Photo-physical, theoretical and photo-cytotoxic evaluation of a new class of lanthanide(iii)-curcumin/diketone complexes for PDT application.

Herein we report the synthesis, characterization, photophysical and photocytotoxicity studies of a new class of curcumin-based lanthanide(iii) complexes of general molecular formula [La(1,10-phen)2(L)(NO3)2] (1-4), where L = 1-phenylbutane-1,3-dione (L1, 1), 1-(anthracen-9-yl)butane-1,3-dione (L2, 2), 1-(3a1,5a1-dihydropyren-1-yl)butane-1,3-dione (L3, 3) and curcumin (L4, 4). Complex 1 was characterized by single-crystal X-ray crystallography and it exhibited the N4O6 coordination of La(iii). The presence of the low-lying and long-lived triplet excited state enabled the luminescent complexes (2-4) to generate singlet oxygen (1O2) in high yield when the complex was activated with visible light (400-700 nm, 10 J cm-2), which could be responsible for the photo-ablation of cancer cells. Complexes (2-4) exhibited remarkable photocytotoxicity in HeLa and MCF-7 cells with photocytotoxicity index 4-50 in the presence of visible light (400-700 nm, 10 J cm-2), while they were non-toxic in the dark with an IC50 value of >100 µM. The significantly lower toxicity (IC50 > 100 µM in the dark; IC50 in visible light ∼60 µM) of the complexes in MCF-10A (normal cells) in the dark and in visible light suggested their potential for targeting anticancer activity. Further studies showed that complex 4 induced caspase-dependent apoptosis through mitochondrial damage, mitochondrial respiration inhibition and reactive oxygen species (ROS) elevation. The cytosolic localization of complex 4 in HeLa cells, having a curcumin moiety as a fluorophore, was proved from the confocal microscopic studies. The photocytotoxicity of the complexes (1-4) was directly correlated to the efficacy of the complexes to generate singlet oxygen, which resulted in the photocytotoxicity order of 4 > 3>2 ≫ 1. Photo-physical studies revealed that the chelation of curcumin by La(iii) facilitated intersystem crossing in curcumin by reducing the energy gap of the singlet to triplet excited state. Therefore, the presence of low-lying and long-lived triplet excited state was responsible for increasing the generation of singlet oxygen and, thereby, photo-cytotoxicity in HeLa and MCF-7 cells. The present study has given an overall (Chemistry to Biology) perspective on the effect of La(iii) on the photo-cytotoxicity of selected photo-active curcumin-based ß-diketonate ligands.

Activatable Photodynamic Therapy with Therapeutic Effect Prediction Based on a Self-correction Upconversion Nanoprobe.

Though emerging as a promising therapeutic approach for cancers, the crucial challenge for photodynamic therapy (PDT) is activatable phototoxicity for selective cancer cell destruction with low "off-target" damage and simultaneous therapeutic effect prediction. Here, we design an upconversion nanoprobe for intracellular cathepsin B (CaB)-responsive PDT with in situ self-corrected therapeutic effect prediction. The upconversion nanoprobe is composed of multishelled upconversion nanoparticles (UCNPs) NaYF4:Gd@NaYF4:Er,Yb@NaYF4:Nd,Yb, which covalently modified with an antenna molecule 800CW for UCNPs luminance enhancement under NIR irradiation, photosensitizer Rose Bengal (RB) for PDT, Cy3 for therapeutic effect prediction, and CaB substrate peptide labeled with a QSY7 quencher. The energy of UCNPs emission at 540 nm is transferred to Cy3/RB and eventually quenched by QSY7 via two continuous luminance resonance energy transfer processes from interior UCNPs to its surface-extended QSY7. The intracellular CaB specifically cleaves peptide to release QSY7, which correspondingly activates RB with reactive oxygen species (ROS) generation for PDT and recovers Cy3 luminance for CaB imaging. UCNPs emission at 540 nm remains unchanged during the peptide cleavage process, which is served as an internal standard for Cy3 luminance correction, and the fluorescence intensity ratio of Cy3 over UCNPs (FI583/FI540) is measured for self-corrected therapeutic effect prediction. The proposed self-corrected upconversion nanoprobe implies significant potential in precise tumor therapy.

BMP-2-Loaded HAp:Ln3+ (Ln = Yb, Er, Gd) Nanorods with Dual-Mode Imaging for Efficient MC3t3-E1 Cell Differentiation Regulation.

Effective bone tissue reconstitution improves the treatment success rate of dental implantation and preserves natural teeth during periodontal tissue repair. Hydroxyapatite (HAp) has received much attention in bone remodeling field because its mineralized structure is similar to that of the natural bone tissue. For this reason, it has been used as a carrier for growth factors. Although HAp possesses outstanding biomedical properties, its capacity of loading and releasing bone growth factors and promoting osteogenesis is not well understood. In this study, Ln3+ (Ln = Yb3+, Er3+, Gd3+)-doped HAp (HAp:Ln3+) nanorods were synthesized by one-step hydrothermal method. To improve its biocompatibility and surface properties, bone morphogenetic protein-2 (BMP-2) was loaded onto the surface of HAp:Ln3+ nanorods. The results showed that BMP-2 incorporation promoted bone formation and enhanced the expression of early bone-related gene and protein (RunX2, SP7, OPN). In addition, Yb3+- and Er3+-doped HAp nanorods were examined by upconversion luminescence with 980 nm near-infrared laser irradiation to monitor the delivery position of BMP-2 protein. Furthermore, due to the positive magnetism correlated with the concentration of Gd3+, HAp:Ln3+ with enhanced contrast brightening can be deemed as T1 MIR contrast agents. These findings indicate that HAp doped with rare-earth ions and loaded with BMP-2 has the potential to promote bone tissue repair and execute dual-mode imaging.

Potential Application of Upconverting Nanoparticles for Brain Photobiomodulation.

Brain photobiomodulation (PBM) describes the use of visible to near-infrared light for modulation or stimulation of the central nervous system in both healthy individuals and diseased conditions. Although the transcranial approach to delivering light to the head is the most common technique to stimulate the brain, delivery of light to deeper structures in the brain is still a challenge. The science of nanoparticle engineering in combination with biophotonic excitation could provide a way to overcome this problem. Upconversion is an anti-Stokes process that is capable of transforming low energy photons that penetrate tissue well to higher energy photons with a greater biological effect, but poor tissue penetration. Wavelengths in the third optical window are optimal for light penetration into brain tissue, followed by windows II, IV, and I. The combination of trivalent lanthanide ions within a crystalline host provides a nanostructure that exhibits the upconversion phenomenon. Upconverting nanoparticles (UCNPs) have been successfully used in various medical fields. Their ability to cross the brain-blood barrier and their low toxicity make them a good candidate for application in brain disorders. It is possible that delivery of UCNPs to the brainstem or deeper parts of the cerebral tissue, followed by irradiation using light wavelengths with good tissue penetration properties, could allow more efficient PBM of the brain.

Photoluminescent organisms: how to make fungi glow through biointegration with lanthanide metal-organic frameworks.

We show that filamentous fungi can emit green or red light after the accumulation of particulate lanthanide metal-organic frameworks over the cell wall. These new biohybrids present photoluminescence properties that are unaffected by the components of the cell wall. In addition, the fungal cells internalise lanthanide metal-organic framework particles, storing them into organelles, thereby making these materials promising for applications in living imaging studies.

A critical comparison of lanthanide based upconversion nanoparticles to fluorescent proteins, semiconductor quantum dots, and carbon dots for use in optical sensing and imaging.

The right choice of a fluorescent probe is essential for successful luminescence imaging and sensing and especially concerning in vivo and in vitro applications, the development of new classes have gained more and more attention in the last years. One of the most promising class are upconversion nanoparticles (UCNPs)-inorganic nanocrystals capable to convert near-infrared light in high energy radiation. In this review we will compare UCNPs with other fluorescent probes in terms of (a) the optical properties of the probes, such as their brightness, photostability and excitation wavelength; (b) their chemical properties such as the dispersibility, stability under experimental or physiological conditions, availability of chemical modification strategies for labelling; and (c) the potential toxicity and biocompatibility of the probe. Thereby we want to provide a better understanding of the advantages and drawbacks of UCNPs and address future challenges in the design of the nanocrystals.

Non-Invasive Optical Guided Tumor Metastasis/Vessel Imaging by Using Lanthanide Nanoprobe with Enhanced Down-Shifting Emission beyond 1500 nm.

Visualization of tumor vessels/metastasis and cerebrovascular architecture is vitally important for analyzing pathological states of brain diseases and a tumor's abnormal blood vessels to improve cancer diagnoses. In vivo fluorescence imaging using second near-infrared emission beyond 1500 nm (NIR-IIb) has emerged as a next generation optical imaging method with significant improvement in imaging sensitivity and spatial resolution. Unfortunately, a highly biocompatible probe capable of generating NIR-IIb emission with sufficient brightness and uniformed size is still scarce. Here, we have proposed the poly(acrylic acid) (PAA)-modified NaLnF4:40Gd/20Yb/2Er nanorods (Ln = Y, Yb, Lu, PAA-Ln-NRs) with enhanced downshifting NIR-IIb emission, high quantum yield (QY), relatively narrow bandwidth (∼160 nm), and high biocompatibility via Ce3+ doping for high performance NIR-IIb bioimaging. The downshifting emission beyond 1500 nm is improved by 1.75-2.2 times with simultaneously suppressing the upconversion (UC) path in Y, Yb, and Lu hosts via Ce3+ doping. Moreover, compared with the traditionally used Y-based host, the QY of NIR-IIb emission in the Lu-based probe in water is improved from 2.2% to 3.6%. The explored bright NIR-IIb emitted PAA-Lu-NRs were used for high sensitivity small tumor (∼4 mm)/metastatic tiny tumor detection (∼3 mm), tumor vessel visualization with high spatial resolution (41 µm), and brain vessel imaging. Therefore, our findings open up the opportunity of utilizing the lanthanide based NIR-IIb probe with bright 1525 nm emission for in vivo optical-guided tumor vessel/metastasis and noninvasive brain vascular imaging.

Time-resolved luminescent biosensing based on inorganic lanthanide-doped nanoprobes.

Time-resolved (TR) photoluminescence (PL) biosensing has been widely adopted in many research and medical institutions. However, commercial molecular TRPL bioprobes like lanthanide (Ln(3+))-chelates suffer from poor photochemical stability and long-term toxicity. Inorganic Ln(3+)-doped nanocrystals (NCs), owing to their superior physicochemical properties over Ln(3+)-chelates, are regarded as a new generation of luminescent nanoprobes for TRPL biosensing. The long-lived PL of Ln(3+)-doped NCs combined with the TRPL technique is able to completely suppress the interference of the short-lived background, resulting in a background-free signal and therefore a remarkable sensitivity for biosensing. In this feature article, we summarize the latest advancements in inorganic Ln(3+)-doped NCs as TRPL nano-bioprobes from their fundamental optical properties to their potential applications for ultrasensitive biodetection and high-resolution bioimaging. Future efforts towards the commercialization of these nanoprobes are also proposed.

Effect of lanthanide ions on dynamic nuclear polarization enhancement and liquid-state T1 relaxation.

In the dynamic nuclear polarization process, microwave irradiation facilitates exchange of polarization from a radical's unpaired electron to nuclear spins at cryogenic temperatures, increasing polarization by >10,000. Doping samples with Gd(3+) ions further increases the achievable solid-state polarization. However, on dissolution, paramagnetic lanthanide metals can be potent relaxation agents, decreasing liquid-state polarization. Here, the effects of lanthanide metals on the solid and liquid-state magnetic properties of [1-(13)C]pyruvate are studied. The results show that in addition to gadolinium, holmium increases not only the achievable polarization but also the rate of polarization. Liquid-state relaxation studies found that unlike gadolinium, holmium minimally affects T(1). Additionally, results reveal that linear contrast agents dissociate in pyruvic acid, greatly reducing liquid-state T(1). Although macrocyclic agents do not readily dissociate, they yield lower solid-state polarization. Results indicate that polarization with free lanthanides and subsequent chelation during dissolution produces the highest polarization enhancement while minimizing liquid-state relaxation.

Lanthanide-binding tags with unnatural amino acids: sensitizing Tb3+ and Eu3+ luminescence at longer wavelengths.

Lanthanide-binding tags (LBTs) are small, genetically encoded, versatile protein fusion partners that selectively bind lanthanide ions with high affinity. The LBT motif features a strategically positioned tryptophan residue that sensitizes Tb3+ luminescence upon excitation at 280 nm. Herein, we describe the preparation of new LBT peptides that incorporate unnatural amino acids in place of tryptophan, and which sensitize both Tb3+ and Eu3+ luminescence at lower energies. We also report the semisynthesis of proteins tagged with these new LBTs using native chemical ligation. This expands the scope of LBTs and will enable their wider use in luminescence applications.

Strategy for photostable proximity bioassays using lanthanides.

We report initial findings for research aimed at creating photostable lanthanide chelate reporters for proximity assays. These reporters take advantage of the nanometer-scale distance dependence of fluorescence enhancement for molecules in the vicinity of noble metal nanoparticles and also capitalize on some unique properties of lanthanide chelates. This approach promises to lead to proximity assays that do not suffer from photobleaching and offer very high on/off enhancement ratios. Results for lanthanide chelates on silver island films and in colloidal suspensions are reported. Enhancement factors range from 1 to 2 orders of magnitude, with larger enhancements for strongly quenched lanthanides.

Wavelength dependent photofragmentation patterns of tris(2,2,6,6-tetramethyl-3,5-heptanedionato)Ln (III) (Ln = Eu, Tb, Gd) in a molecular beam.

Laser photoionization and ligand photodissociation in Ln(thd)(3) (Ln = Eu, Tb, Gd; thd = 2,2,6,6-tetramethyl-3,5-heptanedionato) are studied in a molecular beam via time-of-flight mass spectrometry. The fragmentation patterns are strongly wavelength dependent. With 355 nm excitation, the mass spectrum is dominated by Ln(2+), Ln(+), and LnO(+) fragments. The bare Ln ions are believed to arise from photoionization of neutral Ln atoms. The Ln atoms, in turn, are produced from the Ln(thd)(3) complex in a sequence of Ln reductions (through ligand-to-metal charge-transfer transitions), with each reduction being accompanied by the dissociation of a neutral ligand radical. In contrast, under visible-light (410-450 nm) excitation, a significant Ln(thd)(n)(+) signal is observed (where n = 2,3 for Ln = Tb,Gd and n = 1-3 for Ln = Eu). Thus, with visible excitation, photoionization of Ln(thd)(n) competes effectively with the Ln-reduction/ligand-dissociation sequence that leads to the dominant bare Ln-ion signal seen with 355 nm excitation. The fact that monoligated Ln(thd)(+) is observed only for Ln = Eu is interpreted in terms of the relative accessibility of an excited ligand-to-metal charge-transfer state from the ground electronic state of neutral Ln(thd).

Upconversion from aqueous phase lanthanide chelates.

We have prepared and characterized several lanthanide ion complexes of multidentate ligands or chelates in an effort to develop new upconverting luminescent labels that can be immune to autofluorescence and photobleaching. This study has involved the characterization of various chelates of Nd, Er, and Tm with respect to relative luminescent efficiency and excited-state lifetimes and explored various two-photon stepwise excitation mechanisms. Using peak laser powers near 100 kW, the upconversion emissions of Nd in Nd(EDTA)2(5-) at 386 nm, Er in Er(DPA)3(3-) at 550 nm, and Tm in Tm(DPA)3(3-) at 480 nm, at levels of approximately 10(-12) moles can be detected.

Long-lived near-infrared luminescent lanthanide complexes of imidodiphosphinate "shell" ligands.

Near-infrared emitting complexes of Nd(III), Er(III), and Yb(III) based on hexacoordinate lanthanide ions with an aryl functionalized imidodiphosphinate ligand, tpip, have been synthesized and fully characterized. Three tpip ligands form a shell around the lanthanide with the ligand coordinating via the two oxygens leading to neutral complexes, Ln(tpip)3. In the X-ray crystal structures of Er(III) and Nd(III) complexes there is evidence of CH-pi interactions between the phenyl groups. Photophysical investigations of solution samples of the complexes demonstrate that all complexes exhibit relatively long luminescence lifetimes in nondeuteurated solvents. Luminescence studies of powder samples have also been recorded for examination of the properties of NIR complexes in the solid state for potential material applications. The results underline the effective shielding of the lanthanide by the twelve phenyl groups of the tpip ligands and the reduction of high-energy vibrations in close proximity to the lanthanide, both features important in the design of NIR emitting lanthanide complexes.

Sensitization of lanthanides by nonnatural amino acids.

The sensitization of Eu(III) and Tb(III) by ethylenediaminetetraaceticacid (EDTA)-derivatized tryptophan (Trp), 7-azatryptophan (7AW) and 5-hydroxytryptophan (5HW) has been examined. These Trp analogs were utilized in the present study because they can be incorporated into proteins in place of native Trp residues and because they absorb strongly beyond 305 nm (where Trp absorbance goes to zero), allowing selective excitation of such species in the presence of other Trp-containing proteins. All three indole derivatives were able to sensitize Tb(III) luminescence, with the relative sensitization being in the order Trp > 5HW > 7AW. On the other hand, only the 7AW-EDTA complex was able to sensitize Eu(III) luminescence, likely owing to a better spectral overlap between 7AW emission and Eu(III) absorbance. The sensitized emission of Tb(III) and Eu(II) displayed the expected long emission lifetimes at 545 nm [for Tb(III)] and 617 nm [for Eu(III)], indicating that long-lifetime lanthanide emission could be produced using nonnatural amino-acid donors. Thus, 7AW- and 5HW-sensitized lanthanide emissions should prove to be useful in biophysical studies, such as the use of fluorescence energy transfer to probe biomolecular interactions in vivo.