In-vitro prostate cancer biomarker detection by directed conjugation of anti-PSCA antibody to super paramagnetic iron oxide nanoparticless.

Background: The main property of a successful conjugation of antibodies to nanoparticles is keeping the potency of antibody for binding the antigen, and an oriented conjugation can do that. Under such ground, this study was carried out to explore the efficiency of two conjugation methods in binding iron nanoparticles to an antibody produced against PSCA (prostate stem cell antigen) using in vitro labeling of PC3 cells. Methods: In this experimental study, we conjugated dextran-superparamagnetic iron oxide nanoparticles (dexSPIONs) to anti-PSCA antibody by two different methods, including targeting carbohydrate moieties in FC domain and the free amine group of amino acid side chains. Ultimately, Iron staining was done by anti-PSCA antibody-dexSPIONs in PC3 cells to detect antibody binding to the cells. Results: A strong blue dye was induced by iron staining in conjugated dexSPIONs on the membrane of PC3 cells by the former method than the second one. Moreover, cells treated with 20 nm diameters of dexSPIONs showed higher resolution of blue color than those treated with 100 nm nanoparticles. Conclusion: This oriented conjugation method promoted the efficiency of targeting tumor antigens, and the presence of iron particles might enhance MRI image intensity in vivo by targeting PSCA-overexpressing cells in future studies.

Reduced expression of NGEP is associated with high-grade prostate cancers: a tissue microarray analysis.

New gene expressed in prostate (NGEP) is a newly diagnosed prostate-specific gene that is expressed only in normal prostate and prostate cancer cells. Discovery of tissue-specific markers may promote the development of novel targets for immunotherapy of prostate cancer. In the present study, the staining pattern and clinical significance of NGEP were evaluated in a series of prostate tissues composed of 123 prostate cancer, 19 high-grade prostatic intraepithelial neoplasia and 44 samples of benign prostate tissue included in tissue microarrays using immunohistochemistry. Our study demonstrated that NGEP localized mainly in the apical and lateral membranes and was also partially distributed in the cytoplasm of epithelial cells of normal prostate tissue. All of the examined prostate tissues expressed NGEP with a variety of intensities; the level of expression was significantly more in the benign prostate tissues compared to malignant prostate samples (P value <0.001). Among prostate adenocarcinoma samples, a significant and inverse correlation was observed between the intensity of NGEP expression and increased Gleason score (P = 0.007). Taken together, we found that NGEP protein is widely expressed in low-grade to high-grade prostate adenocarcinomas as well as benign prostate tissues, and the intensity of expression is inversely proportional to the level of malignancy. NGEP could be an attractive target for immune-based therapy of prostate cancer patients as an alternative to the conventional therapies particularly in indolent patients.

The Anti-Proliferative Effect of a Newly-Produced Anti-PSCA-Peptide Antibody by Multiple Bioinformatics Tools, on Prostate Cancer Cells.

BACKGROUND: Prostate Stem Cell Antigen (PSCA) is a small cell surface protein, overexpressed in 90% of prostate cancers. Determination of epitopes that elicit an appropriate response to the antibody generation is vital for diagnostic and immunotherapeutic purposes for prostate cancer treatment. Presently, bioinformatics B-cell prediction tools can predict the location of epitopes, which is uncomplicated, faster, and more cost-effective than experimental methods. OBJECTIVE: We aimed to predict a novel linear peptide for Prostate Stem Cell Antigen (PSCA) protein in order to generate anti-PSCA-peptide (p) antibody and to investigate its effect on prostate cancer cells. METHODS: In the current study, a novel linear peptide for PSCA was predicted using in silico methods that utilize a set of linear B-cell epitope prediction tools. Polyclonal antibody (anti-PSCA-p antibody "Patent No. 99318") against PSCA peptide was generated. The antibody reactivity was determined by the Enzyme-Linked Immunosorbent Assay (ELISA) and its specificity by immunocytochemistry (ICC), immunohistochemistry (IHC), and Western Blotting (WB) assays. The effect of the anti-PSCA-p antibody on PSCA-expressing prostate cancer cell line was assessed by Methylthiazolyldiphenyl- Tetrazolium bromide (MTT) assay. RESULTS: New peptide-fragment of PSCA sequence as "N-CVDDSQDYYVGKKN-C" (PSCA-p) was selected and synthesized. The anti-PSCA-p antibody against the PSCA-p showed immunoreactivity with PSCA-p specifically bound to PC-3 cells. Also, the anti-PSCA-p antibody strongly stained the prostate cancer tissues as compared to Benign Prostatic Hyperplasia (BPH) and normal tissues (P < 0.001). As the degree of malignancy increased, the staining intensity was also elevated in prostate cancer tissue (P < 0.001). Interestingly, the anti-PSCA-p antibody showed anti-proliferative effects on PC-3 cells (31%) with no growth inhibition effect on PSCA-negative cells. CONCLUSION: In this study, we developed a new peptide sequence (PSCA-p) of PSCA. The PSCA-p targeting by anti-PSCA-p antibody inhibited the proliferation of prostate cancer cells, suggesting the potential of PSCA-p immunotherapy for future prostate cancer studies.

Association of human leukocyte antigen polymorphism with outcomes of hepatitis B virus infection.

BACKGROUND AND AIM: Host genetic and environmental factors are viewed as a common basis of the different outcomes of hepatitis B virus (HBV) infection. Human leukocyte antigen (HLA) plays an important role in immunological reaction to HBV infection. In this study, we aimed to determine the association between HBV infection and HLA-A, B, and DRB1 alleles in northern Iran. METHODS: HLA-A, B, and DRB1 alleles in 33 patients with chronic hepatitis B (CHB) and 31 healthy carriers as the persistent group, and 30 subjects who had spontaneously recovered from HBV infection were analyzed by using the polymerase chain reaction (PCR)-sequence-specific primer (PCR-SSP) technique. RESULTS: The frequency of the HLA-A*33 allele was higher in the persistent group than in the recovered group (10.16% vs 0%, P < 0.008); the frequency of the DRB1*13 allele was lower in the persistent group than in the recovered group (3.13% vs 11.67%, P < 0.03). The frequency of the B*52 allele was higher in CHB patients than healthy carriers (7.58% vs 0%, P < 0.05). The logistic regression model showed that the presence of the HLA-DRB1*13 allele was the significant factor associated with protection against the persistency of HBV. There were significant differences between the HBV recovered group, CHB patients, and healthy carriers regarding age, hepatitis B e antigen, and anti-hepatitis B e positivity. CONCLUSION: HLA-A*33 was closely related with susceptibility to persisting hepatitis B infection, and HLA-DRB1*13 was closely related with protection against persisting hepatitis B in an Iranian population. These findings emphasized that the host HLA polymorphism is an important factor in determining the outcome of HBV infection.

Expression of prostate stem cell antigen (PSCA) in prostate cancer: a tissue microarray study of Iranian patients.

Proteins expressed in prostate cancer, including prostate stem cell antigen (PSCA), have been investigated as biomarkers for diagnosis and therapy of prostate cancer. Immunohistochemical analysis of PSCA expression was performed on tissue microarrays of 185 paraffin-embedded tissues of Iranian patients, including 114 prostate cancers (PCa), 21 High Grade Prostatic Intraepithelial Neoplasias (HGPIN) and 50 samples of benign prostate tissue. The level of PSCA expression was compared between benign tissues, HGPIN and PCa. Then the correlations of PSCA expression with clinicopathologic parameters were assessed in PCa. The PSCA expression was detected in the membrane and cytoplasm of epithelial secretory cells in normal prostate tissues, HGPIN and PCa with a variety of intensities. The intensity of PSCA staining was significantly increased in the PCa group as compared with HGPIN and benign prostate tissues (P-value<0.05). Moreover, the level of PSCA expression was increased with higher Gleason score of PCa (P-value=0.036). The data presented here revealed that expression of PSCA as a cell surface marker increased from benign prostate tissues (BPH) and HGPIN to PCa, and its expression in PCa was positively associated with poor cell differentiation, suggesting that PSCA could be considered as a valuable target for diagnosis and therapy of PCa.

Effect of Matricaria chamomilla L. flower essential oil on the growth and ultrastructure of Aspergillus niger van Tieghem.

The antifungal activity of Matricaria chamomilla L. flower essential oil was evaluated against Aspergillus niger with the emphasis on the plant's mode of action at the electron microscopy level. A total of 21 compounds were identified in the plant oil using gas chromatography/mass spectrometry (GC/MS) accounting for 92.86% of the oil composition. The main compounds identified were alpha-bisabolol (56.86%), trans-trans-farnesol (15.64%), cis-beta-farnesene (7.12%), guaiazulene (4.24%), alpha-cubebene (2.69%), alpha-bisabolol oxide A (2.19%) and chamazulene (2.18%). In the bioassay, A. niger was cultured on Potato Dextrose Broth medium in 6-well microplates in the presence of serial two fold concentrations of plant oil (15.62 to 1000 microg/mL) for 96 h at 28 degrees C. Based on the results obtained, A. niger growth was inhibited dose dependently with a maximum of approximately 92.50% at the highest oil concentration. A marked retardation in conidial production by the fungus was noticed in relation to the inhibition of hyphal growth. The main changes of hyphae observed by transmission electron microscopy were disruption of cytoplasmic membranes and intracellular organelles, detachment of plasma membrane from the cell wall, cytoplasm depletion, and complete disorganization of hyphal compartments. In scanning electron microscopy, swelling and deformation of hyphal tips, formation of short branches, and collapse of entire hyphae were the major changes observed. Morphological alterations might be due to the effect on cell permeability through direct interaction of M. chamomilla essential oil with the fungal plasma membrane. These findings indicate the potential of M. chamomilla L. essential oil in preventing fungal contamination and subsequent deterioration of stored food and other susceptible materials.

Fungistatic effects of optical brightener 220 against Trichophyton tonsurans, Aspergillus fumigatus and Candida albicans.

BACKGROUND: Dermatophytes are one of the main causes of dermal infections. Moreover, there are some opportunistic fungi such as Aspergillus fumigatus (mycelial form) and Candida albicans (yeasty form) that in immunosuppressed patients can cause cutaneous disease. OBJECTIVES: The possible effect of optical brightener 220 (OB-220) on the growth of fungi has been evaluated in this study. METHODS: Isolates were grown on agar plates containing OB-220 in concentration between 0.06 and 11.68 mg ml(-1). MICs of OB-220, ketoconazole and fluconazole were obtained by the agar dilution method. Hyphae and yeasts grown with OB-220 were compared with controls by fluorescence and transmission electron microscopy. The cell cytotoxicity of OB-220 was also assessed. RESULTS: The MIC(90) of OB-220 was obtained: 1.17-1.46 mg ml(-1) for A. fumigatus, 0.58-1.17 mg ml(-1) for C. albicans and 0.29 mg ml(-1) for Trichophyton tonsurans. Electron microscopy revealed a thickening and blurred contours of the cell wall by OB-220. OB-220 in concentrations up to 11.68 mg ml(-1) posed no mammalian cell toxicity. CONCLUSION: OB-220 suppresses the growth of fungi by interfering with the formation of normal chitin.