Abstract
BackgroundOvarian cancer is one of the deadliest gynecological malignancies. While the overall survival of ovarian cancer patients has slightly improved in recent years in the developed world, it remains clinically challenging due to its frequent late diagnosis and the lack of reliable diagnostic and/or prognostic markers. The aim of this study was to identify potential new molecular target proteins (NMTPs) responsible for the poor outcomes. When nanoparticles (NP) are exposed to biological fluids, a protein coat, termed the protein corona (PC), forms around the NP, and the PC represents a tool to identify NMTPs. This study investigates the influence of pre-processing conditions, such as lysis conditions and serum/plasma treatment, on the PC composition and the resulting identification of NMTPs.ResultsUsing gel electrophoresis, pre-processing conditions, including cell-lysis techniques and enrichment of low-abundance proteins (LAPs) by immunocentrifugation of serum/plasma, were shown to alter the relative amounts and compositions of proteins. PCs formed when 20 nm gold-NPs (GNPs) were incubated with lysate proteins from either RIPA- or urea lysis. Proteomic analysis of these PCs showed 2–22-fold enrichment of NMTPs in PCs from urea lysates as compared to RIPA lysates. Enriched NMTPs were then classified as cellular components, biological and molecular functions-associated proteins. The impact of enriched LAPs (eLAPs) on both PC composition and NMTP identification was shown by comparative proteomic analysis of original plasma, eLAPs, and PCs derived from eLAPs; eLAPs-PCs enhanced the abundance of NMTPs approximately 13%. Several NMTPs, including gasdermin-B, dermcidin, and kallistatin, were identified by this method demonstrating the potential use of this PC approach for molecular target discovery.ConclusionThe current study showed that the pre-processing conditions modulate PC composition and can be used to enhance identification of NMTPs.
Highlights
Ovarian cancer is one of the deadliest gynecological malignancies
This study examined the influence of pre-processing conditions on the formation and composition of the protein corona (PC) around GNPs with the ultimate goal of facilitating the identification of new molecular targets (Fig. 1)
These results are supported by the report of Marini et al who showed that when the soluble and insoluble fractions of mitochondria from cancer cells are enzymatically digested by trypsin, Glu-C, or chymotrypsin, each lysis condition produces a distinct protein signature (Marini et al 2020; Nierenberg et al 2018)
Summary
Ovarian cancer is one of the deadliest gynecological malignancies. While the overall survival of ovarian cancer patients has slightly improved in recent years in the developed world, it remains clinically challenging due to its frequent late diagnosis and the lack of reliable diagnostic and/or prognostic markers. When nanoparticles (NP) are exposed to biological fluids, a protein coat, termed the protein corona (PC), forms around the NP, and the PC represents a tool to identify NMTPs. This study investigates the influence of pre-processing conditions, such as lysis conditions and serum/plasma treatment, on the PC composition and the resulting identification of NMTPs. Ovarian cancer will account for about 21,750 new cases and 13,940 deaths in the United States in 2020 (Siegel et al 2020). Identification of potentially new molecular targets responsible for poor outcome is the focus of this study; ovarian cancer cell lines and patient plasma samples were investigated as a paradigm for improving the utilization of nanoparticles (NP) to identify proteins of interest (Engelberth et al 2014). The effect of experimental pre-processing conditions on the modulation of PC compositions has not been carefully investigated to date
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