Cytotoxic Imidazolyl-Mesalazine Ester-Based Ru(II) Complexes Reduce Expression of Stemness Genes and Induce Differentiation of Oral Squamous Cell Carcinoma.

The aggressiveness and recurrence of cancer is linked to cancer stem cells (CSCs), but drugs targeting CSCs may not succeed in the clinic due to the lack of a distinct CSC subpopulation. Clinical Pt(II) drugs can increase stemness. We screened 15 RuII or IrIII complexes with mesalazine or 3-aminobenzoate Schiff bases of the general formulas [Ru(p-cym)L]+, [Ru(p-cym)L], and [Ir(Cp*)L]+ (L = L1-L9) and found three complexes (2, 12, and 13) that are active against oral squamous cell carcinoma (OSCC) CSCs. There is a putative oncogenic role of transcription factors (viz. NOTCH1, SOX2, c-MYC) to enhance the stemness. Our work shows that imidazolyl-mesalazine ester-based RuII complexes inhibit growth of CSC-enriched OSCC 3D spheroids at low micromolar doses (2 µM). Complexes 2, 12, and 13 reduce stemness gene expression and induce differentiation markers (Involucrin, CK10) in OSCC 3D cultures. The imidazolyl-mesalazine ester-based RuII complex 13 shows the strongest effect. Downregulating c-MYC suggests that RuII complexes may target c-MYC-driven cancers.

Effect of microchannel geometry and linker molecules on surface immobilization efficiency of proteins in microfluidic devices.

One of the key metrics in the design of biosensors is the presence of an effective capture layer. Surface-immobilized proteins (especially as a part of antibody-antigen combinations) are the most commonly used capture ligands in biosensors. The surface coverage of these proteins in flow-based biosensors are affected by both the linker chemistry used to attach them as well as the microchannel geometry. We used streptavidin as a model protein to compare glutaraldehyde, EDC-NHS, sulfo-SMCC and sulfo-NHS-biotin as linkers inside straight, serpentine and square-wave microchannel geometries. We found that straight microchannels achieve the highest degree of protein immobilization compared to serpentine and square-wave microchannels, irrespective of the linker chemistry used. We also showed that for a given microchannel geometry, sulfo-NHS-biotin leads to the highest immobilization of streptavidin among all the linkers.

Mesoporous carbon structure impregnated with 2D engineered zirconium: A sustainable adsorbent for the removal of dyes from the aqueous solution.

The key designing of new breeds of the adsorbents aimed to improve the physical, chemical and textural morphology along with surface functionalization, selectivity toward the contaminants, and regenerations efficiency. In this aspect, two adsorbents named wet oxidative and ultrasonicated zirconium impregnated composite, have been synthesized through two routes, i.e., wet oxidation and ultrasonication. In wet oxidation method, carbon-based materials are oxidized using an oxidant followed by impregnation, while in ultrasonication assisted route, the impregnation is carried out using acoustic phenomenon. The characterization study revealed that the wet oxidation process is more competent in impregnating zirconium and developing diverse porosity and functionalities. The maximum adsorption capacity of wet oxidative adsorbent was 812 mg/g for Reactive Blue 19 and 203.18 mg/g for Methylene Blue, that accentuated the efficiency of the adsorbent over raw activated carbon. The electrostatic interaction, hydrogen-bonding and ligand exchange phenomenon are the involved adsorption mechanism for dyes. The regeneration study finally asserts that the wet oxidative adsorbent shows an insignificant decrease in its capacity up to the 5th-cycle (i.e., 87.67% removal at 5th cycle) as compared to raw AC (46.71% removal at 5th cycle). Further, a continuous fixed-bed column study revealed a significant correlation between experimental breakthrough data and kinetic data. Thus, the developed adsorbent has a sedulous adsorption capacity to remove the most stubborn toxic dyes and can be used in industrial-scale applications.

Effect of an Imidazole-Containing Schiff Base of an Aromatic Sulfonamide on the Cytotoxic Efficacy of N,N-Coordinated Half-Sandwich Ruthenium(II) p-Cymene Complexes.

Sulfonamides have a broad range of therapeutic applications, which include the inhibition of various isoforms of carbonic anhydrases (CAs). Among the various CA isoforms, CA IX is overexpressed in tumors and regulates the pH of the tumor microenvironment. Herein we present five new ruthenium(II) p-cymene complexes (1-5) of Schiff base ligands (L1-L4) of 4-(2-aminoethyl)benzenesulfonamide by varying the aldehyde to enhance the selective cytotoxicity toward cancer cells. All of the complexes are stable to aquation for the observed period of 24 h except 1, which aquated within 1 h, but the monoaquated species is stable for 24 h. The two imidazole derivatives, 1 and 2, are cytotoxic to the cancer cells MDA-MB-231 and MIA PaCa-2 but not to the noncancerous cells CHO and MDCK. The enhanced toxicity in hypoxia against MDA-MB-231 may be due to the greater expression of CA IX in hypoxia, as per the immunofluorescence data. The most cytotoxic complexes, 1 and 2, are lipophilic, whereas 3-5 show high hydrophilicity and are not cytotoxic up to 200 µM. Complexes 1 and 2 also show a higher cellular accumulation in MDA-MB-231 than the nontoxic yet solution-stable complex 5. The cytotoxic complexes bind with the model nucleobase 9-ethylguanine but have slow reactivity toward cellular tripeptide glutathione. Both 1 and 2 induce apoptosis by depolarizing the mitochondrial membrane potential and arrest the cell cycle in the SubG1 phase.

Hypoxia Active Platinum(IV) Prodrugs of Orotic Acid Selective to Liver Cancer Cells.

Platinum(IV) complexes of orotic acid selectively target liver cancer cells displaying enhanced activity and higher uptake in Hep G2. The comparatively higher expression of Organic Anion Transporter 2 (OAT2) in Hep G2 and decrease in toxicity in the presence of OAT2 inhibitor suggest its involvement in the uptake of the complexes. They are resistant to sequestration by the copper transporter ATP7B, unlike cisplatin and oxaliplatin.

Disruption of the Microtubule Network and Inhibition of VEGFR2 Phosphorylation by Cytotoxic N,O-Coordinated Pt(II) and Ru(II) Complexes of Trimethoxy Aniline-Based Schiff Bases.

Platinum-based complexes are one of the most successful chemotherapeutic agents having a significant ground in cancer chemotherapy despite their side effects. During the past few decades, Ru(II) complexes have been emerging as efficient alternatives owing to their promising activities against platinum-resistant cancer. The pathway of action, lipophilicity, and cytotoxicity of a Pt or Ru complex may be tuned by varying the attached ligands, the coordination mode, and the leaving group. In this work, we report a family of Pt(II) and Ru(II) complexes (1-5) of three N,O and N,N donor-based trimethoxyanilines containing Schiff bases with the general formula [PtII(L)(DMSO)Cl], [RuII(L)(p-cymene)Cl], [RuII(L)(p-cymene)Cl]+, and [PtII(L)Cl2]. All of the complexes are characterized by different analytical techniques. 1H NMR and electrospray ionization mass spectrometry (ESI-MS) data suggest that the N,O-coordinated Pt(II) complexes undergo slower aquation compared to the Ru(II) analogues. The change of the coordination mode to N,N causes the Ru complexes to be more inert to aquation. The N,O-coordinating complexes show superiority over N,N-coordinating complexes by displaying excellent in vitro antiproliferative activity against different aggressive cancer cells, viz., triple-negative human metastatic breast adenocarcinoma MDA-MB-231, human pancreatic carcinoma MIA PaCa-2, and hepatocellular carcinoma Hep G2. In vitro cytotoxicity studies suggest that Pt(II) complexes are more effective than their corresponding Ru(II) analogues, and the most cytotoxic complex 3 is 10-15 times more toxic than the clinical drugs cisplatin and oxaliplatin against MDA-MB-231 cells. Cellular studies show that all of the N,O-coordinated complexes (1-3) initiate disruption of the microtubule network in MDA-MB-231 cells in a dose-dependent manner within 6 h of incubation and finally lead to the arrest of the cell cycle in the G2/M phase and render apoptotic cell death. The disruption of the microtubule network affects the agility of the cytoskeleton rendering inhibition of tyrosine phosphorylation of vascular endothelial growth factor receptor 2 (VEGFR2), a key step in angiogenesis. Complexes 1 and 2 inhibit VEGFR2 phosphorylation in a dose-dependent fashion. Among the Pt(II) and Ru(II) complexes, the former displays higher cytotoxicity, a stronger effect on the cytoskeleton, better VEGFR2 inhibition, and strong interaction with the model nucleobase 9-ethylguanine (9-EtG).

Differences in Stability, Cytotoxicity, and Mechanism of Action of Ru(II) and Pt(II) Complexes of a Bidentate N,O Donor Ligand.

We report [RuII(L)(η6-p-cym)Cl] (1 and 2) and [PtII(L)(DMSO)Cl] (3 and 4) complexes, where L is a chelate imine ligand derived from chloroethylamine and salicylaldehyde (HL1) or o-vanillin (HL2). The complexes were characterized by single-crystal X-ray diffraction and other analytical techniques. The 1H nuclear magnetic resonance data show that both the Ru(II) and Pt(II) complexes start forming the aquated complex within an hour. The aquated complexes are stable at least up to 24 h. The complexes bind to the N7 of the model nucleobase 9-ethylguanine (9-EtG). Interaction with calf thymus (CT) DNA shows moderate binding interactions with binding constants, Kb (3.7 ± 1.2) × 103 M-1 and (4.3 ± 1.9) × 103 M-1 for 1 and 3, respectively. The complexes exhibit significant antiproliferative activity against human pancreas ductal adenocarcinoma (Mia PaCa-2), triple negative metastatic breast adenocarcinoma (MDA-MB-231), hepatocellular carcinoma (Hep G2), and colorectal adenocarcinoma (HT-29) cell lines. The studies show that with the same ligand the Pt(II) complexes are more potent than the Ru(II) complexes. The in vitro potencies of all the complexes toward pancreatic cancer cell line MIA PaCa-2 are more than cisplatin (CDDP). The Pt(II) and Ru(II) complexes show similar binding constants with CT-DNA, but the reactivity of the Pt(II) complex 3 with 9-EtG is faster and their overall cell killing pathways are different. This is evident from the arrest of the cell cycle by the Ru(II) complex 1 in the G2/M phase in contrast to the SubG1 phase arrest by the Pt(II) complex 3. The immunoblot study shows that 3 increases cyclin D and Bcl-2 expression in MDA-MB-231 due to the SubG1 phase arrest where these proteins express in greater quantities. However, both 1 and 3 kill in the apoptotic pathway via dose-dependent activation of caspase 3. Complex 3 depolarizes the mitochondria more efficiently than 1, suggesting its higher preference for the intrinsic pathway of apoptosis. Our work reveals that the same bidentate ligand with a change of the metal center, viz, Pt(II) or Ru(II), imparts significant variation in cytotoxic dosage and pathway of action due to specific intrinsic properties of a metal center (viz, coordination geometry, solution stability) manifested in a complex.

Inhibition of 3D colon cancer stem cell spheroids by cytotoxic RuII-p-cymene complexes of mesalazine derivatives.

The Ru(ii) complex of an imidazole-mesalazine Schiff base is a unique example showing growth inhibition of 3D-colon cancer stem cell spheroids and bulk colon cancer cells at lower dosage than salinomycin or oxaliplatin. Unlike oxaliplatin which increases the expression of stemness genes (SOX2, KLF4, HES1 and Oct4), these complexes maintain a tight regulation.

Effect of N,N Coordination and RuII Halide Bond in Enhancing Selective Toxicity of a Tyramine-Based RuII (p-Cymene) Complex.

Ruthenium compounds are promising anticancer candidates owing to their lower side-effects and encouraging activities against resistant tumors. Half-sandwich piano-stool type RuII compounds of general formula [(L)RuII(η6-arene)(X)]+ (L = chelating bidentate ligand, X = halide) have exhibited significant therapeutic potential against cisplatin-resistant tumor cell lines. In RuII (p-cymene) based complexes, the change of the halide leaving group has led to several interesting features, viz., hydrolytic stability, resistance toward thiols, and alteration in pathways of action. Tyramine is a naturally occurring monoamine which acts as a catecholamine precursor in humans. We synthesized a family of N,N and N,O coordinated RuII (p-cymene) complexes, [(L)RuII(η6-arene)(X)]+ (1-4), with tyramine and varied the halide (X = Cl, I) to investigate the difference in reactivity. Our studies showed that complex 2 bearing N,N coordination with an iodido leaving group shows selective in vitro cytotoxicity against the pancreatic cancer cell line MIA PaCa-2 (IC50 ca. 5 µM) but is less toxic to triple-negative breast cancer (MDA-MB-231), hepatocellular carcinoma (Hep G2), and the normal human foreskin fibroblasts (HFF-1). Complex 2 displays stability toward hydrolysis and does not bind with glutathione, as confirmed by 1H NMR and ESI-HRMS experiments. The inert nature of 2 leads to enhancement of cytotoxicity (IC50 = 5.3 ± 1 µM) upon increasing the cellular treatment time from 48 to 72 h.

Synthesis, Structure, Stability, and Inhibition of Tubulin Polymerization by RuII-p-Cymene Complexes of Trimethoxyaniline-Based Schiff Bases.

Four trimethoxy- and dimethoxyphenylamine-based Schiff base (L1-L4)-bearing RuII-p-cymene complexes (1-4) of the chemical formula [RuII(η6-p-cymene)(L)(Cl)] were synthesized, isolated in pure form, and structurally characterized using single-crystal X-ray diffraction and other analytical techniques. The complexes showed excellent in vitro antiproliferative activity against various forms of cancer that are difficult to cure, viz., triple negative human metastatic breast carcinoma MDA-MB-231, human pancreatic carcinoma MIA PaCa-2, and hepatocellular carcinoma Hep G2. The 1H nuclear magnetic resonance data in the presence of 10% dimethylformamide-d7 or dimethyl sulfoxide-d6 in phosphate buffer (pD 7.4, containing 4 mM NaCl) showed that the complexes immediately generate the aquated species that is stable for at least 24 h. Electrospray ionization mass spectrometry data showed that they do not bind with guanine nitrogen even in the presence of 5 molar equivalents of 9-EtG, during a period of 24 h. The best complex in the series, 1, exhibits an IC50 of approximately 10-15 µM in the panel of tested cancer cell lines. The complexes do not enhance the production of reactive oxygen species in the cells. Docking studies with a tubulin crystal structure (Protein Data Bank entry 1SAO ) revealed that 1 and 3 as well as L1 and L3 have a high affinity for the interface of the α and ß tubulin dimer in the colchicine binding site. The immunofluorescence studies showed that 1 and 3 strongly inhibited microtubule network formation in MDA-MB-231 cells after treatment with an IC20 or IC50 dose for 12 h. The cell cycle analysis upon treatment with 1 showed that the complexes inhibit the mitotic phase because the arrest was observed in the G2/M phase. In summary, 1 and 3 are RuII half-sandwich complexes that are capable of disrupting a microtubule network in a dose-dependent manner. They depolarize the mitochondria, arrest the cell cycle in the G2/M phase, and kill the cells by an apoptotic pathway.

Cytotoxic RuII-p-cymene complexes of an anthraimidazoledione: halide dependent solution stability, reactivity and resistance to hypoxia deactivation.

RuII-(η6-p-cymene) complexes of anthraimidazoldione (PAIDH) based ligand bearing the formula [RuII(η6-p-cymene)(PAIDH)(X)]+ (where, X = Cl, Br and I) showed excellent in vitro antiproliferative activity (IC50 range 1-2 µM) against hepatocellular carcinoma (HepG2), human pancreatic carcinoma (MIA PaCa-2) and triple negative human metastatic breast adenocarcinoma (MDA-MB-231). The ESI-MS and 1H NMR data show that the complexes are stable in aqueous solution at pH 7.4 (4 mM NaCl) with less than 10% hydrolysis in 24 h. However, when the coordinated halide is bromo (2) or iodo (3), the complex exchanges the halide with chloride in solution. The exchange is dependent on chloride concentration. Fastest chloride exchange was observed for the bromo complex 2 and slowest for the iodo complex 3 showing the higher kinetic inertness of the latter. Complex 3 exhibits the weakest interaction with glutathione (GSH) and 9-ethylguanine (9-EtG) in the series. ESI-MS studies of a 20% methanolic solution of 3 in 4 mM aqueous NaCl showed 80% intact complex even after 24 h of incubation with 9-EtG or GSH. 1-3 show similar in vitro cytotoxicity profile, but based on combined results from solution stability and cytotoxicity, the iodo complex 3 seems to be the best one in the series. There is no deterioration of toxicity under hypoxia or by induction of GSH in HepG2 cells. The low cytotoxicity of the complexes against difficult to treat triple negative breast carcinoma viz. MDA-MB-231 in vitro (IC50 = 1.5 ± 0.1 µM) is very encouraging, compared with cytotoxicity of clinical drug cisplatin (IC50 = 37.2 ± 2.5 µM). The complexes can alter mitochondrial membrane potential, arrest the cell cycle in G0/G1 phase and kill cells via apoptosis. They inhibit migration of the metastatic MDA-MB-231 cells at IC20 dose.