A lentiviral-barcoding approach using mouse embryonic cells allows driver-mutation identification.

  • Major Finding: A lentiviral-barcoding approach using mouse embryonic cells allows driver-mutation identification.

  • Concept: One mutation found by the screen is seen in patients with breast cancer and promoted tumorigenesis.

  • Impact: This enables large-scale screening of uncommon breast cancer–associated variants for driver status.

Most mutations and copy-number alterations observed in patients with breast cancer are present at low frequency, and each mutation's status as a disease driver or passenger is not trivial to establish. Complicating matters, the use of mouse mammary epithelium, a useful platform for revealing which genes contribute to breast cancer development, is limited by issues such as lineage specificity (i.e., luminal versus myoepithelial) and gene-targeting challenges. Ying and Beronja developed a method in which lentiviral injection into the intra-amniotic cavity of midgestation mouse embryos can be used to achieve efficient, stable transduction of the adult mammary epithelium. By “barcoding” the lentiviral constructs with 10-base-pair identifier sequences, use of the lentiviral injection method enabled the discovery that approximately 120 ectodermal progenitors with identical potential for growth and homeostasis as well as physiologic expansion and regression give rise to each adult mammary gland. Ectodermal progenitors had approximately equal likelihoods of giving rise to myoepithelial or luminal cells during normal homeostasis and—according to experiments in a breast cancer model driven by oncogenic knock-in of Pik3caH1047R—in the context of oncogenic growth. Further experiments using this in vivo breast cancer model identified candidate driver mutations of Pik3caH1047R-dependent breast tumorigenesis. Among the genes that served as drivers when mutated, Tsc22d1 was selected for further study because mutations in the gene are of unknown driver status and have been found in 2% of patients with breast cancer. After verification that Tsc22d1 depletion promoted tumorigenesis, further experiments showed that this depletion led to a shift to a subtype similar to the HER2-dependent subtype. A mechanistic investigation revealed that the relevant molecular effect of Tsc22d1 loss was p53 depletion mediated by the p53 regulator and E3 ubiquitin–protein ligase MDM2, resulting in HER2-dependent but not HER2-addicted mammary tumors. Collectively, these experiments demonstrate the power of this approach for large-scale screening to identify which uncommon mutations found in patients with breast cancer are true drivers.

Ying Z, Beronja S. Embryonic barcoding of equipotent mammary progenitors functionally identifies breast cancer drivers. Cell Stem Cell 2020 Feb 13 [Epub ahead of print].

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