KSR1 regulates BRCA1 degradation and inhibits breast cancer growth.

Kinase suppressor of Ras-1 (KSR1) facilitates signal transduction in Ras-dependent cancers, including pancreatic and lung carcinomas but its role in breast cancer has not been well studied. Here, we demonstrate for the first time it functions as a tumor suppressor in breast cancer in contrast to data in other tumors. Breast cancer patients (n>1000) with high KSR1 showed better disease-free and overall survival, results also supported by Oncomine analyses, microarray data (n=2878) and genomic data from paired tumor and cell-free DNA samples revealing loss of heterozygosity. KSR1 expression is associated with high breast cancer 1, early onset (BRCA1), high BRCA1-associated ring domain 1 (BARD1) and checkpoint kinase 1 (Chk1) levels. Phospho-profiling of major components of the canonical Ras-RAF-mitogen-activated protein kinases pathway showed no significant changes after KSR1 overexpression or silencing. Moreover, KSR1 stably transfected cells formed fewer and smaller size colonies compared to the parental ones, while in vivo mouse model also demonstrated that the growth of xenograft tumors overexpressing KSR1 was inhibited. The tumor suppressive action of KSR1 is BRCA1 dependent shown by 3D-matrigel and soft agar assays. KSR1 stabilizes BRCA1 protein levels by reducing BRCA1 ubiquitination through increasing BARD1 abundance. These data link these proteins in a continuum with clinical relevance and position KSR1 in the major oncoprotein pathways in breast tumorigenesis.

The regulatory roles of phosphatases in cancer.

The relevance of potentially reversible post-translational modifications required for controlling cellular processes in cancer is one of the most thriving arenas of cellular and molecular biology. Any alteration in the balanced equilibrium between kinases and phosphatases may result in development and progression of various diseases, including different types of cancer, though phosphatases are relatively under-studied. Loss of phosphatases such as PTEN (phosphatase and tensin homologue deleted on chromosome 10), a known tumour suppressor, across tumour types lends credence to the development of phosphatidylinositol 3-kinase inhibitors alongside the use of phosphatase expression as a biomarker, though phase 3 trial data are lacking. In this review, we give an updated report on phosphatase dysregulation linked to organ-specific malignancies.

SILAC-based phosphoproteomics reveals an inhibitory role of KSR1 in p53 transcriptional activity via modulation of DBC1.

BACKGROUND: We have previously identified kinase suppressor of ras-1 (KSR1) as a potential regulatory gene in breast cancer. KSR1, originally described as a novel protein kinase, has a role in activation of mitogen-activated protein kinases. Emerging evidence has shown that KSR1 may have dual functions as an active kinase as well as a scaffold facilitating multiprotein complex assembly. Although efforts have been made to study the role of KSR1 in certain tumour types, its involvement in breast cancer remains unknown. METHODS: A quantitative mass spectrometry analysis using stable isotope labelling of amino acids in cell culture (SILAC) was implemented to identify KSR1-regulated phosphoproteins in breast cancer. In vitro luciferase assays, co-immunoprecipitation as well as western blotting experiments were performed to further study the function of KSR1 in breast cancer. RESULTS: Of significance, proteomic analysis reveals that KSR1 overexpression decreases deleted in breast cancer-1 (DBC1) phosphorylation. Furthermore, we show that KSR1 decreases the transcriptional activity of p53 by reducing the phosphorylation of DBC1, which leads to a reduced interaction of DBC1 with sirtuin-1 (SIRT1); this in turn enables SIRT1 to deacetylate p53. CONCLUSION: Our findings integrate KSR1 into a network involving DBC1 and SIRT1, which results in the regulation of p53 acetylation and its transcriptional activity.

LMTK3 is implicated in endocrine resistance via multiple signaling pathways.

Resistance to endocrine therapy in breast cancer is common. With the aim of discovering new molecular targets for breast cancer therapy, we have recently identified LMTK3 as a regulator of the estrogen receptor-alpha (ERα) and wished to understand its role in endocrine resistance. We find that inhibition of LMTK3 in a xenograft tamoxifen (Tam)-resistant (BT474) breast cancer mouse model results in re-sensitization to Tam as demonstrated by a reduction in tumor volume. A whole genome microarray analysis, using a BT474 cell line, reveals genes significantly modulated (positively or negatively) after LMTK3 silencing, including some that are known to be implicated in Tam resistance, notably c-MYC, HSPB8 and SIAH2. We show that LMTK3 is able to increase the levels of HSPB8 at a transcriptional and translational level thereby protecting MCF7 cells from Tam-induced cell death, by reducing autophagy. Finally, high LMTK3 levels at baseline in tumors are predictive for endocrine resistance; therapy does not lead to alteration in levels, whereas in patient's plasma samples, acquired LMTK3 gene amplification (copy number variation) was associated with relapse while receiving Tam. In aggregate, these data support a role for LMTK3 in both innate (intrinsic) and acquired (adaptive) endocrine resistance in breast cancer.

Aberrant production of soluble costimulatory molecules CTLA-4, CD28, CD80 and CD86 in patients with systemic lupus erythematosus.

OBJECTIVE: The costimulatory interactions of the B7 family molecules CD80 and CD86 on antigen-presenting cells with their T-cell counter-receptors CD28 and CTLA-4 modulate T lymphocyte-mediated immune responses in a reciprocal manner. We investigated the possible aberrant production of soluble (s) forms of the T-cell costimulatory molecules CD80, CD86, CD28 and CTLA-4 in plasma of patients with systemic lupus erythematosus (SLE), an autoimmune disease arising from T-lymphocyte dysregulation. METHODS: Plasma concentration and ex vivo production of soluble costimulatory molecules of 79 SLE patients with or without active disease and 40 sex- and age-matched healthy subjects were measured by enzyme-linked immunosorbent assay. RESULTS: Plasma sCTLA-4, sCD28, sCD80 and sCD86 concentrations of all SLE patients were significantly higher than concentrations in control subjects (all P<0.01). These increases were observed even in patients with inactive disease [SLE Disease Activity Index (SLEDAI) <3]. Plasma sCTLA-4 concentration in all SLE patients correlated significantly with SLEDAI score (r = 0.228, P = 0.043). Upon mitogen treatment of peripheral blood mononuclear cells, the percentage increases in ex vivo production of sCD28 and sCD80 and the percentage decrease in sCTLA-4 release were all significantly smaller in SLE patients with active disease than in healthy subjects (P<0.01, P<0.05 and P<0.0001, respectively). CONCLUSION: The aberrant production of soluble T-cell costimulatory molecules is important in the immunopathogenesis of SLE, which occurs by the dysregulation of T-lymphocyte costimulation. Plasma sCTLA concentration could potentially serve as a surrogate marker of SLE disease activity.

Elevation of plasma osteopontin concentration is correlated with disease activity in patients with systemic lupus erythematosus.

BACKGROUND: Osteopontin (OPN) is an extracellular matrix cell adhesion phosphoprotein with immunological activities including stimulation of macrophage chemotaxis, T-helper type 1 lymphocyte response and B-cell antibody synthesis. Overexpression of OPN has been associated with the development of the autoimmune/lymphoproliferative syndrome. METHODS: We measured the plasma concentration and ex vivo production of OPN, and the plasma proinflammatory IL-18 concentration in 54 SLE patients with or without renal impairment (RSLE group and SLE group, respectively) and 26 sex- and age-matched control (NC) subjects using an enzyme-linked immunoabsorbent assay. RESULTS: Plasma OPN concentrations were significantly higher in RSLE and SLE patients than in the NC group (both P<0.001). Increase in OPN concentration correlated positively and significantly with SLEDAI score in all SLE patients (r = 0.308, P = 0.023). The ex vivo production of OPN upon mitogen activation of peripheral blood mononuclear cells was significantly higher in the RSLE and SLE groups than in the NC group (both P<0.001). In RSLE patients, plasma OPN concentration showed a significant positive correlation with proinflammatory cytokine IL-18 concentration (r = 0.404, P = 0.037). CONCLUSION: The above results suggest that the production of OPN is associated with the inflammatory process and SLE development, and may serve as a potential disease marker of SLE.

Plasma inflammatory cytokines and chemokines in severe acute respiratory syndrome.

Severe acute respiratory syndrome (SARS) is a recently emerged infectious disease caused by a novel coronavirus, but its immunopathological mechanisms have not yet been fully elucidated. We investigated changes in plasma T helper (Th) cell cytokines, inflammatory cytokines and chemokines in 20 patients diagnosed with SARS. Cytokine profile of SARS patients showed marked elevation of Th1 cytokine interferon (IFN)-gamma, inflammatory cytokines interleukin (IL)-1, IL-6 and IL-12 for at least 2 weeks after disease onset, but there was no significant elevation of inflammatory cytokine tumour necrosis factor (TNF)-alpha, anti-inflammatory cytokine IL-10, Th1 cytokine IL-2 and Th2 cytokine IL-4. The chemokine profile demonstrated significant elevation of neutrophil chemokine IL-8, monocyte chemoattractant protein-1 (MCP-1), and Th1 chemokine IFN-gamma-inducible protein-10 (IP-10). Corticosteroid reduced significantly IL-8, MCP-1 and IP-10 concentrations from 5 to 8 days after treatment (all P < 0.001). Together, the elevation of Th1 cytokine IFN-gamma, inflammatory cytokines IL-1, IL-6 and IL-12 and chemokines IL-8, MCP-1 and IP-10 confirmed the activation of Th1 cell-mediated immunity and hyperinnate inflammatory response in SARS through the accumulation of monocytes/macrophages and neutrophils.