Novel 1,4-Diazepane Derivatives as Amyloid Beta (Aβ) Aggregation Inhibitors

Abstract

Alzheimer’s disease (AD) is a progressive and irreversible neurodegenerative disease that represents one of the most pressing medical and social challenges of our time. Characterized by cognitive decline and memory loss, AD is primarily driven by the accumulation of amyloid-β (Aβ) plaques and tau tangles in the brain. The Aβ aggregation is indeed the primary pathological event, playing a key role in the initiation and progression of the disease. Despite extensive research, effective disease-modifying treatments remain elusive, and current therapies offer only symptomatic relief. While monoclonal antibody therapies targeting Aβ have emerged as a potential treatment option, their clinical effectiveness is limited. Small molecules, however, represent a more versatile and economical approach, with the potential for targeted action within the brain. In this regard, this thesis focuses on the design, synthesis, and biological evaluation of small molecules that specifically target Aβ aggregation, aiming to identify effective therapeutic candidates for AD. To this end, the present work explores the use of nitrogen-containing heterocyclics as small-molecule inhibitors of Aβ aggregation. Specifically, small molecules containing a flexible 1,4-diazepane scaffold were investigated for their ability to target the two major isoforms of amyloid-β, Aβ42 and Aβ40. A library of 38 derivatives based on the 1,4-diazepane scaffold was synthesized and evaluated for their ability to inhibit Aβ aggregation. The anti-aggregation activity of these compounds was determined through a combination of fluorescence-based aggregation kinetic assays, transmission electron microscopy (TEM), and computational modelling studies. Additionally, cytotoxicity assessments were performed using mouse hippocampal HT22 neuronal cells, alongside antioxidant assays and blood-brain barrier permeability evaluations. An overview of each chapter is outlined below:

Chapter one provides an in-depth background on AD, addressing its prevalence, clinical diagnosis, and associated brain alterations. It reviews major pathological hypotheses, mainly cholinergic deficits, amyloid-β toxicity, tau pathology, and oxidative stress, and their roles in disease development. The chapter also reviews current therapeutic strategies, including small molecules, and their limitations.

Chapter two provides a rationale for investigating the proposed 1,4-diazepane derivatives in the context of AD drug design by discussing the medicinal chemistry principles and structural features of bioactive natural compounds with amyloid-β inhibition properties. From this rationale, an AD-related hypothesis was formulated, guiding the selection and modification of chemical templates. A design strategy was developed, supported by computational studies, to propose a library of 1,4-diazepane derivatives for evaluation.

Chapter three describes the design, synthesis, and evaluation of the first series of (1,4-diazepan-1-yl)(phenyl)methanone derivatives (4a–n). A library of 14 derivatives was synthesized, featuring varying functional groups at the para-position of the phenyl ring. The design incorporated 3,4-positions of the phenyl ring with known Aβ inhibition pharmacophores and antioxidant moieties, including a masked catechol group to study its effect on Aβ42 and Aβ40 inhibition. The compounds were synthesized by coupling acid chlorides or carboxylic acids with secondary amines. This study identified compounds with moderate to good inhibition of Aβ42 aggregation (32–52%) and enhanced inhibition towards Aβ40 (53–77%). This chapter also reports derivatives showing dual inhibitory effects on both Aβ42 and Aβ40.

Chapter four describes the design, synthesis, and evaluation of a series of symmetric (1,4-diazepane-1,4-diyl)bis(phenylmethanone) derivatives (6a–p). This series contains 16 derivatives and includes derivatives incorporating functional groups at the para-position of the bisphenyl rings. This design strategy also incorporated 3,4-positions of the bis-phenyl rings with known Aβ inhibition pharmacophores and antioxidant moieties, including a masked catechol group, to investigate their impact on the inhibition of Aβ42 and Aβ40 aggregation. Additionally, this design also featured bicyclic aromatic rings, such as naphthyl derivatives. This study identified compounds exhibiting moderated inhibition of Aβ42 aggregation (31– 50%) and better inhibition of Aβ40 (60–63%), along with dual-targeting activity.

Chapter five describes the design, synthesis, and evaluation of a series of (4-substituted-1,4-diazepan-1-yl)(phenyl)methanone derivatives (9a–h). This series comprised eight derivatives, each incorporating alkyl substituents of varying chain lengths at the N4 position of the 1,4-diazepane ring, along with a para-substituent on the phenyl ring. The findings revealed that the N-alkylated 1,4-diazepane derivatives exhibited reduced inhibitory activity toward Aβ42 (34%), in contrast to the derivatives described in Chapters 3 and 4. However, these compounds demonstrated improved inhibition of Aβ40 aggregation (55–67%) compared to those from Chapter 4, with one derivative identified as a dual-targeting agent.

Chapter six focused on the biological evaluation of the most active compounds identified through the biophysical studies conducted in Chapters 3, 4, and 5. The lead 1,4-diazepane derivatives exhibited significant neuroprotective potential and effectively rescued the cells from Aβ42-induced cytotoxicity in HT22 cells (47.9–57.4%) and were not toxic to cells. Furthermore, compounds containing pharmacophores with antioxidant properties demonstrated reactive oxygen species (ROS) scavenging activity (13.2–90.7%). The findings also indicated that these derivatives possessed the ability to cross the blood-brain barrier (BBB).

Chapter seven provides a comprehensive summary of the discovery of 1,4-diazepane derivatives as novel templates to design amyloid aggregation inhibitors and discusses the key findings of this study, including the physicochemical properties of the lead derivatives, the significance of this work, and future research directions to consider.