Germline mutations in the BRCA1, BRCA2, PTEN, or TP53 tumor suppressor genes indicate that 5-10% of breast cancers (BCs) are familial. These genes may accumulate somatic mutations in conjunction with mutations and/or amplifications of PIK3CA and AKT3, as well as deletions or mutations of PTEN, TSC1, and INPP4B in the PI3K/mTOR pathway, although not in the majority of BCs. Indeed, none of them have been directly established as sole driver genes capable of inducing the entire process of transforming epithelial cells into breast cancer.
Regarding the classic RAS family members, unlike pancreatic, lung, and colorectal cancers, activating oncogenic mutations in KRAS, NRAS, and HRAS have not often been found in BC. Indeed, the frequency of mutation of these three genes together is lower than 1% in genome-wide association studies (GWAS) of BC. The RRAS2 member of the RAS-related subfamily, also known as TC21, is a close relative of the classic RAS GTPases and has transforming capacity in vitro similar to or even higher than classic RAS proteins. RRAS2 is, however, found in human cancers, including BC, bearing activating mutations at a very low frequency. However, the wild-type protein has been described as overexpressed in B-cell lymphomas, squamous carcinomas of the oral cavity, esophagus, gastric and lung cancers, as well as in breast cancer. A link between wild-type R-RAS2 protein and BC was established after finding that a polymorphism in the RRAS2 promoter is associated with an unfavorable tamoxifen treatment outcome and that silencing the RRAS2 gene enhances the tamoxifen sensitivity of a BC cell line. These data indicated that RRAS2 is an important gene in the development and clinical outcome of BC.
A peculiarity of the R-RAS2 protein is that it has an intrinsic guanosine nucleotide exchange capacity higher than that of classic RAS proteins, meaning that the wild-type protein might exist in cells in the active GTP-loaded form in a high proportion. We therefore hypothesized that RRAS2 overexpression in the wild-type form might be sufficient to drive the development of cancer. A mouse line conditionally overexpressing human RRAS2 was generated and found to develop a lymphoproliferative disease that was reminiscent of chronic lymphocytic leukemia (CLL) (Hortal et al, 2022; doi: 10.1186/s12943-022-01496-x). CLL appeared in 100% of mice, males and females, and was linked to a lifespan of ~13 months. 
In a recent paper published in Molecular Cancer (Cifuentes et al. 2024; doi: 10.1186/s12943-024-02054-3), we show that female mice overexpressing RRAS2 in the germline all develop breast ductal adenocarcinoma, equivalent to the most frequent human breast cancer, but only if those female mice were breeders. Isolated females that had never been in crossing developed CLL and liver cancer but not breast cancer. These data show that overexpression of wild-type RRAS2 is sufficient to drive the development of breast cancer and that such development is linked to pregnancy. By crossing with breast tissue-specific Cre recombinase-expressing mice, we demonstrate that the driver role of RRAS2 is intrinsic to the mammary epithelial cells.
Having found this effect in mice, we interrogated if RRAS2 overexpression could be behind human breast cancer. We show in the article that the mRNA is overexpressed in the majority of human breast cancers, independent of their molecular type. However, overexpression is more frequent (75%) and highest in the triple-negative type, the breast cancer type developed in the mouse model. Furthermore, looking at the distribution by age, we show that expression is highest in younger women than in older ones and that it is higher in parous than in nulliparous women of the same age, confirming a link to pregnancy in human disease. This aligns with the fact that triple-negative breast cancer is the most frequent type in women during their reproductive age.
Independent of its association with triple-negative breast cancer, the highest RRAS2 expression is associated with reduced life expectancy after diagnosis, with a higher index of proliferating Ki67+ cells in the tumor, and with loss of p53, all indicating poorer prognosis. Therefore, we propose RRAS2 expression as a marker of disease prognosis.
A cause-effect relationship between RRAS2 overexpression and human breast cancer cannot be directly demonstrated. However, we found that an SNP, a polymorphism in the 3’ untranslated region of the RRAS2 mRNA, is associated with higher mRNA expression compared to that of tumors expressing the canonical allele. This SNP is more frequently found in tumor samples than in the blood of healthy volunteers. In addition, we show that the RRAS2 gene is frequently found amplified not only in tumor samples but in the blood of breast cancer patients as well. These two findings strongly suggest a cause-effect relationship between overexpression of RRAS2 and human breast cancer. Furthermore, the SNP frequency and gene amplification in blood point to a pre-existing propensity to develop breast cancer caused by RRAS2 overexpression. This latter finding, of course, requires confirmation by increasing the number of human tissue samples in future studies.