Fusion transcriptome profiling defines the monoclonal origin of multifocal epithelioid haemangioma of bone.

AIMS: Epithelioid haemangioma (EH) of bone remains a highly controversial entity. Indeed, the WHO classifies EHs of soft tissues as benign tumours, whereas bone EHs are considered intermediate-locally aggressive tumours due to common multifocal presentation and local destructive growth. To gain insights into the clinical behaviour and biology of EH of bone we retrospectively analysed 42 patients treated in a single institution from 1978 to 2021. METHODS AND RESULTS: Multifocal presentation was detected in 17 of 42 patients (40%) primarily as synchronous lesions. Patients were treated with curettage (57%), resection (29%) or biopsy, followed by radiotherapy or embolisation (14%). Follow-up (minimum 24 months) was available for 38 patients, with only five local recurrences (13%) and no death of disease. To clarify whether the synchronous bone lesions in multifocal EH represent multicentric disease or clonal dissemination, four cases were profiled by RNA-sequencing. Separate lesions from the same patient, which showed a similar transcriptional profile, expressed the same fusion transcript (involving FOS or FOSB) with identical gene breakpoints. CONCLUSIONS: These results indicate that, in EH of bone, multifocal lesions are clonally related and therefore represent the spread of a same neoplastic clone rather than simultaneous independent tumours. This finding is in apparent contradiction with the benign clinical course of the disease, and suggests that tumour dissemination in bone EH probably reflects a phenomenon of passive spreading, with tumour cells colonising distal sites while maintaining their benign biological nature.

BENZ WS: the Bologna ENZyme Web Server for four-level EC number annotation.

The Bologna ENZyme Web Server (BENZ WS) annotates four-level Enzyme Commission numbers (EC numbers) as defined by the International Union of Biochemistry and Molecular Biology (IUBMB). BENZ WS filters a target sequence with a combined system of Hidden Markov Models, modelling protein sequences annotated with the same molecular function, and Pfams, carrying along conserved protein domains. BENZ returns, when successful, for any enzyme target sequence an associated four-level EC number. Our system can annotate both monofunctional and polyfunctional enzymes, and it can be a valuable resource for sequence functional annotation.

In Silico Identification of a BRCA1:miR-29:DNMT3 Axis Involved in the Control of Hormone Receptors in BRCA1-Associated Breast Cancers.

Germline inactivating mutations in the BRCA1 gene lead to an increased lifetime risk of ovarian and breast cancer (BC). Most BRCA1-associated BC are triple-negative tumors (TNBC), aggressive forms of BC characterized by a lack of expression of estrogen and progesterone hormone receptors (HR) and HER2. How BRCA1 inactivation may favor the development of such a specific BC phenotype remains to be elucidated. To address this question, we focused on the role of miRNAs and their networks in mediating BRCA1 functions. miRNA, mRNA, and methylation data were retrieved from the BRCA cohort of the TCGA project. The cohort was divided into a discovery set (Hi-TCGA) and a validation set (GA-TCGA) based on the platform used for miRNA analyses. The METABRIC, GSE81002, and GSE59248 studies were used as additional validation data sets. BCs were differentiated into BRCA1-like and non-BRCA1-like based on an established signature of BRCA1 pathway inactivation. Differential expression of miRNAs, gene enrichment analysis, functional annotation, and methylation correlation analyses were performed. The miRNAs downregulated in BRCA1-associated BC were identified by comparing the miRNome of BRCA1-like with non-BRCA1-like tumors from the Hi-TCGA discovery cohort. miRNAs:gene-target anticorrelation analyses were then performed. The target genes of miRNAs downregulated in the Hi-TCGA series were enriched in the BRCA1-like tumors from the GA-TCGA and METABRIC validation data sets. Functional annotation of these genes revealed an over-representation of several biological processes ascribable to BRCA1 activity. The enrichment of genes related to DNA methylation was particularly intriguing, as this is an aspect of BRCA1 functions that has been poorly explored. We then focused on the miR-29:DNA methyltransferase network and showed that the miR-29 family, which was downregulated in BRCA1-like tumors, was associated with poor prognosis in these BCs and inversely correlated with the expression of the DNA methyltransferases DNMT3A and DNMT3B. This, in turn, correlated with the methylation extent of the promoter of HR genes. These results suggest that BRCA1 may control the expression of HR via a miR-29:DNMT3:HR axis and that disruption of this network may contribute to the receptor negative phenotype of tumors with dysfunctional BRCA1.

A Glance into MTHFR Deficiency at a Molecular Level.

MTHFR deficiency still deserves an investigation to associate the phenotype to protein structure variations. To this aim, considering the MTHFR wild type protein structure, with a catalytic and a regulatory domain and taking advantage of state-of-the-art computational tools, we explore the properties of 72 missense variations known to be disease associated. By computing the thermodynamic ΔΔG change according to a consensus method that we recently introduced, we find that 61% of the disease-related variations destabilize the protein, are present both in the catalytic and regulatory domain and correspond to known biochemical deficiencies. The propensity of solvent accessible residues to be involved in protein-protein interaction sites indicates that most of the interacting residues are located in the regulatory domain, and that only three of them, located at the interface of the functional protein homodimer, are both disease-related and destabilizing. Finally, we compute the protein architecture with Hidden Markov Models, one from Pfam for the catalytic domain and the second computed in house for the regulatory domain. We show that patterns of disease-associated, physicochemical variation types, both in the catalytic and regulatory domains, are unique for the MTHFR deficiency when mapped into the protein architecture.

Mapping human disease-associated enzymes into Reactome allows characterization of disease groups and their interactions.

According to databases such as OMIM, Humsavar, Clinvar and Monarch, 1494 human enzymes are presently associated to 2539 genetic diseases, 75% of which are rare (with an Orphanet code). The Mondo ontology initiative allows a standardization of the disease name into specific codes, making it possible a computational association between genes, variants, diseases, and their effects on biological processes. Here, we tackle the problem of which biological processes enzymes can affect when the protein variant is disease-associated. We adopt Reactome to describe human biological processes, and by mapping disease-associated enzymes in the Reactome pathways, we establish a Reactome-disease association. This allows a novel categorization of human monogenic and polygenic diseases based on Reactome pathways and reactions. Our analysis aims at dissecting the complexity of the human genetic disease universe, highlighting all the possible links within diseases and Reactome pathways. The novel mapping helps understanding the biochemical/molecular biology of the disease and allows a direct glimpse on the present knowledge of other molecules involved. This is useful for a complete overview of the disease molecular mechanism/s and for planning future investigations. Data are collected in DAR, a database that is free for search and available at https://dar.biocomp.unibo.it .

Pathogenic variation types in human genes relate to diseases through Pfam and InterPro mapping.

Grouping residue variations in a protein according to their physicochemical properties allows a dimensionality reduction of all the possible substitutions in a variant with respect to the wild type. Here, by using a large dataset of proteins with disease-related and benign variations, as derived by merging Humsavar and ClinVar data, we investigate to which extent our physicochemical grouping procedure can help in determining whether patterns of variation types are related to specific groups of diseases and whether they occur in Pfam and/or InterPro gene domains. Here, we download 75,145 germline disease-related and benign variations of 3,605 genes, group them according to physicochemical categories and map them into Pfam and InterPro gene domains. Statistically validated analysis indicates that each cluster of genes associated to Mondo anatomical system categorizations is characterized by a specific variation pattern. Patterns identify specific Pfam and InterPro domain-Mondo category associations. Our data suggest that the association of variation patterns to Mondo categories is unique and may help in associating gene variants to genetic diseases. This work corroborates in a much larger data set previous observations from our group.

Highlighting Human Enzymes Active in Different Metabolic Pathways and Diseases: The Case Study of EC 1.2.3.1 and EC 2.3.1.9.

Enzymes are key proteins performing the basic functional activities in cells. In humans, enzymes can be also responsible for diseases, and the molecular mechanisms underlying the genotype to phenotype relationship are under investigation for diagnosis and medical care. Here, we focus on highlighting enzymes that are active in different metabolic pathways and become relevant hubs in protein interaction networks. We perform a statistics to derive our present knowledge on human metabolic pathways (the Kyoto Encyclopaedia of Genes and Genomes (KEGG)), and we found that activity aldehyde dehydrogenase (NAD(+)), described by Enzyme Commission number EC 1.2.1.3, and activity acetyl-CoA C-acetyltransferase (EC 2.3.1.9) are the ones most frequently involved. By associating functional activities (EC numbers) to enzyme proteins, we found the proteins most frequently involved in metabolic pathways. With our analysis, we found that these proteins are endowed with the highest numbers of interaction partners when compared to all the enzymes in the pathways and with the highest numbers of predicted interaction sites. As specific enzyme protein test cases, we focus on Alpha-Aminoadipic Semialdehyde Dehydrogenase (ALDH7A1, EC 2.3.1.9) and Acetyl-CoA acetyltransferase, cytosolic and mitochondrial (gene products of ACAT2 and ACAT1, respectively; EC 2.3.1.9). With computational approaches we show that it is possible, by starting from the enzyme structure, to highlight clues of their multiple roles in different pathways and of putative mechanisms promoting the association of genes to disease.

Tumor genotype, location, and malignant potential shape the immunogenicity of primary untreated gastrointestinal stromal tumors.

Intratumoral immune infiltrate was recently reported in gastrointestinal stromal tumors (GISTs). However, the tumor-intrinsic factors that dictate GIST immunogenicity are still largely undefined. To shed light on this issue, a large cohort (82 samples) of primary untreated GISTs, representative of major clinicopathological variables, was investigated by an integrated immunohistochemical, transcriptomic, and computational approach. Our results indicate that tumor genotype, location, and malignant potential concur to shape the immunogenicity of primary naive GISTs. Immune infiltration was greater in overt GISTs compared with that in lesions with limited malignant potential (miniGISTs), in KIT/PDGFRA-mutated tumors compared with that in KIT/PDGFRA WT tumors, and in PDGFRA-mutated compared with KIT-mutated GISTs. Within the KIT-mutated subset, a higher degree of immune colonization was detected in the intestine. Immune hot tumors showed expression patterns compatible with a potentially proficient but curbed antigen-specific immunity, hinting at sensitivity to immunomodulatory treatments. Poorly infiltrated GISTs, primarily KIT/PDGFRA WT intestinal tumors, showed activation of Hedgehog and WNT/ß-catenin immune excluding pathways. This finding discloses a potential therapeutic vulnerability, as the targeting of these pathways might prove effective by both inhibiting pro-oncogenic signals and fostering antitumor immune responses. Finally, an intriguing anticorrelation between immune infiltration and ANO1/DOG1 expression was observed, suggesting an immunomodulatory activity for anoctamin-1.

Next-Generation Sequencing Approaches for the Identification of Pathognomonic Fusion Transcripts in Sarcomas: The Experience of the Italian ACC Sarcoma Working Group.

This work describes the set-up of a shared platform among the laboratories of the Alleanza Contro il Cancro (ACC) Italian Research Network for the identification of fusion transcripts in sarcomas by using Next Generation Sequencing (NGS). Different NGS approaches, including anchored multiplex PCR and hybrid capture-based panels, were employed to profile a large set of sarcomas of different histotypes. The analysis confirmed the reliability of NGS RNA-based approaches in detecting sarcoma-specific rearrangements. Overall, the anchored multiplex PCR assay proved to be a fast and easy-to-analyze approach for routine diagnostics laboratories.

Corrigendum: Next-Generation Sequencing Approaches for the Identification of Pathognomonic Fusion Transcripts in Sarcomas: The Experience of the Italian ACC Sarcoma Working Group.

[This corrects the article DOI: 10.3389/fonc.2020.00489.].