CDKN1B mutation and copy number variation are associated with tumor aggressiveness in luminal breast cancer

The CDKN1B gene, encoding for the CDK inhibitor p27kip1 , is mutated in defined human cancer subtypes, including breast, prostate carcinomas and small intestine neuroendocrine tumors (SI-NET). Lessons learned from SI-NET suggests that CDKN1B mutations could be subclonal, raising the question of whether a deeper sequencing approach could lead to the identification of higher numbers of patients with mutations. Here, we addressed this question and analyzed human cancer biopsies from breast (n=396), ovarian (n=110) and head and neck squamous carcinomas (n=202) patients, using an ultra-deep sequencing approach. Notwithstanding this effort, the mutation rate of CDKN1B remained substantially aligned with values from the literature, showing that essentially only hormone-receptor positive breast cancer (HR+, luminal BC) displayed CDKN1B mutations in a relevant number of cases (3%). However, the analysis of copy number variation showed that another fraction of luminal BC displayed loss (8%) or gain (6%) of the CDKN1B gene, further reinforcing the idea that the function of p27kip1 is important in this type of tumor. Intriguingly, an enrichment for CDKN1B alterations was found in samples from pre-menopausal luminal BC patients (n=227, 4%) and in circulating cell-free DNA from metastatic luminal BC patients (n=59, 8.5%), suggesting that CDKN1B alterations could correlate with tumor aggressiveness and/or occur later during disease progression. Notably, many of the identified somatic mutations resulted in p27kip1 protein truncation leading to loss of most of the protein or of its C-terminal domain. Using a gene editing approach in a luminal BC cell line, MCF-7, we observed that the expression of p27kip1 truncating mutants that lose the C-terminal domains failed to rescue most of the phenotypes induced by CDKN1B gene knock-out, indicating that the functions retained by the C-terminal portion are critical for its role as oncosuppressor, at least in luminal BC. This article is protected by copyright. All rights reserved.