Inhibitors of EYA3 Protein in Ewing Sarcoma.

OBJECTIVE: Among sarcomas, Ewing sarcoma (EWS) is characterized as a highly malignant type of bone tumor caused by the fusion of EWS RNA Binding Protein-1 (EWSR1)/ Friend leukemia integration 1 (FLI1) genes. The product of fusion gene gives rise to EWSR1/FLI1 which activates the activity of Eyes absent homolog 3 (EYA3) which causes tumor growth and angiogenesis. EYA3 is now considered as a therapeutic drug target for EWS . The study was designed to gather potential inhibitors for the EYA3 target using medicinal compounds. METHODS: In this study, we have obtained a list of medicinal compounds from the NuBBE database and downloaded their structural information. Then insilico screening analysis of >2,000 medicinal compounds was performed with PyRX virtual drug screening software to discover potential inhibitors for the treatment of EWS. RESULTS: Our investigation revealed that Sorbifolin and 1,7-Dihydroxy-3-methylanthracene-9.10-dione show interactive affinity for EYA3 active residues. Moreover, these compounds have adequate toxicity, can induce cytotoxicity in EWS cells, and are capable of regulating the expression of genes activated by EWSR1/FLI1. CONCLUSION: Our study concluded that Sorbifolin and 1,7-Dihydroxy-3-methylanthracene-9.10-dione are promising drug candidates for the treatment of EWS and should be further subjected to invitro testing.

Characterization of three sialidases from Danio rerio.

Zebrafish encodes several sialidases belonging to the NEU3 group, the plasma membrane-associated member of the family with high specificity toward ganglioside substrates. Neu3.1, Neu3.2 and Neu 3.3 have been expressed in E. coli and purified using the pGEX-2T expression system. Although all the enzymes are expressed by bacterial cells, Neu3.1 formed insoluble aggregates that hampered its purification. Neu3.2 and Neu3.3 formed oligomers as demonstrated by gel filtration chromatography experiments. Actually, the first formed a trimer whereas the second a pentamer. Intriguingly, despite relevant degree of sequence identity and similarity, the two enzymes showed peculiar substrate specificities toward gangliosides other than GM3, two glycoproteins and two forms of sialyllactose. Using molecular modelling and the crystal structure of the human cytosolic sialidase NEU2 as a template, the 3D models of the sialidases from zebrafish have been generated. As expected, the 3D models showed the typical six blade beta-propeller typical of sialidases, with an overall highly conserved active site architecture. The differences among the three zebrafish enzymes and human NEU2 are mainly located in the loops connecting the antiparallel beta strands of the propeller core. These portions of the proteins are probably responsible for the differences observed in substrate specificities, as well as in the different subcellular localization and aggregation features observed in solution. Finally, the in silico analysis of RNA-Seq data evidenced a peculiar expression profile of the three genes during embryogenesis, suggesting different roles of these sialidases during development.

Mitochondrial Adaptations in Elderly and Young Men Skeletal Muscle Following 2 Weeks of Bed Rest and Rehabilitation.

The aim of the study was to evaluate the expression levels of proteins related to mitochondrial biogenesis regulation and bioenergetics in vastus lateralis muscle biopsies from 16 elderly and 7 young people subjected to 14 days of bed-rest, causing atrophy, and subsequent 14 days of exercise training. Based on quantitative immunoblot analyses, in both groups a reduction of two key regulators of mitochondrial biogenesis/remodeling and activity, namely PGC-1α and Sirt3, was revealed during bed-rest, with a subsequent up-regulation after rehabilitation, indicating an involvement of PGC-1α-Sirt3 axis in response to the treatments. A difference was observed comparing the young and elderly subjects as, for both proteins, the abundance in the elderly was more affected by immobility and less responsive to exercise. The expression levels of TOM20 and Citrate Synthase, assayed as markers of outer mitochondrial membrane and mitochondrial mass, showed a noticeable sensitivity in the elderly group, where they were affected by bed-rest and rehabilitation recalling the pattern of PGC-1α. TOM20 and CS remained unchanged in young subjects. Single OXPHOS complexes showed peculiar patterns, which were in some cases dissimilar from PGC-1α, and suggest different influences on protein biogenesis and degradation. Overall, exercise was capable to counteract the effect of immobility, when present, except for complex V, which was markedly downregulated by bed-rest, but remained unaffected after rehabilitation, maybe as result of greater extent of degradation processes over biogenesis. Phosphorylation extent of AMPK, and its upstream activator LKB1, did not change after bed-rest and rehabilitation in either young or elderly subjects, suggesting that the activation of energy-sensing LKB1-AMPK signaling pathway was "missed" due to its transient nature, or was not triggered under our conditions. Our study demonstrates that, as far as the expression of various proteins related to mitochondrial biogenesis/remodeling, adaptations to bed-rest and rehabilitation in the two populations were different. The impact of bed-rest was greater in the elderly subjects, where the pattern (decrease after bed rest and recovery following rehabilitation) was accompanied by changes of mitochondrial mass. Modifications of protein abundance were matched with data obtained from gene expression analyses of four public human datasets focusing on related genes.

In silico discovery of terpenoid metabolism in Cannabis sativa.

Due to their efficacy, cannabis based therapies are currently being prescribed for the treatment of many different medical conditions. Interestingly, treatments based on the use of cannabis flowers or their derivatives have been shown to be very effective, while therapies based on drugs containing THC alone lack therapeutic value and lead to increased side effects, likely resulting from the absence of other pivotal entourage compounds found in the Phyto-complex. Among these compounds are terpenoids, which are not produced exclusively by cannabis plants, so other plant species must share many of the enzymes involved in their metabolism. In the present work, 23,630 transcripts from the canSat3 reference transcriptome were scanned for evolutionarily conserved protein domains and annotated in accordance with their predicted molecular functions. A total of 215 evolutionarily conserved genes encoding enzymes presumably involved in terpenoid metabolism are described, together with their expression profiles in different cannabis plant tissues at different developmental stages. The resource presented here will aid future investigations on terpenoid metabolism in Cannabis sativa.