Clinical and biological significance of RAD51 expression in breast cancer: a key DNA damage response protein

Impaired DNA damage response (DDR) may play a fundamental role in the pathogenesis of breast cancer (BC). RAD51 is a key player in DNA double-strand break repair. In this study, we aimed to assess the biological and clinical significance of RAD51 expression with relevance to different molecular classes of BC and patients’ outcome. The expression of RAD51 was assessed immunohistochemically in a well-characterised annotated series (n = 1184) of early-stage invasive BC with long-term follow-up. A subset of cases of BC from patients with known BRCA1 germline mutations was included as a control group. The results were correlated with clinicopathological and molecular parameters and patients’ outcome. RAD51 protein expression level was also assayed in a panel of cell lines using reverse phase protein array (RPPA). RAD51 was expressed in the nuclei (N) and cytoplasm (C) of malignant cells. Subcellular colocalisation phenotypes of RAD51 were significantly associated with clinicopathological features and patient outcome. Cytoplasmic expression (RAD51C+) and lack of nuclear expression (RAD51 N-) were associated with features of aggressive behaviour, including larger tumour size, high grade, lymph nodal metastasis, basal-like, and triple-negative phenotypes, together with aberrant expression of key DDR biomarkers including BRCA1. All BRCA1-mutated tumours had RAD51C+/N- phenotype. RPPA confirmed IHC results and showed differential expression of RAD51 in cell lines based on ER expression and BRCA1 status. RAD51 N+ and RAD51C+ tumours were associated with longer and shorter breast cancer-specific survival (BCSS), respectively. The RAD51 N+ was an independent predictor of longer BCSS (P<0.0001). Lack of RAD51 nuclear expression is associated with poor prognostic parameters and shorter survival in invasive BC patients. The significant associations between RAD51 subcellular localisation and clinicopathological features, molecular subtype and patients’ outcome suggest that the trafficking of DDR proteins between the nucleus and cytoplasm might play a role in the development and progression of BC

KPNA2 is a nuclear export protein that contributes to aberrant localisation of key proteins and poor prognosis of breast cancer

Background: It is recognised that modulations of the nuclear import of macromolecules have a role in changing cellular phenotypes and carcinogenesis. We and others have noticed that aberrant subcellular localisation of DNA damage response (DDR) proteins in breast cancer (BC) is associated with loss-of-function phenotype. This study aims to investigate the biological and clinical significance of the nucleocytoplasmic transport protein karyopherin a-2 (KPNA2), and its role in controlling DDR proteins subcellular localisation in BC.Methods: A large (n=1494) and well-characterised series of early-stage invasive BC with a long-term follow-up was assessed for KPNA2 protein by using immunohistochemistry.Results: KPNA2 expression was associated with the subcellular localisation of key DDR proteins that showed cytoplasmic expression including BRCA1, RAD51, SMC6L1, gammaH2AX, BARD1, UBC9, PIAS1 and CHK1. High level of KPNA2 was associated not only with cytoplasmic localisation of these proteins but also with their low/negative nuclear expression. Positive KPNA2 expression was associated with negative oestrogen receptor and triple-negative phenotype. Survival analysis showed that KPNA2 was associated with poor outcome (P less than 0.0001), but this effect was not independent of other prognostic variables.Conclusions: This study provides further evidence for the complexity of DDR mechanism in BC, and that KNPA2 has a role in the aberrant subcellular localisation of DDR proteins with subsequent impaired function

Characterization of behavioral, signaling and cytokine alterations in a rat neurodevelopmental model for schizophrenia, and their reversal by the 5-HT₆ receptor antagonist SB-399885

Post-weaning social isolation of rats produces neuroanatomical, neurochemical and behavioral alterations resembling some core features of schizophrenia. This study examined the ability of the 5-HT₆ receptor antagonist SB-399885 to reverse isolation-induced cognitive deficits, then investigated alterations in hippocampal cell proliferation and hippocampal and frontal cortical expression of selected intracellular signaling molecules and cytokines. Male Lister-hooded rats (weaned on post-natal day 21-24 and housed individually or in groups of 3-4) received six i.p. injections of vehicle (1% Tween 80, 1 mL/kg) or SB-399885 (5 or 10 mg/kg) over a two week period starting 40 days post-weaning, on the days that locomotor activity, novel object discrimination (NOD), pre-pulse inhibition of acoustic startle and acquisition, retention and extinction of a conditioned freezing response (CFR) were assessed. Tissue was collected 24 h after the final injection for immunohistochemistry, reverse-phase protein microarray and western blotting. Isolation rearing impaired NOD and cue-mediated CFR, decreased cell proliferation within the dentate gyrus, and elevated hippocampal TNFα levels and Cdc42 expression. SB-399885 reversed the NOD deficit and partially normalized CFR and cell proliferation. These effects were accompanied by altered expression of several members of the c-Jun N-terminal Kinase (JNK) and p38 MAPK signaling pathways (including TAK1, MKK4 and STAT3). Although JNK and p38 themselves were unaltered at this time point hippocampal TAK1 expression and phosphorylation correlated with visual recognition memory in the NOD task. Continued use of this neurodevelopmental model could further elucidate the neurobiology of schizophrenia and aid assessment of novel therapies for drug-resistant cognitive symptoms