In vivo anticancer activity of rhomboidal Pt(II) metallacycles

The development of novel antitumor agents that have high efficacy in suppressing tumor growth, have low toxicity to nontumor tissues, and exhibit rapid localization in the targeted tumor sites is an ongoing avenue of research at the interface of chemistry, cancer biology, and pharmacology. Supramolecular metal-based coordination complexes (SCCs) have well-defined shapes and geometries, and upon their internalization, SCCs could affect multiple oncogenic signaling pathways in cells and tissues. We investigated the uptake, intracellular localization, and antitumor activity of two rhomboidal Pt(II)-based SCCs. Laser-scanning confocal microscopy in A549 and HeLa cells was used to determine the uptake and localization of the assemblies within cells and their effect on tumor growth was investigated in mouse s.c. tumor xenograft models. The SCCs are soluble in cell culture media within the entire range of studied concentrations (1 nM–5 µM), are nontoxic, and showed efficacy in reducing the rate of tumor growth in s.c. mouse tumor xenografts. These properties reveal the potential of Pt(II)-based SCCs for future biomedical applications as therapeutic agents.Molecular assemblies of nanoscale-size and well-defined geometries have recently emerged as an interesting new paradigm in drug design and drug delivery. To date, liposomes, the self-assembled lipid nanoparticles held together by weak interactions, are among the most widely studied and clinically successful nanoparticle-based drug carriers. Their use allows the drug to achieve sustained plasma levels while encapsulated, with the size preventing the fast clearance by the kidneys that often occurs with the free drug. However, liposomes themselves do not produce a therapeutic effect and their application as drug carriers for medical purposes has often been hindered by poor loading capacity (<5 wt %) and the inability to pass through biological barriers (1, 2). Inorganic and hybrid porous materials, such as molecular organic frameworks (MOFs), have also shown promise due to their higher loading capacities (>25 wt %) (3–5), but MOFs have poor hydrolytic stability (6, 7). Recent studies on materials from Institut Lavoisier (MIL)-100(Cr) and MIL-100(Fe), however, suggest that MOFs can persist in biologically relevant environments and can act as vehicles for some anticancer and antiviral agents (8–10). These early findings have prompted further investigations into the biomedical applications of supramolecular coordination complexes (SCCs) (11–24). SCCs preserve the attractive properties of MOFs, such as building block modularity (22, 23, 25), yet afford an increased solubility in the biological milieu and lend themselves to small-molecule characterization techniques due to their well-defined structure.Although development of SCCs for biomedical applications is in its infancy, some SCCs, such as trigonal prisms self-assembled from p-cymene and ruthenium-based metal fragments with pyridyl donors, have shown the ability to act as effective carriers of some chemotherapeutic agents (26–28). Moreover, a library of cytotoxic to cancer cells p-cymene ruthenium-based polygons and cages has also been developed (11). For biomedical applications, the information about the cellular uptake, delivery of a guest, and metabolism of the drug delivery vehicle is critical, although currently the fate of SCCs in biological environments is not well understood. In a rare report, a systematic investigation of the structural stability of a water-soluble, hexacationic ruthenium-based trigonal prism was performed; however, it was determined that the ruthenium-based trigonal prisms decompose in the presence of amino acids histidine, lysine, and arginine (29).An intriguing approach is the design of tumor-targeted modalities that combine detection and treatment through the self-assembly of emissive, metal-based coordination complexes. Such modalities can be especially valuable as they often do not require photoexcitation to elicit cytotoxicity. Recently Gray, Gross, and Medina-Kauwe and coworkers reported HerGa, a self-assembled tumor-targeted particle that bears the Ga(III)-metalated derivative of the sulfonated corrole (30, 31). The particle, which contained Ga(III)-corrole noncovalently bound to the tumor-targeting cell penetration protein HerPBK10, provided both tumor detection and elimination. Systemic injection of this protein–corrole complex resulted in tumor accumulation, which can be visualized in vivo due to the red corrole fluorescence. Cytotoxic and cytostatic properties of these targeted Ga(III) corroles were found to be cell-line dependent, with the ability to induce late M-phase arrest in several cancer cell lines (32).Despite the well-known cytotoxic properties of mono- and multinuclear platinum complexes (33–35), studies of the antitumor properties of platinum-based SCCs are rare (17, 36). Moreover, recent reports have demonstrated that platinum-based SCCs can act as effective hosts for guests and have interesting photophysical properties (37–42). In particular, highly emissive rhomboids based on aniline-containing donors and Pt-based metal acceptors have been developed that display different photophysical properties from those of their constituent subunits (40). These assemblies are interesting targets to investigate the cytotoxicity of organoplatinum SCCs, whereas their emission spectra could be used for interrogating the structural integrity in vitro. Here, for the first time to our knowledge, we report the uptake of SCCs in vitro in cell-based assays, determined by using laser-scanning confocal microscopy, and an in vivo assessment of the anticancer activity of SCCs in mouse s.c. tumor xenograft models.     

Molecular outcomes of neuromyelitis optica (NMO)-IgG binding to aquaporin-4 in astrocytes

The astrocytic aquaporin-4 (AQP4) water channel is the target of pathogenic antibodies in a spectrum of relapsing autoimmune inflammatory central nervous system disorders of varying severity that is unified by detection of the serum biomarker neuromyelitis optica (NMO)-IgG. Neuromyelitis optica is the most severe of these disorders. The two major AQP4 isoforms, M1 and M23, have identical extracellular residues. This report identifies two novel properties of NMO-IgG as determinants of pathogenicity. First, the binding of NMO-IgG to the ectodomain of astrocytic AQP4 has isoform-specific outcomes. M1 is completely internalized, but M23 resists internalization and is aggregated into larger-order orthogonal arrays of particles that activate complement more effectively than M1 when bound by NMO-IgG. Second, NMO-IgG binding to either isoform impairs water flux directly, independently of antigen down-regulation. We identified, in nondestructive central nervous system lesions of two NMO patients, two previously unappreciated histopathological correlates supporting the clinical relevance of our in vitro findings: (i) reactive astrocytes with persistent foci of surface AQP4 and (ii) vacuolation in adjacent myelin consistent with edema. The multiple molecular outcomes identified as a consequence of NMO-IgG interaction with AQP4 plausibly account for the diverse pathological features of NMO: edema, inflammation, demyelination, and necrosis. Differences in the nature and anatomical distribution of NMO lesions, and in the clinical and imaging manifestations of disease documented in pediatric and adult patients, may be influenced by regional and maturational differences in the ratio of M1 to M23 proteins in astrocytic membranes.     

Vitamin C modulates DNA damage induced by hydrogen peroxide in human colorectal adenocarcinoma cell lines (HT29) estimated by comet assay in vitro

Introduction

Cancer cells, compared to normal cells, are under increased oxidative stress associated with oncogenic transformation, alterations in metabolic activity, and increased generation of reactive oxygen species.

Material and methods

We investigated the ability of vitamin C to reduce the damage induced by hydrogen peroxide, in human colorectal adenocarcinoma cells in vitro by the comet assay. Additionally, we measured the kinetics and efficacy of the repair of DNA damage after incubation with vitamin C in the presence of H₂O₂.

Results

The obtained results showed that 1 h pre-incubation with vitamin C and exposure to H₂O₂ for the last 10 min of incubation caused a statistically significant (p < 0.05) increase in DNA migration in comet tails in all experimental series. For the 10 µM, 25 µM, 50 µM, 100 µM vitamin C concentrations the levels of DNA damage were as follows: 18.6%, 21.1%, 25.3% and 27.2%, respectively, as compared to the untreated cells (3.26%). However, in comparison with H₂O₂ alone (29.1%), we observed a statistically significant (p < 0.05) decrease of the genotoxic effect in HT29 cells induced by H₂O₂ for the two lowest of concentrations of vitamin C: 10 µM and 25 µM. The HT29 cells were able to achieve effective repair of the damaged DNA within 60 and 120 min after incubation with the tested compounds. All the values obtained in the test were statistically significant (p < 0.05).

Conclusions

Vitamin C caused a weaker DNA damaging effect of hydrogen peroxide and positively influences the level of oxidative DNA damage in HT29 cells (decrease ∼ 30%). We noted that DNA damage was effectively repaired during 120 min postincubation in the tested cells and that oxidative damage was the major type of damage.     

Using high-voltage electrical stimulation in the treatment of recalcitrant pressure ulcers: results of a randomized, controlled clinical study

The use of electrical stimulation has been studied in a variety of wounds emphasizing different variables with regard to provision of therapy. The purpose of this prospective, randomized, controlled clinical study was to evaluate the effect of high-voltage electrical stimulation (HVES) on nonhealing, lower-extremity, Stage II and Stage III pressure ulcers. Patients admitted for care and eligible to participate in the study received standard supportive care and topical treatments covered with wet-to-moist dressings. Patients assigned to the treatment arm of the study also received HVES (100 V; 100 μs; 100 Hz) continuously for 50 minutes once daily, five times per week. Patients were followed until healing for a maximum of 6 weeks. Wound tracings and measurements were obtained weekly. Over a 4-year period, 26 patients were enrolled in the treatment and 24 in the control group. Ulcers had existed for an average of 3.17 and 2.83 months in the treatment and control groups, respectively. Most were classified as Stage II (17 in the treatment and 16 in the control group) with an average baseline size of 4.54 cm2 and 3.97 cm2, respectively. Wound areas and linear measurements decreased significantly in both groups (P <0.05), but increases in granulation tissue were significant in the treatment group only (P = 0.006). Wound area, linear measurement, wound volume, and granulation tissue changes were statistically significantly greater in the treatment than in the control group starting in the second week of treatment. Week 6 surface area change was 88.9% (SD 14) in the treatment and 44.4% (SD 63.1) in the control group (P = 0.00003). Correlation coefficients between changes in wound surface area, longest length, and longest width were R = 0.96 and R = 0.98 in the treatment and R = 0.94 and R = 0.89 in the control group. HVES improved the healing rate of recalcitrant Stage II and Stage III pressure ulcers. Research to compare the effectiveness of using cathodic and anodal stimulation combined or alone and to determine the optimal duration of these two types of electrical stimulation is warranted.     

IGFBP-3 Mediates Metabolic Homeostasis During Hyperosmolar Stress in the Corneal Epithelium

PurposeThe insulin-like growth factor binding protein-3 (IGFBP-3) is a multifunctional secretory protein with well-known roles in cell growth and survival. Data in our laboratory suggest that IGFBP-3 may be functioning as a stress response protein in the corneal epithelium. The purpose of this study is to determine the role of IGFBP-3 in mediating the corneal epithelial cell stress response to hyperosmolarity, a well-known pathophysiological event in the development of dry eye disease.MethodsTelomerase-immortalized human corneal epithelial (hTCEpi) cells were used in this study. Cells were cultured in serum-free media with (growth) or without (basal) supplements. Hyperosmolarity was achieved by increasing salt concentrations to 450 and 500 mOsM. Metabolic and mitochondrial changes were assessed using Seahorse metabolic flux analysis and assays for mitochondrial calcium, polarization and mtDNA. Levels of IGFBP-3 and inflammatory mediators were quantified using ELISA. Cytotoxicity was evaluated using a lactate dehydrogenase assay. In select experiments, cells were cotreated with 500 ng/mL recombinant human (rh)IGFBP-3.ResultsHyperosmolar stress altered metabolic activity, shifting cells towards a respiratory phenotype. Hyperosmolar stress further altered mitochondrial calcium levels, depolarized mitochondria, decreased levels of ATP, mtDNA, and expression of IGFBP-3. In contrast, hyperosmolar stress increased production of the proinflammatory cytokines IL-6 and IL-8. Supplementation with rhIGFBP-3 abrogated metabolic and mitochondrial changes with only marginal effects on IL-8.ConclusionsThese findings indicate that IGFBP-3 is a critical protein involved in hyperosmolar stress responses in the corneal epithelium. These data further support a new role for IGFBP-3 in the control of cellular metabolism.     

Gender differences in clinicopathological features and survival in surgically treated patients with renal cell carcinoma: an analysis of the multicenter CORONA database

Purpose

To investigate gender differences in clinicopathological features and to analyze the prognostic impact of gender in renal cell carcinoma (RCC) patients undergoing surgery.

Methods

A total of 6,234 patients (eleven centers; Europe and USA) treated by radical or partial nephrectomy were included in this retrospective study (median follow-up 59 months; IQR 30–106). Gender differences in clinicopathological parameters were assessed. Multivariable Cox regression models were applied to determine the influence of parameters on disease-specific survival (DSS) and overall survival (OS).

Results

A total of 3,751 patients of the study group were male patients (60.2 %), who were significantly younger at diagnosis and received more frequently NSS than women. Significantly, more often high-grade tumors and simultaneous metastasis were present in men. Whereas tumor size and pTN stages did not differ between genders, clear-cell and chromophobe RCC was diagnosed less frequently, but papillary RCC more often in men. Gender also independently influenced DSS (HR 0.75, p < 0.001) and OS (HR 0.80, p < 0.001) with a benefit for women. However, inclusion of gender in multivariable models did not significantly gain predictive accuracies (PA) for DSS (0.868–0.870, p = 0.628) and OS (0.775–0.777, p = 0.522). Furthermore, no significantly different DSS and OS rates were found in patients undergoing NSS.

Conclusions

This study demonstrates important gender differences in clinicopathological features and outcome of RCC patients with improved DSS and OS for women compared to men, even if solely patients with clear-cell RCC or M0-stage are taken into evaluation. However, inclusion of gender in multivariable models does not significantly gain PA of multivariable models.     

Common trace elements alleviate pain in an experimental mouse model

Trace elements represent a group of essential metals or metaloids necessary for life, present in minute amounts. Analgesic adjuvants can enhance the effect of other pain drugs or be used for pain control themselves. Previous studies on the effects of trace elements on nociception and their potential use as analgesic adjuvants have yielded conflicting results. In this study, we tested the hypothesis that three vital trace elements (Zn²⁺, Mg²⁺, Cu²⁺) have direct antinociceptive effects. Groups of eight Swiss mice were intraperitoneally (i.p) injected with incremental concentrations of Zn²⁺ sulfate (0.5, 2.0 mg/kg), Zn²⁺ citrate (0.125, 0.5 mg/kg), Mg²⁺ chloride (37.5, 75, 150 mg/kg), Cu²⁺ chloride (0.5, 1.0, 2.0 mg/kg), and Cu²⁺ sulfate (0.5, 1.0 mg/kg) or saline (control). Evaluations were made by hot plate (HP) and tail flick (TF) tests for central antinociceptive effect, writhing test (WT) for visceral antinociceptive effect, and activity cage (AC) test for spontaneous behavior. Zn²⁺ induced pain inhibition in HP/TF tests (up to 17%) and WT (up to 25%), with no significant differences among the salts used. Mg²⁺ salts induced pain inhibition for all performed tests (up to 85% in WT). Cu²⁺ salts showed antinociceptive effects for HP/TF (up to 28.6%) and WT (57.28%). Only Mg²⁺ and Cu²⁺ salts have displayed significant effects in AC (Mg²⁺ anxiolytic/depressant effect; Cu²⁺ anxiolytic effect). We interpret these data to mean that all tested trace elements induced antinociceptive effects in central and visceral pain tests. Our data indicate the potential use of these cheap adjuvants in pain therapy. © 2013 Wiley Periodicals, Inc.     

Influence of inorganic filler content on the radiopacity of dental resin cements

Digital radiography was used to measure the radiopacity of 18 resin cements to determine the influence of inorganic filler content on radiopacity. Four disk specimens (n=4) of each light-curing cement were digitally radiographed alongside an aluminum step wedge using an intraoral sensor (XIOS Plus, Sirona, Germany), and their mean gray value measured. Percentage of filler by weight was determined using an analytical combustion furnace. Data were statistically analyzed using one-way ANOVA and Tukey's test (α=0.05). All materials were more radiopaque than dentin and 12 materials were more radiopaque than enamel. Filler percentage ranged between 17.36 to 53.56 vol% and radiopacity between 1.02 to 3.40 mm Al. There were no statistically significant differences in inorganic filler percentage and radiopacity among the different shades of the same material (p>0.05), but the highest radiopacity was measured for the material which contained a higher percentage of filler.