NMR-Based Prostate Cancer Metabolomics Euceda, Leslie R; Andersen, Maria K; Tessem, May-Britt ...
Methods in molecular biology (Clifton, N.J.),
01/2018, Letnik:
1786
Journal Article
Odprti dostop
Prostate cancer is the second most common malignancy, and the fifth leading cause of cancer-related death among men, worldwide. A major unsolved clinical challenge in prostate cancer is the ability ...to accurately distinguish indolent cancer types from the aggressive ones. Reprogramming of metabolism is now a widely accepted hallmark of cancer development, where cancer cells must be able to convert nutrients to biomass while maintaining energy production. Metabolomics is the large-scale study of small molecules, commonly known as metabolites, within cells, biofluids, tissues, or organisms. Nuclear magnetic resonance (NMR) spectroscopy is commonly applied in metabolomics studies of cancer. This chapter provides protocols for NMR-based metabolomics of cell cultures, biofluids (serum and urine), and intact tissue, with concurrent advice for optimal biobanking and sample preparation procedures.
•quantification of Annexin A3 in biological samples at ng/mL levels;•novel Protein Imprinted Material based on electropolimerization of caffeic acid;•successful use of the sensor on the analysis of ...Annexin A3 in urine samples;•good sensitivity, selectivity and sensor stability was achieved.
The development of fast and reliable methods for protein determination are of great relevance to a diversity of areas from industry to diagnostics. Molecular Imprinted Materials (MIM) has proved to be an interesting methodology for protein analysis however further studies of the effect of the experimental parameters and starting materials in the performance of the MIM are still required. Caffeic acid (CAF) is employed for the first time as a monomer to tailor a synthetic receptor for a protein target. This was done by bulk-electropolymerization, applying a constant potential of +2.0V, for 30s, on a carbon screen-printed electrode, immersed in a solution of protein and CAF prepared in phosphate buffer. Annexin A3 (ANXA3) was selected as protein target due to the fact that this is an emerging biomarker in prostate cancer.
The assembly of the protein imprinted material (PIM) was followed by Electrochemical Impedance Spectroscopy (EIS) and Raman Spectroscopy. A non-imprinted material (NIM) was prepared in parallel as control.
Square wave voltammetry (SWV) was used to monitor the electrochemical signal of the Fe(CN)63−/Fe(CN)64− redox for the quantification of ANXA3.
The optimized PIM-based device showed average detection limits (LOD) of 0.095ng/mL, a linear behavior against log (concentration) between 0.10, and 200ng/mL and good selectivity. The NIM-based device showed random behavior against protein concentration. Finally, the PIM-sensor was successfully applied to the analysis of ANXA3 in spiked urine samples.
Prostate cancer is one of the major causes of cancer-related deaths in men and there is a growing interest in identifying natural compounds for its management. We analyzed bioactive withanolides in
...Withania coagulans
from 11 different sites in Pakistan and evaluated the antiprostate cancer activities of leaf extracts from two sites with the greatest amounts. Total withanolide concentration differed by ~ 17-fold between sites, ranging from 1.01 ± 0.01 mg/g dry weight (mean ± SE) at Jand to 16.83 ± 0.02 mg/g at Mohmand Agency. Different tissues varied in their total withanolide content with roots having the least (0.42 ± 0.07 mg/g dry weight) and leaves the most (2.45 ± 0.45 mg/g). We found strong inverse correlations between site annual precipitation versus withanolide amounts in fruits (
r
= − 0.84,
P
= 0.001), leaves (
r
= − 0.88,
P
< 0.001), roots (
r
= − 0.91,
P
< 0.001), and total (
r
= − 0.89,
P
< 0.001), but not stems (
r
= − 0.20,
P
= 0.556). Extracts made from Mianwali and Mohmand Agency leaves possessed high anticancer activity in terms of increased induction of apoptosis and decreased cell viability, cell proliferation, invasion, and migration of different prostate cancer cell lines. These results are useful for the selection of withanolide-rich germplasm with potent anticancer properties.
The development of integrated instrumentation for universal bioassay systems serves as a key goal for the lab-on-a-chip community. The programmable bio-nano-chip (p-BNC) system is a versatile ...multiplexed and multiclass chemical- and bio-sensing system for bioscience and clinical measurements. The system is comprised of two main components, a disposable cartridge and a portable analyzer. The customizable single-use plastic cartridges, which now can be manufactured in high volumes using injection molding, are designed for analytical performance, ease of use, reproducibility, and low cost. These labcard devices implement high surface area nano-structured biomarker capture elements that enable high performance signaling and are index-matched to real-world biological specimens. This detection modality, along with the convenience of on-chip fluid storage in blisters and self-contained waste, represents a standard process to digitize biological signatures at the point-of-care. A companion portable analyzer prototype has been developed to integrate fluid motivation, optical detection, and automated data analysis, and it serves as the human interface for complete assay automation. In this report, we provide a systems-level perspective of the p-BNC universal biosensing platform with an emphasis on flow control, device integration, and automation. To demonstrate the flexibility of the p-BNC, we distinguish diseased and non-case patients across three significant disease applications: prostate cancer, ovarian cancer, and acute myocardial infarction. Progress towards developing a rapid 7 minute myoglobin assay is presented using the fully automated p-BNC system.
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•Stattic enhances the efficacy of docetaxel by changing the cell cycle distribution of DU-145 prostate cancer cells.•Co-treatment of docetaxel and stattic has a strong synergistic ...effect.•Combination treatment with two agents (stattic and docetaxel) increase apoptotic cell death in comparison to single treatment.•Comparison of single treatment, combinatorial treatment with stattic and docetaxel leads to decreases the expression of key anti-apoptotic genes (Bcl-2, Bcl-xl).
We investigated the effects of Stattic, an important signal transducer and activator of transcription 3 (STAT3) inhibitor, on enhancing the anti-proliferative and apoptotic effects of docetaxel in lung A549 and prostate cancer DU145 cells. Cell proliferation, cellular apoptosis and expression of STAT3 target genes were evaluated by DAPI staining, Annexin V/PI staining, cell cycle analysis and Real Time PCR. The anticancer effects of docetaxel and Stattic combination therapy induced synergistic effects in A549 and DU145 cells with combination indices of 0.71 and 0.52, respectively. Annexin V/PI demonstrated that there was a 2-fold increase in the proportion of apoptotic cells in cells treated with docetaxel and Stattic in A549 and DU145 cell lines, compared with control groups. Compared with cells treated with either docetaxel or Stattic alone, the results of the current study identified a significant decrease in the transcript levels of the anti-apoptotic proteins Bcl-2 and Bcl-xl and a marked increase in the level of the pro-apoptotic protein, Bax in cells that underwent combination therapy with docetaxel and Stattic combination (P < 0.05). These results suggest that combination of a STAT3 inhibitor and taxan derivatives may be developed as a promising therapeutic strategy to treat patients with lung and prostate cancer.
The physical and mechanical properties of the tumor microenvironment are crucial for the growth, differentiation and migration of cancer cells. However, such microenvironment is not found in the ...geometric constraints of 2D cell culture systems used in many cancer studies. Prostate cancer research, in particular, suffers from the lack of suitable in vitro models. Here a 3D superporous scaffold is described with thick pore walls in a mechanically stable and robust architecture to support prostate tumor growth. This scaffold is generated from the cryogelation of poly(ethylene glycol) diacrylate to produce a defined elastic modulus for prostate tumor growth. Lymph node carcinoma of the prostate (LNCaP) cells show a linear growth over 21 d as multicellular tumor spheroids in such a scaffold with points of attachments to the walls of the scaffold. These LNCaP cells respond to the growth promoting effects of androgens and demonstrate a characteristic cytoplasmic‐nuclear translocation of the androgen receptor and androgen‐dependent gene expression. Compared to 2D cell culture, the expression or androgen response of prostate cancer specific genes is greatly enhanced in the LNCaP cells in this system. This scaffold is therefore a powerful tool for prostate cancer studies with unique advantages over 2D cell culture systems.
Superporous poly(ethylene glycol) diacrylate cryogels mimicking the stiffness of malignant prostate tissues have been engineered for studies of prostate cancer cell growth and function. Cells grow in an anchorage‐dependent manner in this scaffold for three weeks and respond to androgen and antiandrogen treatment. Compared to 2D cell culture system, androgen‐dependent prostate target gene expression is highly regulated in this system.
Objective
Magnetic resonance imaging with hyperpolarized contrast agents can provide unprecedented in vivo measurements of metabolism, but yields images that are lower resolution than that achieved ...with proton anatomical imaging. In order to spatially localize the metabolic activity, the metabolic image must be interpolated to the size of the proton image. The most common methods for choosing the unknown values rely exclusively on values of the original uninterpolated image.
Methods
In this work, we present an alternative method that uses the higher-resolution proton image to provide additional spatial structure. The interpolated image is the result of a convex optimization algorithm which is solved with the fast iterative shrinkage threshold algorithm (FISTA).
Results
Results are shown with images of hyperpolarized pyruvate, lactate, and bicarbonate using data of the heart and brain from healthy human volunteers, a healthy porcine heart, and a human with prostate cancer.
•Androgens and β-agonists are growth promoters illegally used in farm animals.•Testes, sex accessory glands, skeletal muscle are target organs of these molecules.•Myosin heavy chain of Longissimus ...dorsi muscle of treated animals is up-regulated.•Indirect and direct methods of analysis can be flanked to detect illicit treatment.
The present study describes different effects of the selective androgen receptor modulator (SARM) nandrolone phenylpropionate (Nandrosol) and the β-agonist ractopamine administration in veal calves, and it investigates different strategies applied to trace these molecules.
Morphological changes of gonads and accessory glands attributed to androgen effects, such as testicular atrophy, seminiferous tubule diameter reduction and hyperplasia of prostate epithelium, were detected, although SARMs are not described to cause these lesions. The gene expression analysis showed an anabolic activity of Nandrosol in Longissimus dorsi muscle, where myosin heavy chain (MYH) was significantly up-regulated. An IGF1 increase was weakly significant only in Vastus lateralis muscle.
In conclusion, the anatomo-histopathological observations and the MYH mRNA up-regulation in Longissimus dorsi muscle confirm the androgenic treatment in experimental animals. The biosensor assay was not enough sensitive to detect residues in urines and only the direct chemical analysis of urine samples confirmed both β-agonist and SARM treatment.
In biological tissues, radiation causes the formation of reactive oxygen species (ROS), some of which lead to sequential oxidation of certain protein cysteine residues. Resultant cysteinyl radicals ...are subject to post-translational modification through S-glutathionylation. The present clinical trial was designed to determine if S-glutathionylated serine protease inhibitors (serpins) in blood could be used as biomarkers of exposure to radiation. 56 male prostate cancer patients treated with radiotherapy were enrolled in the trial and levels of S-glutathionylated serpins A1 and A3 were assessed by immunoblotting. Patients were classified into three groups: (1) external beam radiation therapy (EBRT); (2) brachytherapy (BT); (3) both EBRT and BT. Prior to treatment, baseline plasma levels of both unmodified and S-glutathionylated serpins were similar in each group. We identified elevated plasma levels of S-glutathionylated serpin A1 monomer, trimer and serpin A3 monomer in patient blood following radiation. Maximal increased levels of these S-glutathionylated serpins were correlated with increased duration of radiotherapy treatments. We conclude that it is practical to quantify patient plasma S-glutathionylated serpins and that these post-translationally modified proteins are candidate biomarkers for measuring radiation exposure. This provides a platform for use of such biomarkers in trials with the range of drugs that, like radiation, produce ROS.
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