Design and Synthesis of Cyanine-based Near-infrared Fluorescent Probes and Their Application in Bioimaging

Abstract

NIR probes have become increasingly popular tools in the field of bioimaging. It possesses two distinct advantages over visible light emission fluorescent probes. First, NIR light penetrates deeper into biological tissues than visible light does. It therefore allows imaging of biological analytes in deeper tissues. Second, the low absorption of biological molecules in the NIR region leads to dramatically reduced levels of auto-fluorescence, which enables the in vivo imaging of important bio-molecules. The mian objective of this dissertation is to design and synthesize small molecular-based near-infrated (NIR) fluorescent probes, which can find application in imaging biologically important small molecules or enzymes.

In chapter 1, an introduction to NIR fluorescent probes was provided, with a focus on the strategies for design of NIR fluorescent probes and their bio-imaging application.

In chapter 2, two novel NIR probes for imaging HNO were designed and prepared. One of the probes was found to display high sensitivity towards HNO, with up to 67-fold of fluorescence increment after reaction with HNO. The detection limit of this probe was determined to be as low as 0.043 μM. In addition, the probe displayed high selectivity towards HNO over other biologically related species including metal ions, reactive oxygen species, reactive nitrogen species and reactive sulfur species. Furthermore, the probe was shown to be suitable for imaging of exogenous and endogenous HNO in living cells. Interestingly, the probe was found to be mainly localized in lysosomes. We envision that the new NIR probe described in this chapter will serve as a useful tool for further elucidation of the intricate roles of HNO in living cells.

In chapter 3, we developed the first reaction based NIR probe (DHXP) for sensitive detection of alkaline phosphatase activity both in vitro and in vivo. Our studies demonstrated that the probe displayed an up to 66-fold fluorescence increment upon incubation with alkaline phosphatases, and the detection limit of our probe was determined to be 0.07 U/L, which is lower than that of most of alkaline phosphatase probes reported in literature. Furthermore, we demonstrated that the probe can be applied to detect alkaline phosphatase activity in cells and mice. In addition, our probe possesses excellent biocompatibility and rapid cell-internalization ability.

In chapter 4, we constructed a number of NIR probes for NQO1, which is overexpressed in human cancer cells. In vitro experiments demonstrated that one of the probe utilizing QCy7 as the latent NIR reporter displayed excellent properties such as fast response rate (5 min), good water solubility (without any organic co-solvent), high sensitivity, and superb selectivity. We proved that the probe can be applied to detect NQO1 activity in living cells. In addition, application of the probe to image tumor xenografts in mice is under study.

In chapter 5, we designed and synthesized an NIR probe for human granzyme B. The probe can detect granzyme B in NK cell lysates. However, the cell imaging experiment was unsuccessful due to the diffusion of the NIR reporter. Due to the lack of time, imaging granzyme B in living cells was not successful. Our lab is currently investigating on using cell-tracking NIR dye as the reporter and decorating the probe with cell-penetrating peptide.

Finally, chapter 6 provides a brief summary of this dissertation, pointing out the drawbacks of NIR probes for bio-imaging applications. At the same time, it also highlights that much efforts should be put in designing NIR fluorescent probes with good photo and chemical stability, appropriate balance of lipophilicity and hydrophilicity, and superb signal to noise ratio. In additon, future effrots also include designing and preparing NIR probes with fast cell internalization ability and released fluorophore would precipitate or immobilized at the reaction sites, which would provide better tools for real time information of the bio-analytes.

Date of Award	26 Jan 2018
Original language	English
Awarding Institution	City University of Hong Kong
Supervisor	Hongyan SUN (Supervisor)