Application of Conjugated Polyelectrolyte in Biosensor

Ph.DDOCTOR OF PHILOSOPH

Nanoparticle-assisted optical tethering of endosomes reveals the cooperative function of dyneins in retrograde axonal transport

Dynein-dependent transport of organelles from the axon terminals to the cell bodies is essential to the survival and function of neurons. However, quantitative knowledge of dyneins on axonal organelles and their collective function during this long-distance transport is lacking because current technologies to do such measurements are not applicable to neurons. Here, we report a new method termed nanoparticle-assisted optical tethering of endosomes (NOTE) that made it possible to study the cooperative mechanics of dyneins on retrograde axonal endosomes in live neurons. In this method, the opposing force from an elastic tether causes the endosomes to gradually stall under load and detach with a recoil velocity proportional to the dynein forces. These recoil velocities reveal that the axonal endosomes, despite their small size, can recruit up to 7 dyneins that function as independent mechanical units stochastically sharing load, which is vital for robust retrograde axonal transport. This study shows that NOTE, which relies on controlled generation of reactive oxygen species, is a viable method to manipulate small cellular cargos that are beyond the reach of current technology.National Institutes of Health (U.S.) (DP2-NS082125)National Science Foundation (U.S.) (Award 1055112)National Science Foundation (U.S.) (Award 1344302

Development of optical nanoprobes for molecular imaging of reactive oxygen and nitrogen species

Reactive oxygen and nitrogen species (RONS) play important roles in cell signal transduction. However, overproduction of RONS is associated with a series of pathological processes and may disrupt cellular homeostasis, causing oxidative and nitrosative stress. Accurate methods to selectively and specifically monitor RONS in living systems are required to further elucidate the biological functions of these species. Optical imaging possesses high sensitivity, high spatiotemporal resolution, and real-time imaging capability. These qualities are advantageous for the detection of RONS in living systems. This review summarizes the development of optical nanoprobes with near-infrared (NIR) fluorescent, upconversion luminescent, chemiluminescent, or photoacoustic signals for molecular imaging of RONS in living systems. In this review, we discuss the design principles and advantages of RONS-responsive activatable nanoprobes, as well as applications of these optical imaging modalities in different disease models.MOE (Min. of Education, S’pore)Accepted versio

Semiconducting polymer nanomaterials as near-infrared photoactivatable protherapeutics for cancer

Cancer therapy is routinely performed in the clinic to cure cancer and control its progression, wherein therapeutic agents are generally used. To reduce side effects, protherapeutic agents that can be activated by overexpressed cancer biomarkers are under development. However, these agents still face certain extent of off-target activation in normal tissues, stimulating the interest to design external-stimuli activatable protherapeutics. In this regard, photoactivatable protherapeutic agents have been utilized for cancer treatments. However, because of the intrinsic features of photolabile moieties, most photoactivatable protherapeutic agents only respond to ultraviolet-visible light, limiting their in vivo applications. Thus, protherapeutic agents that can be activated by near-infrared (NIR) light with minimal phototoxicity and increased tissue penetration are highly desired.In this Account, we summarize our semiconducting polymer nanomaterials (SPNs) as NIR photoactivatable protherapeutic agents for cancer treatment. SPNs are transformed from π-conjugated polymers that efficiently convert NIR light into heat or singlet oxygen (¹O₂). With photothermal and photodynamic properties, SPNs can be directly used as photomedicine or serve as light transducers to activate heat or ¹O₂₋ responsive protherapeutic agents.The heat-activatable SPN-based protherapeutic agents are developed by loading or conjugating of SPNs with therapeutic agents (e.g., agonist, gene, and enzyme). For instance, photothermally triggered release of agonists specifically activates certain protein ion channels on the cellular membrane, leading to ion overinflux induced mitochondria dysfunction and consequently apoptosis of cancer cells. Moreover, photothermal activation of temperature-sensitive bromelain can promote the in situ degradation of collagens (the major components of extracellular matrix), resulting in an improved accumulation of agents in tumor tissues and thus amplified therapeutic outcome.The¹O₂₋ activatable SPN-based protherapeutic agents are constructed through covalent conjugation of SPNs with caged therapeutic agents via hypoxia- or ¹O₂₋ cleavable linkers. Upon NIR photoirradiation, SPNs consume oxygen to generate ¹O₂, which leads to photodynamic therapy (PDT), and meanwhile breaks hypoxia- or ¹O₂₋ cleavable linkers for on-demand release and in situ activation of caged protherapeutic molecules (e.g., chemodrug, enzyme, and inhibitor). Such remote activation of SPN-based protherapeutic agents can be applied to induce DNA damage, ribonucleic acid degradation, inhibition of protein biosynthesis, or immune system activation in tumors of living animals. By synergizing PDT with NIR photoactivation of those biological actions, these protherapeutic agents effectively eliminate tumors and even fully inhibit tumor metastasis.This Account highlights the potential of SPNs for construction of versatile NIR photoactivatable protherapeutics to treat cancer at designated times and locations with high therapeutic outcome and precision.Ministry of Education (MOE)Nanyang Technological UniversityK.P. thanks Nanyang Technological University (Start-Up Grant: M4081627) and Singapore Ministry of Education Academic Research Fund Tier 1 (2017-T1-002-134, RG147/17) and Academic Research Fund Tier 2 (MOE2016-T2-1-098&MOE2018-T2-2-042) for the financial support

Multimodal biophotonics of semiconducting polymer nanoparticles

Biophotonics as an interdisciplinary frontier plays an increasingly important role in modern biomedical science. Optical agents are generally involved in biophotonics to interpret biomolecular events into readable optical signals for imaging and diagnosis or to convert photons into other forms of energy (such as heat, mechanical force, or chemical radicals) for therapeutic intervention and biological stimulation. Development of new optical agents including metallic nanoparticles, quantum dots, up-conversion nanoparticles, carbon dots, and silica nanoparticles has contributed to the advancement of this field. However, most of these agents have their own merits and demerits, making them less effective as multimodal biophotonic platforms. In this Account, we summarize our recent work on the development of near-infrared (NIR) semiconducting polymer nanoparticles (SPNs) as multimodal light converters for advanced biophotonics. SPNs are composed of π-electron delocalized semiconducting polymers (SPs) and often possess the advantages of good biocompatibility, high photostability, and large absorption coefficients. Because the photophysical properties of SPNs are mainly determined by the molecular structures of the precursor polymers, molecular engineering allows us to fine tune their photophysical processes to obtain different optical responses, even to light in the second NIR window (1000-1700 nm). Meanwhile, the facile nanoformulation methods of SPNs enable alteration of their outer and inner structures for diverse biological interactions. The unique photophysical properties of SPNs have brought about ultrasensitive deep-tissue molecular imaging. NIR-absorbing SPNs with strong charge-transfer backbones can convert photoenergy into mechanical acoustic waves, permitting photoacoustic imaging that bypasses the issue of light scattering and reaches the centimeter tissue penetration depth. Differently, phenylenevinylene-containing SPNs can store photon energy via chemical defects and emit long-NIR afterglow luminescence with a half-life of ∼6 min after cessation of light excitation. Such an afterglow process avoids tissue autofluorescence, giving rise to ultrahigh signal-to-background ratios. So far, SPN-based molecular photoacoustic or afterglow probes have been developed to image disease tissues (tumors), biomarkers (biothiols and reactive oxygen species), and physiological indexes (pH and temperature) in different preclinical animal models. The synthetic flexibility of SPNs further permits light-modulated biological and therapeutic interventions. Till now, SPNs with high photothermal conversion efficiencies have been shaped into photothermal transducers to remotely regulate biological events including protein ion channels, enzymatic activity, and gene expression. In conjunction with the desired biodistribution and tumor-homing ability, SPNs have been doped or coated with other inorganic agents for amplified photothermal or self-regulated photodynamic cancer therapy. This Account thus demonstrates that SPNs serve as a multimodal biophotonic nanoplatform to provide unprecedented opportunities for molecular imaging, noninvasive bioactivation, and advanced therapy.MOE (Min. of Education, S’pore)Accepted versio

Molecular and nanoengineering approaches towards activatable cancer immunotherapy

Cancer immunotherapy is an emerging treatment strategy that modulates the immune system to fight against cancer. Although several immunotherapeutic agents have been utilized in the clinic for cancer treatment, low patient response rates and potential immune-related adverse events remain two major challenges. With the merits of delivery controllability and modular flexibility, nanomedicines provide opportunities to facilitate immunotherapies for clinical translation in a safe and effective manner. In this review, we discuss the convergence of nanomedicine with immunotherapy with a focus on molecular and nanoengineering approaches towards activatable cancer immunotherapy. These activatable nanoagents exert immunotherapeutic action only in response to internal or external stimuli. This allows them to locally reprogram the tumor microenvironment and activate antitumor immunity while reducing the incidence of immune-related adverse events. The category of activatable immunotherapeutic nanoagents are discussed along with an overview of their clinical translation prospects and challenges.Ministry of Education (MOE)Nanyang Technological UniversityK. P. thanks Nanyang Technological University (Start-up Grant No. NTU-SUG: M4081627.120) and Singapore Ministry of Education, Academic Research Fund Tier 1 (2017-T1-002-134; 2019-T1-002-045) and Academic Research Fund Tier 2 (MOE2018-T2-2-042) for the financial support

Molecular fluorescence and photoacoustic imaging in the second near-infrared optical window using organic contrast agents

Noninvasive near-infrared (NIR) light ranging from 650 to 1000 nm (NIR-I) is widely employed in fundamental research and clinical applications; however, a recently discovered second NIR (NIR-II) window from 1000 to 1700 nm exhibits even better deep-tissue imaging capability due to reduced photon scattering, minimized tissue autofluorescence, and increased applicable power at longer wavelengths. This review focuses on recent advances of organic contrast agents developed for in vivo fluorescence and photoacoustic imaging in the NIR-II optical window. The superiority of the NIR-II over the NIR-I window for molecular imaging is first discussed in detail, followed by discussion of fluorescence and photoacoustic imaging of cancer, vasculature, and the brain using organic contrast agents in the NIR-II window. At last, challenges and perspectives of organic contrast agents for NIR-II in vivo imaging are suggested.Accepted versio

Development of organic semiconducting materials for deep-tissue optical imaging, phototherapy and photoactivation

Biophotonics as a highly interdisciplinary frontier often requires the assistance of optical agents to control the light pathways in cells, tissues and living organisms for specific biomedical applications. Organic semiconducting materials (OSMs) composed of π-conjugated building blocks as the optically active components have recently emerged as a promising category of biophotonic agents. OSMs possess the common features including excellent optical properties, good photostability and biologically benign composition. This review summarizes the recent progress in the development of OSMs based on small-molecule fluorophores, aggregation-induced emission (AIE) dyes and semiconducting oligomer/polymer nanoparticles (SONs/SPNs) for advanced biophotonic applications. OSMs have been exploited as imaging agents to transduce biomolecular interactions into second near-infrared fluorescence, afterglow or photoacoustic signals, enabling deep-tissue ultrasensitive imaging of biological tissues, disease biomarkers and physiological indexes. By finetuning the molecular structures, OSMs can also convert light energy into cytotoxic free radicals or heat, allowing for effective cancer phototherapy. Due to their instance light response and efficient light-harvesting properties, precise regulation of biological activities using OSMs as the remote transducers has been demonstrated for protein ion channels, gene transcription and protein activation. In addition to highlighting OSMs as a multifunctional platform for a wide range of biomedical applications, current challenges and perspectives of OSMs in biophotonics are discussed.MOE (Min. of Education, S’pore)Accepted versio

Molecular probes for autofluorescence-free optical imaging

Optical imaging is an indispensable tool in clinical diagnostics and fundamental biomedical research. Autofluorescence-free optical imaging, which eliminates real-time optical excitation to minimize background noise, enables clear visualization of biological architecture and physiopathological events deep within living subjects. Molecular probes especially developed for autofluorescence-free optical imaging have been proven to remarkably improve the imaging sensitivity, penetration depth, target specificity, and multiplexing capability. In this Review, we focus on the advancements of autofluorescence-free molecular probes through the lens of particular molecular or photophysical mechanisms that produce long-lasting luminescence after the cessation of light excitation. The versatile design strategies of these molecular probes are discussed along with a broad range of biological applications. Finally, challenges and perspectives are discussed to further advance the next-generation autofluorescence-free molecular probes for in vivo imaging and in vitro biosensors.Agency for Science, Technology and Research (A*STAR)Ministry of Education (MOE)K.P. thanks Singapore Ministry of Education, Academic Research Fund Tier 1 (2019-T1-002-045 and RG125/19), Academic Research Fund Tier 2 (MOE2018-T2-2-042), and A*STAR SERC AME Programma