Probes for INS

Background Germline mutations in breast cancer susceptibility gene 1 (BRCA1) increase the risk of breast and ovarian cancers. However, no association between BRCA1 germline mutation and glioblastoma malignancy has ever been highlighted. Here we report two cases of BRCA1 mutated patients who developed a glioblastoma multiform (GBM). Cases presentation: Two patients diagnosed with triple negative breast cancer (TNBC) were screened for BRCA1 germline mutation. They both carried a pathogenic mutation introducing a premature STOP codon in the exon 11 of the BRCA1 gene. Few years later, both patients developed a glioblastoma and a second breast cancer. In an attempt to clarify the role played by a mutated BRCA1 allele in the GBM development, we investigated the BRCA1 mRNA and protein expression in breast and glioblastoma tumours for both patients. The promoter methylation status of this gene was also tested by methylation specific PCR as BRCA1 expression is also known to be lost by this mechanism in some sporadic breast cancers. Conclusion: Our data show that BRCA1 expression is maintained in glioblastoma at the protein and the mRNA levels, suggesting that loss of heterozygosity (LOH) did not occur in these cases. The protein expression is tenfold higher in the glioblastoma of patient 1 than in her first breast carcinoma, and twice higher in patient 2. In agreement with the high protein expression level in the GBM, BRCA1 promoter methylation was not observed in these tumours. In these two cases, despite of a BRCA1 pathogenic germline mutation, the tumour-suppressor protein expression is maintained in GBM, suggesting that the BRCA1 mutation is not instrumental for the GBM development.

The integrity of lens epithelial cells (LECs) lays the foundation for lens function and transparency. By contrast, epithelial-mesenchymal transition (EMT) of LECs leads to lens fibrosis, such as anterior subcapsular cataracts (ASC) and fibrotic forms of posterior capsule opacification (PCO). However, the underlying mechanisms remain unclear. Here, we aimed to explore the role of long non-coding RNA (lncRNA) H19 in regulating TGF-β2-induced EMT during lens fibrosis, revealing a novel lncRNA-based regulatory mechanism. In this work, we identified that lncRNA H19 was highly expressed in LECs, but downregulated by exposure to TGF-β2. In both human lens epithelial explants and SRA01/04 cells, knockdown of H19 aggravated TGF-β2-induced EMT, while overexpressing H19 partially reversed EMT and restored lens epithelial phenotypes. Semi-in vivo whole lens culture and H19 knockout mice demonstrated the indispensable role of H19 in sustaining lens clarity through maintaining LEC features. Bioinformatic analyses further implied a potential H19-centered regulatory mechanism via Smad-dependent pathways, confirmed by in vitro experiments. In conclusion, we uncovered a novel role of H19 in inhibiting TGF-β2-induced EMT of the lens by suppressing Smad-dependent signaling, providing potential therapeutic targets for treating lens fibrosis.

Exercise training is well established as the most effective way to enhance muscle performance and muscle building. The composition of skeletal muscle fiber type affects systemic energy expenditures, and perturbations in metabolic homeostasis contribute to the onset of obesity and other metabolic dysfunctions. Long noncoding RNAs (lncRNAs) have been demonstrated to play critical roles in diverse cellular processes and diseases, including human cancers; however, the functional importance of lncRNAs in muscle performance, energy balance, and obesity remains elusive. We previously reported that the lncRNA H19 regulates the poly-ubiquitination and protein stability of dystrophin (DMD) in muscular dystrophy.Here, we identified mouse/human H19-interacting proteins using mouse/human skeletal muscle tissues and liquid chromatography-mass spectrometry (LC-MS). Human induced pluripotent stem-derived skeletal muscle cells (iPSC-SkMC) from a healthy donor and Becker Muscular Dystrophy (BMD) patients were utilized to study DMD post-translational modifications and associated proteins. We identified a gain-of-function (GOF) mutant of H19 and characterized the effects on myoblast differentiation and fusion to myotubes using iPSCs. We then conjugated H19 RNA gain-of-function oligonucleotides (Rgof) with the skeletal muscle enrichment peptide agrin (referred to as AGR-H19-Rgof) and evaluated AGR-H19-Rgof's effects on skeletal muscle performance using wild-type (WT) C57BL/6 J mice and its anti-obesity effects using high-fat diet (HFD)- and leptin deficiency-induced obese mouse models.We demonstrated that both human and mouse H19 associated with DMD and that the H19 GOF exhibited enhanced interaction with DMD compared to WT H19. DMD was found to associate with serine/threonine-protein kinase MRCK alpha (MRCKα) and α-synuclein (SNCA) in iPSC-SkMC derived from BMD patients. Inhibition of MRCKα and SNCA-mediated phosphorylation of DMD antagonized the interaction between H19 and DMD. These signaling events led to improved skeletal muscle cell differentiation and myotube fusion. The administration of AGR-H19-Rgof improved the muscle mass, muscle performance, and base metabolic rate of WT mice. Furthermore, mice treated with AGR-H19-Rgof exhibited resistance to HFD- or leptin deficiency-induced obesity.Our study suggested the functional importance of the H19 GOF mutant in enhancing muscle performance and anti-obesity effects.

Purpose: Poly(ADP-Ribose) Polymerase (PARP) inhibitors are promising targeted treatment options for hereditary breast tumors with a Homologous Recombination (HR) deficiency caused by BRCA1 or BRCA2 mutations. However, the functional consequence of BRCA gene mutations is not always known and tumors can be HR deficient for other reasons than BRCA gene mutations. Therefore, we aimed to develop a functional test to determine HR activity in tumor samples to facilitate selection of patients eligible for PARP inhibitor treatment. Experimental design: We obtained 54 fresh primary breast tumor samples from patients undergoing surgery. We determined their HR capacity by studying the formation of ionizing radiation induced foci (IRIF) of the HR protein RAD51 after ex vivo irradiation of these organotypic breast tumor samples. Tumors showing impaired RAD51 IRIF formation were subjected to genetic and epigenetic analysis. Results: Five out of 45 primary breast tumors with sufficient numbers of proliferating tumor cells were RAD51 IRIF formation deficient (11%, 95%CI: 5%-24%). This HR defect was significantly associated with Triple Negative Breast Cancer (OR:57, 95%CI: 3.9-825, p=0.003). Two out of five HR deficient tumors were not caused by mutations in the BRCA genes, but by BRCA1 promoter hypermethylation. Conclusion: The functional RAD51 IRIF assay faithfully identifies HR deficient tumors and has clear advantages over gene sequencing. It is a relatively easy assay that can be performed on biopsy material, making it a powerful tool to select patients with an HR-deficient cancer for PARP inhibitor treatment in the clinic.