Biphenyl ethers conjugated CdSe/ZnS core/shell quantum dots and interpretation of the mechanism of amyloid fibril disruption

Abstract

The biphenyl ethers (BPEs) are the potent inhibitors of TTR fibril formation and are efficient fibril disrupter. However, the mechanism by which the fibril disruption occurs is yet to be fully elucidated. To gain insight into the mechanism, we synthesized and used a new QD labeled BPE to track the process of fibril disruption. Our studies showed that the new BPE-QDs bind to the fiber uniformly and has affinity and specificity for TTR fiber and disrupted the pre-formed fiber at a relatively slow rate. Based on these studies we put forth the probable mechanism of fiber disruption by BPEs. Also, we show here that the BPE-QDs interact with high affinity to the amyloids of Aβ₄₂, lysozyme and insulin. The potential of BPE-QDs in the detection of senile plaque in the brain of transgenic Alzheimer’s mice has also been explored.

Introduction

It is well documented that proteins frequently alter their native conformation in response to different external stimuli. Some of these conformations may lead to pathogenic states. Under the conditions which destabilize their native structure, a number of proteins aggregate into the characteristic β-sheet rich fibrillar assemblies known as amyloid fibers. The extracellular deposition of these amyloid fibrils is a hallmark of many devastating diseases which include Alzheimer’s disease, prion disease, dialysis-related amyloidosis, familial amyloid polyneuropathy and type II diabetes. Currently, ∼40 different proteins and peptides are known to cause human amyloid disease [1], [2], [3], [4], [5], [6]. Although these diseases have been identified long ago, their cure still remains elusive.

Transthyretin, a tetrameric serum protein has been implicated in the systemic senile amyloidosis (SSA), familial amyloid polyneuropathy (FAP) and familial amyloid cardiopathy (FAC) [7], [8], [9], [10], [11]. In our previous report we designed some potent biphenyl ether (BPE) based molecules as inhibitors of TTR fibrillogenesis and fibril disrupters [12]. We proposed the mechanism of inhibition by these potential BPE inhibitors. However, the mechanism of fiber disruption by these compounds still remains under-explored. We had hypothesized that these small molecules bind to the fiber and thus are able to disrupt them; but the pattern/site and density of binding on fiber remained unanswered. As the process of fibril disruption is very slow, a stable probe is required. To address this issue we have designed and synthesized a new biphenyl ether derivative P8 and coupled them with Quantum dots (QDs). QDs were chosen as models for our studies as they display very interesting optical properties [13], [14], [15], [16], [17] which can be tailored readily according to the end application and unusual target binding properties. These properties make quantum dots very attractive candidates for biolabeling for studies that require longer duration (Fig. 1).

Here, we report mechanism of TTR fibril disruption by BPEs as well as simple and easy-to-perform detection assay for amyloid using a two-step-synthesized, water-soluble, BPE-conjugated core–shell CdSe–ZnS quantum dots. To compare the specificity of these conjugates, the unconjugated QDs and sugar-conjugated QDs (S-QDs) [18] have been used. In this report, we highlight the potential for BPE-conjugated QD (BPE-QD) nanoparticles to disrupt amyloid fibrils in vitro.