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Apart from cytotoxicity, inhibitors of the COX-2 enzyme have demonstrated additional effects important for cancer treatment (such as radiosensitization of tumor cells and cell antimigratory effects); however, the relationship between the inhibition of other inflammation-related enzyme 5-LOX inhibitors and anticancer activity is still not well understood. In our study, the cytotoxicity of thirteen COX-2 and 5-LOX inhibitors previously presented by our group (1-13) was tested on three cancer cell lines (HCT 116, HT-29 and BxPC-3) and one healthy cell line (MRC-5). Compounds 3, 5, 6 and 7 showed moderate cytotoxicity, but good selectivity towards cancer cell lines. IC50 values were in the range of 22.99-51.66 µM (HCT 116 cell line), 8.63-41.20 µM (BxPC-3 cell line) and 24.78-81.60 µM (HT-29 cell line; compound 7 > 100 µM). In comparison to tested, commercially available COX-2 and 5-LOX inhibitors, both cytotoxicity and selectivity were increased. The addition of compounds 6 and 7 to irradiation treatment showed the most significant decrease in cell proliferation of the HT-29 cell line (p < 0.001). The antimigratory potential of the best dual COX-2 and 5-LOX inhibitors (compounds 1, 2, 3 and 5) was tested by a wound-healing assay using the SW620 cell line. Compounds 1 and 3 were singled out as compounds with the most potent effect (relative wound closure was 3.20% (24 h), 5,08% (48 h) for compound 1 and 3.86% (24 h), 7.68% (48 h) for compound 3). Considering all these results, compound 3 stood out as the compound with the most optimal biological activity, with the best dual COX-2 and 5-LOX inhibitory activity, good selectivity towards tested cancer cell lines, significant cell antimigratory potential and a lack of toxic effects at therapeutic doses.

Chemotherapy is one of the most effective approaches used to treat cancer, a leading cause of death worldwide.[1] Ruthenium-based complexes have emerged as a powerful alternative to state-of-the-art platinum chemotherapeutics due to their reduced toxicity, higher selectivity and anti-metastatic potential.[2] Here, we employ mass spectrometry and spectroscopic techniques to characterize a novel cis-[Ru(II)(dcbpy)2Cl2] complex and investigate its interaction with cellular lipids. In addition, the cytotoxic effect and mode of action on four standard cancer cell lines, namely melanoma A375, cervical cancer HeLa and pancreas ductal adenocarcinoma MIA PaCa-2 and PANC-1, were elucidated. Our results show a dose-dependent cytotoxicity of the complex on A375, MIA PaCa-2 and PANC-1. Further, the Ru(II) complex reduces the clonogenic potential caused by G1 arrest and increases ROS production in all tested cell lines. Lipid peroxidation is also observed in HeLa and MIA PaCa-2 cells, possibly indicating ferroptosis induction. Synchrotron radiation Fourier Transform Infrared Spectroscopy was applied to study the overall interaction of a Ru-complex with cellular biomolecules, and our results demonstrated a shift in bands in the lipid fingerprint area of most cell lines, confirming the lipid rearrangement process induced by the interaction with Ru(II) complex. The complex significantly increases the percentage of apoptotic A375, HeLa and MIA PaCa-2, while a significant decrease of p62 protein, a marker for autophagy induction, is detected solely in A375 cells. The wound healing assay revealed that the Ru(II) complex reduces the metastatic potential of HeLa and MIA PaCa-2 cells by reducing their migration ability. Overall, our results indicate the existence of a general mode of action of the tested Ru(II) complex that involves growth arrest and ROS production, as well as cancer cell line-specific effects. These involve dual role trough inducing programmed cell death - apoptosis and

Small, carbon-based nanoparticles, carbon dots (CDs), are investigated as photosensitizers (PSs) for photodynamic therapy of cancers. The results presented here show biochemical changes in cancer cells treated with surface-modified CDs, which are augmented by illumination, indicating a high potential of CDs as selective drug carriers and potent PSs. In addition, we will show that CDs produced from black carrot extract (biomass) can be exploited for image-guided therapy and be cytotoxic for cancer cells, thanks to their fluorescence properties. Finally, since biomass-fabricated CDs interact strongly with nucleic acids, they can be further developed for cell cycle sensing purposes.

In recent years researchers have developed new strategies to enhance the effectiveness of wound healing by combining nanoparticles and infra red (IR) light. For example, some studies have shown that nanoparticles can be used to enhance the absorption of near-infrared laser (NIR) light by tissues, leading to increased healing rates [1]. The influence of NIR light on proliferation, collagen production, and wound healing was tested on: keratocytes (HaCaT) and fibroblasts (MRC-5) cells that are used as model systems of human skin equivalents that comprise an epidermal and a dermal compartment of skin. Also, these cells were treated with carbon quantum dots/silver-based metal-organic framework composites (Ag-MoFs-NCDs and Ag-MoFs-SCDs), which previously showed high antibacterial activity [2], without and with laser light. Firstly, we have found the most convenient and effective CW laser intensity (16 mW/cm2) and illumination time (3 minutes), which is not too high and short enough to influence human cells' proliferation and metabolism positively. Additional chemical treatment with Ag-MoFs-NCDs and Ag-MoFs-SCDs results in a further increase in human cell viability. Our measurements showed that the proliferation index in laser-illuminated cells and cells treated with Ag-MoFs-SCDs was at the level of the untreated control. Furthermore, Ag-MoFs-SCDs treatment and laser illumination induced a mild, insignificant increase in cellular proliferation. On the other hand, Ag-MoFs-NCDs treatment led to a more pronounced, albeit not significant increase, in cellular proliferation, while Ag-MoFs-NCDs treatment combined with laser illumination significantly increased proliferation. Also, we have detected a mild change in collagen level estimated by hydroxyproline assay, which may indicate a positive outcome of combined laser illumination and treatment, taking into account that after 48 hours, a change in cell's response to the treatment could be noticed. Finally, based on migration ass

Carbon Dots (CDs) are biocompatible, fluorescent, water-soluble, and stable nanoparticles with a high potential to be used for vast biomedical applications [1,2]. We explore the application of CDs produced from natural sources, black carrots, as anti-cancer and imaging agents. These nanoparticles suppress cell growth of three different cancer cell lines, cervical (HeLa), pancreatic (PANC-1), and melanoma (A375) cell lines in vitro. However, the cytotoxic effect against A375 cells stands out, with only 20% of viable cells left after treatment (Fig.1(a)), antimetastatic potential, and a selectivity index higher than two, which indicates that the efficacy against melanoma cells is significantly greater than the toxicity against non-malignant cells (MRC-5). Furthermore, after the cellular uptake, green fluorescence was visible in the cytosol of A375 cells (Fig. 1 (b)). On the other hand, the DAPI stain for DNA was visible as a blue light in the cell nucleus. Moreover, cells with a higher intensity of green fluorescence in the nucleus, Fig. 1 (c) indicated with arrows, were the cells with condensed chromatin in the mitotic phase of the cell cycle (Fig. 1 (d) and (e)), which indicates that CDs interact with chromatin and that they could be used as a marker of cells mitosis and proliferation. In summary, we have demonstrated the great anti-cancer potential of black carrot CDs, for image-guided anti-cancer therapy of melanoma that can be used to recognize cell proliferation.

Treating cancer remains a major challenge, despite the development of many therapies and advances in general knowledge about the disease. The treatments commonly used are invasive and non-selective, leading to severe side effects and unsatisfactory long-term outcomes. Nevertheless, external stimuli activating therapeutic agents in the affected area can be more beneficial than these aggressive therapies. Photodynamic therapy (PDT) is a minimally invasive, selective treatment that uses photosensitizer (PS) to damage cancer cells. The PS is activated by light, triggering a series of processes that produce reactive oxygen species (ROS), ultimately leading to cancer cell death. Numerous types of nanomaterial possess the capability to act as PS, one of which is TiO2 [1]. Although nanostructured TiO2 is biocompatible in the absence of light, its valence band electrons can be stimulated only by ultraviolet (UV) light irradiation. Since the penetration of UV light into tissue is limited, for application in PDT, nanostructured TiO2 can be doped with heteroatoms like N or C to allow visible light responsiveness [2,3]. This work evaluated the PS properties of unmodified nanostructured TiO2 (spherical nanoparticles TiO2 NPs and prolate nanospheroids, TiO2 PNSs) and doped TiO2 (N- and C-TiO2 NPs). After the synthesis, the size of TiO2 was confirmed to be in the nanoscale range (5-104 nm) by transmission electron microscopy [3,4]. The doped TiO2 was found to absorb visible light, as demonstrated by UV-Vis spectroscopy and bandgap calculations. Additionally, hydroxyl radicals were detected in water suspensions of TiO2 PNSs by electron paramagnetic resonance (EPR) spectroscopy, both with and without UV light illumination [4]. However, this radical was observed only with blue light stimulation of the water suspensions of N- and C-TiO2 NPs [3]. Cell experiments further revealed the internalization process of nanostructured TiO2 within cells, their cytotoxicity profiles, and the differ

Background: Cancer nanomedicine is a rapidly developing fi eld that uses nanoparƟ cles (NPs) for the diagnosis and treatment of cancer. Currently, many nanomaterials with diff erent shapes, sizes, structures, and composiƟ ons have been invesƟ gated to produce eff ecƟ ve anƟ cancer NPs. The interest in the biomedical applicaƟ ons of bismuth-containing nanoparƟ cles, such as bismuth ferrite (BFO-NP) is a result of their promising properƟ es such as cost-effecƟ veness, chemical inertness, high stability, and simplicity of funcƟ onalizaƟ on. Material and Methods: A375 human melanoma and HeLa cervical carcinoma cells were used to study the anƟ tumor acƟ vity of BFO-NP. Clonogenicity of treated cells was analyzed by colony forming assay, while cell death was examined using fl ow cytometry. DCF-DA fl uorescent assay was applied to measure ROS producƟ on. Protein expression of p62 and TfR1 was detected by Western blot. Cell migraƟ on was analyzed using a wound scratch assay, while an SRB assay was used to assess cell adhesion. Results: BFO-NP (200 ng/L) signifi cantly reduced the clonogenicity of A375 and HeLa cells by 46 and 60%, respecƟ vely. Detected ROS producƟ on was increased considerably, especially for A375 melanoma cells, and amounted to 400%. The number of late apoptoƟ c and/or necroƟ c cells increased by 10-12%, compared to the control. Signifi cantly increased expression of autophagy-related protein p62 was observed in both cell lines aŌ er BFO-NP treatment. Ferroptosis-related transferrin receptor (TfR1) expression was slightly increased in treated A375 end HeLa cells (~14%). The noƟ ced increase in cell adhesion ranged from 20-30% followed by a decrease in cell migraƟ on. Conclusion:BFO-NP is a promising anƟ tumor agent with a signifi cant inhibitory eff ect on A375 and HeLa cell growth and metastaƟ c potenƟ al. Molecular mechanisms involved in these processes include ROS producƟ on and increased p62 expression. Reduced metastaƟ c potenƟ al resulted from the

Nanocomposite system formulated from surface-modified S-doped carbon dot (S-CD) nanoparticle with a potential metallodrug, palladium(II) complex, dichloro(1,2-diaminocyclohexane)palladium(II), [Pd(dach)Cl2] (Pd@S-CD), was investigated as a model system for the treatment of cervical carcinoma (HeLa) cells. To examine the intracellular biochemical effects induced by the Pd@S-CD, we used Synchrotron Radiation-based Fourier-transform infrared spectroscopy (SR FTIR). SR FTIR spectroscopy was employed to investigate the alterations in cellular components’ biochemical composition and secondary structure upon exposure to Pd@S-CD. Spectral analysis, complemented by statistical techniques, revealed changes in biomolecules, lipids, proteins, nucleic acids, and carbohydrates caused by the treatment with Pd@CDs. These results and the increased cytotoxicity of the system demonstrate its high anti-cervical cancer therapeutic potential.

Benefitni efekat crvenog vina na zdravlje ljudi od davnina je poznat. Među alkoholnim pićima crveno vino pokazuje najjači efekat protiv pojave oboljenja povezanih sa oksidativnim stresom, kao što su kardiovaskularna, neurodegenerativna oboljenja, dijabetes i kancer.1 U okviru ove studije ispitan je citotoksični efekat crvenog vina sorte merlo (komercijalno dostupno vino i vina dobijena od klonova VCR1 i VCR101) poreklom iz Crne Gore, berba 2011, u odnosu na ćelijsku liniju kancera pankreasa (PANC-1) i ćelijsku liniju melanoma (A375). Citotoksični efekat vina je određen SRB metodom kroz praćenje ćelijskog preživljavanja kancerskih ćelija 48 sati nakon tretmana sa tri različita zapreminska procenta analiziranih vina (5, 10 i 20%). Dobijeni rezultati su pokazali citotoksični efekat svih analiziranih vina na obe ćelijske linije. Ćelijsko preživljavanje nakon tretmana vinima se kretalo od 36 do 64%. Citotoksični efekat svih vina u odnosu na A375 ćelije je bio veći nego na PANC-1 ćelijsku liniju, što je od posebnog značaja ako se uzme u obzir rezistentnost melanomskih ćelija na postojeće terapeutske tretmane. Takođe, klonska sortna vina su pokazala veći citotoksični efekat na A375 ćelijsku liniju od komercijalno dostupnog vina. Dobijeni rezultati su ukazali da umereno konzumiranje crvenog vina sorte merlo sa specifičnim geografskim poreklom predstavlja dobar izvor bioaktivnih komponenata sa pozitivnim biološkim efektom, što je od posebne važnosti za klonska sortna vina.

Positive effects of moderate wine consumption in individuals with cardiovascular disease, hypertension, diabetes, and cancers have been shown in numerous epidemiological and clinical studies. This research examined the phenolic content of commercial and two clonal Merlot wines as well as their biological potential. The obtained results indicated that all analyzed samples were a good source of phenolic compounds. Cytotoxicity assay on melanoma (A375) and cervical (HeLa) cancer cell lines have shown that all analyzed wines inhibited the growth of human cancer cells in vitro with differing susceptibility among tested cell lines. Clonal wines in the volume ratio of 10 and 20% showed to be more efficient anti-proliferative agents than commercial wine regarding the A375 cells. This could be connected with higher total phenolic content in clonal wines. The effect of all analyzed samples on the A375 cells was greater compared to HeLa cell line.

The development of new types of nanoparticles has become the focus of biomedical research in recent years. Cerium oxide nanoparticles (CONP) have shown particularly promising results as antitumor agents with a selective effect on tumor and normal cells. On the other side, melanoma and pancreatic carcinoma are among the most aggressive types of cancer with no satisfactory therapy1,2 . Considering that, they represent important model systems for studying new treatment approaches. In this study, the antitumor potential of CONP (size below 10 nm) was studied on human A375 melanoma and PANC-1 pancreatic carcinoma cells. The obtained results indicated that analyzed CONP significantly inhibited clonogenic survival, with the number of colonies reduced on ~30% in A375 cells, while treated PANC-1 cells didn’t form colonies. Growth inhibition was followed by G 2 cell cycle arrest (9% for A375, 17% for PANC-1). Percent of apoptotic PANC-1 cells was 38%, whereas ROS production increased for 78%. CONP significantly reduced metastatic potential through the decrease in cell migration and the increase in cell adhesiveness (up to 30 and 40% for A375 and PANC-1 respectively). These findings emphasize the significant CONP antitumor potential, based on the increase in ROS production, as well as a reduction of A375 and PANC-1 metastatic potential.

Existing therapies for the treatment of pancreatic cancer are insufficiently effective and accompanied by a large number of side effects. Ruthenium complexes have shown promising antitumor properties in the previous studies 1,2 . Thus, in this investigation, anticancer effects of cis-dichlorobis (2,2'-bipyridyl-4,4'dicarboxylic acid)ruthenium(II) (Ru(II) complex) were evaluated using human pancreatic carcinoma cell lines MIA PaCa-2 and PANC-1 in vitro. Cell viability estimated with SRB assay showed significant antitumor activity of Ru(II) complex on MIA PaCa-2 (~55% of control) 48 and 72 h after treatment. On the other hand, PANC-1 cell viability was decreased only 72 h after treatment with the highest concentration of Ru(II) complex (~70% of control). Seven days after the treatment, analysis of cell survival using clonogenic assay showed a significant decrease in cell growth in both cell lines. Ru(II) complex also caused G 1 cell cycle arrest of ~13% in both cell lines. The highest percentage of apoptotic MIA PaCa-2 cells was obtained 48 h after treatment. In addition, the intracellular level of reactive oxygen species (ROS) was significantly increased, whereas cell migration was reduced in both cell lines. Summarized, Ru(II)complex demonstrates antitumor properties mediated by increased oxidative stress and also implies the antimetastatic potential, which deserves further study.

Platinum-based complexes represent the mainstay of treatment for various cancer types. However, their usage is often restricted by numerous side effects or intrinsic and acquired resistance. Therefore, significant research efforts have focused on developing therapeutics based on other transition metals, such as ruthenium 1,2. In this study, effects of transition metal complex, cis-dichlorobis (2,2ʹ-bipyridyl-4,4ʹ-dicarboxylic acid)ruthenium(II) (Ru(II) complex) were analyzed on A375 human melanoma and HeLa cell growth, adhesion ability andmigration. Cell viability assay indicated significant antitumor activity of Ru(II) complex on A375 (~60% of control) up to 72 h after treatment, but not on HeLa cells. However, analysis by clonogenic assay showed that growth of both cell lines was decreased 7 days after treatment. Growth inhibition was followed by G1 phase cell cycle arrest (5–10% G1 increase for A375 and 5–8% for HeLa cells compared to control). Moreover, Ru(II) complex increased adhesivity of A375 and HeLa cells by 11 and 16 % respectively and decreased cell migration, as shown by scratch assay. The obtained results indicate that the analyzed Ru(II) complex is a promising metallodrug, as itinduced growth inhibition of A375 and HeLa cells through induction of G1 arrest and decreased metastatic potential of these cells through the increase ofadhesivity and decrease of cell migration.

Melanoma is responsible for 75% of all deaths from skin cancer. Its lethality arises from its rapid progression, easy metastasis and drug-resistance as well. Red wine is a natural product rich in polyphenolic compounds with potent anticancer activities. It seems that in cancer cells these compounds behave as prooxidants initiating reactive oxygen species mediated cellular DNA breakage and consequent cell death. The aim of this study was to investigate prooxidative activity of red wine samples (Merlot variety, commercial as well as VCR1 and VCR101 clonal wines) in melanoma A375 cells, through measuring the relationship of reduced and oxidized form of glutathione (GSH/GSSG) and comparison with the GSH/GSSG ratio in control (untreated melanoma cells). The data obtained showed that tested red wine samples decrease GSH/GSSG ratio in A375 cells compared to control (4.6 ± 0), with the largest decrease noticed in treatment with VCR101 wine (0.66 ± 0.05).

Objects of the present study are improved fullerene C60 drug carrier properties trough encapsulation by microbial polysaccharides, levan (LEV), pullulan (PUL), and their hydrophobized cholesterol-derivatives (CHL and CHP), that show better interaction with cancer cells. The zeta potential, polydispersity index, and the diameter of particles were determined, and their cytotoxicity against three cancer cell lines were tested. Biochemical changes in HeLa cells are analyzed by synchrotron radiation (SR) FTIR spectro-microscopy combined with the principal component analysis (PCA). The most significant changes occur in HeLa cells treated with LEV-C60 and correspond to the changes in the protein region, i.e. Amide I band, and the changes in the structure of lipid bodies and membrane fluidity are evident. The highest cytotoxicity was also induced by LEV-C60. In HeLa cells, cytotoxicity could not be strictly associated with biochemical changes in lipids, proteins and nucleic acids, but these findings are significant contribution to the study of the mechanism of interaction of C60-based nanoparticles with cellular biomolecules. In conclusion, LEV, PUL, CHL, and CHP enhanced fullerene C60 potential to be used as target drug delivery system with the ability to induce specific intracellular changes in HeLa cancer cells.

Aim: Newly synthesized platinum(IV) complexes with ethylenediamine-N,N’-diacetate ligands (EDDA-type) (butyl-Pt and pentyl-Pt) were investigated against two cancer (A549 lung, and HTB 140 melanoma) and one non-cancerous (MRC-5 embryonic lung fibroblast) human cell lines. Materials and Methods: The effects of these agents were compared with those of cisplatin after 6-, 24- and 48-h treatment. Sulforhodamine-B (SRB) assay was performed to estimate the cytotoxic effect, while the inhibitory effect on cell proliferation was measured using 5-bromo-2,-deoxyuridine (BrdU) incorporation assay. Cell cycle analysis was performed by flow cytometry. Type of cell death induced by these agents was determined by electrophoretic analysis of DNA, flow cytometry and by western blot analysis of proteins involved in induction of apoptosis. The effects of gamma irradiation, alone and in combination with platinum-based compounds, were examined by clonogenic and SRB assays. Results: All examined platinum-based compounds had inhibitory and antiproliferative effects on A549 cells, but not on HTB140 and MRC-5 cells. Butyl-Pt, pentyl-Pt and cisplatin arrested the cell cycle in the S-phase and induced apoptotic cell death via regulation of expression of B-cell lymphoma 2 (BCL2) and BCL2-associated X (BAX) proteins. Platinum-based compounds increased the sensitivity of A549 cells to gamma irradiation. Butyl-Pt and pentyl-Pt showed better antitumour effects against A549 cells than did cisplatin, by interfering in cell proliferation and the cell cycle, and by triggering apoptosis. Conclusion: The effects of gamma irradiation on tumour cells may be amplified by pre-treatment of cells with platinum-based compounds.

Study of the interaction between nanoparticles and human cells is usually performed using customized biochemical assays that mostly offer measurements of a single quantity/property and use labels. Raman spectroscopy on the other hand offers integral insight into complex information on biomolecular composition and molecule conformation inside cells by measuring vibrational spectra from the entire cell [1]. Furthermore, it does not require dyes nor other labels and sample preparation is very simple, which reduces time consumation and possibility of cell damage during preparation. Cerium-dioxide (CeO2) nanoparticles are known for their controversial dual activity in numerous studied cancer cell lines: while protecting some cell types from oxidative damage, their cytotoxic effect in other cell lines is also reported [2, 3]. Here, effects of two types of CeO2 nanoparticles: uncoated and dextran-coated, were studied in HeLa cells, a cervical carcinoma derived cell line. Nanoparticle-treated cells were probed by routinely used biological assays for cell growth and viability, based on dying with Sulforhodamine B and Trypan Blue, respectively [3]. The tests have shown that the nanoparticles have more prominent effect on cell growth than on viability. In the light of this information Raman spectroscopy was employed in order to investigate the changes in biomolecular content of the cervical cancer cells after treatment with nanoparticles and find connection between these changes and the resulting cell status. Raman spectra of nanoparticletreated and control (untreated) cells were obtained using 532 nm laser line as an excitation probe. From each experimental group, at least 250 cell spectra were measured. Principal component analysis (PCA) covering the spectral regions (700-1800) cm-1 and (2800-3200) cm-1 has extracted the differences between vibrational spectra features of nanoparticle-treated and control cells, but also between spectra of cells treated with uncoated and coat

We studied the colloidal TiO2 nanoparticles as a carrier for controlled delivery of the ruthenium complex to the melanoma cell line. The system demonstrated slower complex release upon visible and increased release rate upon UV light illumination. Accordingly, the light-dependent cytotoxicity of the system was demonstrated on amelanotic melanoma cancer line. The cell death is enhanced by UV and reduced by red light in the presence of investigated nanocomposite system. Both components of the system may act as photosensitizers, by generating reactive oxygen species, which promote cell death. Thus, the system might act dually, as photodynamic therapeutic agent and as the light tunable system for metallo-drug delivery and it might be of interest for development of new more efficient drug delivery approaches by using a light as external stimulus. (C) 2017 Elsevier B.V. All rights reserved.

Lečenje kancera u današnje vreme uključuje primenu različitih terapija koje se uglavnom zasnivaju na primeni kompleksa prelaznih metala. Kada se razmatraju nove strategije u lečenju kancera, ističe se fotodinamička terapija. Ovaj vid terapije koristi svetlo kao aktivator leka ili nosača leka na mestu obolelog tkiva. Uzimajući u obzir navedeno, testirali smo potencijal prethodno sintetisanog fotoaktivnog nanokompozitnog sistema i njegovih komponenti za primenu u fotodinamičkoj terapiji. Pomenuti sistem se sastoji od nanočestica TiO2 kao nosača leka i kompleksa [Ru(II)(dcbpy)2Cl2], kao potencijalnog citostatika, vezanog za površinu nanočestica. Rutenijumov kompleks iskazuje svoje citotosično delovanje vezivanjem za ciljne biomolekule, dok dosač, TiO2, vezujući kompleks može smanjiti njegovu reaktivnost sa zdravim ćelijama dok ne dođe do tumorskog tkiva. Pomenuti nanokompozitni sistem i njegove komponente su testirane na ćelijama melanoma. Rezultati ukazuju na to da dejstvom UV svetla, nakon tretmana sa obe komponente sistema (kompleksom i TiO2) dolazi do značajnog smanjenja u broju tumorskih ćelija, dok je nanokompozitni sistem pokazao sinergistički efekat i dovodio do smanjenja broja tumorskih ćelija ispod 50%. Stoga, ovaj sistem ostavlja mesta za dalja izučavanja jer pruža mogućnost razvoja efiksnije terapije u kojoj bi doziranje leka bilo kontrolisanano svetlom., The treatment of cancer involves different therapies mainly based on transition metal complexes. When exploring new strategies for the treatment of cancer, one possibility is the use of photodynamic therapy. Photodynamic therapy utilizes light to activate a drug or a drug/carrier system when they are localized in the tumor tissue. Therefore, we have tested the potential of previously synthesized photoactive nanocomposite system and its components as a candidate for photodynamic therapy. The nanocomposite system consists of TiO2 nanoparticles as drug/carrier and ruthenium.complex [Ru(II)(dcbpy)2Cl2] bound to its surface as a potential antitumor drug. This Ru complex have been shown to exert cytotoxic effect through binding to target biomolecules, while TiO2 carrier can serve to minimize side reactivity in the bloodstream and healthy cells until it reaches tumor cells. The nanocomposite system and its components are successfully applied on melanoma tumor cell lines. It was demonstrated that the illumination by UV light, after treatment either with Ru-complex or TiO2 nanoparticles significantly decreases melanoma cell survival rate, while NCS shown synergistic effect and have reduced the number of melanoma cells under 50%. Hence, this system might be of interest from the standpoint of development of more efficient, light controllable drug delivery approaches.

Background and objectives: The main goal when treating malignancies with radiation is to deprive tumour cells of their reproductive potential. One approach is to induce tumour cell apoptosis. This study was conducted to evaluate the ability of carbon ions (C-12) to induce apoptosis and cell cycle arrest in human HTB140 melanoma cells. Methods: In this in vitro study, human melanoma HTB140 cells were irradiated with the 62 MeV/n carbon (C-12) ion beam, having two different linear energy transfer (LET) values: 197 and 382 keV/mu m. The dose range was 2 to 16 Gy. Cell viability was estimated by the sulforhodamine B assay seven days after irradiation. The cell cycle and apoptosis were evaluated 48 h after irradiation using flow cytometry. At the same time point, protein and gene expression of apoptotic regulators were estimated using the Western blot and q-PCR methods, respectively. Results: Cell viability experiments indicated strong anti-tumour effects of C-12 ions. The analysis of cell cycle showed that C-12 ions blocked HTB140 cells in G2 phase and induced the dose dependent increase of apoptosis. The maximum value of 21.8 per cent was attained after irradiation with LET of 197 keV/mu m at the dose level of 16 Gy. Pro-apoptotic effects of C-12 ions were confirmed by changes of key apoptotic molecules: the p53, Bax, Bcl-2, poly ADP ribose polymerase (PARP) as well as nuclear factor kappa B (NF kappa B). At the level of protein expression, the results indicated significant increases of p53, NF kappa B and Bax/Bcl-2 ratio and PARP cleavage. The Bax/Bcl-2 mRNA ratio was also increased, while no change was detected in the level of NF kappa B mRNA. Interpretation and conclusions: The present results indicated that anti-tumour effects of C-12 ions in human melanoma HTB140 cells were accomplished through induction of the mitochondrial apoptotic pathway as well as G2 arrest.

Ionizing radiation induces DNA double strand breaks (DSBs) that trigger phosphorylation of the histone protein H2AX (gamma H2AX). Immunofluorescent staining visualizes formation of gamma H2AX foci, allowing their quantification. This method, as opposed to Western blot assay and Flow cytometry, provides more accurate analysis, by showing exact position and intensity of fluorescent signal in each single cell. In practice there are problems in quantification of gamma H2AX. This paper is based on two issues: the determination of which technique should be applied concerning the radiation dose, and how to analyze fluorescent microscopy images obtained by different microscopes. HTB140 melanoma cells were exposed to gamma-rays, in the dose range from 1 to 16 Gy. Radiation effects on the DNA level were analyzed at different time intervals after irradiation by Western blot analysis and immunofluorescence microscopy. Immunochemically stained cells were visualized with two types of microscopes: AxioVision (Zeiss, Germany) microscope, comprising an ApoTome software, and AxioImagerA1 microscope (Zeiss, Germany). Obtained results show that the level of gamma H2AX is time and dose dependent. Immunofluorescence microscopy provided better detection of DSBs for lower irradiation doses, while Western blot analysis was more reliable for higher irradiation doses. AxioVision microscope containing ApoTome software was more suitable for the detection of gamma H2AX foci.