Quantitative Proteomics Analysis of FFPE Tumor Samples Reveals the Influences of NET-1 siRNA Nanoparticles and Sonodynamic Therapy on Tetraspanin Protein Involved in HCC

Abstract

Hepatocellular carcinoma (HCC) poses a severe threat to human health. The NET-1 protein has been proved to be strongly associated with HCC proliferation and metastasis in our previous study. Here, we established and validated the NET-1 siRNA nanoparticles system to conduct targeted gene therapy of HCC xenograft in vivo with the aid of sonodynamic therapy. Then, we conducted a label-free proteome mass spectrometry workflow to analyze formalin-fixed and paraffin-embedded HCC xenograft samples collected in this study. The result showed that 78 proteins were differentially expressed after NET-1 protein inhibited. Among them, the expression of 17 proteins upregulated and the expression of 61 proteins were significantly downregulated. Of the protein abundance, the vast majority of Gene Ontology enrichment terms belong to the biological process. The KEGG pathway enrichment analysis showed that the 78 differentially expressed proteins significantly enriched in 45 pathways. We concluded that the function of the NET-1 gene is not only to regulate HCC but also to participate in a variety of biochemical metabolic pathways in the human body. Furthermore, the protein–protein interaction analysis indicated that the interactions of differentially expressed proteins are incredibly sophisticated. All the protein–protein interactions happened after the NET-1 gene has been silenced. Finally, our study also provides a useful proposal for targeted therapy based on tetraspanin proteins to treat HCC, and further mechanism investigations are needed to reveal a more detailed mechanism of action for NET-1 protein regulation of HCC.

Role of Tetraspanins in Hepatocellular Carcinoma

“…Based on the fact that TSPAN1 is highly expressed in HCC cells and positively regulating HCC cell proliferation, Wu et al developed a gene therapy method to specifically silence TSPAN1 gene expression in HCC cells (39,40,42,43). They used targeted nanobubbles to deliver TSPAN1 siRNA to HCC cells, with the aid of ultrasound exposure to increase transfection efficiency.…”

Advances in Multi-Omics Applications in HBV-Associated Hepatocellular Carcinoma

“…Based on iTRAQ quantitative comparative proteomics, researchers have utilized liquid chromatograph-mass spectrometer/mass spectrometer (LC-MS/MS) to recognize and quantitate differential proteins in HepG2 cell lines stably containing different functional domains of HBx, and p90 ribosomal S6 kinase 2 (RSK2) has been identified as a new host protein that plays a key role in HBx enhancing HBV replication ( 140 ). Plasma fibronectin was demonstrated to be related with serum clearance of HBsAg and may be a potential predictor of “functional cure” of CHB by iTRAQ-based quantitative proteomics ( 141 ), meanwhile the TMT isobaric labeling-based technology was used to quantitatively characterize the renal proteome of HBV transgenic mice, and to elucidate the pathogenesis of HBV-associated glomerulonephritis (HBV-GN) ( 142 ). Additionally, proteomic analyses of formalin-fixed paraffin-embedded (FFPE) HCC graft samples, conducted using a label-free proteome mass spectrometry workflow, were used to characterize the global quantitative analysis of protein expression profiles after gene therapy and to identify differentially expressed proteins ( 143 ).…”

Advanced Omics and Radiobiological Tissue Archives: The Future in the Past

“…Not included in this review are repositories held at clinics and hospitals for their own purposes (e.g., [44,45]), as it is not certain whether FFPE blocks from these institutions will be made available to interested scientists from outside. Moreover, the authors are well aware that the number of archives listed here might be incomplete.…”