Value of the loss of heterozygosity to BRCA1 variant classification.

At least 10% of the BRCA1/2 tests identify variants of uncertain significance (VUS) while the distinction between pathogenic variants (PV) and benign variants (BV) remains particularly challenging. As a typical tumor suppressor gene, the inactivation of the second wild-type (WT) BRCA1 allele is expected to trigger cancer initiation. Loss of heterozygosity (LOH) of the WT allele is the most frequent mechanism for the BRCA1 biallelic inactivation. To evaluate if LOH can be an effective predictor of BRCA1 variant pathogenicity, we carried out LOH analysis on DNA extracted from 90 breast and seven ovary tumors diagnosed in 27 benign and 55 pathogenic variant carriers. Further analyses were conducted in tumors with PVs yet without loss of the WT allele: BRCA1 promoter hypermethylation, next-generation sequencing (NGS) of BRCA1/2, and BRCAness score. Ninety-seven tumor samples were analyzed from 26 different BRCA1 variants. A relatively stable pattern of LOH (65.4%) of WT allele for PV tumors was observed, while the allelic balance (63%) or loss of variant allele (15%) was generally seen for carriers of BV. LOH data is a useful complementary argument for BRCA1 variant classification.

Newly recognized recessive syndrome characterized by dysmorphic features, hypogonadotropic hypogonadism, severe microcephaly, and sensorineural hearing loss maps to 3p21.3.

We present a newly recognized, likely autosomal recessive, pleiotropic disorder seen in four individuals (three siblings and their nephew) from a consanguineous family of Pakistani origin. The condition is characterized by hypogonadotropic hypogonadism, severe microcephaly, sensorineural deafness, moderate learning disability, and distinctive facial dysmorphic features. Autozygosity mapping using SNP array genotyping defined a single, large autozygous region of 13.1 Mb on chromosome 3p21 common to the affected individuals. The critical region contains 227 genes and initial sequence analysis of a functional candidate gene has not identified causative mutations.

Prox1 maintains muscle structure and growth in the developing heart.

Impaired cardiac muscle growth and aberrant myocyte arrangement underlie congenital heart disease and cardiomyopathy. We show that cardiac-specific inactivation of the murine homeobox transcription factor Prox1 results in the disruption of expression and localisation of sarcomeric proteins, gross myofibril disarray and growth-retarded hearts. Furthermore, we demonstrate that Prox1 is required for direct transcriptional regulation of the genes encoding the structural proteins alpha-actinin, N-RAP and zyxin, which collectively function to maintain an actin-alpha-actinin interaction as the fundamental association of the sarcomere. Aspects of abnormal heart development and the manifestation of a subset of muscular-based disease have previously been attributed to mutations in key structural proteins. Our study reveals an essential requirement for direct transcriptional regulation of sarcomere integrity, in the context of enabling foetal cardiomyocyte hypertrophy, maintenance of contractile function and progression towards inherited or acquired myopathic disease.

Thymosin beta-4 is essential for coronary vessel development and promotes neovascularization via adult epicardium.

Ischemic heart disease leading to myocardial infarction causes irreversible cell loss and scarring and is a major cause of morbidity and mortality in humans. Significant effort in the field of cardiovascular medicine has been invested in the search for adult cardiac progenitor cells that may replace damaged muscle cells and/or contribute to new vessel formation (neovascularization) and in the identification of key factors, which may induce such progenitor cells to contribute to myocardial repair and collateral vessel growth. We recently demonstrated that the actin monomer-binding protein, thymosin beta-4 (Tbeta-4), when secreted from the myocardium provides a paracrine stimulus to the cells of the epicardium-derived cells (EPDCs) to promote their inward migration and differentiation into endothelial and smooth muscle cells to form the coronary vasculature. Translating this essential role for Tbeta-4 in coronary vessel development to the adult, we found that treatment of cultured adult explants with Tbeta-4 stimulated extensive outgrowth of epicardin-positive epicardial cells, which, as they migrated away from the explant, differentiated into procollagen type I, SMalphaA, and Flk1-positive cells indicative of fibroblasts, smooth muscle, and endothelial cells; thus releasing the adult epicardium from a quiescent state and restoring pluripotency. The ability of Tbeta-4 to promote coronary vessel development and potentially induce new vasculature in the adult is essential for cardiomyocyte survival and could contribute significantly toward the reported Tbeta4-induced cardioprotection and repair in the adult heart. Tbeta-4 is currently subject to multicenter phase 1 clinical trials for treatment of cardiovascular disease (http://www.regenerx.com), therefore, insight into the repair mechanism(s) induced by Tbeta-4 is an essential step toward harnessing therapeutic survival, migration, and repair properties of the peptide in the context of acute myocardial damage.

Thymosin beta4 induces adult epicardial progenitor mobilization and neovascularization.

Cardiac failure has a principal underlying aetiology of ischaemic damage arising from vascular insufficiency. Molecules that regulate collateral growth in the ischaemic heart also regulate coronary vasculature formation during embryogenesis. Here we identify thymosin beta4 (Tbeta4) as essential for all aspects of coronary vessel development in mice, and demonstrate that Tbeta4 stimulates significant outgrowth from quiescent adult epicardial explants, restoring pluripotency and triggering differentiation of fibroblasts, smooth muscle cells and endothelial cells. Tbeta4 knockdown in the heart is accompanied by significant reduction in the pro-angiogenic cleavage product N-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP). Although injection of AcSDKP was unable to rescue Tbeta4 mutant hearts, it significantly enhanced endothelial cell differentiation from adult epicardially derived precursor cells. This study identifies Tbeta4 and AcSDKP as potent stimulators of coronary vasculogenesis and angiogenesis, and reveals Tbeta4-induced adult epicardial cells as a viable source of vascular progenitors for continued renewal of regressed vessels at low basal level or sustained neovascularization following cardiac injury.