Functional Analysis of Breast Cancer Susceptibility Genes

            Our laboratory uses mouse genetics to study the biological function(s) of BRCA1 and BRCA2, to understand how loss of these functions leads to early onset breast and ovarian cancer, and to generate animal models of the human disease.

Germline mutations in the BRCA1 and BRCA2 genes account for 70-80% of hereditary breast and ovarian cancers. Both are thought to be tumor suppressor genes as the wild-type alleles are lost in tumors of heterozygous carriers. BRCA1 and BRCA2 encode novel proteins, but primary amino acid sequences yield little information about their normal function. Extensive research efforts from all disciplines contributed a plethora of data suggesting a role for both, BRCA1 and BRCA2 in embryonic proliferation, homologous recombination and DNA repair pathways, cell cycle checkpoint control and, transcriptional regulation. However, it is unclear which function(s) specifically suppress tumorigenesis. TP53 is mutated in a large number of BRCA-associated tumors consistent with the hypothesis that inactivation of a cell cycle checkpoint is a necessary step and may precede BRCA loss during tumorigenesis.

The early embryonic lethality associated with global ablation of Brca1 and Brca2 precluded an analysis of the phenotypic consequences in adult mice. To bypass the lethality and to study tumor development, we introduced by gene targeting in embryonic stem cells hypomorphic and conditional mutations in Brca1, Brca2 and p53.

In contrast to the embryonic lethality of Brca1 nullizygotes, mice with a hypomorphic, truncating Brca1 mutation survive fully, but only by enrichment in particular genetic backgrounds. Cultured fibroblasts from Brca1^tr/tr embryos proliferate poorly and spontaneously accumulate chromosomal aberrations. Most of the Brca1^tr/tr mutants develop lymphomas, sarcomas and carcinomas, including breast cancer.

Mammary gland-specific inactivation of a conditional Brca2 allele results in a high incidence (77%) of breast tumors that develop after long latency. These invasive adenocarcinomas are histologically quite uniform, exhibiting predominantly a solid nodular pattern, in striking contrast to the morphological heterogeneity of mammary tumors in Brca1^tr/tr mutant mice. Thus, it appears that in both mice and humans, mutations in the breast cancer susceptibility genes Brca1 and Brca2 can augment cancer risk.

Brca1, Brca2 and p53 conditional mutant mice are currently used in a breeding program to evaluate, in comparison with controls, incidence and latency of mammary tumor development. In parallel, we are examining the phenotypic effects of the mutations at the cellular level in primary cells established from control and mutant mice. Furthermore, we have generated mutations in the Brca1-interacting proteins Bard1 and CtIP, which may function in Brca1-mediated tumor suppression and are currently analyzing their phenotypes.

Selected publications

Ludwig, T., Chapman D.L., Papaioannou, V.E., and Efstratiadis, A. (1997). Targeted mutations of breast cancer susceptibility gene homologs in mice: lethal phenotypes of Brca1, Brca2, Brca1/Brca2, Brca1/p53 and Brca2/p53 nullizygous embryos. Genes & Dev. 11, 1226-1241.

Ludwig, T., Fisher, P., Ganesan, S., and Efstratiadis, A. (2001). Tumorigenesis in mice carrying a truncating Brca1 mutation. Genes & Devel. 15, 1188-1193.

Ludwig, T., Fisher, P., Murty, V., and Efstratiadis, A. (2001). Development of mammary adenocarcinomas by tissue-specific knockout of Brca2 in mice. Oncogene 20, 3937-3948.

Baer, R., and Ludwig, T. (2002) The BRCA1/BARD1 heterodimer: a tumor suppressor complex with ubiquitin E3 ligase activity. Curr. Opin. Genet. Dev. 12, 86-91.