Development of amphiphilic compounds as therapeutic and diagnostic agents targeting amyloid aggregates for Alzheimer’s Disease

Yu, Zhengxin

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https://hdl.handle.net/2142/121230 Copy

Description

Title

Development of amphiphilic compounds as therapeutic and diagnostic agents targeting amyloid aggregates for Alzheimer’s Disease

Author(s)

Yu, Zhengxin

Issue Date

2023-07-12

Director of Research (if dissertation) or Advisor (if thesis)

Mirica, Liviu

Doctoral Committee Chair(s)

Mirica, Liviu

Committee Member(s)

• Murphy, Catherine
• Chan, Jefferson
• Dobrucki, Wawrzyniec

Department of Study

Chemistry

Discipline

Chemistry

Degree Granting Institution

University of Illinois at Urbana-Champaign

Degree Name

Ph.D.

Degree Level

Dissertation

Keyword(s)

• Amphiphilic compounds
• Alzheimer’s Disease
• Amyloid-β (Aβ)
• Near-infrared (NIR) fluorescence imaging
• Positron emission tomography (PET) imaging

Abstract

Neurodegenerative diseases have garnered significant attention in the 21st century due to the projected substantial growth of the global aging population. Among these diseases, Alzheimer's Disease (AD) stands out as the most prevalent, severely impacting patients' daily lives through memory loss and cognitive decline. Unfortunately, effective treatments remain scarce. Multiple pathological factors have been implicated in AD, with proteopathy being a prominent mechanism. Encouragingly, two monoclonal antibodies (aducanumab and lecanemab) have been approved by the U.S. Food and Drug Administration (FDA) as the first disease-modifying therapies (DMTs). These antibodies have demonstrated the ability to reduce amyloid-β (Aβ) aggregates and slow cognitive decline in early-stage AD patients. These recent breakthroughs reinforce the notion that targeting Aβ aggregates will continue to play a vital role in the development of therapeutic and diagnostic agents for AD. In our pursuit of developing small molecule therapeutics for AD, we have generated a diverse library of small molecules comprising various hydrophobic and hydrophilic fragments, aiming to target Aβ aggregates. The resulting amphiphilic compounds demonstrate notable binding affinity towards both Aβ plaques and oligomers, with six compounds exhibiting selective binding specifically to Aβ oligomers. Immunofluorescence staining of brain sections from transgenic AD mice confirms the ability of these amphiphilic compounds to label native Aβ species. Encouragingly, four of the amphiphilic compounds exhibit the capacity to alleviate Cu2+-Aβ induced toxicity as demonstrated in cell viability assays. Furthermore, confocal fluorescence imaging studies reveal that two compounds, namely ZY-15-MT and ZY-15-OMe, effectively disrupt the interactions between Aβ oligomers and the cell membranes of human neuroblastoma SH-SY5Y cells. Collectively, these investigations strongly indicate that the development of compounds possessing amphiphilic properties, capable of targeting Aβ oligomers and modulating their interactions with cell membranes, holds substantial potential as an effective strategy for the development of small molecule therapeutics for AD. In addition to the development of potential therapeutic agents for AD, our endeavours also encompass the advancement of imaging agents for the early diagnosis of AD. The presymptomatic stage of AD can extend over decades, preceding a stage of mild cognitive impairment (MCI) during which progressive impairment develops. By utilizing reliable and readily available detection methodologies, our aim is to diagnose patients with AD at an early stage, facilitating prompt intervention that can significantly enhance their quality of life. Near-Infrared (NIR) and positron emission tomography (PET) imaging are advanced molecular imaging techniques extensively employed for AD diagnosis. The development of multimodal imaging agents that offer cooperative advantages is highly desirable. Through rational molecular design, we present a novel near-infrared probe, DCM-OH-2-DT, based on the dicyanomethylene-4H-pyran (DCM) platform, capable of detecting both soluble and insoluble Aβ species. DCM-OH-2-DT exhibits high binding affinity towards Aβ oligomers and excellent blood brain barrier (BBB) permeability. In vivo imaging demonstrates significantly higher NIR fluorescence signal of DCM-OH-2-DT in 7-month 5xFAD AD mice compared to age-matched wild type mice. Additionally, the Cu-chelating groups, 2,4-dimethyl-1,4,7-triazacyclononane (Me2TACN), integrated into DCM-OH-2-DT exhibit strong Cu binding ability, highlighting its potential as 64Cu positron emission tomography (PET) imaging agent. The developed probe, which can be employed for both NIR and PET imaging, holds promise for monitoring therapeutic effects in preclinical studies or routine clinical applications. Furthermore, our investigation reveals that the incorporation of macrocyclic Me2TACN groups onto the DCM scaffold facilitates excited-state intramolecular transfer (ESIPT) and enables emission in the NIR window. We anticipate that this macrocycle promoted ESIPT phenomenon can find valuable applications in biomedical imaging. While a few PET imaging agents based on 11C (t1/2 = 20.4 min) and 18F (t1/2 = 109.7 min) are available for clinical AD diagnosis, their limited half-life poses a challenge for broader applications. In this regard, 64Cu (t1/2 = 12.7 h), with its longer half-life, holds promise for brain imaging in AD diagnosis. However, 64Cu PET imaging agents have been associated with in vivo reduction and unproductive demetallation. To enhance the in vivo stability of 64Cu-based PET imaging tracers for AD, novel ligand design strategies are required. Therefore, we developed sulfur-containing (N2S2) bifunctional chelators (BFCs) as well as the conventional all nitrogen-based (N4tBu) BFCs to directly compare their abilities to chelate Cu and target Aβ aggregates. Our results demonstrate that both N2S2-based and N4tBu-based BFCs exhibit high stability constants towards Cu(II). Notably, the N2S2-based BFCs show over 10 orders of magnitude higher binding affinity towards Cu(I) compared to their N4tBu-based counterparts. Aβ binding experiments were conducted to study the structure-affinity relationship, and fluorescence microscopy imaging studies confirmed the selective labelling of the BFCs and their copper complexes. Furthermore, we investigated the potential of these ligands for 64Cu-based PET imaging of AD through radiolabelling and autoradiography studies. We believe our findings provide molecular insights into the design of bifunctional Cu chelators that can effectively stabilize both Cu(II) and Cu(I). These insights can greatly contribute to the development of 64Cu PET imaging for the diagnosis of AD.

Graduation Semester

2023-08

Type of Resource

Thesis

Copyright and License Information

Copyright 2023 Zhengxin Yu

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