Cobalt(III) Complexes for Light-Activated Delivery of Acetylacetonate-BODIPY, Cellular Imaging, and Photodynamic Therapy.

Cobalt(III) complexes [Co(TPA)(L1)](ClO4)2 (1), [Co(4-COOH-TPA)(L1)](ClO4)2 (2), [Co(TPA)(L2)]Cl2 (3), and [Co(4-COOH-TPA)(L2)]Cl2 (4) having acetylacetonate-linked boron-dipyrromethene ligands (L1, acac-BODIPY; L2, acac-diiodo-BODIPY) were prepared and characterized, and their utility as bioimaging and phototherapeutic agents was evaluated (TPA, tris-(2-pyridylmethyl)amine; 4-COOH-TPA, 2-((bis-(2-pyridylmethyl)amino)methyl)isonicotinic acid). HL1, HL2, and complex 1 were structurally characterized by X-ray crystallography. Complexes 1 and 2 on photoactivation or in a reducing environment (excess GSH, ascorbic acid, and 3-mercaptopropionic acid) released the acac-BODIPY ligand. They exhibited strong absorbance near 501 nm (ε ∼ (5.2-5.8) × 104 M-1 cm-1) and emission bands near 513 nm (ΦF ∼ 0.13, λex = 490 nm) in dimethyl sulfoxide (DMSO). Complexes 3 and 4 with absorption maxima at ∼536 and ∼538 nm (ε ∼ (1.2-1.8) × 104 M-1 cm-1), respectively, afforded high singlet oxygen quantum yield (ΦΔ âˆ¼ 0.79) in DMSO. Complexes 1-4 showed Co(III)-Co(II) redox responses near -0.2 V versus saturated calomel electrode (SCE) in dimethylformamide (DMF)-0.1 M tetrabutylammonium perchlorate (TBAP). The photocleavage of pUC19 DNA by complex 4 revealed the formation of both singlet oxygen and superoxide anion radicals as the reactive oxygen species (ROS). Confocal fluorescence microscopy showed the selective accumulation of complex 1 in the endoplasmic reticulum (ER) in A-549 cells. Complex 4 exhibited a high phototherapeutic index value (PI > 7000) in HeLa cancer cells (IC50 ∼ 0.007 µM in visible light of 400-700 nm, total dose ∼5 J cm-2). The ancillary ligands in the complexes demonstrated a structure-activity relationship and modulated the Co(III)-Co(II) redox potential, the complex solubility, acac-BODIPY ligand release kinetics, and phototherapeutic efficacy.

BODIPY-Tagged Platinum(II) Curcumin Complexes for Endoplasmic Reticulum-Targeted Red Light PDT.

[Pt(RB)(Cur)]NO3 (RBC), [Pt(IRB)(Cur)]NO3 (IRBC), and [Pt(L)(Cur)]NO3 (PBC), where HCur is curcumin, L is 1-benzyl-2-(2-pyridyl)benzimidazole, and RB and IRB are red-light-active non-iodo and diiodo-BODIPY tagged to L, respectively, were synthesized and characterized, and their anticancer activities were studied (BODIPY, boron-dipyrromethene). RBC and IRBC displayed BODIPY-centered absorption bands within 615-635 nm along with the respective curcumin bands at 445 and 492 nm in 10% dimethyl sulfoxide (DMSO)-Dulbecco's phosphate-buffered saline (DPBS). Emission bands were observed at 723 and 845 nm for RBC and IRBC, respectively, in 10% DMSO-DPBS. RBC (ΦΔ, 0.27) and IRBC (ΦΔ, 0.40) generated singlet oxygen in red light (λ = 642 nm) as evidenced from 1,3-diphenylisobenzofuran (DPBF) titrations. The formation of 1O2 from BODIPY and HO• from the curcumin was evidenced from the mechanistic pUC19 DNA photocleavage studies. The BODIPY complexes showed photocytotoxicity in A549, HeLa, and MDA-MB-231 cells while being less toxic in the dark [IC50: 1.3-6.9 µM, red light; 7.2-12.8 µM, 400-700 nm visible light]. The emissive RBC displayed localization in the endoplasmic reticulum (ER). Apoptotic cell death was evidenced from the Annexin-V/fluorescein isothiocyanate (FITC)/propidium iodide (PI) assay and green fluorescence in red light in the Fluo-4 AM assay due to ER stress, and mitochondrial dysfunction was evidenced from the 5,5,6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolylcarbocyanine iodide (JC-1) assay in A549 cells.

Genome-wide DNA methylation changes in oral submucous fibrosis.

OBJECTIVE: Oral submucous fibrosis (OSF) is a debilitating potentially malignant condition of the buccal cavity characterized by extensive extracellular matrix deposition resulting in stiffness and trismus. As OSF is a progressive disease, we hypothesized that there would be extensive epigenetic changes in OSF tissues. MATERIALS AND METHODS: Using the Infinium HumanMethylation450 BeadChip Array, we analyzed gross DNA methylation changes in seven OSF tissues compared to five controls. Comparison with transcriptomic data and pathway analyses was conducted to find commonly regulated genes. RESULTS: A total of 3,294 differentially methylated regions mapping to 857 genes were identified. Comparison with transcriptome data revealed 38 downregulated-hypermethylated genes and 55 hypomethylated-upregulated genes. Using methylation-specific and qRT-PCR, aberrant hypomethylation and increased expression of FGF13, RPS6KA3, and ACSL4 genes were confirmed. Pathways involved in insulin signaling, ubiquitin-mediated proteolysis, nicotine addiction, and RAS/MAPK pathways were dysregulated, among others. Intriguingly, numerous genes located on the X chromosome were dysregulated in OSF tissues as the transcript for XIST gene was downregulated due to hypermethylation of the XIST promoter. CONCLUSIONS: This study highlights global epigenetic dysregulation of tissues of the oral cavity in OSF patients and hints at possible X chromosomal dysregulation, previously not implicated in the pathogenesis of OSF.

BODIPY-Ruthenium(II) Bis-Terpyridine Complexes for Cellular Imaging and Type-I/-II Photodynamic Therapy.

A series of multichromophoric ruthenium(II) complexes with the formulation [Ru(tpy-BODIPY)(tpy-R)]Cl2 (1-4), having a heteroleptic Ru(II)-bis-tpy (tpy = 4'-phenyl-2,2':6',2″-terpyridine) moiety covalently linked to a boron-dipyrromethene (BODIPY) pendant, have been prepared and characterized and their application as a phototherapeutic and photodetection agent in cancer therapy has been explored. Ligand L1 with a terpyridine-BODIPY moiety and complex 1 as its PF6 salt (1a) have been structurally characterized by a single-crystal X-ray diffraction study. Complex 1a has a distorted-octahedral RuN6 core with a Ru(II)-bis-terpyridine unit that is covalently linked to one photoactive BODIPY unit. The complexes exhibit strong absorbance near 502 nm (ε ≈ (3.7-7.8) × 104 M-1 cm-1) and high singlet oxygen sensitization ability, giving singlet oxygen quantum yield (ΦΔ) values ranging from 0.57 to 0.75 in DMSO. An emission-based study using complex 4 and Singlet Oxygen Sensor Green (SOSG) displays the formation of singlet oxygen inside the cells and also in the buffer medium upon light irradiation. DNA (pUC19) photocleavage experiments using ROS scavengers/stabilizers reveal photoinduced generation of singlet oxygen by a type-II process and of the superoxide anion radical by a type-I process. Complex 4 having a pendant biotin moiety as a cancer cell targeting group shows high photocytotoxicity with a remarkable phototherapeutic index (PI) value of >1400 in HeLa cancer cells with a low light dose activation (400-700 nm, 2.2 J cm-2). The complexes display reduced activity in noncancerous HPL1D cells. The emission property of the complexes is used for cellular imaging, thus making them suitable as next-generation theranostic PDT agents.

Maloplatin-B, a Cisplatin-Based BODIPY-Tagged Mito-Specific "Chemo-PDT" Agent Active in Red Light.

Maloplatin-B, a cisplatin-based complex, namely [Pt(A-BOD)(NH3)2](NO3) (Pt-A-BOD) with a pendant boron-dipyrromethene (BODIPY) moiety, where HA-BOD is a methyl malonyl chloride derived monostyryl BODIPY ligand, was designed and developed as near-IR light (600-720 nm) organelle-targeting photodynamic therapy agent. The complex [Pt(acac)(NH3)2](NO3) (Pt-Ac) was used as a control. Pt-A-BOD displayed an absorption band at 616 nm (ε = 2.9 × 104 M-1 cm-1) in 10% dimethyl sulfoxide/Dulbecco's Modified Eagle's Medium (DMSO/DMEM, pH 7.2). This complex displayed a broad emission band within 650-850 nm with a λem value of 720 nm in 10% DMSO-DMEM (pH 7.2) upon excitation (λex) at 615 nm with a large Stokes shift. The fluorescence quantum yield (ΦF) value for Pt-A-BOD is 0.032 and for the ligand HA-BOD is 0.24. The BODIPY complex and ligand showed the formation of singlet oxygen as the ROS (reactive oxygen species) on irradiation with near-IR red light of 660 nm, as evidenced from a 1,3-diphenylisobenzofuran (DPBF) assay. The complex displayed remarkable apoptotic NIR light-induced PDT activity with half-maximum inhibitory concentration values (IC50) of 1.6-2.4 µM in A549 lung and HeLa cervical cancer cells, while it was less active in the dark. The cellular ROS generation by the complex in red light was ascertained by a DCFDA (2',7'-dichlorofluorescein diacetate) assay. Cellular imaging showed its localization primarily in the mitochondria of A549 cancer cells. The JC1 and Annexin-V FITC/PI assays carried out for A549 cancer cells treated with the BODIPY complex showed the alteration of mitochondrial membrane potential and apoptotic cell death on near-IR red light (600-720 nm) irradiation, respectively.

Low mitochondrial DNA copy number is associated with poor prognosis and treatment resistance in glioblastoma.

INTRODUCTION: Mitochondrial DNA (mtDNA) content in several solid tumors was found to be lower than in their normal counterparts. However, there is paucity of literature on the clinical significance of mtDNA content in glioblastoma and its effect on treatment response. Hence, we studied the prognostic significance of mtDNA content in glioblastoma tumor tissue and the effect of mtDNA depletion in glioblastoma cells on response to treatment. MATERIALS AND METHODS: 130 newly diagnosed glioblastomas, 32 paired newly diagnosed and recurrent glioblastomas and 35 non-neoplastic brain tissues were utilized for the study. mtDNA content in the patient tumor tissue was assessed and compared with known biomarkers and patient survival. mtDNA was chemically depleted in malignant glioma cell lines, U87, LN229. The biology and treatment response of parent and depleted cells were compared. RESULTS: Lower range of mtDNA copy number in glioblastoma was associated with poor overall survival (p = 0.01), progression free survival (p = 0.04) and also with wild type IDH (p = 0.02). In recurrent glioblastoma, mtDNA copy number was higher than newly diagnosed glioblastoma in the patients who received RT (p = 0.01). mtDNA depleted U87 and LN229 cells showed higher survival fraction post radiation exposure when compared to parent lines. The IC50 of TMZ was also higher for mtDNA depleted U87 and LN229 cells. The depleted cells formed more neurospheres than their parent counterparts, thus showing increased stemness of mtDNA depleted cells. CONCLUSION: Low mtDNA copy number in glioblastoma is associated with poor patient survival and treatment resistance in cell lines possibly by impacting stemness of the glioblastoma cells.

A snapshot of reproductive and sexual health and rights of women living with HIV in Asia.

OBJECTIVE: A brief review of status of sexual and reproductive health and rights of women living with human immunodeficiency virus (HIV) in Asia with a focus on their access to related services. SUBJECT(S) AND METHODS: The content has primarily emerged from several meetings, workshops and research studies that regional networks of women living with HIV have conducted in the region. We used published and unpublished literature in English after the year 2000. RESULTS: We found many hindrances to realization of sexual and reproductive rights by HIV-positive women and their access to related health services. Rampant stigma and discrimination by service providers take the form of denial of contraceptive and maternal health services, forced sterilization, breach of confidentiality and disclosure of status to others without the consent of the woman. Families blaming women for bringing HIV and male partners objecting to using condoms affect the well-being of HIV-positive women. Sexual rights specifically aspects of positive sexuality such as sexual pleasure have not received attention. CONCLUSION: Findings suggest the need to enhance capacities of HIV-positive women's organizations to meaningfully participate in decisions on positive women's sexual, reproductive health and rights and sensitize service providers to adequately address HIV-positive women's concerns.

Ruthenium(II) Conjugates of Boron-Dipyrromethene and Biotin for Targeted Photodynamic Therapy in Red Light.

The ruthenium(II) complexes [RuCl(L1)(L3)]Cl (1), [RuCl(L1)(L4)]Cl (2), [RuCl(L2)(L4)]Cl (3), [RuCl(L1)(L5)]Cl (4), and [RuCl(L2)(L5)]Cl (5) of NNN-donor dipicolylamine (dpa) bases (L4, L5) having BODIPY (boron-dipyrromethene) moieties, NN-donor phenanthroline derivatives (L1, L2), and benzyldipicolylamine (bzdpa, L3) were prepared and characterized by spectroscopic techniques and their cellular localization/uptake and photocytotoxicity studied. Complex 1, as its PF6 salt (1a), has been structurally characterized with help of a single-crystal X-ray diffraction technique. It has a RuN5Cl core with the Cl bonded trans to the amine nitrogen atom of bzdpa. The complexes showed intense absorption spectral bands near 500 nm (ε ≈ 58000 M-1 cm-1) in 2 and 3 and 654 nm (ε ≈ 80000 M-1 cm-1) in 4 and 5 in 1/1 DMSO/DPBS (v/v). Complex 5 having biotin and PEGylated-disteryl BODIPY gave a singlet oxygen quantum yield (ΦΔ) of ∼0.65 in DMSO. Complex 5 exhibited remarkable PDT (photodynamic therapy) activity (IC50 ≈ 0.02 µM) with a photocytotoxicity index (PI) value of >5000 in red light of 600-720 nm in A549 cancer cells. The biotin-conjugated complexes showed better photocytotoxicity in comparison to nonbiotinylated analogues in A549 cells. The complexes displayed less toxicity in HPL1D normal cells in comparison to A549 cancer cells. The emissive BODIPY complexes 3 and 5 (ΦF ≈ 0.07 in DMSO) showed significant mitochondrial localization.

Serum biomarkers identification by iTRAQ and verification by MRM: S100A8/S100A9 levels predict tumor-stroma involvement and prognosis in Glioblastoma.

Despite advances in biology and treatment modalities, the prognosis of glioblastoma (GBM) remains poor. Serum reflects disease macroenvironment and thus provides a less invasive means to diagnose and monitor a diseased condition. By employing 4-plex iTRAQ methodology, we identified 40 proteins with differential abundance in GBM sera. The high abundance of serum S100A8/S100A9 was verified by multiple reaction monitoring (MRM). ELISA and MRM-based quantitation showed a significant positive correlation. Further, an integrated investigation using stromal, tumor purity and cell type scores demonstrated an enrichment of myeloid cell lineage in the GBM tumor microenvironment. Transcript levels of S100A8/S100A9 were found to be independent poor prognostic indicators in GBM. Medium levels of pre-operative and three-month post-operative follow-up serum S100A8 levels predicted poor prognosis in GBM patients who lived beyond median survival. In vitro experiments showed that recombinant S100A8/S100A9 proteins promoted integrin signalling dependent glioma cell migration and invasion up to a threshold level of concentrations. Thus, we have discovered GBM serum marker by iTRAQ and verified by MRM. We also demonstrate interplay between tumor micro and macroenvironment and identified S100A8 as a potential marker with diagnostic and prognostic value in GBM.