time along northern Patagonia active margin, changing the Cretaceous ?NPB orogenic? setting to a Palaeogene ?Munro transitional? tectono-magmatic setting. Near the contacts, the tonalite contains abundant enclaves of igneous appearance and variable size from several cm to dm, described as autoliths. The study of autoliths and host tonalite reveals interesting results on the processes of fractionation in a thermally zoned magma chamber. Autoliths, and in a large extent the host tonalite, represent disguised cumulates from which a hydrous silicic liquid was extracted. Barometry calculations from mineral chemistry in both autoliths and tonalites record a shallow pressure of emplacement of 0.5 kbar. Rhyolite-dacite flows and ignimbrites, surrounding the northern contact of the Cerro Munro tonalite, may represent the exsolved liquid from the plutonic cumulates. The study by cathodoluminiscence and electron backscattered diffraction techniques from a rhyolite-hosted quartz supports this protracted history of the Cerro Munro magma chamber.Facultad de Ciencias Naturales y MuseoCentro de Investigaciones GeológicasConsejo Nacional de Investigaciones Científicas y Técnica

Geochemical and metallogenetical study of the pegmatites and associated granites from southern Comechingones pegmatitic field, Córdoba

El distrito pegmatítico Comechingones, ubicado en el faldeo oriental de la sierra homónima, en la provincia de Córdoba, involucra pegmatitas graníticas correspondientes a la clase de Elementos Raros, tipo berilo, subtipo berilo-columbita-fosfatos, algunas en transición a la clase muscovítica, con mineralizaciones de Be-Nb- Ta-U y minerales industriales. Dos tipos de pegmatitas graníticas han sido descriptas en el sector sur del distrito: pegmatitas tipo I, con tamaños que en total pueden alcanzan los 1000 metros de longitud y superar los 50 de ancho, internamente zonadas y portadoras de Be, Nb-Ta y U; y pegmatitas tipo II, de menores dimensiones, no zonadas, ricas en cuarzo de alta pureza, carentes de mineralizaciones metalíferas, y asociadas espacial y genéticamente con leucogranitos aplíticos. En este trabajo se presentan y discuten los datos geoquímicos preliminares de ambos tipos de pegmatitas y granitos asociados. Los datos geoquímicos obtenidos, apoyados con descripciones de campo y petrográficas, permiten establecer que las dos tipologías de pegmatitas corresponden a dos eventos magmáticos diferentes (muy probablemente diacrónicos). El primero generó las pegmatitas tipo I, las cuales de sur a norte presentan un aumento en el grado de fraccionamiento desde pegmatitas poco evolucionadas y sin mineralizaciones de elementos raros, hasta pegmatitas evolucionadas con depósitos metalíferos de interés económico. El segundo evento dio origen a las pegmatitas tipo II y a los granitos aplíticos, y carece de especialización metalogenética, evidenciado por los indicadores de diferenciación magmática sistemáticamente inferiores a los de las pegmatitas tipo I y a la carencia de mineralizaciones metalíferas.The Comechingones pegmatitic field (CPF) is located in the eastern flank of the Sierra de Comechingones, Córdoba province. It is composed of granite pegmatites belonging to the Rare-Element class, beryl type, beryl-columbite-phosphate subtype; some of them are transitional into the Muscovite class. Beryllium, Nb, Ta and U deposits, as well as high-quality industrial mineral deposits, are frequently associated with these pegmatites. In the southern part of the CPF two different pegmatite types have been described. Type I pegmatites constitute large zoned bodies with up to 1000 m long and 50 m thick, and may constitute rare element deposits, whereas type II pegmatites occur as small, unzoned quartz-rich dykes, without metalliferous mineralizations, spatial and genetically associated with aplitic leucogranites. Preliminary geochemical data from both pegmatites types and granites are presented and discussed in this contribution. Geochemical evidences, supported by field and petrographic observations, suggest that the two types of pegmatites identified in the study area represent two different, probably diachronic, magmatic stages. Type I pegmatites display a geochemical gradation in a S-N direction, from barren pegmatites in the south to fractionated pegmatites in the northern part of the study area, and are the lithological product of the first magmatic stage. The second stage lead to the crystallization of aplitic granites and barren type II pegmatites, geochemically less fractionated than type II pegmatites.Centro de Investigaciones Geológica

A Single-Molecule Bioelectronic Portable Array for Early Diagnosis of Pancreatic Cancer Precursors

A cohort of 47 patients is screened for pancreatic cancer precursors with a portable 96-well bioelectronic sensing-array for single-molecule assay in cysts fluid and blood plasma, deployable at point-of-care (POC). Pancreatic cancer precursors are mucinous cysts diagnosed with a sensitivity of at most 80% by state-of-the-art cytopathological molecular analyses (e.g., KRASmut DNA). Adding the simultaneous assay of proteins related to malignant transformation (e.g., MUC1 and CD55) is deemed essential to enhance diagnostic accuracy. The bioelectronic array proposed here, based on single-molecule-with-a-large-transistor (SiMoT) technology, can assay both nucleic acids and proteins at the single-molecule limit-of-identification (LOI) (1% of false-positives and false-negatives). It comprises an enzyme-linked immunosorbent assay (ELISA)-like 8 × 12-array organic-electronics disposable cartridge with an electrolyte-gated organic transistor sensor array, and a reusable reader, integrating a custom Si-IC chip, operating via software installed on a USB-connected smart device. The cartridge is complemented by a 3D-printed sensing gate cover plate. KRASmut, MUC1, and CD55 biomarkers either in plasma or cysts-fluid from 5 to 6 patients at a time, are multiplexed at single-molecule LOI in 1.5 h. The pancreatic cancer precursors are classified via a machine-learning analysis resulting in at least 96% diagnostic-sensitivity and 100% diagnostic-specificity. This preliminary study opens the way to POC liquid-biopsy-based early diagnosis of pancreatic-cancer precursors in plasma

Characterisation and radioimmunotherapy of L19-SIP, an anti-angiogenic antibody against the extra domain B of fibronectin, in colorectal tumour models

Angiogenesis is a characteristic feature of tumours and other disorders. The human monoclonal antibody L19- SIP targets the extra domain B of fibronectin, a marker of angiogenesis expressed in a range of tumours. The aim of this study was to investigate whole body distribution, tumour localisation and the potential of radioimmunotherapy with the L19-small immunoprotein (SIP) in colorectal tumours. Two colorectal tumour models with highly different morphologies, the SW1222 and LS174T xenografts, were used in this study. Localisation and retention of the L19-SIP antibody at tumour vessels was demonstrated using immunohistochemistry and Cy3-labelled L19-SIP. Whole body biodistribution studies in both tumour models were carried out with 125I-labelled L19-SIP. Finally, 131I-labelled antibody was used to investigate the potential of radioimmunotherapy in SW1222 tumours. Using immunohistochemistry, we confirmed extra domain B expression in the tumour vasculature. Immunofluorescence demonstrated localisation and retention of injected Cy3-labelled L19-SIP at the abluminal side of tumour vessels. Biodistribution studies using a 125I-labelled antibody showed selective tumour uptake in both models. Higher recorded values for localisation were found in the SW1222 tumours than in the LS174T (7.9 vs 6.6 %ID g−1), with comparable blood clearance for both models. Based on these results, a radioimmunotherapy study was performed in the SW1222 xenograft using 131I-Labelled L19-SIP (55.5 MBq), which showed selective tumour uptake, tumour growth inhibition and improved survival. Radio- and fluorescence-labelled L19-SIP showed selective localisation and retention at vessels of two colorectal xenografts. Furthermore, 131I-L19-SIP shows potential as a novel treatment of colorectal tumours, and provides the foundation to investigate combined therapies in the same tumour models