Copper(ii) curcumin complexes for endoplasmic reticulum targeted photocytotoxicity.

Three copper(ii) complexes viz. [Cu(cur)(L)(ClO4)] (1-3), where Hcur is curcumin and L is 1,10-phenanthroline (phen, 1), dipyrido[3,2-d:2',3'-f]quinoxaline (dpq, 2), or dipyrido[3,2-a:2',3'-c]phenazine (dppz, 3) were synthesized, fully characterized by various physicochemical methods and evaluated for their light-assisted chemotherapeutic potential. The complexes [Cu(acac)(L)(ClO4)] (4-6), where Hacac is acetylacetone and L is phen (in 4), dpq (in 5) and dppz (in 6), were synthesized and used as controls. The solid state structures of complexes 4 and 5 were determined by single crystal X-ray diffraction. The curcumin complexes (1-3) were redox inactive at the copper centre, whereas the acetylacetonato complexes (4-6) displayed a Cu(ii)/Cu(i) couple at ∼0.1 V vs. Ag/AgCl reference electrode in DMF. Complexes 1-3 showed an intense curcumin-based band at ∼440 nm in DMF-Tris-HCl buffer (pH = 7.2) (1 : 9 v/v) which masks the copper based d-d band. The complexes bind to human serum albumin (HSA) with moderate efficacy. They also displayed significant binding affinity for calf-thymus (CT) DNA. The lipophilic curcumin complexes show remarkable visible light induced cytotoxicity (IC50 = ∼4 µM) with high phototoxic indices (PI) with low dark toxicity in human cervical carcinoma (HeLa) and human lung carcinoma (A549) cells. The corresponding acetylacetonato controls (4-6) did not show significant cytotoxicity in the dark or light. DCFDA and annexin V-FITC/PI assays using flow cytometry confirm the induction of significant apoptosis in cancer cells via generation of cytotoxic reactive oxygen species upon photoactivation. Confocal microscopic images using complex 3 demonstrate localization of the complexes predominantly in the endoplasmic reticulum of HeLa cells.

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.

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.

Iron(III) Complex-Functionalized Gold Nanocomposite as a Strategic Tool for Targeted Photochemotherapy in Red Light.

Iron(III)-phenolate/carboxylate complexes exhibiting photoredox chemistry and photoactivated reactive oxygen species (ROS) generation at their ligand-to-metal charge-transfer (LMCT) bands have emerged as potential strategic tools for photoactivated chemotherapy. Herein, the synthesis, in-depth characterization, photochemical assays, and remarkable red light-induced photocytotoxicities in adenocarcinomic human immortalized human keratinocytes (HaCaT) and alveolar basal epithelial (A549) cells of iron(III)-phenolate/carboxylate complex of molecular formula, [Fe(L1)(L2)] (1), where L1 is bis(3,5 di-tert-butyl-2-hydroxybenzyl)glycine and L2 is 5-(1,2-dithiolan-3-yl)-N-(1,10-phenanthroline-5-yl)pentanamide, and the gold nanocomposite functionalized with complex 1 (1-AuNPs) are reported. There was a significant red shift in the UV-visible absorption band on functionalization of complex 1 to the gold nanoparticles (λmax: 573 nm, 1; λmax: 660 nm, 1-AuNPs), rendering the nanocomposite an ideal candidate for photochemotherapeutic applications. The notable findings in our present studies are (i) the remarkable cytotoxicity of the nanocomposite (1-AuNPs) to A549 (IC50: 0.006 µM) and HaCaT (IC50: 0.0075 µM) cells in red light (600-720 nm, 30 J/cm2) while almost nontoxic (IC50 > 500 µg/mL, 0.053 µM) in the dark, (ii) the nontoxicity of 1-AuNPs to normal human diploid fibroblasts (WI-38) or human peripheral lung epithelial (HPL1D) cells (IC50 > 500 µg/mL, 0.053 µM) both in the dark and red light signifying the target-specific anticancer activity of the nanocomposite, (iii) localization of 1-AuNPs in mitochondria and partly nucleus, (iv) remarkable red light-induced generation of reactive oxygen species (ROS: 1O2, •OH) in vitro, (v) disruption of the mitochondrial membrane due to enhanced oxidative stress, and (vi) caspase 3/7-dependent apoptosis. A similar cytotoxic profile of complex 1 was another key finding of our studies. Overall, our current investigations show a new red light-absorbing iron(III)-phenolate/carboxylate complex-functionalized gold nanocomposite (1-AuNPs) as the emerging next-generation iron-based photochemotherapeutic agent for targeted cancer treatment modality.

Photocytotoxic cancer cell-targeting platinum(ii) complexes of glucose-appended curcumin and biotinylated 1,10-phenanthroline.

Mixed-ligand platinum(ii) complexes, [Pt(phen)(pacac)](NO3) (1), [Pt(phen)(cur)](NO3) (2), [Pt(bt-phen)(cur)](NO3) (3) and [Pt(phen)(scur)](NO3) (4), where phen is 1,10-phenanthroline, bt-phen is 5-biotin-1,10-phenanthroline, pacac is 1,3-diphenyl-1,3-propanedioate anion, Hcur is curcumin and Hscur is diglucosylcurcumin, were prepared, characterized and their anticancer activity studied. Complexes 2-4 showed absorption bands within 410-430 nm (ε, 2.1 × 104 to 2.8 × 104 M-1 cm-1) in 10% DMSO-DPBS (Dulbecco's phosphate-buffered saline) and emission bands near 530 nm (λex = 410-430 nm) with a fluorescence quantum yield (ΦF) value of ∼0.02. The curcumin complexes showed stability over a study period of 48 h. The photocytotoxicity was studied using human cervical HeLa, human liver HepG2, human breast cancer MDA-MB 231 and human lung adenocarcinoma A549 cancer cells along with human immortalized lung epithelial HPL1D as normal cells. Complexes 2-4 showed apoptotic photo-induced cell death in light of wavelength 400-700 nm (IC50, half maximal inhibitory concentration: 6-28 µM) by reactive oxygen species (ROS), while remaining inactive in the dark (IC50: 43-95 µM). The selectivity of the complexes 3 and 4 was enhanced significantly towards the cancer cells than towards the normal cells, thus making them targeted photochemotherapeutic agents. The ROS formation and mode of cell death were studied from 2',7'-dichlorofluorescein diacetate (DCFDA) and annexin-V/FITC (fluorescein isothiocyanate)-PI assays, respectively. Preferential nuclear and mitochondrial localization was evidenced from inductively coupled plasma mass spectrometry (ICP-MS) studies.

Diplatinum(II) Catecholate of Photoactive Boron-Dipyrromethene for Lysosome-Targeted Photodynamic Therapy in Red Light.

The binuclear platinum(II) boron-dipyrromethene (BODIPY) complex [{Pt(dach)}2(µ-Dcrb)] (DP), where dach is 1,2-diaminocyclohexane and H4Dcrb is a morpholine-conjugated BODIPY-linked dicatechol bridging ligand, was prepared for lysosome organelle targeting and near-IR (NIR) light (600-720 nm) induced photocytotoxic activity. The platinum complex [Pt(dach)(cat)] (CP), where H2cat is catechol, was synthesized and used as a control complex without bearing the BODIPY unit. The complex DP displayed a band at 660 nm (ε = 2.1 × 104 M-1 cm-1) in the red region of the UV-visible spectrum recorded in 10% dimethyl sulfoxide/Dulbecco's Modified Eagle's Medium (DMSO/DMEM, pH 7.2). The complex DP and the BODIPY ligand displayed emission in 10% DMSO-DMEM (pH 7.2) giving an λem value of 668 nm (λex = 650 nm) with a ΦF value of 0.02 for DP and 0.16 for H4Dcrb (ΦF, fluorescence quantum yield). Titration experiments using 1,3-diphenylisobenzofuran (DPBF) indicated that the complex DP and H4Dcrb on irradiation with near-IR light of 600-720 nm generated singlet oxygen (1O2) as the ROS (reactive oxygen species). The complex DP showed significant lysosomal localization and remarkable apoptotic photodynamic therapy (PDT) effects, giving half-maximal inhibitory concentration values (IC50) within 0.6-3.4 µM in HeLa cervical cancer, A549 lung cancer, and MDA-MB231 multidrug resistant cancer cells, while being essentially nontoxic in the dark and in the HPL1D immortalized lung epithelial normal cells. The acridine orange assay using A549 cells showed lysosomal membrane permeabilization by the complex DP under near-IR light (600-720 nm). This complex on near-IR light (600-720 nm) activation in A549 cells induced apoptotic cell death, as observed from an Annexin-V FITC assay.

Glucose-Appended Platinum(II)-BODIPY Conjugates for Targeted Photodynamic Therapy in Red Light.

Platinum(II) complexes [Pt(L1)(R-BODIPY)]Cl (1) and [Pt(L2)(R-BODIPY)]Cl (2), where R-BODIPY is 8-(4-ethynylphenyl)-distyryl-4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene-3, L1 is 4'-phenyl-2,2':6',2″-terpyridine, and L2 is (2,2':6',2″-terpyridin-4'-oxy)ethyl-ß-d-glucopyranoside, were synthesized and characterized, and their photocytotoxicity was studied. The phenylacetylide complex [Pt(L1)(C≡CPh)]Cl (3) was prepared and used as a control. Complexes 1 and 2 showed near-IR absorption bands at 713 nm (ε = 3.47 × 104 M-1 cm-1) and 715 nm (3.2 × 104 M-1 cm-1) in 10% dimethyl sulfoxide (DMSO)-Dulbecco's Modified Eagle's Medium (DMEM) (pH 7.2). The BODIPY complexes are emissive in 10% DMSO-DMEM at pH 7.2 with λem (λex, Φf) = 822 nm (710 nm, 0.022) for complex 1 and λem (λex, Φf) = 825 nm (710 nm, 0.026) for complex 2. They generated singlet oxygen (1O2) in red light as evidenced from 1,3-diphenylisobenzofuran (DPBF) titration experiments. The singlet oxygen quantum yield (ΦΔ) values for 1 and 2 were ∼0.6 signifying their photosensitizing ability. They were remarkably photodynamic therapy (PDT) active in red light showing significant red light-induced cytotoxicity in cervical HeLa, lung cancer A549, and breast cancer MCF-7 cells (IC50: 2.3-24.7 µM in light) with negligible dark toxicity (IC50 > 100 µM). A significant enhancement in cellular uptake was observed for 2 having glucose-appended terpyridine ligand compared to 1. The confocal microscopy showed significant mitochondrial localization of the complexes as evidenced from the JC-1 assay. The complexes released the photoactive R-BODIPY ligand upon red light-irradiation as evidenced from the mass and 1H NMR spectral studies. Complex 2 is remarkable in satisfying the essential requirements of targeted PDT in red light.

Photochemotherapy of Infrared Active BODIPY-Appended Iron(III) Catecholates for in Vivo Tumor Growth Inhibition.

Iron(III) catecholates of BODIPY (boron-dipyrromethene)-conjugated dipicolylamine ligands, viz. [Fe(L1)(cat)Cl] (1) and [Fe(L2)(cat)Cl] (2) (H2cat = catechol), with a ligand-to-metal charge transfer band at ∼800 nm were studied for their in vivo activity in dark and infrared light in luciferase-expressing human breast adenocarcinoma (BT474luc) cells. Complex 2 displayed in vitro photocytotoxicity in BT474luc cells (IC50: 6 µM) in infrared light of 600-720 nm with moderate dark toxicity (IC50: 18 µM) as evidenced from the MTT and Annexin-V FITC/PI staining assays. The mitochondria-localizing complexes showed apoptotic cell death involving reactive oxygen species whose generation was evidenced from 2,7-dichlorofluorescein diacetate assay. In vivo studies showed tumor growth inhibition in mice with an optimized complex 2 dose of 5 mg per kg body weight on exposure to infrared light of 685 nm (dose of 20 mW/cm2). The in vivo results exemplify complex 2 as a unique iron-based infrared-active photochemotherapeutic agent.