Investigating the Role of RECQL in Breast Cancer Susceptibility

RECQL was first proposed to be a candidate breast cancer susceptibility gene in 2015 by our group (Cybulski et al, Nature Genetics 2015) and by a Chinese group (Sun et al, PLOS Genetics 2015) independently. RECQL helps to overcome stalled DNA replication forks, and thereby prevents double-strand DNA breaks. Cells with defective RECQL, accumulate more double-strand DNA breaks than normal cells. In our initial analysis of French-Canadian and Polish breast cancer patients, we identified two rare but recurrent RECQL mutations (one in each population); each was significantly more frequent among breast cancer patients than among controls in the respective population. In Quebec, seven of 1,013 high-risk breast cancer patients carried the c.643C>T (p.Arg215*) variant (NM_002907.3), compared to only one of 7,136 newborns in Quebec (OR = 49.3; p < 10‑5). In Poland, 30 of 13,136 unselected breast cancer cases carried the c.1667_1667+3delAGTA (p.K555delinsMYKLIHYSFR) variant, compared to two of 4,702 controls in Poland (OR = 5.4; p = 0.008). Since then several other studies have been done to study the role of RECQL mutation in association with higher risk for breast cancer that have been reviewed by our team (Ahmed et al, Nature Genetics 2018).

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The next step – and the goal of this study – is to explore the significance of RECQL mutations in breast cancer on a broader level.

To get a better estimate of the RECQL mutation frequency and the breast cancer risk associated with carrying a RECQL pathogenic mutation, we are screening 5,000 patients and 1,000 healthy women from Ontario for mutations on the entire coding region of the gene. In addition, we determine whether or not any of the observed RECQL missense mutations are pathogenic using in vitro functional assays, such as complementing RECQL siRNA/shRNA/CRISPR cell lines with RECQL-Flag wild-type and mutant constructs. This part of the project is done in collaboration with Drs. Masson’s laboratory at Laval University.

From our previous work, we also found that the wild-type allele of RECQL is intact in the tumour cells of the RECQL germline mutation carriers, suggesting that RECQL haploinsufficiency is probably enough to increase the risk to cancer. To further investigate this, we examine RECQL allele-specific expression level in the tumour cells of 40 mutation carriers and 40 matched non-carriers. Also, since previous studies have demonstrated that RECQL function is necessary for cancer cells to survive, our team will also study how RECQL-insufficient cancer cells cope with the low expression of RECQL. It is possible that other genes with overlapping functions with RECQL are over expressed in haploinsufficient RECQL cells.

In addition, we have established the association of RECQL mutations with breast cancer risk, however, we still do not know whether a dysfunctional RECQL can result in breast cancer directly. To address this we will use Recql knockout mice to see whether or not these mice are more prone to mammary tumours compared to wild-type mice. We will treat a group of wild-type, Recql +/-, and Recql -/- mice with DMBA (7, 12-dimethylbenz(alpha)anthracene), a mammary carcinogen, and observe tumor incidence. Mouse work are done at Dr. Sharma’s lab in Howard University.