Fluorescent Probes for Disease Diagnosis.

The identification and detection of disease-related biomarkers is essential for early clinical diagnosis, evaluating disease progression, and for the development of therapeutics. Possessing the advantages of high sensitivity and selectivity, fluorescent probes have become effective tools for monitoring disease-related active molecules at the cellular level and in vivo. In this review, we describe current fluorescent probes designed for the detection and quantification of key bioactive molecules associated with common diseases, such as organ damage, inflammation, cancers, cardiovascular diseases, and brain disorders. We emphasize the strategies behind the design of fluorescent probes capable of disease biomarker detection and diagnosis and cover some aspects of combined diagnostic/therapeutic strategies based on regulating disease-related molecules. This review concludes with a discussion of the challenges and outlook for fluorescent probes, highlighting future avenues of research that should enable these probes to achieve accurate detection and identification of disease-related biomarkers for biomedical research and clinical applications.

Tracking the cellular uptake and phototoxicity of Ru(ii)-polypyridyl-1,8-naphthalimide Tröger's base conjugates.

Ruthenium(ii) complexes are attracting significant research attention as a promising class of photosensitizers (PSs) in photodynamic therapy (PDT). Having previously reported the synthesis of two novel Ru(ii)-polypyridyl-1,8-naphthalimide Tröger's base compounds 1 and 2 with interesting photophysical properties, where the emission from either the Ru(ii) polypyridyl centres or the naphthalimide moieties could be used to monitor binding to nucleic acids, we sought to use these compounds to investigate further and in more detail their biological profiling, which included unravelling their mechanism of cellular uptake, cellular trafficking and cellular responses to photoexcitation. Here we demonstrate that these compounds undergo rapid time dependent uptake in HeLa cells that involved energy dependent, caveolae and lipid raft-dependent mediated endocytosis, as demonstrated by confocal imaging, and transmission and scanning electron microscopy. Following endocytosis, both compounds were shown to localise to mostly lysosomal and Golgi apparatus compartments with some accumulation in mitochondria but no localisation was found to the nucleus. Upon photoactivation, the compounds increased ROS production and induced ROS-dependent apoptotic cell death. The photo-activated compounds subsequently induced DNA damage and altered tubulin, but not actin structures, which was likely to be an indirect effect of ROS production and induced apoptosis. Furthermore, by changing the concentration of the compounds or the laser used to illuminate the cells, the mechanism of cell death could be changed from apoptosis to necrosis. This is the first detailed biological study of Ru(ii)-polypyridyl Tröger's bases and clearly suggests caveolae-dependent endocytosis is responsible for cell uptake - this may also explain the lack of nuclear uptake for these compounds and similar results observed for other Ru(ii)-polypyridyl complexes. These conjugates are potential candidates for further development as PDT agents and may also be useful in mechanistic studies on cell uptake and trafficking.

Nanoscale assembly of enantiomeric supramolecular gels driven by the nature of solvents.

Understanding the key parameters that control the self-assembly process is critical to predict self-assembly modes in multi-component systems, which will lead to the development of nanofibrous materials with tuneable properties. Enantiomeric amino acid-based low-molecular-weight gelators (LMWGs) were mixed in polar (polar protic) and aromatic apolar (aromatic) solvents and compared to their individual counterparts to probe the effect of solvent polarity on the self-assembly process. Scanning electron microscopy (SEM) reveals that xerogels of individual components display hollow needles in polar protic solvents, while chiral coils are observed in aromatic solvents. In contrast, the multi-component gel displays hollow needle morphologies in both solvents, indicating similar morphologies in polar protic solvents but an entirely different nanostructure for the individual gel networks in aromatic solvents. PXRD experiments performed on the dried gels showed that the nature of the solvents plays a vital role in the co-assembly process of multi-component gels. The self-assembly modes and the gel state structure of the gels are analysed by wide-angle X-ray diffraction (WAXS) and small-angle neutron diffraction (SANS), which reveals that the mixed gel undergoes different co-assembly modes depending on the nature of the solvent systems. This study shows that different co-assembly modes can be achieved for structurally similar components by varying the solvent polarity, demonstrating the importance of solvent choice in the self-assembly process of multi-component gels.

Ligand Chirality Transfer from Solution State to the Crystalline Self-Assemblies in Circularly Polarized Luminescence (CPL) Active Lanthanide Systems.

The synthesis of a family of chiral and enantiomerically pure pyridyl-diamide (pda) ligands that upon complexation with europium [Eu(CF3SO3)3] result in chiral complexes with metal centered luminescence is reported; the sets of enantiomers giving rise to both circular dichroism (CD) and circularly polarized luminescence (CPL) signatures. The solid-state structures of these chiral metallosupramolecular systems are determined using X-ray diffraction showing that the ligand chirality is transferred from solution to the solid state. This optically favorable helical packing arrangement is confirmed by recording the CPL spectra from the crystalline assembly by using steady state and enantioselective differential chiral contrast (EDCC) CPL Laser Scanning Confocal Microscopy (CPL-LSCM) where the two enantiomers can be clearly distinguished.

Correction: Time-resolved infra-red studies of photo-excited porphyrins in the presence of nucleic acids and in HeLa tumour cells: insights into binding site and electron transfer dynamics.

Correction for 'Time-resolved infra-red studies of photo-excited porphyrins in the presence of nucleic acids and in HeLa tumour cells: insights into binding site and electron transfer dynamics' by Páraic M. Keane et al., Phys. Chem. Chem. Phys., 2022, 24, 27524-27531, https://doi.org/10.1039/D2CP04604K.

Synthesis of 3,4-dihydroxy-1,8-naphthalimides with inbuilt catechol and crown ether functionalities.

Herein the synthesis of 1,8-naphthalimides functionalised as the 3,4-dihydroxy-1,8-naphthalimide (catechol, Nap-Cat) and the corresponding 15-crown-5 (Nap-Crown) is reported. These compounds represent the first examples where these two recognition groups are directly incorporated into the 1,8-naphthalimide ring system. Both Nap-Cat and Nap-Crown were evaluated for their capacity to respond to analytes such as H2O2 (a mimic for cellular oxidation) and metal ions (as elements of environmental and physiological interest). While slow oxidation was observed for Nap-Cat upon prolonged exposure to H2O2, no significant changes in photophysical properties were observed upon treatment of Nap-Crown with metal ions.

Formation of lanthanide luminescent di-metallic helicates in solution using a bis-tridentate (1,2,3-triazol-4-yl)-picolinamide (tzpa) ligand.

The chiral bis-tridentate (1,2,3-triazol-4-yl)-picolinamide (tzpa) ligand 1 was used in the formation of lanthanide di- and triple stranded di-metallic helicates in acetonitrile solution, where the changes in the ground and the Tb(III) excited state properties were used to monitor the formation of these supramolecular structures in situ under kinetic control.

Photoinduced electron transfer (PeT) based fluorescent probes for cellular imaging and disease therapy.

Typical PeT-based fluorescent probes are multi-component systems where a fluorophore is connected to a recognition/activating group by an unconjugated linker. PeT-based fluorescent probes are powerful tools for cell imaging and disease diagnosis due to their low fluorescence background and significant fluorescence enhancement towards the target. This review provides research progress towards PeT-based fluorescent probes that target cell polarity, pH and biological species (reactive oxygen species, biothiols, biomacromolecules, etc.) over the last five years. In particular, we emphasise the molecular design strategies, mechanisms, and application of these probes. As such, this review aims to provide guidance and to enable researchers to develop new and improved PeT-based fluorescent probes, as well as promoting the use of PeT-based systems for sensing, imaging, and disease therapy.

Generating water/MeOH-soluble and luminescent polymers by grafting 2,6-bis(1,2,3-triazol-4-yl)pyridine (btp) ligands onto a poly(ethylene-alt-maleic anhydride) polymer and cross-linking with terbium(III).

The synthesis of two new polymers made from P(E-alt-MA) (poly(ethylene-alt-maleic anhydride) and possessing 2,6-bis(1,2,3-triazol-4-yl)pyridine (btp) ligand side chains in 3 and 6 mol%, respectively (P1 and P2, respectively) is described. These polymers were shown to be soluble in MeOH solution and, in the case of P1, also in water, while P2 needed prolonged heating to enable water dissolution. Btp ligands are known for coordinating both d- and f-metal ions and so, herein, we demonstrate by using both UV-Vis absorption, fluorescence emission, as well as time-gated phosphorescence spectroscopies, that both P1 and P2 can bind to Tb(III) ions to give rise to luminescent polymers. From the analysis of the titration data, which demonstrated large changes in the emission intensity properties of the polymer upon Tb(III) binding (ground state changes were also clearly observed, with the absorption being red-shifted at lower energy), we show that the dominant stoichiometry in solution is 1 : 2 (M : L; Tb(III) : btp ratio) which implies that two btp ligands from the polymer background are able to crosslink through lanthanide coordination and that the backbone of the polymer is very likely to aid in coordinating the ions.

Glycosidase activated prodrugs for targeted cancer therapy.

In this review glycosidase activated prodrugs that target cancer cells are discussed. Glycosylated prodrugs undergo enzymatic bioconversion, cleaving the prodrug to release the anticancer drug at the desired site of action, hence minimising the toxic side effects associated with many current anticancer drugs. In addition, the presence of the carbohydrate moiety increases the aqueous solubility of the drugs, allowing for a more effective treatment. In the past decade, significant advancements have been made in this field that have led to the development of many novel carbohydrate-based prodrugs - ranging from simple glycoconjugates to complex self-assemblies and materials, which are discussed in detail herein.

Time-resolved infra-red studies of photo-excited porphyrins in the presence of nucleic acids and in HeLa tumour cells: insights into binding site and electron transfer dynamics.

Cationic porphyrins based on the 5,10,15,20-meso-(tetrakis-4-N-methylpyridyl) core (TMPyP4) have been studied extensively over many years due to their strong interactions with a variety of nucleic acid structures, and their potential use as photodynamic therapeutic agents and telomerase inhibitors. In this paper, the interactions of metal-free TMPyP4 and Pt(II)TMPyP4 with guanine-containing nucleic acids are studied for the first time using time-resolved infrared spectroscopy (TRIR). In D2O solution (where the metal-free form exists as D2TMPyP4) both compounds yielded similar TRIR spectra (between 1450-1750 cm-1) following pulsed laser excitation in their Soret B-absorption bands. Density functional theory calculations reveal that vibrations centred on the methylpyridinium groups are responsible for the dominant feature at ca. 1640 cm-1. TRIR spectra of D2TMPyP4 or PtTMPyP4 in the presence of guanosine 5'-monophosphate (GMP), double-stranded {d(GC)5}2 or {d(CGCAAATTTGCG)}2 contain negative-going signals, 'bleaches', indicative of binding close to guanine. TRIR signals for D2TMPyP4 or PtTMPyP bound to the quadruplex-forming cMYC sequence {d(TAGGGAGGG)}2T indicate that binding occurs on the stacked guanines. For D2TMPyP4 bound to guanine-containing systems, the TRIR signal at ca. 1640 cm-1 decays on the picosecond timescale, consistent with electron transfer from guanine to the singlet excited state of D2TMPyP4, although IR marker bands for the reduced porphyrin/oxidised guanine were not observed. When PtTMPyP is incorporated into HeLa tumour cells, TRIR studies show protein binding with time-dependent ps/ns changes in the amide absorptions demonstrating TRIR's potential for studying light-activated molecular processes not only with nucleic acids in solution but also in biological cells.

Hook, Line, and Sinker! Spectroscopic Studies of Bi-Modular Mono- and Bis-1,8-naphthalimide-Ru(bpy)3-conjugates as DNA "Light Switches".

Bi-chromophoric ruthenium polypyridyl complexes comprising one or two nitro-1,8-naphthalimide groups are shown to be effective DNA binders with off-on light switching properties. The binding to DNA was investigated using a combination of studies such as UV-visible absorption and emission titrations, thermal denaturation, and circular dichroism spectroscopy. The DNA affinity was shown to be sensitive to both the linker length and the number of naphthalimides (one vs two) contained in these systems and binding constants ranging from 106 to 107 M-1 for salmon testes DNA. The strong DNA binding is attributed to the combination of naphthalimide intercalation and the electrostatic interaction of the ruthenium complex. Large emission enhancements from the metal to ligand charge transfer (MLCT) emission arising from the metal complex were observed upon DNA binding, which was attributed to the interruption of intramolecular electron transfer quenching processes. Moving the nitro substitution from the 4-position to the 3-position is found to result in modification of the DNA binding and the resulting optical properties. The off-on light switch phenomena reported demonstrate the potential of these complexes to act as DNA probes.

Hierarchical supramolecular co-assembly formation employing multi-component light-harvesting charge transfer interactions giving rise to long-wavelength emitting luminescent microspheres.

Charge transfer (CT) interaction induced formation of a hierarchical supramolecular assembly has attracted attention due to its wide diversity of structural and functional characteristics. In the present work, we report the generation of green luminescent microspheres from the charge transfer interaction induced co-assembly of a bis-naphthyl dipicolinic amide (DPA) derivative with tetracyanobenzene (TCNB) for the first time. The properties of these self-assemblies were studied both in solution and the solid-state using spectroscopic and a variety of microscopy techniques. The X-ray crystal structure analysis showed a mixed stack arrangement of DPA and TCNB. The molecular orbital and energy level calculations confirm the charge transfer complex formation between DPA and TCNB. Furthermore, energy transfer was observed from the green luminescent CT complex to a red-emitting dye, pyronin Y, in the microsphere matrix, leading to the formation of a light-harvesting tri-component self-assembly.

Design, Synthesis, and Anticancer Studies of a p-Cymene-Ru(II)-Curcumin Organometallic Conjugate Based on a Fluorescent 4-Amino-1,8-naphthalimide Tröger's Base Scaffold.

A unique V-shaped "chiral" supramolecular scaffold, N-(4-pyridyl)-4-amino-1,8-naphthalimide Tröger's base (TBNap), was synthesized in good yield from a precursor N-(4-pyridyl)-4-amino-1,8-naphthalimide (Nap). TBNap was characterized using different spectroscopic methods and the molecular structure was elucidated by diffraction analysis. A new p-cymene-Ru(II)-curcumin conjugate (TB-Ru-Cur) was designed by reacting TBNap dipyridyl donor and ruthenium-curcuminato acceptor [RuCur = (p-cymene)Ru-(curcuminato)Cl] in the presence of silver triflate. TB-Ru-Cur was isolated in quantitative yield and characterized using Fourier transform infrared (FT-IR), NMR (1H, 13C, and 19F), and electrospray ionization mass spectrometry (ESI-MS), and the molecular structure has been predicted using a computational study. Both TBNap and TB-Ru-Cur exhibited intramolecular charge transfer (ICT)-based fluorescence emission. Furthermore, the anticancer properties of TBNap, Ru-Cur, and TB-Ru-Cur were assessed in different cancer cell lines. Gratifyingly, the conjugate TB-Ru-Cur displayed fast-cellular internalization and good cytotoxicity against HeLa, HCT-116, and HepG2 cancer cells and the estimated IC50 value was much lower than that of the precursors (TBNap and Ru-Cur) and the well-known chemotherapeutic drug cisplatin.

Real-Time Multi-Photon Tracking and Bioimaging of Glycosylated Theranostic Prodrugs upon Specific Enzyme Triggered Release.

Real-time tracking of prodrug uptake, delivery and activation in vivo represents a major challenge for prodrug development. Herein, we demonstrate the use of novel glycosylated theranostics of the cancer pharmacophore Amonafide in highly-selective, enzymatic triggered release. We show that the use of endogenous enzymes for activated release of the therapeutic component can be observed, in real time, and monitored using one and two-photon bioimaging, offering unique insight into the prodrug pharmacokinetic profile. Furthermore, we demonstrate that the potent cytotoxicity of Amonafide is preserved using this targeted approach.

Macrocyclic vs. [2]catenane btp structures: influence of (aryl) substitution on the self templation of btp ligands in macrocyclic synthesis.

The synthesis of four 2,6-bis(1,2,3-triazol-4-yl)pyridine (btp) olefin based ligands 3, 4, 11 and 12 is described and their attempted use to form mechanically interlocked molecules using ring closing metatheses (RCM) reactions. The btp ligands were modified in two ways, in 3 and 4 the aryl substitution pattern was changed from 4th position to 3rd position and in the case of 11 and 12, the arms were replaced with aliphatic chains. Our study demonstrates that for all four ligands, the RCM reactions only result in the formation of macrocyclic structures, which in three of the cases, were structurally characterised in both solution (using NMR and HRMS) and in the solid-state using X-ray crystallography. NMR studies were also carried out to investigate if these ligands could preorganise in solution via hydrogen bonding interactions. This study provides a handle of how such precursor substitution can be used to direct the formation of macrocycles or mechanically interlocked structures.

Squaramide-Based Self-Associating Amphiphiles for Anion Recognition.

The synthesis and characterisation of two novel self-assembled amphiphiles (SSAs) SQS-1 and SQS-2 are reported. Both compounds, based on the squaramide motif, were fully soluble in a range of solvents and were shown to undergo self-assembly through a range of physical techniques. Self-assembly was shown to favour the formation of crystalline domains on the nanoscale but also fibrillar film formation, as suggested by SEM analysis. Moreover, both SQS-1 and SQS-2 were capable of anion recognition in DMSO solution as demonstrated using 1 H NMR and UV/Vis absorption spectroscopy, but displayed lower binding affinities for various anions when compared against other squaramide based receptors. In more competitive solvent mixtures SQS-1 gave rise to a colourimetric response in the presence of HPO42- that was clearly visible to the naked eye. We anticipate that the observed response is due to the basic nature of the HPO42- anion when compared against other biologically relevant anions.

Fluorescent 4-amino-1,8-naphthalimide Tröger's bases (TBNaps) possessing (orthogonal) 'α-amino acids', esters and di-peptides and their solvent dependent photophysical properties.

The synthesis of fifteen luminescent bis-naphthalimide based Tröger's bases (TBNaps) derived from 4-amino-1,8-naphthalimide (4-Amino-Nap) precursors is described; these scaffolds possess α-amino acids, esters or di-peptides conjugated at the imide site and show minor fluorescence in aqueous solution while being highly emissive in organic solvents. The investigation shows that these TBNaps possessing ICT excited state properties are capable of generating either positive or negative solvatochromic effects in response to changes in polarity and/or the hydrogen bonding capabilities of the medium.

Mechanically Interlocked Chiral Self-Templated [2]Catenanes from 2,6-Bis(1,2,3-triazol-4-yl)pyridine (btp) Ligands.

We report the efficient self-templated formation of optically active 2,6-bis(1,2,3-triazol-4-yl)pyridine (btp) derived homocircuit [2]catenane enantiomers. This represents the first example of the enantiopure formation of chiral btp homocircuit [2]catenanes from starting materials consisting of a classical chiral element; X-ray diffraction crystallography enabled the structural characterization of the [2]catenane. The self-assembly reaction was monitored closely in solution facilitating the characterization of the pseudo-rotaxane reaction intermediate prior to mechanically interlocking the pre-organised system via ring-closing metathesis.

[Re(CO)3(5-PAN)Cl], a rhenium(I) naphthalimide complex for the visible light photocatalytic reduction of CO2.

A rhenium(i) naphthalimide complex [Re(CO)3(5-PAN)Cl] (Re(5-PAN); 5-PAN = 1-(1,10-phenanthroline)-4-nitro-naphthalimide) was synthesized, characterized, and evaluated as a photocatalyst for CO2 reduction. Characterization included use of MALDI-ToF mass spectrometry, FT-IR, RAMAN, 1H and 13C NMR, elemental analysis, electronic absorption and emission spectroscopy, single crystal X-ray diffraction, DFT and cyclic voltammetry. Photocatalytic (406 nm) reduction of 13CO2 to formate (H13COO) in the presence of this catalyst was tracked via13C NMR. Results support Re5-PAN (φ = 0.021) functioning as a catalyst for the reduction of CO2 (maximum turn-over 48-50 at 300 equiv. triethylamine as the sacrificial electron donor).