ABSTRACT
Boron-dipyrromethene (BODIPY) based photosensitizers as porphyrinoids and curcumin as natural product possess exciting photophysical features suitable for theranostic applications, namely, imaging and photodynamic therapy (PDT). Limited aqueous solubility and insufficient physiological stability, however, reduce their efficacy significantly. We have designed a novel strategy to deliver these two unusable cytotoxins simultaneously in cancer cells and herein, report the synthesis, characterization and imaging-assisted photocytotoxicity of three zinc(II) complexes containing N3-donor dipicolylamine (dpa) ligands (L1-3) and O,O-donor curcumin (Hcur) viz. [Zn(L1)(cur)]Cl (1), [Zn(L2)(cur)]Cl (2) and [Zn(L3)(cur)]Cl (3), where L2 and L3 have pendant fluorescent BODIPY and non-emissive di-iodo-BODIPY moieties. Metal chelation imparted remarkable biological stability (pH â¼7.4) to the respective ligands and induces significant aqueous solubility. These ternary complexes could act as replacements of the existing metalloporphyrin-based PDT photosensitizers as their visible-light photosensitizing ability is reinforced by the dual presence of blue light absorbing curcumin and green light harvesting BODIPY units. Complex 2 having emissive BODIPY unit L2 and curcumin, showed mitochondria selective localization in HeLa, MCF-7 cancer cells and complex 3, the di-iodinated analogue of complex 2, exhibited type-I/II PDT activity via inducing apoptosis through mitochondrial membrane disruption in cancer cells while being significantly nontoxic in dark and to the healthy cells.
Subject(s)
Antineoplastic Agents/pharmacology , Coordination Complexes/pharmacology , Light , Optical Imaging , Photochemotherapy , Photosensitizing Agents/pharmacology , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/chemistry , Boron Compounds/chemistry , Boron Compounds/pharmacology , Cell Line , Cell Proliferation/drug effects , Coordination Complexes/chemical synthesis , Coordination Complexes/chemistry , Curcumin/chemistry , Curcumin/pharmacology , Dose-Response Relationship, Drug , Humans , Molecular Structure , Photosensitizing Agents/chemical synthesis , Photosensitizing Agents/chemistry , Structure-Activity Relationship , Zinc/chemistry , Zinc/pharmacologyABSTRACT
Dipicolylamine (dpa) based cis-dichlorido zinc(ii) complexes [Zn(L1-3)Cl2] (1-3), where L2 and L3 are non-iodo and di-iodo BODIPY-appended dpa in 2 and 3, and L1 is dpa in control complex 1, were prepared and characterized and their photocytotoxicity was studied. Complexes 2 and 3 were developed as potential substitutes for zinc(ii)-porphyrins/phthalocyanines that are photodynamic therapeutic agents with moderate activity owing to their inherent hydrophobicity and aggregation-induced deactivation mechanism. In our approach, we strategically designed hybrid inorganic-organic zinc-BODIPY conjugates as theranostic photosensitizers. The structurally characterized diamagnetic Zn(ii) cis-dichlorido complexes mimic cisplatin and serve as new-generation photosensitizers with enhanced aqueous solubility and mito-DNA targeting propensity while imparting significant physiological stability to the heavy atom tethered BODIPY ligand, L3. The BODIPY complexes showed a visible band near 500 nm (ε⼠34 000-44 000 dm3 mol-1 cm-1) and an emission band at 507 nm for 2 in 1% DMSO-Dulbecco's phosphate buffered saline. The labile chlorido ligands (ΛMâ¼ 200 S m2 mol-1 in 9 : 1 H2O-DMSO) generated positively charged complexes inside the cellular medium enabling them to cross the mitochondrial membrane for this organelle-selective localization and singlet oxygen-mediated apoptotic photocytotoxicity at nanomolar concentrations for 3 in HeLa and MCF-7 cells in light (400-700 nm), while being less active in the dark.
Subject(s)
Boron Compounds , Fluorescent Dyes , Photosensitizing Agents , Zinc , Boron Compounds/chemistry , Boron Compounds/pharmacology , Cell Survival/drug effects , Cell Survival/radiation effects , DNA, Mitochondrial/metabolism , Fluorescent Dyes/chemistry , Fluorescent Dyes/pharmacology , HeLa Cells , Humans , Light , MCF-7 Cells , Microscopy, Confocal , Mitochondria/metabolism , Photosensitizing Agents/chemistry , Photosensitizing Agents/pharmacology , Precision Medicine , Singlet Oxygen/metabolism , Solubility , Zinc/chemistry , Zinc/pharmacologyABSTRACT
Tumor necrosis factor-α (TNFα), one of the major proinflammatory cytokines, plays a key role in an effective immune response. However, the chronic presence of TNFα can lead to several inflammatory disorders, such as rheumatoid arthritis, psoriasis, Crohn's disease, etc. Inhibition of TNFα by pharmacological inhibitors or antibodies has proven to be effective in palliative treatment to some extent. The aim of this study was to develop an anti-TNFα antibody, which may be used as a therapeutic option to inhibit TNFα-mediated cytotoxicity. We characterized several hybridoma clones secreting monoclonal antibodies (mAbs) to human-TNFα. Four mAbs rescued L929 fibroblast cells from TNFα-triggered cell death and one of these, namely C8, was found to have the highest affinity. To gain insights into the mechanism by which mAb C8 inhibits human TNFα-mediated toxicity, the epitope corresponding to the mAb was delineated. The antigenic determinant was found to comprise of the stretch of amino acids 99-120, of which, 102-104 (glutamine, arginine, glutamic acid) form the core epitope. The observation was supported by bioinformatics analyses of an antigen/antibody complex model. In addition, the binding affinity of mAb C8 to TNFα was found to be comparable with that of infliximab, which is a commercially available anti-TNFα mAb.
Subject(s)
Antibodies, Monoclonal, Humanized/immunology , Fibroblasts/immunology , Hybridomas/immunology , Immunoglobulin G/immunology , Recombinant Proteins/immunology , Tumor Necrosis Factor-alpha/antagonists & inhibitors , Animals , Antibodies, Monoclonal, Humanized/biosynthesis , Antibodies, Monoclonal, Humanized/pharmacology , Antibody Formation , Cells, Cultured , Female , Fibroblasts/cytology , Humans , Mice , Mice, Inbred BALB CABSTRACT
Copper(ii) acetylacetonates of N,N,N-donor dipicolylamine (dpa) ligands, viz. [Cu(L1)(acac)]ClO4 (1), [Cu(L2)(acac)]ClO4 (2) and [Cu(L3)(acac)]ClO4 (3), where L1 is benzyldipicolylamine (bzdpa), L2 and L3 are non-iodinated and diiodinated BODIPY (borondipyrromethene) ligands and Hacac is acetylacetone, were synthesized and characterized and their photocytotoxicity was studied. The BODIPY complex 2, structurally characterized by X-ray crystallography, has copper(ii) in a distorted square-pyramidal geometry (degree of trigonality, τ5 = 0.28). The one-electron paramagnetic and redox active copper(ii) complexes displayed 1 : 1 electrolytic behaviour in polar organic solvents. The BODIPY complexes 2 and 3 showed respective visible bands at 498 and 539 nm in 5% DMSO-phosphate buffered saline (PBS). Complex 2 displayed an emission band at 511 nm in 5% DMSO-PBS (λex = 465 nm) with a fluorescence quantum yield (ΦF) value of 0.15. Cellular imaging using this complex showed significant mitochondrial localization in HeLa and MCF-7 cancer cells. Complex 3 with a diiodo-BODIPY moiety was non-emissive (ΦF = 0.01) but acted as an efficient photosensitizer with a singlet oxygen quantum yield value of 0.59 (ΦΔ). Complex 3 showed a remarkable PDT effect with apoptotic cell death due to singlet oxygen giving IC50 values within 0.04-0.06 µM in HeLa and MCF-7 cells using visible light (400-700 nm), while being less toxic in the dark.
ABSTRACT
Poly (ADP-ribose) polymerase 1 (PARP1) inhibitors are actively under clinical trials for the treatment of breast and ovarian cancers that arise due to mutations in BRCA1 and BRCA2. The RAD51 paralog RAD51C has been identified as a breast and ovarian cancer susceptibility gene. The pathological RAD51C mutants that were identified in cancer patients are hypomorphic with partial repair function. However, targeting cancer cells that express hypomorphic mutants of RAD51C is highly challenging. Here, we report that RAD51C-deficient cells can be targeted by a 'synthetic lethal' approach using PARP inhibitor and this sensitivity was attributed to accumulation of cells in the G2/M and chromosomal aberrations. In addition, spontaneous hyperactivation of PARP1 was evident in RAD51C-deficient cells. Interestingly, RAD51C-negative cells exhibited enhanced recruitment of non-homologous end joining (NHEJ) proteins onto chromatin and this accumulation correlated with increased activity of error-prone NHEJ as well as genome instability leading to cell death. Notably, inhibition of DNA-PKcs or depletion of KU70 or Ligase IV rescued this phenotype. Strikingly, stimulation of NHEJ by low dose of ionizing radiation (IR) in the PARP inhibitor-treated RAD51C-deficient cells and cells expressing pathological RAD51C mutants induced enhanced toxicity 'synergistically'. These results demonstrate that cancer cells arising due to hypomorphic mutations in RAD51C can be specifically targeted by a 'synergistic approach' and imply that this strategy can be potentially applied to cancers with hypomorphic mutations in other homologous recombination pathway genes.