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Germline mutations in the BRCA genes are associated with a higher risk of carcinogenesis, which is linked to an increased mutation rate and loss of the second unaffected BRCA allele (loss of heterozygosity, LOH). However, the mechanisms triggering mutagenesis are not clearly understood. The BRCA genes contain high numbers of repetitive DNA sequences. We detected replication forks stalling, DNA breaks, and deletions at these sites in haploinsufficient BRCA cells, thus identifying the BRCA genes as fragile sites. Next, we found that stalled forks are repaired by error-prone pathways, such as microhomology-mediated break-induced replication (MMBIR) in haploinsufficient BRCA1 breast epithelial cells. We detected MMBIR mutations in BRCA1 tumor cells and noticed deletions-insertions (>50 bp) at the BRCA1 genes in BRCA1 patients. Altogether, these results suggest that under stress, error-prone repair of stalled forks is upregulated and induces mutations, including complex genomic rearrangements at the BRCA genes (LOH), in haploinsufficient BRCA1 cells. [Display omitted] • Replication forks stall at BRCA1 and 2 genes in haploinsufficient BRCAmut/+ cells • BRCA genes are early-replicating fragile sites that break in BRCAmut/+ cells • Stalled forks are repaired by MMBIR, an error-prone non-HR pathway in BRCA1mut/+ cells • Increase in error-prone repair leaves mutations and explains LOH in BRCA1 patients Deshpande et al. discovered an increase in replication fork stalling and deletions at the BRCA1 and 2 genes in stressed haploinsufficient BRCAmut/+ cells. The stalled forks are repaired increasingly by error-prone DNA repair pathways, such as MMBIR, which leave mutations, thus elevating mutagenesis and LOH in BRCA1 patients. [ABSTRACT FROM AUTHOR]

The prognosis of patients with glioblastoma (GBM) remains dismal, with a median survival of approximately 15 months. Current preclinical GBM models are limited by the lack of a "normal" human microenvironment and the inability of many tumor cell lines to accurately reproduce GBM biology. To address these limitations, we have established a model system whereby we can retro-engineer patient-specific GBMs using patient-derived glioma stem cells (GSCs) and human embryonic stem cell (hESC)-derived cerebral organoids. Our cerebral organoid glioma (GLICO) model shows that GSCs home toward the human cerebral organoid and deeply invade and proliferate within the host tissue, forming tumors that closely phenocopy patient GBMs. Furthermore, cerebral organoid tumors form rapidly and are supported by an interconnected network of tumor microtubes that aids in the invasion of normal host tissue. Our GLICO model provides a system for modeling primary human GBM ex vivo and for high-throughput drug screening., (Copyright © 2019 The Author(s). Published by Elsevier Inc. All rights reserved.)