Synthesis of methoxy-X04 derivatives and their evaluation in Alzheimer's disease pathology.

BACKGROUND: Alzheimer's disease is characterized by two notorious protein aggregates in the brain: extracellular senile plaques mainly consisting of amyloid-ß peptides and tau-protein-derived intracellular paired helical filaments. The diagnosis of Alzheimer's disease is impaired by insufficient sensitivity and specificity of diagnostic methods to visualize these pathological hallmarks over all disease stages. OBJECTIVE: The established fluorescence marker methoxy-X04 stains plaques, tau tangles and amyloid-derived angiopathies with good specificity, yet it is limited by slow elimination in vivo. Since the need for new markers is high, we prepared methoxy-X04 derivatives and evaluated their potential as imaging agents in Alzheimer's disease pathology. METHODS AND RESULTS: In this study, we describe an improved synthesis for methoxy-X04 and its derivatives and their affinity determination for the respective protein targets by immunohistology and a displacement assay. CONCLUSION: This resulted in the identification of new derivatives of methoxy-X04 with improved binding affinity.

Bis(arylvinyl)pyrazines, -pyrimidines, and -pyridazines as imaging agents for tau fibrils and ß-amyloid plaques in Alzheimer's disease models.

The in vivo diagnosis of Alzheimer's disease (AD) is of high socioeconomic interest and remains a demanding field of research. The biopathological hallmarks of the disease are extracellular plaques consisting of aggregated ß-amyloid peptides (Aß) and tau protein derived intracellular tangles. Here we report the synthesis and evaluation of fluorescent pyrazine, pyrimidine,and pyridazine derivatives in vitro and in vivo aiming at a tau-based diagnosis of AD. The probes were pre-evaluated on human brain tissue by fluorescence microscopy and were found to label all known disease-related alterations at high contrast and specificity. To quantify the binding affinity, a new thiazine red displacement assay was developed and selected candidates were toxicologically profiled. The application in transgenic mouse models demonstrated bioavailability and brain permeability for one compound. In the course of histological testing, we discovered an AD-related deposition of tau aggregates in the Bowman's glands of the olfactory epithelium, which holds potential for an endoscopic diagnosis of AD in the olfactory system.

NSAID-derived γ-secretase modulation requires an acidic moiety on the carbazole scaffold.

Modulation of γ-secretase activity holds potential for the treatment of Alzheimer's disease. Most NSAID-derived γ-secretase modulators feature a carboxylic acid, which may impair blood-brain barrier permeation. The structure activity relationship of 33 carbazoles featuring diverse carboxylic acid isosteres or metabolic precursors thereof was established in a cellular amyloid secretion assay. The modulatory activity was observed for acidic moieties and metabolically labile esters only, which supports our hypothesis of an acid-lysine interaction to be relevant for this type of γ-secretase modulators.

In vivo multiphoton imaging reveals gradual growth of newborn amyloid plaques over weeks.

The kinetics of amyloid plaque formation and growth as one of the characteristic hallmarks of Alzheimer's disease (AD) are fundamental issues in AD research. Especially the question how fast amyloid plaques grow to their final size after they are born remains controversial. By long-term two-photon in vivo imaging we monitored individual methoxy-X04-stained amyloid plaques over 6 weeks in 12 and 18 months old Tg2576 mice. We found that in 12 months old mice, newly appearing amyloid plaques were initially small in volume and subsequently grew over time. The growth rate of plaques was inversely proportional to their volume; thus amyloid plaques that were already present at the first imaging time point grew over time but slower compared to new plaques. Additionally, we analyzed 18 months old Tg2576 mice in which we neither found newly appearing plaques nor a significant growth of pre-existing plaques over 6 weeks of imaging. In conclusion, newly appearing amyloid plaques are initially small in size but grow over time until plaque growth can not be detected anymore in aged mice. These results suggest that drugs that target plaque formation should be most effective early in the disease, when plaques are growing.