All-photonic kinase inhibitors: light-controlled release-and-report inhibition.

Light-responsive molecular tools targeting kinases affords one the opportunity to study the underlying cellular function of selected kinases. In efforts to externally control lymphocyte-specific protein tyrosine kinase (LCK) activity, the development of release-and-report LCK inhibitors is described, in which (i) the release of the active kinase inhibitor can be controlled externally with light; and (ii) fluorescence is employed to report both the release and binding of the active kinase inhibitor. This introduces an unprecedented all-photonic method for users to both control and monitor real-time inhibitory activity. A functional cellular assay demonstrated light-mediated LCK inhibition in natural killer cells. The use of coumarin-derived caging groups resulted in rapid cellular uptake and non-specific intracellular localisation, while a BODIPY-derived caging group predominately localised in the cellular membrane. This concept of release-and-report inhibitors has the potential to be extended to other biorelevant targets where both spatiotemporal control in a cellular setting and a reporting mechanism would be beneficial.

Photoswitchable optoelectronic properties of 2D MoSe2/diarylethene hybrid structures.

The ability to modulate optical and electrical properties of two-dimensional (2D) semiconductors has sparked considerable interest in transition metal dichalcogenides (TMDs). Herein, we introduce a facile strategy for modulating optoelectronic properties of monolayer MoSe2 with external light. Photochromic diarylethene (DAE) molecules formed a 2-nm-thick uniform layer on MoSe2, switching between its closed- and open-form isomers under UV and visible irradiation, respectively. We have discovered that the closed DAE conformation under UV has its lowest unoccupied molecular orbital energy level lower than the conduction band minimum of MoSe2, which facilitates photoinduced charge separation at the hybrid interface and quenches photoluminescence (PL) from monolayer flakes. In contrast, open isomers under visible light prevent photoexcited electron transfer from MoSe2 to DAE, thus retaining PL emission properties. Alternating UV and visible light repeatedly show a dynamic modulation of optoelectronic signatures of MoSe2. Conductive atomic force microscopy and Kelvin probe force microscopy also reveal an increase in conductivity and work function of MoSe2/DAE with photoswitched closed-form DAE. These results may open new opportunities for designing new phototransistors and other 2D optoelectronic devices.

Sub-picosecond collapse of molecular polaritons to pure molecular transition in plasmonic photoswitch-nanoantennas.

Molecular polaritons are hybrid light-matter states that emerge when a molecular transition strongly interacts with photons in a resonator. At optical frequencies, this interaction unlocks a way to explore and control new chemical phenomena at the nanoscale. Achieving such control at ultrafast timescales, however, is an outstanding challenge, as it requires a deep understanding of the dynamics of the collectively coupled molecular excitation and the light modes. Here, we investigate the dynamics of collective polariton states, realized by coupling molecular photoswitches to optically anisotropic plasmonic nanoantennas. Pump-probe experiments reveal an ultrafast collapse of polaritons to pure molecular transition triggered by femtosecond-pulse excitation at room temperature. Through a synergistic combination of experiments and quantum mechanical modelling, we show that the response of the system is governed by intramolecular dynamics, occurring one order of magnitude faster with respect to the uncoupled excited molecule relaxation to the ground state.

Diarylethene Isomerization by Using Triplet-Triplet Annihilation Photon Upconversion.

Green-to-blue triplet-triplet annihilation photon upconversion with the well-studied upconversion pair 9,10-diphenylanthracene (DPA)/platinum octaethylporphyrin (PtOEP) was used to reversibly drive the photoisomerization of diarylethene (DAE) photoswitches by using visible light. By carefully selecting the kinetic and spectral properties of the molecular system as well as the experimental geometry, a single green light source can be used to selectively trigger both the ring-opening and the ring-closing reactions, whilst also inducing fluorescence from the colored closed isomer that can be used as a readout to monitor the isomerization process in situ. The upconversion solution and the DAE solution are kept physically separated, allowing them to be characterized both concomitantly and individually without further separation processes. The ring-closing reaction using upconverted photons was quantified and compared to the efficiency of direct isomerization with ultraviolet light.

Stimuli-Responsive Membrane Anchor Peptide Nanofoils for Tunable Membrane Association and Lipid Bilayer Fusion.

Self-assembled peptide nanostructures with stimuli-responsive features are promising as functional materials. Despite extensive research efforts, water-soluble supramolecular constructs that can interact with lipid membranes in a controllable way are still challenging to achieve. Here, we have employed a short membrane anchor protein motif (GLFD) and coupled it to a spiropyran photoswitch. Under physiological conditions, these conjugates assemble into ∼3.5 nm thick, foil-like peptide bilayer morphologies. Photoisomerization from the closed spiro (SP) form to the open merocyanine (MC) form of the photoswitch triggers rearrangements within the foils. This results in substantial changes in their membrane-binding properties, which also varies sensitively to lipid composition, ranging from reversible nanofoil reformation to stepwise membrane adsorption. The formed peptide layers in the assembly are also able to attach to various liposomes with different surface charges, enabling the fusion of their lipid bilayers. Here, SP-to-MC conversion can be used both to trigger and to modulate the liposome fusion efficiency.

Optically Switchable NIR Photoluminescence of PbS Semiconducting Nanocrystals using Diarylethene Photoswitches.

Precisely modulated photoluminescence (PL) with external control is highly demanded in material and biological sciences. However, it is challenging to switch the PL on and off in the NIR region with a high modulation contrast. Here, we demonstrate that reversible on and off switching of the PL in the NIR region can be achieved in a bicomponent system comprised of PbS semiconducting nanocrystals (NCs) and diarylethene (DAE) photoswitches. Photoisomerization of DAE to the ring-closed form upon UV light irradiation causes substantial quenching of the NIR PL of PbS NCs due to efficient triplet energy transfer. The NIR PL fully recovers to an on state upon reversing the photoisomerization of DAE to the ring-open form with green light irradiation. Importantly, fully reversible switching occurs without obvious fatigue, and the high PL on/off ratio (>100) outperforms all previously reported assemblies of NCs and photoswitches.

Basic-to-acidic reversible pH switching with a merocyanine photoacid.

The application of merocyanine photoacids has previously been largely limited to neutral and acidic pH values. Here we introduce a new merocyanine photoacid with superior pH switching qualities. By increasing the pKa in the dark (pKdarka) and the solubility we increased the reversible visible light induced pH jump to 3.5 units. Moreover, it is the first demonstration of a merocyanine photoacid able to generate a significant pH drop from a basic (pH 8.3) to an acidic (pH 5.2) environment.

Design and development of photoswitchable DFG-Out RET kinase inhibitors.

REarranged during Transfection (RET) is a transmembrane receptor tyrosine kinase that is required for development of multiple human tissues, but which is also an important contributor to human cancers. RET activation through rearrangement or point mutations occurs in thyroid and lung cancers. Furthermore, activation of wild type RET is an increasingly recognized mechanism promoting tumor growth and dissemination of a much broader group of cancers. RET is therefore an attractive therapeutic target for small-molecule kinase inhibitors. Non-invasive control of RET signaling with light offers the promise of unveiling its complex spatiotemporal dynamics in vivo. In this work, photoswitchable DFG-out RET kinase inhibitors based on heterocycle-derived azobenzenes were developed, enabling photonic control of RET activity. Based on the binding mode of DFG-out kinase inhibitors and using RET kinase as the test model, we developed a photoswitchable inhibitor with a quinoline "head" constituting the azoheteroarene. This azo compound was further modified by three different strategies to increase the difference in biological activity between the E-isomer and the light enriched Z-isomer. Stilbene-based derivatives were used as model compounds to guide in the selection of substituents that could eventually be introduced to the corresponding azo compounds. The most promising quinoline-based compound showed more than a 15-fold difference in bioactivity between the two isomers in a biochemical assay. However, the same compound showed a decreased Z/E (IC50) ratio in the cellular assay, tentatively assigned to stability issues. The corresponding stilbene compound gave a Z/E (IC50) ratio well above 100, consistent with that measured in the biochemical assay. Ultimately, a 7-azaindole based photoswitchable DFG-out kinase inhibitor was shown to display more than a 10-fold difference in bioactivity between the two isomers, in both a biochemical and a cell-based assay, as well as excellent stability even under reducing conditions.

Large, Tunable, and Reversible pH Changes by Merocyanine Photoacids.

Molecular photoswitches capable of generating precise pH changes will allow pH-dependent processes to be controlled remotely and noninvasively with light. We introduce a series of new merocyanine photoswitches, which deliver reversible bulk pH changes up to 3.2 pH units (pH 6.5 to pH 3.3) upon irradiation with 450 nm light, displaying tunable and predictable timescales for thermal recovery. We present models to show that the key parameters for optimizing the bulk pH changes are measurable: the solubility of the photoswitch, the acidity of the merocyanine form, the thermal equilibrium position between the spiropyran and the merocyanine isomers, and the increased acidity under visible light irradiation. Using ultrafast transient absorption spectroscopy, we determined the quantum yields for the ring-closing reaction and found that the lifetimes of the transient cis-merocyanine isomers ranged from 30 to 550 ns. Quantum yields did not appear to be a limitation for bulk pH switching. The models we present use experimentally determined parameters and are, in principle, able to predict the change in pH obtained for any related merocyanine photoacid.

Design and development of a photoswitchable DFG-out kinase inhibitor.

We report the synthesis and characterisation of a photoswitchable DFG-out kinase inhibitor. Photocontrol of the target kinase in both enzymatic and living cell assays is demonstrated.

Rapid amplitude-modulation of a diarylethene photoswitch: en route to contrast-enhanced fluorescence imaging.

A water soluble diarylethene (DAE) derivative that displays exceptionally intense fluorescence from the colorless open form has been synthesized and characterized using UV/vis spectroscopy and fluorescence microscopy. We show that the bright emission from the open form can be rapidly switched using amplitude modulated red light, that is, by light at wavelengths longer than those absorbed by the fluorescent species. This is highly appealing in any context where undesired background fluorescence disturbs the measurement, e.g., the autofluorescence commonly observed in fluorescence microscopy. We show that this scheme is conveniently applicable using lock-in detection, and that robust amplitude modulation of the probe fluorescence is indeed possible also in cell studies using fluorescence microscopy.

Improving Fatigue Resistance of Dihydropyrene by Encapsulation within a Coordination Cage.

Photochromic molecules undergo reversible isomerization upon irradiation with light at different wavelengths, a process that can alter their physical and chemical properties. For instance, dihydropyrene (DHP) is a deep-colored compound that isomerizes to light-brown cyclophanediene (CPD) upon irradiation with visible light. CPD can then isomerize back to DHP upon irradiation with UV light or thermally in the dark. Conversion between DHP and CPD is thought to proceed via a biradical intermediate; bimolecular events involving this unstable intermediate thus result in rapid decomposition and poor cycling performance. Here, we show that the reversible isomerization of DHP can be stabilized upon confinement within a PdII6L4 coordination cage. By protecting this reactive intermediate using the cage, each isomerization reaction proceeds to higher yield, which significantly decreases the fatigue experienced by the system upon repeated photocycling. Although molecular confinement is known to help stabilize reactive species, this effect is not typically employed to protect reactive intermediates and thus improve reaction yields. We envisage that performing reactions under confinement will not only improve the cyclic performance of photochromic molecules, but may also increase the amount of product obtainable from traditionally low-yielding organic reactions.

Toward Two-Photon Absorbing Dyes with Unusually Potentiated Nonlinear Fluorescence Response.

The combination of two two-photon-induced processes in a Förster resonance energy transfer (FRET)-operated photochromic fluorene-dithienylethene dyad lays the foundation for the observation of a quartic dependence of the fluorescence signal on the excitation light intensity. While this photophysical behavior is predicted for a four-photon absorbing dye, the herein proposed approach opens the way to use two-photon absorbing dyes, reaching the same performance. Hence, the spatial resolution limit, being a critical parameter for applications in fluorescence imaging or data storage with common two-photon absorbing dyes, is dramatically improved.

A simplicity-guided cocktail approach toward multicolor fluorescent systems.

A molecular cocktail containing two photochromic diarylethene derivatives that displays multicolor emission spanning blue-green to orange in a color-correlated fashion has been devised. The function does not rely on excited state communication such as energy transfer reactions, which is the typical case for similar systems. Instead, harnessing the intrinsic fluorescent and photochromic properties of the two individual diarylethene derivatives run in parallel is enough to realize the color changes. This offers an extremely flexible situation as for the choice of the fluorophores and their respective concentrations. The function is conveniently demonstrated in bulk solution at µM concentrations, where a single light source serves as the color changing stimulus.

Multi-color emission with orthogonal input triggers from a diarylethene pyrene-OTHO organogelator cocktail.

We report on a pyrene-decorated supramolecular gelator based on an oxotriphenylhexanoate (OTHO) that can switch emission profiles between the solution and gel phase. A cocktail of the gelator and a photochromic diarylethene derivative enables four distinct emissive states to be obtained, which are modulated with light and heat as orthogonal input triggers.

Visible-Light Photoswitching by Azobenzazoles.

Three visible-light responsive photoswitches are reported, azobis(1-methyl-benzimidazole) (1), azobis(benzoxazole) (2) and azobis(benzothiazole) (3). Photostationary distributions are obtained upon irradiation with visible light comprising approximately 80 % of the thermally unstable isomer, with thermal half-lives up to 8 min and are mostly invariant to solvent. On protonation, compound 1H+ has absorption extending beyond 600 nm, allowing switching with yellow light, and a thermal half-life just under 5 minutes. The two isomers have significantly different pKa values, offering potential as a pH switch. The absorption spectra of 2 and 3 are insensitive to acid, although changes in the thermal half-life of 3 indicate more basic intermediates that significantly influence the thermal barrier to isomerization. These findings are supported by high-level ab initio calculations, which validate that protonation occurs on the ring nitrogen and that the Z isomer is more basic in all cases.

An all-photonic full color RGB system based on molecular photoswitches.

On-command changes in the emission color of functional materials is a sought-after property in many contexts. Of particular interest are systems using light as the external trigger to induce the color changes. Here we report on a tri-component cocktail consisting of a fluorescent donor molecule and two photochromic acceptor molecules encapsulated in polymer micelles and we show that the color of the emitted fluorescence can be continuously changed from blue-to-green and from blue-to-red upon selective light-induced isomerization of the photochromic acceptors to the fluorescent forms. Interestingly, isomerization of both acceptors to different degrees allows for the generation of all emission colors within the red-green-blue (RGB) color system. The function relies on orthogonally controlled FRET reactions between the blue emitting donor and the green and red emitting acceptors, respectively.

A Fluorescent Kinase Inhibitor that Exhibits Diagnostic Changes in Emission upon Binding.

The development of a fluorescent LCK inhibitor that exhibits favourable solvatochromic properties upon binding the kinase is described. Fluorescent properties were realised through the inclusion of a prodan-derived fluorophore into the pharmacophore of an ATP-competitive kinase inhibitor. Fluorescence titration experiments demonstrate the solvatochromic properties of the inhibitor, in which dramatic increase in emission intensity and hypsochromic shift in emission maxima are clearly observed upon binding LCK. Microscopy experiments in cellular contexts together with flow cytometry show that the fluorescence intensity of the inhibitor correlates with the LCK concentration. Furthermore, multiphoton microscopy experiments demonstrate both the rapid cellular uptake of the inhibitor and that the two-photon cross section of the inhibitor is amenable for excitation at 700 nm.

Light-driven control of the composition of a supramolecular network.

The composition of a supramolecular network, constituted by several cucurbituril receptors and guests, can be controlled by the reversible and all-photonic switching of a dithienylethene guest.

Diastereoselective Control of Tetraphenylethene Reactivity by Metal Template Self-Assembly.

The reaction of 4,4',4'',4'''-(ethene-1,1,2,2-tetrayl)tetraaniline with 2-pyridinecarboxaldehyde and iron(II) chloride resulted, after aqueous workup, in the diastereoselective formation of an [Fe2 L3 ]4+ triple-stranded helicate structure, irrespective of the stoichiometry employed. The helicate structure was characterized in solution by multinuclear NMR spectroscopy, and in the solid state by single-crystal X-ray crystallography. The reaction of iron(II) tetrafluoroborate or iron(II) bistriflimide with the tetraaniline and 2-pyridinecarboxaldehyde allowed the formation of an [Fe8 L6 ]16+ cube when the appropriate stoichiometry was used, but these structures were unstable with respect to hydrolysis. The pendant amine groups on the helicate can be functionalized by reaction with acid chlorides or anhydrides, and the resulting functionalized tetraphenylethene (TPE) units were isolated by the reaction of the helicate with tris(2-aminoethyl)amine. The emission properties of the TPE units were studied in THF/water mixtures, and they were found by dynamic light scattering to self-assemble into large (av. diameter 250 nm) structures.