Search

A perceived drawback of ruthenium(II) polypyridyl photosensitizers for phototherapeutic applications is their inadequate absorption in the visible region. Ru(II) dyads attached to light‐harvesting organic chromophores are studied to address this issue. Biotin‐appended compounds having one chromophore, namely BODIPY (boron‐dipyrromethene) (1) and Ru(II)‐(tpy)2 (2), or a dyad system containing BODIPY linked to Ru(II)‐(tpy)2 via a diphenylacetylene linker (3) were prepared and studied as photodetection agent and photosensitizes for Photodynamic Therapy (PDT) applications. The bichromophoric 3 with a strong absorption profile (ϵ≈71000 M−1 cm−1 at λmax=503 nm) and high singlet oxygen quantum yield (ΦΔ=0.63 in DMSO) was studied for PDT activity. This complex produced a superoxide anion radical via type‐I and singlet oxygen via type‐II photosensitization processes on light exposure, as evidenced from DNA photo‐cleavage experiments and in vitro DCFDA assay using reactive oxygen species scavengers/quenchers. Dyad 3 displayed an apoptotic photo‐cytotoxic effect against HeLa and H1299 cancer cells with a photocytotoxicity index (PI) value of >625 in HeLa cells with a PDT efficacy much superior to those of its monochromophoric analogs 1 and 2. The intrinsic emission of complex 3, utilized for cellular imaging, showed selectivity towards lysosomes. Finally, the remarkable potential of complex 3 was evidenced using a clinically relevant 3D multicellular tumor spheroid model. [ABSTRACT FROM AUTHOR]

Hydrogen sulfide (H2S) is recognized as an endogenous gaseous signaling agent in many biological activities. Lysosomes are the main metabolic site and play a pivotal role in cells. Herein, we designed and synthesized two new fluorescent probes BDP‐DNBS and BDP‐DNP with a BODIPY core to distinguish H2S. The sensing mechanism is based on the inhibition‐recovery of the photo‐induced electron transfer (PET) process. Through comparing the responsive behaviors of the two probes toward H2S, BDP‐DNBS showed a fast response time (60 s), low limit of detection (LOD, 51 nM), high sensitivity and selectivity. Moreover, the reaction mechanism was demonstrated by mass spectrometry and fluorescence off‐on mechanism was proved by density functional theory (DFT). Significantly, confocal fluorescence imaging indicated that BDP‐DNBS was successfully used to visualize H2S in lysosomes in living HeLa cells. [ABSTRACT FROM AUTHOR]

Platinum(II)‐BODIPY (boron‐dipyrromethene) complexes of 1,2‐diaminocyclohexane (dach), [Pt(dach)(ML)] (MP) and [Pt(dach)(IML)] (IMP), where H2ML is morpholine‐appended O,O‐donor mono‐styryl BODIPY‐appended catecholate and H2IML is di‐iodinated morpholine‐conjugated O,O‐donor mono‐styryl BODIPY linked catecholate, were prepared for photodynamic therapy in near‐IR light (600–720 nm) by targeting lysosomes. Complex MP exhibited a band at 596 nm (ϵ=2.9×104 M−1 cm−1) and IMP at 710 nm (ϵ=2.6×104 M−1 cm−1) within PDT window in dimethyl sulfoxide (10 %)/Dulbecco's Modified Eagle's Medium (DMSO/DMEM, pH 7.2). Complexes MP and IMP showed red emission (λem) at 615 nm (λex=580 nm, ΦF=0.04) and 718 nm (λex=685 nm) (ΦF, fluorescence quantum yield) in DMSO (10 %)‐DMEM (pH 7.2). Emissive complex MP showed lysosome‐specific localization in cellular imaging study. Complex IMP showed photocytotoxicity in near‐IR light of 600–720 nm (IC50: 0.86–1.2 μM) with apoptotic cell death forming singlet oxygen in a type‐II pathway. [ABSTRACT FROM AUTHOR]

ClC‐7 is a chloride‐proton antiporter of the CLC protein family. In complex with its accessory protein Ostm‐1, ClC‐7 localizes to lysosomes and to the osteoclasts' ruffled border, where it plays a critical role in acidifying the resorption lacuna during bone resorption. Gene inactivation in mice causes severe osteopetrosis, neurodegeneration, and lysosomal storage disease. Mutations in the human CLCN7 gene are associated with diverse forms of osteopetrosis. The functional evaluation of ClC‐7 variants might be informative with respect to their pathogenicity, but the cellular localization of the protein hampers this analysis. Here we investigated the functional effects of 13 CLCN7 mutations identified in 13 new patients with severe or mild osteopetrosis and a known ADO2 mutation. We mapped the mutated amino acid residues in the homology model of ClC‐7 protein, assessed the lysosomal colocalization of ClC‐7 mutants and Ostm1 through confocal microscopy, and performed patch‐clamp recordings on plasma‐membrane‐targeted mutant ClC‐7. Finally, we analyzed these results together with the patients' clinical features and suggested a correlation between the lack of ClC‐7/Ostm1 in lysosomes and severe neurodegeneration. © 2020 American Society for Bone and Mineral Research (ASBMR). [ABSTRACT FROM AUTHOR]

Mouse Apolipoprotein L9 is a 34-kDa phosphatidylethanolamine (PE)-binding protein. The gene is present only in mouse and rat genomes; hence it is restricted to two species. To understand why, it is essential to uncover details about its functions in cellular processes. Here we show that ApoL9 interacts with the proteins of the LC3 and GABARAP subfamilies, which are key players in macroautophagy. In vitro binding studies show a strong association with GABARAP, and in amino acid-starved cells it preferentially interacts with lipidated LC3B, likely by binding to its PE moiety through its lipid-binding domain. On treatment with autophagy inhibitors bafilomycin A1 and chloroquine, ApoL9 is found near swollen mitochondria and on lysosomes/LAMP1-positive compartments. However, ApoL9 itself does not seem to be degraded as a result of autophagy, suggesting that it is not an autophagy cargo receptor. Deletions in a putative transmembrane region between amino acids 110 and 145 abolish binding to PE. In addition, ApoL9 can redistribute to stress granules, can homo-oligomerize, and is a microtubule-associated protein. In short, its distribution in the cell is quite widespread, suggesting that it could have functions at the intersection of membrane binding and reorganization, autophagy, cellular stress and intracellular lipid transport. This article has an associated First Person interview with the first author of the paper.

Many invasive cancers emerge through a years-long process of somatic evolution, characterized by an accumulation of heritable genetic and epigenetic changes and the emergence of increasingly aggressive clonal populations. In solid tumors, such as breast ductal carcinoma, the extracellular environment for cells within the nascent tumor is harsh and imposes different types of stress on cells, such as hypoxia, nutrient deprivation, and cytokine inflammation. Acidosis is a constant stressor of most cancer cells due to its production through fermentation of glucose to lactic acid in hypoxic or normoxic regions (Warburg effect). Over a short period of time, acid stress can have a profound effect on the function of lysosomes within the cells exposed to this environment, and after long term exposure, lysosomal function of the cancer cells can become completely dysregulated. Whether this dysregulation is due to an epigenetic change or evolutionary selection has yet to be determined, but understanding the mechanisms behind this dysregulation could identify therapeutic opportunities.

To overcome the drawbacks associated with chemotherapeutic and porphyrin-based photodynamic therapy (PDT) agents, the use of BODIPY (boron-dipyrromethene) scaffold has gained prominence in designing a new generation of photosensitizers-cum-cellular imaging agents. However, their poor cell permeability and limited solubility in aqueous medium inhibits the in-vitro application of their organic form. This necessitates the development of metal-BODIPY conjugates with improved physiological stability and enhanced therapeutic efficacy. We have designed two iron(III)-BODIPY conjugates, [Fe(L1/2)(L3)Cl] derived from benzyl-dipicolylamine and its glycosylated analogue along with a BODIPY-tagged catecholate. The complexes showed intense absorption bands (ε ∼ 55,000 M−1 cm−1) and demonstrated apoptotic PDT activity upon red-light irradiation (30 J/cm2, 600–720 nm). The complex with singlet oxygen quantum yield value of ∼0.34 gave sub-micromolar IC 50 (half-maximal inhibitory concentration) value (∼0.08 μM) in both HeLa and H1299 cancer cells with a photocytotoxicity index value of >1200. Both the complexes were found to have significantly lower cytotoxic effects in non-cancerous HPL1D (human peripheral lung epithelial) cells. Singlet oxygen was determined to be the prime reactive oxygen species (ROS) responsible for cell damage from pUC19 DNA photo-cleavage studies, 1,3-diphenylisobenzofuran and SOSG (Singlet Oxygen Sensor Green) assays. Cellular imaging studies showed excellent fluorescence from complex 2 within 4 h, with localization in lysosomes. Significant drug accumulation into the core of 3D multicellular tumor spheroids was observed within 8 h from intense in-vitro emission. The complexes exemplify iron-based targeted PDT agents and show promising results as potential transition metal-based drugs for ROS mediated red light photocytotoxicity with low dosage requirement. Iron(III) complexes of catecholate-BODIPY (boron-dipyrromethene) and dipicolylamine-glucose ligands showed remarkable PDT (photodynamic therapy) activity against cancer cells in red-light, forming singlet oxygen as the reactive oxygen species from BODIPY and radicals from iron(III)-catecholate metal-based charge transfer band. Lysosome selective apoptotic cell death was observed with negligible toxicity in non-tumorous cells. [Display omitted] • Glycosylated iron(III) BODIPY-catecholates showed apoptotic PDT activity in red light. • Confocal cellular imaging microscopy showed localization of complex 2 in lysosomes. • Glycosylation enhanced cellular uptake and PDT activity of the iron(III) complex 2. • Iron(III) BODIPY complex forms singlet oxygen as the prime reactive oxygen species. • The complexes are highly PDT active in cancer cells but much less toxic in normal cells. [ABSTRACT FROM AUTHOR]

Lysosomal localization of mammalian target of rapamycin complex 1 (mTORC1) is a critical step for activation of the molecule. Rag GTPases are essential for this translocation. Here, we demonstrate that Nudix-type motif 2 (NUDT2) is a novel positive regulator of mTORC1 activation. Activation of mTORC1 is impaired in NUDT2-silenced cells. Mechanistically, NUDT2 binds to Rag GTPase and controls mTORC1 translocation to the lysosomal membrane. Furthermore, NUDT2-dependent mTORC1 regulation is critical for proliferation of breast cancer cells, as NUDT2-silenced cells arrest in G0/G1 phases. Taken together, these results show that NUDT2 is a novel complex formation enhancing factor regulating mTORC1-Rag GTPase signaling that is crucial for cell growth control. [ABSTRACT FROM AUTHOR]

The green fluorescent protein (GFP) chromophore has been widely applied as a biological marker or viscosity sensor. However, modulation of GFP chromophore for photodynamic therapy (PDT) has rarely been reported. Inspired by this, through the unique phenothiazine-anchored strategy, a near-infrared emission photosensitizer Lys-PtzFP was designed and synthesized to achieve this process. The maximum absorption wavelength of Lys-PtzFP was 460 nm and the corresponding molar extinction coefficient was 6.60 × 104 M−1 cm−1 in MeOH. The maximum emission wavelengths of Lys-PtzFP was at 725 nm. The photosensitizer Lys-PtzFP showed a high singlet oxygen quantum yield (Φ Δ = 0.42) and low fluorescence quantum yield (φ F = 0.002) in MeOH. Besides, Lys-PtzFP possessed high specificity for lysosomes with negligible dark cytotoxicity (MTT assay >94.6%), good photostability and low half-maximal inhibitory concentration (IC 50) of 0.93 μM. Finally, the intracellular photodynamic therapy experiments revealed that Lys-PtzFP can produce singlet oxygen in lysosomes and induce apoptosis under irradiation. In conclusion, Lys-PtzFP could be a promising photosensitizer for PDT and two-photon imaging applications. This type of photosensitizer can inspire the development of new photosensitizers for further explore highly efficient PDT. • Heavy atom-free photosensitizer, Φ Δ = 0.42, IC 50 = 0.93 μM. • Phenothiazine-GFP chromophore with near-Infrared emission of 725 nm. • Two-photon fluorescent imaging. [ABSTRACT FROM AUTHOR]

Many invasive cancers emerge through a years-long process of somatic evolution, characterized by an accumulation of heritable genetic and epigenetic changes and the emergence of increasingly aggressive clonal populations. In solid tumors, such as breast ductal carcinoma, the extracellular environment for cells within the nascent tumor is harsh and imposes different types of stress on cells, such as hypoxia, nutrient deprivation, and cytokine inflammation. Acidosis is a constant stressor of most cancer cells due to its production through fermentation of glucose to lactic acid in hypoxic or normoxic regions (Warburg effect). Over a short period of time, acid stress can have a profound effect on the function of lysosomes within the cells exposed to this environment, and after long term exposure, lysosomal function of the cancer cells can become completely dysregulated. Whether this dysregulation is due to an epigenetic change or evolutionary selection has yet to be determined, but understanding the mechanisms behind this dysregulation could identify therapeutic opportunities. [ABSTRACT FROM AUTHOR]

Mouse Apolipoprotein L9 is a 34-kDa phosphatidylethanolamine (PE)-binding protein. The gene is present only in mouse and rat genomes; hence it is restricted to two species. To understand why, it is essential to uncover details about its functions in cellular processes. Here we show that ApoL9 interacts with the proteins of the LC3 and GABARAP subfamilies, which are key players in macroautophagy. In vitro binding studies show a strong association with GABARAP, and in amino acid-starved cells it preferentially interacts with lipidated LC3B, likely by binding to its PE moiety through its lipid-binding domain. On treatment with autophagy inhibitors bafilomycin A1 and chloroquine, ApoL9 is found near swollen mitochondria and on lysosomes/LAMP1-positive compartments. However, ApoL9 itself does not seem to be degraded as a result of autophagy, suggesting that it is not an autophagy cargo receptor. Deletions in a putative transmembrane region between amino acids 110 and 145 abolish binding to PE. In addition, ApoL9 can redistribute to stress granules, can homo-oligomerize, and is a microtubule-associated protein. In short, its distribution in the cell is quite widespread, suggesting that it could have functions at the intersection of membrane binding and reorganization, autophagy, cellular stress and intracellular lipid transport.This article has an associated First Person interview with the first author of the paper., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2019. Published by The Company of Biologists Ltd.)

We disclose herein the first example of stable monodispersed hybrid nanoparticles (termed MelaSil-NPs) made up of eumelanin biopolymer intimately integrated into a silica nanoscaffold matrix and endowed with high antioxidant and cytoprotective effects associated with a specific subcellular localization. MelaSil-NPs have been fabricated by an optimized sol-gel methodology involving ammonia-induced oxidative polymerization of a covalent conjugate of the eumelanin building block 5,6-dihydroxyindole-2-carboxylic acid (DHICA) with 3-aminopropyltriethoxysilanes (APTS). They displayed a round-shaped (ca. 50-80 nm) morphology, exhibited the typical electron paramagnetic resonance signal of eumelanin biopolymers, and proved effective in promoting decomposition of hydrogen peroxide under physiologically relevant conditions. When administered to human ovarian cancer cells (A2780) or cervical cancer cells (HeLa), MelaSil-NPs were rapidly internalized and colocalized with lysosomes and exerted efficient protecting effects against hydrogen peroxide-induced oxidative stress and cytotoxicity.