Human Epidermoid Carcinoma Research Articles

The otc gene of Streptococcus suis plays an important role in biofilm formation, adhesion, and virulence in a murine model

Streptococcus suis (S. suis) is an emerging zoonotic pathogen that can cause meningitis, arthritis, pneumonia, and sepsis. It poses a serious threat to the swine industry and public health worldwide. Ornithine carbamoyltransferase (OTC) is involved in the arginine deiminase system. OTC, which is a widely distributed enzyme in microorganisms, mammals, and higher plants, catalyzes the conversion of ornithine to citrulline. The present study showed that the otc gene plays an important role in the pathogenesis of S. suis infections. The ability of an otc-deficient mutant (Δotc) to form a biofilm was significantly reduced compared to the wild-type (WT) strain, as determined by crystal violet staining. Confocal laser scanning microscopy and scanning electron microscopy observations showed that the weakening of biofilm formation by the Δotc strain is related to a decrease in the extracellular matrix. In addition, compared to the WT strain, the Δotc strain had a reduced capacity to adhere to human laryngeal epidermoid carcinoma (HEp-2) cells compared to the WT strain. A real-time PCR analysis showed that the expression of adhesion-related genes by the Δotc strain was also lower than that of the WT strain. The virulence of the Δotc strain was significantly lower than that of the WT strain in a murine infection model. In addition, a histological analysis showed that the pathogenicity of the Δotc strain was lower than that of the WT strain, causing only slight inflammatory lesions in lung, liver, spleen, and kidney tissues. No significant differences were observed between the complemented mutant (CΔotc) and WT strains with respect to biofilm formation, adhesion, gene expression, and virulence. The present study provided evidence that the otc gene plays a pivotal role in the regulation of S. suis adhesion and biofilm formation. It also suggested that the otc gene is indirectly involved in the pathogenesis of S. suis serotype 2 infections.

Ethanolic extracts of South African plants, Buddleja saligna Willd. and Helichrysum odoratissimum (L.) Sweet, as multifunctional ingredients in sunscreen formulations

Exposure to solar ultraviolet (UV) radiation is a major contributing factor to the increasing number of skin cancer cases. Interest has grown to use plant extracts as natural ingredients in cosmetic formulations due to their photoprotective effect, antioxidant and anti-inflammatory activity, as well as other biological activities. The aim of this study was to evaluate the biological activity of two South African plant extracts, Helichrysum odoratissimum (L.) Sweet. and Buddleja saligna Willd., and to successfully incorporate these extracts into sunscreen formulations (o/w emulsions) due to their reported biological activity. Ethanolic extracts were prepared from the leaves and stems of H. odoratissimum and B. saligna and evaluated for their antioxidant activity, mutagenic potential and antiproliferative activity against human dermal fibroblasts (MRHF). The extracts were further characterized using gas chromatography-mass spectrometry (GC-MS). Thereafter, the extracts were incorporated into separate sunscreen formulations to evaluate the in vivo dermal irritancy potential, in vivo sun protection factor, in vitro UVA protection, photostability and long term stability of the formulation, to confirm that by incorporating the extracts, the stability or photoprotective effect of the sunscreen formulation was not reduced and that these formulation were considered safe for topical application. Three separate sunscreen formulations were prepared; the base sunscreen formulation (formulation A), the base sunscreen formulation containing B. saligna (formulation B) and H. odoratissimum (formulation C) respectively. Both extracts showed significant radical scavenging activity using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay with a fifty percent inhibitory concentration (IC50) of 5.13 ± 0.07 and 8.16 ± 0.34 µg/mL for H. odoratissimum and B. saligna respectively. No mutagenic activity was observed when the extracts were tested in the Ames assay using Salmonella typhimurium (TA98 and TA100). The PrestoBlue® cell viability assay was used to determine the antiproliferative activity of the extracts against MRHF cells, both extracts showed an IC50 value >90 µg/mL. Photoprotective activity was measured using in vivo sun protection factor (SPF) test method according to South African (SANS 1557) and International (ISO 24444) standards as well as the in vitro UVA SPF testing procedure (ISO 24443). The SPF results showed that the formulations had broad-spectrum UV protection with SPF values of 15.8±0.41, 16.1±0.66 and 16.0±0.49 and UVAPF values of 6.47±0.06, 6.45±0.06 and 6.47±0.07 for formulation A, B and C respectively. Furthermore, the formulations remained stable under normal and extreme conditions and the plant extracts did not affect the photoprotective effect of the sunscreen formulations and contributed towards the formulations stability. Additionally, each of the formulations were photostable, whereas the formulations with the addition of the extracts showed an incremental increase in photostability when compared to the base formulation. Both these extracts have been previously reported to display antiproliferative activity against skin cancer cell lines (previously published data), with an IC50 value of 31.80 ± 0.35 µg/mL (human malignant melanoma, UCT-MEL-1) for B. saligna and IC50 values of 15.50 ± 0.20 (human epidermoid carcinoma, A431) and 55.50 ± 6.60 µg/mL (human malignant melanoma, A375) for H. odoratissimum, contributing towards the medicinal benefit of using these extracts as ingredients into sunscreen formulations. Therefore, Helichrysum odoratissimum and Buddleja saligna could be considered as useful and viable additives to sunscreen formulations due to their reported biological activity.

Anti-skin cancer activity of Alpinia calcarata ZnO nanoparticles: Characterization and potential antimicrobial effects

The skin, being the human organ most exposed to the polluted environment, is susceptible to the worst effects of different biological, physical and chemical hazards that give rise to infections and skin cancer, among others. Besides, conventional treatments often have a number of undesirable side effects so nanotechnology is emerging as an ideal alternative. In particular, green synthesized nanoparticles (NPs) ensure proper permeation and sustained release with great physicochemical stability. On the other hand, rhizomes of Alpinia calcarata are endowed with rich medicinal properties which along with nano form of zinc oxide can produce synergistic effects in treatments of skin based ailments. In the current research work, zinc oxide NPs (ZnONPs) were synthesized from the extract of rhizomes of Alpinia calcarata and their biotherapeutic properties were characterized. These ZnONPs were widely characterized using UV–Vis spectrophotometer, SEM, EDAX, PXRD and FTIR to assess their morphology and metallic nature. The presence of ZnONPs was confirmed by UV–Vis spectrophotometry since it exhibits a peak at 350 nm. Morphology of the ZnONPs was irregular and their diameter varies in the range 70–80 nm as examined through SEM. Our results demonstrate that ZnONPs reveal a great biocidal activity against E. coli and C. albicans, among other microorganisms. Treatment with ZnONPs showed a dose-dependent reduction in human epidermoid carcinoma A431 cell viability by MTT assay and distinct morphological changes related with apoptosis by AO/EB staining. According to our results, A. calcarata ZnONPs can serve as a multifaceted agent for various skin based ailments.

Zinc Oxide Nanoparticle Induces Apoptosis in Human Epidermoid Carcinoma Cells Through Reactive Oxygen Species and DNA Degradation.

Zinc oxide nanoparticles (ZnO-NPs) are used immensely in technology and medicine, but very less is known about toxicity mechanism to human epidermal cells. The objective of this study was to evaluate possible anticancer properties of ZnO-NPs on human epidermoid carcinoma cells using MTT assay, measurement of reactive oxygen species, DNA fragmentation, and nuclear condensation. ZnO-NPs were synthesized by sol-gel method using zinc acetate dihydrate, ethylene glycol, and 2-propyl alcohol. Numerous characterization techniques such as UV-visible spectroscopy, X-ray powder diffraction, transmission electron microscopy, and dynamic light scattering spectroscopy were used to confirm synthesis, purity, optical, and surface characteristics, size, shape, and distribution of ZnO-NPs. Our finding showed that ZnO-NPs considerably decreased cell viability of human epidermoid carcinoma A431 cells with a parallel increase in nuclear condensation and DNA fragmentation in a dose dependent manner. Moreover, real time PCR expression study showed that treatment of human epidermoid carcinoma cells with ZnO-NPs trigger increased expression of tumor suppressor gene p53, bax, and caspase-3 while downregulate antiapoptotic gene bcl-2. Thus ZnO-NPs induce apoptosis in A431 cells through DNA degradation and generation of reactive oxygen species via p53, bax/bcl-2, and caspase pathways.

Topical nanostructured lipid carrier gel of quercetin and resveratrol: Formulation, optimization, in vitro and ex vivo study for the treatment of skin cancer.

The objective of this investigation was to develop dual drug-loaded nanostructured lipid carrier (NLC) gel of quercetin and resveratrol to enhance their disposition in dermal and epidermal layers. The optimization of the lipidic phase, i.e., liquid lipid and solid lipid was done on the basis of the solubility of quercetin & resveratrol in lipids in the preformulation stage. NLC formulation was optimized by central composite rotatable design (CCRD). The NLC formulation contained lipid binary mixture (1.0% w/w) and Cremophor RH40 (5% w/v) as a surfactant and had a particle size of 191nm±5.20, polydispersity index (PDI) of 0.33±0.01, zeta potential (ZP) of -10.00mV±0.30 and entrapment efficiency (EE) of 92.85±0.25% (quercetin), 89.05±0.18% (resveratrol) respectively. The flux and permeability coefficient of quercetin and resveratrol from NLC gel were found to be 14.09µg/cm2/h, 3.70µg/cm2/h and 7.21×10-2cm/h, 4.69×10-2cm/h respectively. Dermatokinetic studies revealed that there was a significant increase in the CSkin max and AUC0-8h in skin treated with NLC gel as compared to skin treated with conventional gel, which was prepared using carbopol 934 (1.5% w/w). Further, all claims of dermatokinetic studies were proved by confocal microscopic (CLMS) studies, which revealed that the disposition of combinatorial NLC gel was higher (~3 folds) as compared to the conventional gel. Furthermore, skin treated with NLC gel and untreated skin were analysed by FTIR and DSC spectra to understand the permeation dynamics of NLC gel. The cytotoxic effect of combinatorial NLC gel and the conventional gel assessed in human epidermoid carcinoma (A431) cell line by MTT assay, revealed that IC50 of NLC gel and the conventional gel was 86.50µM and 123.64µM respectively. Thus, these results disclosed that NLC gel could be used as a potential carrier for the delivery of quercetin & resveratrol into deeper layers of the skin and can serve as a promising formulation for treatment of skin cancer.

The Role of miR-143, miR-145, and miR-590 in Expression Levels of CD44 and Vascular Endothelial Cadherin in Oral Squamous Cell Carcinoma

Background: Oral squamous cell carcinoma (OSCC) is the most common head and neck cancer with a continuing rise of incidence in developing countries. Despite the improvement in the clinical outcome of OSCC, the overall 5-year survival rate of patients is still disappointing. MicroRNAs (miRNAs) regulate gene expression in the post-transcriptional stage by targeting mRNAs. Advances in knowledge of the pathogenesis and molecular events lead to the improvement of OSCC treatments. Materials and Methods: Human oral epidermoid carcinoma cells (KB), HGF1, and HEK 293T cell line were cultured. Lentiviral vectors were constructed and Western Blot analysis was performed. Then, immunocytochemistry staining was performed by using CD44 and vascular endothelial cadherin (VE-Cadherin) antibodies. All data were analyzed using the REST 2009 software. P values of ≤0.05 were considered statistically significant. Results: MiR-143, miR-145, and miR 590 were down-regulated in the oral cancer cell line. Following transfection of Lv miR 143, Lv-miR-145, and Lv-miR-590, the expression of CD44 was markedly decreased in KB cells. In addition, transfection of miR-143 and miR- 590 mimics into the oral cancer cell line significantly decreased the expresion level of VE-Cadherin; however, transfection of miR-145 mimic had no significant effect on the expression of VE-Cadherin. Western blot analysis and immunocytochemistry staining confirmed the results. Conclusion: This study revealed that miR-143, miR-145, and miR-590 negatively regulate CD44 and VE-Cadherin (except miR-145) expression, which might play a crucial role in the induction of cancer stem cells proliferation and angiogenesis in OSCC cells. The knowledge about the involved factors may provide new insights for the clinical use of miR- 143, mR-145, and miR-590 in the treatment of patients with OSCC.

Cisplatin treatment modulates Annexin A1 and inhibitor of differentiation to DNA 1 expression in cervical cancer cells

Cisplatin (Cis) is a choice chemotherapy approach to cervical cancer by inducing DNA adducts and subsequent apoptosis. We have investigated the effects of Cis on Annexin A1 (ANXA1) and inhibitor of DNA binding 1 (ID1) proteins expression to elucidate further mechanisms of Cis actions. Human cervical tissue samples from twenty-four patients, with Cervical Intraepithelial Neoplasia (CIN, stage I, II and III), were evaluated to quantified ANXA1 and ID1 expressions. In vitro, human epidermoid carcinoma of the cervix (SiHa cell line) were treated with Annexin A1 peptide (ANXA12−26), Cis or Cis + ANXA12−26 to evaluate cell proliferation and migration, cytotoxicity of treatments as well as ANXA1 and ID1 modulations by mRNA and protein expression. Our findings showed expression of ID1 and ANXA1 proteins in tissue samples from Cervical Intraepithelial Neoplasia (CIN) patients, with intense immunological identification of ID1 in the CIN III stage. In SiHa cells, treatments with Cis alone or Cis + ANXA12−26, increase mRNA expressions of the ANXA1 and reduced the ID1. In agreement, Cis + ANXA12−26 enhanced ANXA1 protein expression and Cis or Cis + ANXA12−26 abolished ID1 protein expression. Cell proliferation was reduced after treatment with ANXA12−26 peptide and more significant after Cis or Cis + ANXA12−26 treatments. These two last treatments reduced cell viability, by inducing late apoptosis, and impaired cell migration. Together, our data highlight endogenous ANXA1 is involved in Cis therapy for cervical cancer.

Synthesis, Cytotoxic Activity and Molecular Docking Studies of New Condensed Thieno[2,3-d]Pyrimidines as Antitumor Agents

A series of twenty molecules based on thienopyrimidine nucleus fused with 5-, 6-, and 7-membered lactam rings as well as triazole were synthesized via 4-step reactions. The target compounds were obtained in good yield and confirmed for their structural integrity. The cytotoxic activity against A431 human epidermoid carcinoma and H9c2 rat cardiomyocyte cells were studied by MTT assay. Most of the synthesized compounds except 8a showed 85% cell death in both human A341 and rat H9c2 cell lines, indicating potential anticancer activity of these compounds. Interestingly, compound 8a showed a reversal in trend by enhancing the growth of both A341 and H9c2 cells.

Oncolytic activity of a new group of non-classified rotavirus of Reoviridae family on human epidermoid carcinoma A431 growth.

e15272 Background: Significant increase of malignant tumors` incidence all over the world in spite of vast arsenal of anticancer drugs and their combinations challenges the oncologists to develop new approaches to tumor treatment. Application of oncolytic virus is considered to be rather promising. Here we studied two strains of the new group of non-classified rotavirus of Reoviridae family (RVK): 100 and 228 ( http://jbks.ru/archive/issue-10/article-6 ). RVK are attenuated non-pathogenic virus growing on pig embryo kidney cell culture with concentration 5·109 per 1 ml. The aim of the study was to assess the effect of two RVK strains (100 and 228) on the growth of human epidermoid carcinoma A431 transplanted to nude mice in vivo. Methods: Mice Balb/cNude 22-24 g body weight (n = 13) were injected with 4х106 cells of human epidermoid carcinoma A431 subcutaneously in SPF-area of vivarium. After formation of palpable tumors (1 week after transplantation) administration of alive RVK strains 100 and 228 was performed. The 3rd group was the control one and received 0.85% NaCl. The injections were performed once a week 0.3 ml. Tumor growth rate and its` volume were measured. 1 week after the completion of the course mice were sacrificed, tumors were weighted and their morphology was studied. Results: In mice injected with RVK tumor growth inhibition developed in early date after the injection and was statistically significant within 30 - 40 days of monitoring only after the administration of strain 100. In these animals tumor weight was 2.6 times lower than in control mice (6.3±3.0 и 16.6±2.3 g respectively, р < 0.05), in mice having received strain 228 it was 1.7 times less tan in controls. Layers of high-grade sqamous carcinoma cells without keratinization with inflammatory and necrotic foci were observed in tumors of control mice while in tumors inhibited by RVK dystrophic changes and fragmentation, inflammation in places severe and necrotic foci were seen. Implication of the strain 100 resulted in 2-3 time reduction of tumor size, decrease of tumor cells` layers, mononuclear infiltration. Conclusions: In the in vivo model of human epidermoid carcinoma A431 transplanted to nude mice the inhibition of tumor growth under RVK administration was established. Our results confirm the oncolytic activity in RVK, particularly in strain 100.

Evaluation of the Molluscicidal, Artemicidal and Cytotoxic Activities of the Lectin from Opuntia ficus-indica Cladodes (OfiL)

Background: Biomphalaria glabrata is an intermediate host for the larvae of Schistosoma mansoni, which is distributed widely in the tropics. B. glabrata control is important to minimize the spread of schistosomiasis and natural compounds have been sought for use against this disease. The Artemia salina bench-top bioassay has been used to investigate the ecotoxicity of many natural compounds, and its results also correlate well with the in vitro cytotoxicity of natural compounds to tumor cells.
 Aims: To evaluate deleterious effects of the Opuntia ficus-indica lectin (OfiL) on B. glabrata, A. salina and human cancer cell lines.
 Methods: OfiL was isolated following a previously established protocol. The effects of OfiL on B. glabrata were investigated by determining survival of adults as well as development and hatching of embryos. The concentration required to kill 50% (LC50) of A. salina nauplii was determined. The cytotoxicity was determined using the human cell lines Hep-2 (human larynx epidermoid carcinoma), NCI-H292 (human lung mucoepidermoid carcinoma) and K562 (chronic myelocytic leukemia).
 Results: The development of most embryos (92.5–97.5%) treated with 1, 10 and 100 µg/mL of OfiL was found to be delayed, and dead (2.2–3.3%) and malformed (0.3–5.2%) embryos were also observed. OfiL did not kill B. glabrata adults, but a high percentage (30–45%) of the embryos generated by snails incubated with the lectin exhibited malformations. OfiL exhibited toxicity against A. salina (LC50: 61.02 µg/mL) but did not display cytotoxicity against the tumor cell lines evaluated.
 Conclusion: In conclusion, this study showed that OfiL can be a tool for schistosomiasis control that acts by impairing the viability of B. glabrata eggs and the fecundity of adult snails.

Impaired actin filaments decrease cisplatin sensitivity via dysfunction of volume-sensitive Cl- channels in human epidermoid carcinoma cells.

Cisplatin is a widely used platinum-based anticancer drug in the chemotherapy of numerous human cancers. However, cancer cells acquire resistance to cisplatin. So far, functional loss of volume-sensitive outwardly rectifying (VSOR) Cl- channels has been reported to contribute to cisplatin resistance of cancer cells. Here, we analyzed protein expression patterns of human epidermoid carcinoma KB cells and its cisplatin-resistant KCP-4 cells. Intriguingly, KB cells exhibited higher β-actin expression and clearer actin filaments than KCP-4 cells. The β-actin knockdown in KB cells decreased VSOR Cl- currents and inhibited the regulatory volume decrease (RVD) process after cell swelling. Consistently, KB cells treated with cytochalasin D, which depolymerizes actin filaments, showed smaller VSOR Cl- currents and slower RVD. Cytochalasin D also inhibited cisplatin-triggered apoptosis in KB cells. These results suggest that the disruption of actin filaments cause the dysfunction of VSOR Cl- channels, which elicits resistance to cisplatin in human epidermoid carcinoma cells.

Synthesis and Cytotoxicity Studies of Stilbene Long-Chain Fatty Acid Conjugates.

A series of 16 conjugates of the tubulin polymerization inhibitor combretastatin A4 (CA-4) and other functionally related stilbene with four 18-carbon fatty acids, namely, stearic, oleic, linoleic, and linolenic acids, have been synthesized in good yields. These new derivatives have been evaluated against the KB-3-1 (human epidermoid carcinoma), NCI-H460 (human lung cancer), HEK293 (human embryonic kidney), and MCF-7 (human breast adenocarcinoma) cell lines for antiproliferative activity, with the exhibited cytotoxic activities comparable with those of CA-4 and colchicine. Compounds 22 and 23, CA-4 conjugates of linoleic and linolenic acids, respectively, were determined to have exhibited the most active in vitro assays, with compound 23 exhibiting very similar activity to the parent compound against the NCI-H460 cell line. Our studies further delineated the structurally required Z-geometry of the stilbene moiety and that conjugation of the less active E-stilbenes with the most active fatty acid had minimal or no improvement in their respective activities.

Antiproliferative and apoptotic effects of anti-human HB-EGF neutralizing polyclonal antibodies in vitro.

To obtain HB-EGF neutralizing polyclonal antibodies and test their anti-proliferative properties in vitro. Lab rabbits and mice were used for immunization with recombinant HB-EGF. The effect of generated polyclonal antibodies on viability and apoptosis of human epidermoid carcinoma derived A431cell line was assessed using MTT and Annexin V-propidium iodide assays. Rabbit polyclonal anti-HB-EGF serum could block binding of soluble HB-EGF to epidermal growth factor receptor/human epidermal growth factor receptor. Also, anti-HB-EGF antibodies could bind to surface of A431cells which express abnormally high levels of membrane bound proHB-EGF and its receptor. It has been shown that immune serum with polyclonal antibodies against HB-EGF was able to block the mitogenic activation of the cells with HB-EGF and cause apoptotic cell death. Inhibition of HB-EGF activity with neutralizing polyclonal antibodies can effectively inhibit mitogenic activation and cause apoptosis of cancer cells with significant epidermal growth factor receptor overexpression.

Targeted Delivery of 111In Into the Nuclei of EGFR Overexpressing Cells via Modular Nanotransporters With Anti-EGFR Affibody.

Since cell nucleus is one of the most vulnerable compartments, the maximum therapeutic effect from a variety of locally acting agents, such as photosensitizers, alfa-emitters, Auger electron emitters, will be expected when they get there. Therefore, the targeted delivery of these agents into the nuclei of target tumor cells is necessary for their anticancer effects and minimization of side effects. Modular nanotransporters (MNT) are artificial polypeptides comprising several predefined modules that recognize target cell, launching their subsequent internalization, escape from endosomes, and transport the drug load to the nucleus. This technology significantly enhances the cytotoxicity of locally acting drugs in vitro and in vivo. Epidermal growth factor receptors (EGFR) are useful molecular targets as they are overexpressed in glioblastoma, head-and-neck cancer, bladder cancer, and other malignancies. Here, we examined the possibility of using internalizable anti-EGFR affibody as an EGFR-targeting MNT module for drug transport into the cancer cell nuclei. It binds to both murine and human EGFR facilitating preclinical studies. We showed that MNT with affibody on the N-terminus (MNTN-affibody) effectively delivered the Auger electron emitter 111In to target cell nuclei and had pronounced cytotoxic efficacy against EGFR-overexpressing human A431 epidermoid carcinoma cells. Using EGFR-expressing human adenocarcinoma MCF-7 cells, we demonstrated that in contrast to MNT with N-terminal epidermal growth factor (EGF), MNTN-affibody and MNT with EGF on the C-terminus did not stimulate cancer cell proliferation.

Low-frequency mechanical vibration induces apoptosis of A431 epidermoid carcinoma cells.

Cancer research is increasingly focused on discovering strategies to induce cancer cell apoptosis without affecting surrounding normal cells. One potential biocompatible method is mechanical vibration, which has been developed as part of the emerging field of mechanomedicine. Previous studies of mechanical vibration have employed high‐frequency vibration, which damages healthy cells. In this study, we examined the effects of brief (1 h) low‐frequency (20 Hz) mechanical vibration on glucose consumption and survival (apoptosis, necrosis, HMGB1 release) of the human epidermoid carcinoma cell line A431. We found that apoptosis, but not necrosis, was significantly increased at 48 h after mechanical vibration compared with cells maintained in static culture. In keeping with this, extracellular release of HMGB1, a necrosis marker, was lower in cultures of A431 cells subjected to mechanical vibration compared with control cells. Glucose consumption was increased in the first 24 h after mechanical vibration but returned to control levels before the onset of apoptosis. Although the precise intracellular mechanisms by which low‐frequency mechanical vibration triggers apoptosis of A431 cells is unknown, these results suggest a possible role for metabolic pathways. Mechanical vibration may thus represent a novel application of mechanomedicine to cancer therapy.

Escherichia coli-based synthesis of cadmium sulfide nanoparticles, characterization, antimicrobial and cytotoxicity studies.

The present research describes the synthesis of cadmium sulfide (CdS) nanoparticles from Escherichia coli under the influence of bacterial enzyme sulphate reductase and study on their cytotoxicity for applications in cancer therapy. Escherichia coli cells were used to synthesize CdS nanoparticles under different concentrations of cadmium chloride and sodium sulfide. The morphology of the nanoparticles was analysed using scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDX) was used for elemental analysis of nanoparticles. Fourier-transform infrared spectroscopy analysis (FTIR) was performed to assess the functional groups of the nanoparticles. Crystalline nature of nanoparticles was assessed using powder X-ray diffraction (XRD). Antibacterial studies of CdS nanoparticles were carried out on foodborne pathogens and cytotoxicity studies were carried out on Mus musculus skin melanoma (B16F10) and human epidermoid carcinoma (A431) cell lines. CdS nanoparticle showed more cytotoxic effect on cancer cells compared with standard 5-aminolevulinic acid (5-ALA). The Escherichia coli-synthesized CdS nanoparticles showed highest zone of inhibition in the ratio 4:1 of cadmium chloride and sodium sulfide on all tested bacterial strains. The nanoparticles were also tested for haemolytic activity on RBC cells, which exhibited lower cytotoxicity than sodium dodecyl sulphate which was used as positive control. The cytotoxicity of CdS nanoparticles assessed on A431 cells showed an inhibition of 81.53% at 100μM concentration while the cytotoxicity assessed on B16F10 cells showed an inhibition of 75.71% at 200μM concentration which was much efficient than 5-ALA which showed an inhibition of 31.95% at a concentration against B16F10 cells and 33.45% against A431 cells at a concentration of 1mM. Cadmium sulfide nanoparticles were thus found to be highly toxic on cancer cells compare with standard anticancerous drug 5-ALA.

Pegylated carbon nitride nanosheets for enhanced reactive oxygen species generation and photodynamic therapy under hypoxic conditions

The application of photodynamic therapy (PDT) is of ever-increasing importance in the treatment of malignant tumors; however, there are several major constraints that make it impossible to achieve optimal therapeutic effects. Our objective is to develop a novel photosensitizing drug for skin cancer. In the experiment, we fabricated four-arm-poly ethylene glycol modified amino-rich graphite phase carbon nitride nanosheets (AGCN-PEG), which have good stability in physiological solution and show selective accumulation in tumor cells. Under hypoxic conditions, the AGCN-PEG induced PDT can effectively inhibit growth on A431 human epidermoid carcinoma cells in vivo and in vitro. What's more, after being combined with TMPyP4, the therapeutic effect of AGCN-PEG was greatly improved.

The PI3K subunits, P110\u03b1 and P110\u03b2 are potential targets for overcoming P-gp and BCRP-mediated MDR in cancer

BackgroundPI3K/AKT is a vital signaling pathway in humans. Recently, several PI3K/AKT inhibitors were reported to have the ability to reverse cancer multidrug resistance (MDR); however, specific targets in the PI3K/AKT pathways and the mechanisms associated with MDR have not been found because many of the inhibitors have multiple targets within a large candidate protein pool. AKT activation is one presumed mechanism by which MDR develops during cancer treatment.MethodsThe effects of inhibiting PI3K 110α and 110β by BAY-1082439 treatment and CRISPR/Cas9 knockout were examined to determine the possible functions of BAY-1082439 and the roles of PI3K 110α and 110β in the reversal of MDR that is mediated by the downregulation of P-gp and BCRP. Inhibition of AKT with GSK-2110183 showed that the downregulation of P-gp and BCRP is independent of generalized AKT inactivation. Immunofluorescence, immunoprecipitation, MTT, flow cytometry and JC-1 staining analyses were conducted to study the reversal of MDR that is mediated by P-gp and BCRP in cancer cells. An ATPase assay and a structural analysis were also used to analyze the potential mechanisms by which BAY-1082439 specifically targets PI3K 110α and 110β and nonspecifically influences P-gp and BCRP.ResultsBy inhibiting the activation of the PI3K 110α and 110β catalytic subunits through both the administration of BAY-1082439 and the CRISPR/Cas9 deletion of Pik3ca and Pik3cb, the ATP-binding cassette transporters P-gp/ABCB1 and BCRP/ABCG2 were downregulated, thereby reestablishing the drug sensitivity of human epidermoid carcinoma and non-small cell lung cancer (NSCLC) MDR cells. Inhibition of AKT did not reverse the MDR mediated by P-gp or BCRP. The ABC family proteins and AKT may play MDR-enhancing roles independently.ConclusionsThe reversal of the dual functions of ABC-transporter-mediated and AKT-activation-enhanced MDR through the inhibition or knockout of PI3K 110α or 110β promises to improve current strategies based on combined drug treatments to overcome MDR challenges.

Evaluation of In vitro Cytotoxic Activity of Methanolic leaves Extract of Chloris barbata

The present research involves cytotoxic evaluation of Chloris barbata leaves by employing various in vitro models. Extraction of active constituents of the leaves of Chloris barbata was done by using soxhlet apparatus with polar and non-polar solvents. Further the extract was subjected to phytochemical screening and the methanolic extract of the leaf reveals the presence of flavonoids, glycosides, alkaloids, saponins and triterpinoids. Invitro cytotoxic efficacy of the Chloris barbata leaf methanolic extract was assessed with Human epidermoid larynx carcinoma (Hep 2), African green monkey kidney normal (Vero), Dalton's Ascitic Lymphoma (DAL) cell lines by using tryphan blue dye exclusion test, MTT, Sulphorodamine B (SRB) and Neutral Red Uptake (NRU)assay. In Tryphan blue dye exclusion assay it was found that there is an increase in percentage of dead vero (92±0.49), DAL (76±0.47) and Hep-2 cells (82±0.36) with chloris barbata leaf extract 200μg/mlwhen compared to control. Standard 5-Fluorouracil (5μg/ml) has shown an increase in percentage of dead vero, DAL and Hep-2 cells to an extent of 100±1.49, 94±1.35 and 99±0.21 when compared to control. In MTT assay the percentage viability of DAL, Hep-2 and vero cells has been significantly reduced by chloris barbata leaf extract at 200μg/ml (DAL 0.941±0.95, Hep-2 0.714±1.62 and vero cells 1.364±1.48) when compared to control (% viability is 100). In SRB assay, the percentage growth inhibition of cell lines by methanolic leaf extract was found to be 56.362 (vero cells), 69.128 (DAL cells) and 61.354 (Hep-2 cells) when compared to control group. NRU assay has shown the IC50 values 24.11±3.82 (vero), 47.16±0.14 (DAL) and 31.26±2.15 (Hep-2 cells) with leaf extract when compared to control. The above results signify that the methanolic extract of Chloris barbata leaves have remarkable cytotoxic potential. Further isolation of active constituents and in vivo cytotoxic studies has to be carried out to establish the exact mechanism involved in anticancer activity of Chloris barbata leaf extract.

Pharmacological inhibition of mTORC1 increases CCKBR-specific tumor uptake of radiolabeled minigastrin analogue [177Lu]Lu-PP-F11N.

Rationale: A high tumor-to-healthy-tissue uptake ratio of radiolabeled ligands is an essential prerequisite for safe and effective peptide receptor radionuclide therapy (PRRT). In the present study, we searched for novel opportunities to increase tumor-specific uptake of the radiolabeled minigastrin analogue [177Lu]Lu-DOTA-(DGlu)6-Ala-Tyr-Gly-Trp-Nle-Asp-Phe-NH2 ([177Lu]Lu-PP-F11N), that targets the cholecystokinin B receptor (CCKBR) in human cancers.Methods: A kinase inhibitor library screen followed by proliferation and internalization assays were employed to identify compounds which can increase uptake of [177Lu]Lu-PP-F11N in CCKBR-transfected human epidermoid carcinoma A431 cells and natural CCKBR-expressing rat pancreatic acinar AR42J cells. Western blot (WB) analysis verified the inhibition of the signaling pathways and the CCKBR level, whereas the cell-based assay analyzed arrestin recruitment. Biodistribution and SPECT imaging of the A431/CCKBR xenograft mouse model as well as histological analysis of the dissected tumors were used for in vivo validation.Results: Our screen identified the inhibitors of mammalian target of rapamycin complex 1 (mTORC1), which increased cell uptake of [177Lu]Lu-PP-F11N. Pharmacological mTORC1 inhibition by RAD001 and metformin increased internalization of [177Lu]Lu-PP-F11N in A431/CCKBR and in AR42J cells. Analysis of protein lysates from RAD001-treated cells revealed increased levels of CCKBR (2.2-fold) and inhibition of S6 phosphorylation. PP-F11N induced recruitment of β-arrestin1/2 and ERK1/2 phosphorylation. In A431/CCKBR-tumor bearing nude mice, 3 or 5 days of RAD001 pretreatment significantly enhanced tumor-specific uptake of [177Lu]Lu-PP-F11N (ratio [RAD001/Control] of 1.56 or 1.79, respectively), whereas metformin treatment did not show a significant difference. Quantification of SPECT/CT images confirmed higher uptake of [177Lu]Lu-PP-F11N in RAD001-treated tumors with ratios [RAD001/Control] of average and maximum concentration reaching 3.11 and 3.17, respectively. HE staining and IHC of RAD001-treated tumors showed a significant increase in necrosis (1.4% control vs.10.6% of necrotic area) and the reduction of proliferative (80% control vs. 61% of Ki67 positive cells) and mitotically active cells (1.08% control vs. 0.75% of mitotic figures). No significant difference in the tumor vascularization was observed after five-day RAD001 or metformin treatment.Conclusions: Our data demonstrates, that increased CCKBR protein level by RAD001 pretreatment has the potential to improve tumor uptake of [177Lu]Lu-PP-F11N and provides proof-of-concept for the development of molecular strategies aimed at enhancing the level of the targeted receptor, to increase the efficacy of PRRT and nuclear imaging.