Integrating trans-omics, cellular experiments and clinical validation to identify ILF2 as a diagnostic serum biomarker and therapeutic target in gastric cancer.

BACKGROUND: Gastric cancer (GC) lacks serum biomarkers with clinical diagnostic value. Multi-omics analysis is an important approach to discovering cancer biomarkers. This study aimed to identify and validate serum biomarkers for GC diagnosis by cross-analysis of proteomics and transcriptomics datasets. METHODS: A cross-omics analysis was performed to identify overlapping differentially expressed genes (DEGs) between our previous aptamer-based GC serum proteomics dataset and the GC tissue RNA-Seq dataset in The Cancer Genome Atlas (TCGA) database, followed by lasso regression and random forest analysis to select key overlapping DEGs as candidate biomarkers for GC. The mRNA levels and diagnostic performance of these candidate biomarkers were analyzed in the original and independent GC datasets to select valuable candidate biomarkers. The valuable candidate biomarkers were subjected to bioinformatics analysis to select those closely associated with the biological behaviors of GC as potential biomarkers. The clinical diagnostic value of the potential biomarkers was validated using serum samples, and their expression levels and functions in GC cells were validated using in vitro cell experiments. RESULTS: Four candidate biomarkers (ILF2, PGM2L1, CHD7, and JCHAIN) were selected. Their mRNA levels differed significantly between tumor and normal tissues and showed different diagnostic performances for GC, with areas under the receiver operating characteristic curve (AUROCs) of 0.629-0.950 in the TCGA dataset and 0.736-0.840 in the Gene Expression Omnibus (GEO) dataset. In the bioinformatics analysis, only ILF2 (interleukin enhancer-binding factor 2) gene levels were associated with immune cell infiltration, some checkpoint gene expression, chemotherapy sensitivity, and immunotherapy response. Serum levels of ILF2 were higher in GC patients than in controls, with an AUROC of 0.944 for the diagnosis of GC, and it was also detected in the supernatants of GC cells. Knockdown of ILF2 by siRNA significantly reduced the proliferation and colony formation of GC cells. Overexpression of ILF2 significantly promotes the proliferation and colony formation of gastric cancer cells. CONCLUSIONS: Trans-omics analysis of proteomics and transcriptomics is an efficient approach for discovering serum biomarkers, and ILF2 is a potential diagnostic biomarker and therapeutic target of gastric cancer.

The Index sAGP is Valuable for Distinguishing Atypical Hepatocellular Carcinoma from Atypical Benign Focal Hepatic Lesions.

Purpose: The differential diagnosis of atypical hepatocellular carcinoma (aHCC) and atypical benign focal hepatic lesions (aBFHL) usually depends on pathology. This study aimed to develop non-invasive approaches based on conventional blood indicators for the differential diagnosis of aHCC and aBFHL. Patients and Methods: Hospitalized patients with pathologically confirmed focal hepatic lesions and their clinical data were retrospectively collected, in which patients with HCC with serum alpha-fetoprotein (AFP) levels of ≤200 ng/mL and atypical imaging features were designated as the aHCC group (n = 224), and patients with benign focal hepatic lesions without typical imaging features were designated as the aBFHL group (n = 178). The performance of indexes (both previously reported and newly constructed) derived from conventional blood indicators by four mathematical operations in distinguishing aHCC and aBFHL was evaluated using the receiver operating characteristic (ROC) curve and diagnostic validity metrics. Results: Among ten previously reported derived indexes related to HCC, the index GPR, the ratio of γ-glutamyltransferase (GGT) to platelet (PLT), showed the best performance in distinguishing aHCC from aBFHL with the area under ROC curve (AUROC) of 0.853 (95% CI 0.814-0.892), but the other indexes were of little value (AUROCs from 0.531 to 0.700). A new derived index, sAGP [(standardized AFP + standardized GGT)/standardized PLT], was developed and exhibited AUROCs of 0.905, 0.894, 0.891, 0.925, and 0.862 in differentiating overall, BCLC stage 0/A, TNM stage I, small, and AFP-negative aHCC from aBFHL, respectively. Conclusion: The sAGP index is an efficient, simple, and practical metric for the non-invasive differentiation of aHCC from aBFHL.

Analysis of aptamer-target binding and molecular mechanisms by thermofluorimetric analysis and molecular dynamics simulation.

Introduction: Aptamers are valuable for bioassays, but aptamer-target binding is susceptible to reaction conditions. In this study, we combined thermofluorimetric analysis (TFA) and molecular dynamics (MD) simulations to optimize aptamer-target binding, explore underlying mechanisms and select preferred aptamer. Methods: Alpha-fetoprotein (AFP) aptamer AP273 (as the model) was incubated with AFP under various experimental conditions, and melting curves were measured in a real-time PCR system to select the optimal binding conditions. The intermolecular interactions of AP273-AFP were analysed by MD simulations with these conditions to reveal the underlying mechanisms. A comparative study between AP273 and control aptamer AP-L3-4 was performed to validate the value of combined TFA and MD simulation in selecting preferred aptamers. Results: The optimal aptamer concentration and buffer system were easily determined from the dF/dT peak characteristics and the melting temperature (Tm) values on the melting curves of related TFA experiments, respectively. A high Tm value was found in TFA experiments performed in buffer systems with low metal ion strength. The molecular docking and MD simulation analyses revealed the underlying mechanisms of the TFA results, i.e., the binding force and stability of AP273 to AFP were affected by the number of binding sites, frequency and distance of hydrogen bonds, and binding free energies; these factors varied in different buffer and metal ion conditions. The comparative study showed that AP273 was superior to the homologous aptamer AP-L3-4. Conclusion: Combining TFA and MD simulation is efficient for optimizing the reaction conditions, exploring underlying mechanisms, and selecting aptamers in aptamer-target bioassays.

Bioguided Fraction and Isolation of the Antitumor Components from Phyllanthus niruri L.

Phyllanthus niruri L., a well-known medicinal plant, has been used as a folk antitumor remedy in the worldwide scale. However, the antitumor components in P. niruri have not been reported. In order to verify the antitumor components of P. niruri and the plants which have the high content of these components, we isolated the antitumor components with bioguided fraction and isolation, by different chromatographic methods from the ethyl acetate fraction of P. niruri., and identified them as ethyl brevifolincarboxylate and corilagin by 1H-NMR, 13C-NMR, 2D-NMR, and mass spectrometric analyses. Cell cytotoxicity assays showed that corilagin has broad-spectrum antitumor activity, a better antitumor potential, and lower toxicity in normal cells. Besides, the coefficient of drug interaction (CDI) of 10 µM corilagin and 20 µM cDDP reached up to 0.77, which means corilagin can promote the antitumor activity of cDDP. Furthermore, by the extensive screening among 10 species of plants reported to contain corilagin, we found that Dimocarpus longan Lour. has the maximum content of corilagin. In conclusion, corilagin is the major active antitumor composition in P. niruri. L. on HCC cells and has high content in D. longan.

Compound K-induced apoptosis of human hepatocellular carcinoma MHCC97-H cells in vitro.

An intestinal bacterial metabolite of ginseng protopanaxadiol saponin, 20-O-(ß-D-glucopyranosyl)-20(S)-protopanaxadiol (compound K), has been reported to induce apoptosis in a variety of cancer cells. However, the precise mechanisms induced by compound K in human hepatocellular carcinoma (HCC) cells remain unclear. In order to examine possible apoptotic mechanisms, we investigated the anticancer effect of compound K in MHCC97-H. MTT assay showed that compound K inhibited the proliferation of MHCC97-H cells with a relatively low toxicity in normal hepatoma cells. Cell cycle progression and cell staining showed an increase in apoptotic sub-G1 fraction. Treatment of MHCC97-H with compound K also induced a reduction in mitochondrial membrane potential (Δψm) and DNA damage. Further study showed that compound K upregulated Fas, FasL, Bax/Bcl-2 ratio and downregulated pro-caspase-9, pro-caspase-3 in a dose-dependent manner, and it also inhibited Akt phosphorylation. These results suggest that compound K significantly inhibits cell proliferation and induces apoptosis in MHCC97-H cells through Fas- and mitochondria-mediated caspase-dependent pathways in human HCC cells.

Two new p-terphenyl derivatives from the marine fungal strain Aspergillus sp. AF119.

Two new p-terphenyl derivatives (1, 2), together with six known ones (3-8), have been isolated from the marine fungal strain Aspergillus sp. AF119. The structures for terphyl acid (1) and terphyl diacid (2) were determined on the basis of HR Q-TOF-MS, and 1D- and 2D-NMR spectroscopic data. The in vitro cytotoxic activities of compounds 1-8 were tested against human tumor cell lines HeLa, HepG-2 and MDA-MB-435; only compounds 5-8 exhibited inhibitory activity against the tested cell lines with IC50 values < 20 microM. Moreover, compound 5 showed a mechanism of inducing cell cycle arrest and apoptosis mediated by the generation of ROS and subsequent DNA double-strand break.