Objective:Examine the relationship between scalp EEG biomarkers of hyperexcitability in Alzheimer’s disease (AD) and determine how these electrical biomarkers relate to the clinical expression of seizures in AD.Methods:In this cross-sectional study, we performed 24-hour ambulatory scalp EEGs on 43 cognitively normal elderly healthy controls (HC), 41 early-stage AD participants with no history or risk factors for epilepsy (AD-NoEp), and 15 early-stage AD participants with late-onset epilepsy related to AD (AD-Ep). Two epileptologists, blinded to diagnosis, visually reviewed all EEGs and annotated all potential epileptiform abnormalities. A panel of 9 epileptologists, blinded to diagnosis, was then surveyed to generate a consensus interpretation of epileptiform abnormalities in each EEG.Results:Epileptiform abnormalities were seen in 53% of AD-Ep, 22% of AD-NoEp, and 4.7% of HC participants. Specific features of epileptiform discharges, including high frequency, robust morphology, right temporal location, and occurrence during wakefulness and REM, were associated with clinical seizures in AD. Multiple electrical biomarkers concordantly demonstrated a pattern of left temporal lobe hyperexcitability in early stages of AD, whereas clinical seizures in AD were often associated with bi-temporal hyperexcitability. Frequent small sharp spikes were specifically associated with epileptiform EEGs and thus identified as a potential biomarker of hyperexcitability in AD.Conclusion:Epileptiform abnormalities are common in AD, but not all equivalent. Specific features of epileptiform discharges are associated with clinical seizures in AD. Given the difficulty recognizing clinical seizures in AD, these EEG features could provide guidance on which AD patients are at high risk for clinical seizures.

Transcription mediated by hypoxia-inducible factor (HIF-1) contributes to tumor angiogenesis and metastasis but is also involved in activation of cell-death pathways and normal physiological processes. Given the complexity of HIF-1 signaling, it could be advantageous to target a subset of HIF-1 effectors rather than the entire pathway. We compare the genome-wide effects of three molecules that each interfere with the HIF-1-DNA interaction: a polyamide targeted to the hypoxia response element, small interfering RNA targeted to HIF-1alpha, and echinomycin, a DNA-binding natural product with a similar but less specific sequence preference than the polyamide. The polyamide affects a subset of hypoxia-induced genes consistent with its binding site preferences. For comparison, HIF-1alpha siRNA and echinomycin each affect the expression of nearly every gene induced by hypoxia. Remarkably, the total number of genes affected by either polyamide or HIF-1alpha siRNA over a range of thresholds is comparable. The data show that polyamides can be used to affect a subset of a pathway regulated by a transcription factor. In addition, this study offers a unique comparison of three complementary approaches towards exogenous control of endogenous gene expression.

Regulation of endogenous genes by DNA-binding polyamides requires effective nuclear localization. Previous work employing confocal microscopy to study uptake of fluorophore-labeled polyamides has demonstrated the difficulty of predicting a priori the nuclear uptake of a given polyamide. The data suggest that dye identity influences uptake sufficiently such that a dye-conjugate cannot be used as a proxy for unlabeled analogs. Polyamides capable of nuclear localization unaided by fluorescent dyes are desirable due to size and other limitations of fluorophores. Recently, a polyamide-fluorescein conjugate targeted to the hypoxia response element (HRE) was found to inhibit vascular endothelial growth factor (VEGF) expression in cultured HeLa cells. The current study uses inhibition of VEGF expression as a biological read-out for effective nuclear localization of HRE-targeted polyamides. We synthesized a focused library of non-fluorescent, HRE-targeted polyamides in which the C-terminus ‘tail’ has been systematically varied. Members of this library bind the HRE with affinities comparable or superior to that of the fluorescein-labeled analog. Although most library members demonstrate modest or no biological activity, two non-fluorescent polyamides are reported with activity rivaling that of the previously reported fluorescein-labeled polyamide. We also show evidence that promoter occupancy by HIF-1, the transcription factor that binds the HRE, is inhibited by HRE-targeted polyamides.