Recent Developments in Positron Emission Tomography Tracers for Proteinopathies Imaging in Dementia

An early detection and intervention for dementia represent tremendous unmet clinical needs and priorities in society. A shared feature of neurodegenerative diseases causing dementia is the abnormal accumulation and spreading of pathological protein aggregates, which affect the selective vulnerable circuit in a disease-specific pattern. The advancement in positron emission tomography (PET) biomarkers has accelerated the understanding of the disease mechanism and development of therapeutics for Alzheimer’s disease and Parkinson’s disease. The clinical utility of amyloid-β PET and the clinical validity of tau PET as diagnostic biomarker for Alzheimer’s disease continuum have been demonstrated. The inclusion of biomarkers in the diagnostic criteria has introduced a paradigm shift that facilitated the early and differential disease diagnosis and impacted on the clinical management. Application of disease-modifying therapy likely requires screening of patients with molecular evidence of pathological accumulation and monitoring of treatment effect assisted with biomarkers. There is currently still a gap in specific 4-repeat tau imaging probes for 4-repeat tauopathies and α-synuclein imaging probes for Parkinson’s disease and dementia with Lewy body. In this review, we focused on recent development in molecular imaging biomarkers for assisting the early diagnosis of proteinopathies (i.e., amyloid-β, tau, and α-synuclein) in dementia and discussed future perspectives.

Diazaborines Oxidize Slowly with H2O2 but Rapidly with Peroxynitrite in Aqueous Buffer

Reactive oxygen species (ROS) such as H2O2 and peroxynitrite (ONOO-) oxidize arylboronic acids to their corresponding phenols. When used in molecular imaging probes and in ROS-responsive molecules, however, simple arylboronic...

Red-Shift (2-Hydroxyphenyl)-Benzothiazole Emission by Mimicking the Excited-State Intramolecular Proton Transfer Effect

Novel strategies to optimize the photophysical properties of organic fluorophores are of great significance to the design of imaging probes to interrogate biology. While the 2-(2-hydroxyphenyl)-benzothiazole (HBT) fluorophore has attracted considerable attention in the field of fluorescence imaging, its short emission in the blue region and low quantum yield restrict its wide application. Herein, by mimicking the excited-state intramolecular proton transfer (ESIPT) effect, we designed a series of 2-(2-hydroxyphenyl)-benzothiazole (HBT) derivatives by complexing the heteroatoms therein with a boron atom to enhance the chance of the tautomerized keto-like resonance form. This strategy significantly red-shifted the emission wavelengths of HBT, greatly enhanced its quantum yields, and caused little effect on molecular size. Typically, compounds 12B and 13B were observed to emit in the near-infrared region, making them among the smallest organic structures with emission above 650 nm.

Functionalized Nanomaterials as Tailored Theranostic Agents in Brain Imaging

Functionalized nanomaterials of various categories are essential for developing cancer nano-theranostics for brain diseases; however, some limitations exist in their effectiveness and clinical translation, such as toxicity, limited tumor penetration, and inability to cross blood–brain and blood-tumor barriers. Metal nanomaterials with functional fluorescent tags possess unique properties in improving their functional properties, including surface plasmon resonance (SPR), superparamagnetism, and photo/bioluminescence, which facilitates imaging applications in addition to their deliveries. Moreover, these multifunctional nanomaterials could be synthesized through various chemical modifications on their physical surfaces via attaching targeting peptides, fluorophores, and quantum dots (QD), which could improve the application of these nanomaterials by facilitating theranostic modalities. In addition to their inherent CT (Computed Tomography), MRI (Magnetic Resonance Imaging), PAI (Photo-acoustic imaging), and X-ray contrast imaging, various multifunctional nanoparticles with imaging probes serve as brain-targeted imaging candidates in several imaging modalities. The primary criteria of these functional nanomaterials for translational application to the brain must be zero toxicity. Moreover, the beneficial aspects of nano-theranostics of nanoparticles are their multifunctional systems proportioned towards personalized disease management via comprising diagnostic and therapeutic abilities in a single biodegradable nanomaterial. This review highlights the emerging aspects of engineered nanomaterials to reach and deliver therapeutics to the brain and how to improve this by adopting the imaging modalities for theranostic applications.

Applications and Biological Functions of Exosomes: A Comprehensive Review

Exosomes are also known as extracellular vesicles (EVs) which is bounded by a membrane mostly seen in eukaryotic cells secreted within the endosomal compartment along with some of the selected composition of RNA, proteins, lipids and DNA. They are capable of transferring signals among cells therefore it is used as a mediator for cell-to-cell communication. Exosomes helps in the excretion of cellular waste from the body. Exosomes possess various widespread activity in many of the biological functions such as transferring the biomolecules like enzymes, proteins, ribonucleic acid, lipids and also in the regulation of various pathological and physiological process in various diseases. Exosomes are released in to the in vitro growth medium with the help of cultured cells. They are said to be identified in coined matrix and tissue matrix. They are also identified in some of the biological fluids such as cerebrospinal fluid, urine, blood. Exosomes are considered as promising biomarkers in identification and treatment of many diseases as they contribute a lot in the diagnosis of various therapies. The efficacy and stability of imaging probes and therapeutics are enhanced by its biocompatible nature. Exosomes play a major role because of their use in the field of clinical application. It is important to understand the molecular mechanism behind their function and transport in order to explore more about exosomes. Here we discuss about the review and advancement done in the field of exosomes along with their biomedical applications, isolation techniques and biological functions.

Discovery and directed evolution of nitroreductase enzymes for activation of prodrugs and PET imaging compounds

<p><b>Bacterial nitroreductase enzymes, which exhibit the capacity to reduce a wide range of nitroaromatic drugs, antibiotics and environmental pollutants, have shown promise in the activation of prodrugs such as CB1954 and PR-104A. Use of these prodrugs in gene-directed enzyme prodrug therapy (GDEPT) cancer treatment would allow for targeted chemotherapy in tumour cells following specific delivery of nitroreductases to these cancerous tissues, using specialised bacterial or viral vectors. However, one key limitation in nitroreductase-based GDEPT is the current inability to rapidly and non-invasively determine vector localisation and gene delivery prior to systemic administration of prodrug. Dual-purpose nitroreductases that exhibit the ability to activate both GDEPT prodrugs and radioisotope-labelled PET imaging probes, in a manner that renders them temporarily cell-entrapped for detection using a PET scanner, would facilitate clinical development of this treatment.</b></p> <p>Previous attempts to repurpose hypoxia-activated 2-nitroimidazole PET imaging probes for nitroreductase detection have suffered from relatively high background activation under hypoxia alone. The design of nextgeneration 5-nitroimidazole PET imaging probes, by our collaborators at the Auckland Cancer Society Research Centre (ACSRC), has resulted in much lower levels of hypoxia activation in vivo.</p> <p>This thesis describes attempts to generate improved nitroreductases that can activate a bespoke 5-nitroimidazole PET-capable imaging probe, S33. A 58-membered library of nitroreductase candidates, including enzymes from many different bacterial species and oxidoreductase families, was heterologously over-expressed in E. coli screening strains. Microplate-based screening strategies were then used to identify enzymes that exhibited the most activity with S33, based on the ability of high levels of activated S33 to induce DNA damage and (at very high levels) E. coli cell death. Following this, site-targeted libraries of two different promising nitroreductase NfsA homologues were screened for S33 activity, with selected variants from eachlibrary showing improvement in S33 activation over the parent nitroreductase. In parallel I performed error-prone PCR mutagenesis of a top NfsA variant and top NfsB variant, subjecting each to two rounds of random mutagenesis, and selecting improved variants using a specialised E. coli screening strain and fluorescence-activated cell sorting (FACS). Selected variants from the NfsB (but not NfsA) nitroreductase candidate library showed substantially improved capacity to activate S33 over the parent enzyme.</p> <p>As an alternative means for developing improved nitroreductase variants, two different nitroaromatic ‘anti-prodrugs’, the anthelmintic niclosamide and the antibiotic chloramphenicol, whose cytotoxic effects on E. coli can be mitigated by the presence of an over-expressed active nitroreductase, were used to select for improved S33-activating enzymes from a site-targeted NfsA library. Variants were discovered that exhibited improved ability to active S33 as well as other nitroaromatic substrates of interest. Finally, attempts to discover novel nitroreductases from nature through the screening of cloned soil metagenomic fragments, were made utilising a novel cloning strategy to improve expression of the cloned gene fragments in E. coli. Screening and selection of nitroreductase gene ragments was conducted using niclosamide as well as nitroaromatic compounds that change colour upon activation.</p>

Discovery and directed evolution of nitroreductase enzymes for activation of prodrugs and PET imaging compounds

<p><b>Bacterial nitroreductase enzymes, which exhibit the capacity to reduce a wide range of nitroaromatic drugs, antibiotics and environmental pollutants, have shown promise in the activation of prodrugs such as CB1954 and PR-104A. Use of these prodrugs in gene-directed enzyme prodrug therapy (GDEPT) cancer treatment would allow for targeted chemotherapy in tumour cells following specific delivery of nitroreductases to these cancerous tissues, using specialised bacterial or viral vectors. However, one key limitation in nitroreductase-based GDEPT is the current inability to rapidly and non-invasively determine vector localisation and gene delivery prior to systemic administration of prodrug. Dual-purpose nitroreductases that exhibit the ability to activate both GDEPT prodrugs and radioisotope-labelled PET imaging probes, in a manner that renders them temporarily cell-entrapped for detection using a PET scanner, would facilitate clinical development of this treatment.</b></p> <p>Previous attempts to repurpose hypoxia-activated 2-nitroimidazole PET imaging probes for nitroreductase detection have suffered from relatively high background activation under hypoxia alone. The design of nextgeneration 5-nitroimidazole PET imaging probes, by our collaborators at the Auckland Cancer Society Research Centre (ACSRC), has resulted in much lower levels of hypoxia activation in vivo.</p> <p>This thesis describes attempts to generate improved nitroreductases that can activate a bespoke 5-nitroimidazole PET-capable imaging probe, S33. A 58-membered library of nitroreductase candidates, including enzymes from many different bacterial species and oxidoreductase families, was heterologously over-expressed in E. coli screening strains. Microplate-based screening strategies were then used to identify enzymes that exhibited the most activity with S33, based on the ability of high levels of activated S33 to induce DNA damage and (at very high levels) E. coli cell death. Following this, site-targeted libraries of two different promising nitroreductase NfsA homologues were screened for S33 activity, with selected variants from eachlibrary showing improvement in S33 activation over the parent nitroreductase. In parallel I performed error-prone PCR mutagenesis of a top NfsA variant and top NfsB variant, subjecting each to two rounds of random mutagenesis, and selecting improved variants using a specialised E. coli screening strain and fluorescence-activated cell sorting (FACS). Selected variants from the NfsB (but not NfsA) nitroreductase candidate library showed substantially improved capacity to activate S33 over the parent enzyme.</p> <p>As an alternative means for developing improved nitroreductase variants, two different nitroaromatic ‘anti-prodrugs’, the anthelmintic niclosamide and the antibiotic chloramphenicol, whose cytotoxic effects on E. coli can be mitigated by the presence of an over-expressed active nitroreductase, were used to select for improved S33-activating enzymes from a site-targeted NfsA library. Variants were discovered that exhibited improved ability to active S33 as well as other nitroaromatic substrates of interest. Finally, attempts to discover novel nitroreductases from nature through the screening of cloned soil metagenomic fragments, were made utilising a novel cloning strategy to improve expression of the cloned gene fragments in E. coli. Screening and selection of nitroreductase gene ragments was conducted using niclosamide as well as nitroaromatic compounds that change colour upon activation.</p>