Search

Objectives: The receptor for hyaluronan-mediated motility (RHAMM, intracellular hyaluronan binding protein [IHABP], CD168), whose over-expression in tumors is associated with poor prognosis and early age at diagnosis, localizes to the centrosome, interacts with microtubules, functions in the maintenance of spindle pole stability and is required for centrosomal targeting. A previous study has shown that genetic variation at the HMMR locus (encoding for RHAMM) modifies breast cancer risk among BRCA1 mutation carriers and interplay between BRCA1, AURKA (aurora kinase A), RHAMM and phosphorylated RHAMM (pRHAMM) plays an important role in epithelial apicobasal polarization and breast carcinogenesis. Aim of this study was to investigate mechanisms by which BRCA1 regulates the function of RHAMM. Methods: Four pairs of shBRCA1 oligonucleotides containing target sequences against BRCA1 transcript were designed and synthesized. After annealing, these four doublestrand shBRCA1 fragments were inserted into pLVTH vector and stably transfected into MCF-12A cells for knock-down of BRCA1 expression. The BRCA1 knock-out breast cancer cell line HCC1937 with homozygous BRCA1 c.5382insC mutation was transfected with the full length wt-BRCA1 to restore BRCA1 function. HCC1937 cells were also transfected with the BRCA1-delExon11 isoform that lacks BRCA1 exon 11 in its entirety. Western blotting analysis and immunohistochemical staining with anti-BRCA1, anti-RHAMM, anti-pRHAMM and anti-AURKA antibodies were performed. Real-time PCR was used to quantify RHAMM and AURKA expression in these different cell lines and transfectants. Results: Stable transfection of MCF-12A cells with the different shRNAs resulted in a significant decrease of BRCA1 transcript and protein levels. Real-time PCR showed increased RHAMM expression in MCF-12A cells transfected with shRNA BRCA1 knockdown constructs. Increased RHAMM gene expression was also determined in HCC1937 cells lacking BRCA1 function due to homozygous deleterious BRCA1 mutation c.5382insC. Restored wt-BRCA1 expression in HCC1937 cells transfected with the full length BRCA1 cDNA results in was significantly reduced RHAMM expression. Inhibition of RHAMM gene expression was even more pronounced in HCC1937 cells transfected with the BRCA1- delExon11variant lacking 1,143 amino acids of the BRCA1 protein. On the protein level loss of BRCA1 function affects nuclear localization of pT703-RHAMM the product of AURKA activity: pT703-RHAMM staining was revealed to be strong at the nuclear envelope in MCF12A BRCA1 knock-down transfectants compared to the homogenous and less intense but clearly nuclear pT703-RHAMM staining in MCF12A wt-BRCA1 cells. Conclusions: BRCA1 negatively regulates RHAMM gene expression and might be involved in the nuclear localization of pRHAMM. Interestingly on the transcription level the BRCA1delexon11 isoform of BRCA1 displayed a stronger RHAMM repressor function than BRCA1 wild type. Taken together, our data provide insight into new functions of BRCA1 regulating RHAMM expression and mediating pRHAMM function. [ABSTRACT FROM AUTHOR]

Pathogenic sequence variants (PSV) in BRCA1 or BRCA2 ( BRCA1/2 ) are associated with increased risk and severity of prostate cancer. We evaluated whether PSVs in BRCA1/2 were associated with risk of overall prostate cancer or high grade (Gleason 8+) prostate cancer using an international sample of 65 BRCA1 and 171 BRCA2 male PSV carriers with prostate cancer, and 3,388 BRCA1 and 2,880 BRCA2 male PSV carriers without prostate cancer. PSVs in the 3' region of BRCA2 (c.7914+) were significantly associated with elevated risk of prostate cancer compared with reference bin c.1001-c.7913 [HR = 1.78; 95% confidence interval (CI), 1.25-2.52; P = 0.001], as well as elevated risk of Gleason 8+ prostate cancer (HR = 3.11; 95% CI, 1.63-5.95; P = 0.001). c.756-c.1000 was also associated with elevated prostate cancer risk (HR = 2.83; 95% CI, 1.71-4.68; P = 0.00004) and elevated risk of Gleason 8+ prostate cancer (HR = 4.95; 95% CI, 2.12-11.54; P = 0.0002). No genotype-phenotype associations were detected for PSVs in BRCA1 . These results demonstrate that specific BRCA2 PSVs may be associated with elevated risk of developing aggressive prostate cancer. SIGNIFICANCE: Aggressive prostate cancer risk in BRCA2 mutation carriers may vary according to the specific BRCA2 mutation inherited by the at-risk individual., (©2019 American Association for Cancer Research.)