Effect of Brownian motion on reduced agglomeration of nanostructured metal oxide towards development of efficient cancer biosensor.

We report results of the studies relating to fabrication of nanostructured metal oxide (NMO) based cancer biosensor. With the help of 2D electroactive reduced graphene oxide (RGO), we successfully inhibited the Brownian motion of NMO that led to reduced agglomeration of NMO. The nanostructured hafnium oxide (nHfO2) was used as a model NMO. The reduced agglomeration of nHfO2 was achieved through controlled hydrothermal synthesis and investigated via nanoparticles tracking analysis (NTA). X-ray diffraction (XRD), scanning electron microscopy (SEM) and transmission electron microscope (TEM) techniques were used for phase identification as well as morphological analysis of the synthesized nanohybrid (nHfO2@RGO) material. The 3-aminopropyl triethoxysilane (APTES) was used for the functionalization of nHfO2@RGO and electrophoretic deposition (EPD) technique was used for its deposition onto ITO coated glass electrode. Further, antibodies of cancer biomarker (anti-CYFRA-21-1) were immobilized via EDC-NHS chemistry and Bovine serum albumin (BSA) was used for blocking of the non-specific binding sites. The electrochemical response studies of fabricated immunoelectrode (BSA/anti-CYFRA-21-1/APTES/nHfO2@RGO/ITO) revealed higher sensitivity (18.24µAmLng-1), wide linear detection range (0 to 30ngmL-1), with remarkable lower detection limit (0.16ngmL-1). The obtained results showed good agreement with the concentration of CYFRA-21-1 obtained through enzyme linked immunosorbent assay (ELISA) in saliva samples of oral cancer patients.

Identification of Keratin 19-Positive Cancer Stem Cells Associating Human Hepatocellular Carcinoma Using 18F-Fluorodeoxyglucose Positron Emission Tomography.

Purpose: The current lack of tools for easy assessment of cancer stem cells (CSC) prevents the development of therapeutic strategies for hepatocellular carcinoma (HCC). We previously reported that keratin 19 (K19) is a novel HCC-CSC marker and that PET with 18F-fluorodeoxyglucose (18F-FDG) is an effective method for predicting postoperative outcome in hepatocellular carcinoma. Herein, we examined whether K19+ HCC-CSCs can be tracked using 18F-FDG-PET.Experimental Design: K19 and glucose transporter-1 (GLUT1) expression was evaluated by IHC in 98 hepatocellular carcinoma patients who underwent 18F-FDG-PET scans before primary tumor resection. Standardized uptake values (SUV) for primary tumors and tumor-to-nontumor SUV ratios (TNR) were calculated using FDG accumulation levels, and values were compared among K19+/K19- patients. Using hepatocellular carcinoma cell lines encoding with a K19 promoter-driven enhanced GFP, 18F-FDG uptake and GLUT1 expression were examined in FACS-isolated K19+/K19- cells.Results: In hepatocellular carcinoma patients, K19 expression was significantly correlated with GLUT1 expression and FDG accumulation. ROC analyses revealed that among preoperative clinical factors, TNR was the most sensitive indicator of K19 expression in hepatocellular carcinoma tumors. In hepatocellular carcinoma cells, FACS-isolated K19+ cells displayed significantly higher 18F-FDG uptake than K19- cells. Moreover, gain/loss-of-function experiments confirmed that K19 regulates 18F-FDG uptake through TGFß/Smad signaling, including Sp1 and its downstream target GLUT1.Conclusions:18F-FDG-PET can be used to predict K19 expression in hepatocellular carcinoma and should thereby aid in the development of novel therapeutic strategies targeting K19+ HCC-CSCs. Clin Cancer Res; 23(6); 1450-60. ©2016 AACR.

L-cysteine capped lanthanum hydroxide nanostructures for non-invasive detection of oral cancer biomarker.

In this paper, we present the result of studies related to the in situ synthesis of amino acid (L-Cysteine) capped lanthanum hydroxide nanoparticles [Cys-La(OH)3 NPs] towards the fabrication of efficient immunosensor for non-invasive detection of oral cancer. The characterization of Cys-La(OH)3 NPs was carried out by different techniques including X-ray diffraction, scanning electron microscopy, transmission electron microscopy, fourier transform infrared spectroscopy and electrochemical techniques. These Cys-La(OH)3 NPs were electrophoretically deposited onto an indium-tin-oxide glass substrate and used for immobilization of anti-cytokeratin fragment-21-1 (anti-Cyfra-21-1) for the electrochemical detection of Cyfra-21-1. This immunosensor shows a broad detection range of 0.001-10.2ngmL-1, the low detection limit of 0.001ngmL-1, and high sensitivity of 12.044µA (ng per mL cm-2)-1 with a response time of 5min. This immunosensor was found to be more advanced in terms of high sensitivity and low detection limit as compared to previously reported biosensors and commercially available ELISA kit (Kinesis DX).

Improvement of the cytological diagnostic accuracy of follicular thyroid lesions by the use of the Ki-67 proliferative index in addition to cytokeratin-19 and HBME-1 immunomarkers: a study of 61 cases of liquid-based FNA cytology with histological controls.

OBJECTIVES: To evaluate HBME-1, cytokeratin-19 (CK-19) and Ki-67 immunomarkers in order to increase the diagnostic accuracy of preoperative thyroid fine needle aspiration (FNA) cytology. METHODS: Immunocytochemistry against HBME-1, CK-19 and Ki-67 was performed on 123 thyroid FNAs processed by liquid-based cytology (LBC). Statistical analysis was carried out on 61 cases with histological control and sufficient material for one or more of the three markers. The Bethesda System was used for cytological diagnosis. RESULTS: Taking into account all the cytological categories, with a cut-off of 30% of positive cells, HBME-1 (n = 47) and CK-19 (n = 53) showed a sensitivity for malignancy of 66.7% (95% confidence interval, 53.2-80.1) and 90.5% (82.6-98.4) and a specificity of 90.6% (82.3-99) and 75% (63.3-86.7), respectively. For Ki-67 (n = 54) with a cut-off of 1% of positive cells, the sensitivity was 85.0% (75.5-94.5) and the specificity 70.6% (58.4-82.7). In the follicular neoplasm/suspicious for follicular neoplasm (FN/SFN) category (n = 37), which was the focus of the study, papillary thyroid carcinomas (PTCs) were less numerous (four cases, three of which were the follicular variant), the positivity of the three immunomarkers combined showed an overall accuracy of 91% (21/23). The mean percentage of Ki-67-positive cells was increased in malignant lesions, with the exception of follicular variant PTCs: 16% ± 15.6% in two follicular carcinomas, 4.8% ± 3.2% in 13 classical PTCs, 1% ± 1.2% in five follicular variant PTCs and 0.5% ± 1.9% in 34 non-malignant lesions. CONCLUSIONS: Immunocytochemistry using HBME-1, CK-19 and the Ki-67 proliferative index increased the diagnostic accuracy of FNA in the FN/SFN category of the Bethesda System, which may help to distinguish lesions in this category with a low or high risk of malignancy. Thus, clinical management would be improved.

Correlating purity by microdissection with gene expression in gastric cancer tissue.

Microdissection is a feasible tool for the purification of target cells from heterogeneous tissue components. However, the extent to which cells need to be purified by microdissection for use in gene expression analysis has not been determined. In the present study, we obtained diffuse-type gastric cancer tissues at varying purities, and evaluated the corresponding expression of a cancer-specific gene, KRT19, by quantitative real-time PCR. The relationship between the degree of purity and gene expression was confirmed by using 60-mer oligonucleotide microarray analysis. Cancer-specific gene expression was stable in tissues of 10-50% purity, but at 60% or greater purity the slope of the graph was much steeper, indicating a correlation between tissue purity and increased gene expression. Tissues of 70% purity for cancer cells, acquired by microdissection, were therefore deemed to be of sufficient quality to distinguish between gene expression profiles from microdissected and non-microdissected specimens.