Longitudinal CSF proteomics identifies NPTX2 as a prognostic biomarker of Alzheimer's disease.

INTRODUCTION: Biomarkers that reflect pathologic processes affecting neuronal function during preclinical and early stages of Alzheimer's disease (AD) are needed to aid drug development. METHODS: A targeted, stable isotope, quantitative mass spectrometry-based investigation of longitudinal changes in concentrations of previously identified candidate biomarkers was performed in cerebrospinal fluid (CSF) of Alzheimer's Disease Neuroimaging Initiative participants who were classified as cognitively normal (CN; n = 76) or with mild cognitive impairment (MCI; n = 111) at baseline. RESULTS: Of the candidate biomarkers, the CSF concentration of neuronal pentraxin 2 (NPTX2), a protein involved in synaptic function, exhibited rates of change that were significantly different between three comparison groups (i.e., CN vs. MCI participants; AD pathology positive vs. negative defined by phosphorylated tau181/amyloid beta1-42 ratio; and clinical progressors vs. non-progressors). The rate of change of NPTX2 also significantly correlated with declining cognition. DISCUSSION: CSF NPTX2 concentration is a strong prognostic biomarker candidate of accelerated cognitive decline with potential use as a therapeutic target.

Inhibition of BET proteins impairs estrogen-mediated growth and transcription in breast cancers by pausing RNA polymerase advancement.

Estrogen (E2)-induced transcription requires coordinated recruitment of estrogen receptor α (ER) and multiple factors at the promoter of activated genes. However, the precise mechanism by which this complex stimulates the RNA polymerase II activity required to execute transcription is largely unresolved. We investigated the role of bromodomain (BRD) containing bromodomain and extra-terminal (BET) proteins, in E2-induced growth and gene activation. JQ1, a specific BET protein inhibitor, was used to block BET protein function in two different ER-positive breast cancer cell lines (MCF7 and T47D). Real-time PCR and ChIP assays were used to measure RNA expression and to detect recruitment of various factors on the genes, respectively. Protein levels were measured by Western blotting. JQ1 suppressed E2-induced growth and transcription in both MCF7 and T47D cells. The combination of E2 and JQ1 down-regulated the levels of ER protein in MCF7 cells but the loss of ER was not responsible for JQ1-mediated inhibition of E2 signaling. JQ1 did not disrupt E2-induced recruitment of ER and co-activator (SRC3) at the E2-responsive DNA elements. The E2-induced increase in histone acetylation was also not altered by JQ1. However, JQ1 blocked the E2-induced transition of RNA polymerase II from initiation to elongation by stalling it at the promoter region of the responsive genes upstream of the transcription start site. This study establishes BET proteins as the key mediators of E2-induced transcriptional activation. This adds another layer of complexity to the regulation of estrogen-induced gene activation that can potentially be targeted for therapeutic intervention.

Cyclin dependent kinase-9 mediated transcriptional de-regulation of cMYC as a critical determinant of endocrine-therapy resistance in breast cancers.

Endocrine therapy resistance in estrogen receptor alpha positive (ERα+) breast cancers remains a major obstacle for maintaining efficacy of targeted therapies. We investigated the significance and the mechanisms involved in cMYC over-expression in a MCF7 derived panel of ERα+ breast cancer cells which can proliferate in the absence of estrogen with different sensitivities to anti-hormone therapies. We show that all the resistant cell lines tested over-express cMYC as compared to parental MCF7 cells and its inhibition lead to the differential blocking of estrogen-independent proliferation in resistant cells. Further investigation of the resistant cell line, MCF7:5C, suggested transcriptional de-regulation of cMYC gene was responsible for its over-expression. Chromatin immuno-precipitation assay revealed markedly higher recruitment of phosphorylated serine-2 carboxy-terminal domain (CTD) of RNA polymerase-II at the proximal promoter of cMYC gene, which is responsible for transcriptional elongation of the cMYC RNA. The level of CDK9, a factor responsible for the phosphorylation of serine-2 of RNA polymerase II CTD, was found to be elevated in all the resistant cell lines. Pharmacological inhibition of CDK9 not only reduced the transcripts and the protein levels of cMYC in MCF7:5C cells but also selectively inhibited the estrogen-independent growth of all the resistant cell lines. This study describes the up-stream molecular events involved in the transcriptional over-expression of cMYC gene in breast cancer cells proliferating estrogen-independently and identifies CDK9 as a potential novel drug target for therapeutic intervention in endocrine-resistant breast cancers.

Snapshot of spatio-temporal cytokine responses to single and co-infections with helminths and bacteria.

Cytokines play a key role in maintaining communication between organs and in so doing modulate the interaction between concurrent infections. The extent of these effects depends on the properties of the organ infected and the intensity and type of infections. To determine systemic bystander effects among organs, IFN-γ, IL-4 and IL-10 gene expression was quantified at 7 days post-challenge in directly infected and uninfected organs during single and co-infections with the respiratory bacterium Bordetella bronchiseptica and the gastrointestinal helminths Graphidium strigosum and Trichostrongylus retortaeformis. Results showed that cytokine expression in a specific organ was influenced by the type of infection occurring in another organ, and this bystander effect was more apparent in some organs than others. Within the same organ the relative cytokine expression was consistent across infections, although some cytokines were more affected by bystander effects than others. For the infected gastrointestinal tract, a stronger cytokine response was observed in the tissue that harbored the majority of helminths (i.e. duodenum and fundus). Overall, co-infections altered the intensity but to a lesser extent the relative cytokine profile against the focal infection, indicating clear bystander effects and low organ compartmentalization. However, organs appear to actively modulate cytokine expression to avoid potential immuno-pathological consequences.