Invasiveness is a very challenging clinical problem in nonfunctional pituitary adenomas (NFPAs), and currently, there are no effective invasiveness-related molecular biomarkers. The post-neurosurgery treatment is much different as for invasive and noninvasive NFPAs. The aim of this study was to integrate phosphoproteomics and transcriptomics data to reveal phosphorylation-mediated molecular events for invasive characteristics of NFPAs to achieve a potential tool for patient stratification, and prognostic/predictive assessment to discriminate invasive from noninvasive NFPAs for personalized attitude.
The 6-plex tandem mass tag (TMT) labeling reagents coupled with TiO2 enrichment of phosphopeptides and liquid chromatography-tandem mass spectrometry (LC-MS/MS) were used to identify and quantify each phosphoprotein and phosphosite in NFPAs and controls. Differentially expressed genes (DEGs) between invasive NFPA and control tissues were obtained from the Gene Expression Omnibus (GEO) database. The overlapping analysis was performed between phosphoprotiens and invasive DEGs. Gene Ontology (GO) enrichment, the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway, and protein–protein interaction (PPI) analyses were used to analyze these overlapped molecules.
In total, 1035 phosphoproteins with 2982 phosphorylation sites were identified in NFPAs vs. controls, and 2751 DEGs were identified in invasive NFPAs vs. controls. Overlapping analysis of these phosphoproteins and DEGs exposed 130 overlapped molecules (phosphoproteins; invasive DEGs). GO enrichment and KEGG pathway analyses of 130 overlapped molecules revealed multiple biological processes and signaling pathway network alterations, including cell–cell adhesion, platelet activation, GTPase signaling pathway, protein kinase signaling, calcium signaling pathway, estrogen signaling pathway, glucagon signaling pathway, cGMP–PKG signaling pathway, GnRH signaling pathway, inflammatory mediator regulation of TRP channels, vascular smooth muscle contraction, and Fc gamma R-mediated phagocytosis, which were obviously associated with tumor invasive characteristics. For 130 overlapped molecules, PPI network-based molecular complex detection (MCODE) identified 10 hub molecules, namely SLC2A4, TSC2, AKT1, SCG3, ALB, APOL1, ACACA, SPARCL1, CHGB, and IGFBP5. These hub molecules are involved in multiple signaling pathways and represent potential predictive/prognostic markers in NFPA patients as well as they represent potential therapeutic targets.
This study provided the first large-scale phosphoprotein profiling and phosphorylation-related signaling pathway network alterations in human NFPA tissues. Further, overlapping analysis of phosphoproteins and invasive DEGs revealed the phosphorylation-mediated signaling pathway network changes in invasive NFPAs. These findings are the precious resource for in-depth insight into the molecular mechanisms of NFPAs, as well as for the discovery of effective phosphoprotein biomarkers and therapeutic targets for invasive NFPAs.
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acetyl-CoA carboxylase 1
automatic gain control
protein kinase B
RAC-alpha serine/threonine-protein kinase
AMP-activated protein kinase
activating transcription factor 2
cyclic nucleotide cGMP
differentially expressed gene
differentially expressed protein
- r-ERG channel:
rat ether-à-go-go-related (ERG) channel
extracellular signal–regulated kinase
endosomal sorting complex required for transport
electrospray ionization-ion trap
false discovery rate
fibroblast growth factor-2
functional pituitary adenoma
Gene Expression Omnibus
pituitary growth hormone 3
high-energy collision dissociation
high-performance liquid chromatography
insulin growth factor-1
insulin-like growth factor-binding protein 5
Kyoto Encyclopedia of Genes and Genomes
mitogen-activated protein kinase
molecular complex detection
tandem mass spectrometry
mammalian target of rapamycin
National Center for Biotechnology Information
nuclear factor kappa-B
nonfunctional pituitary adenoma
nuclear magnetic resonance
pituitary adenylyl cyclase activating polypeptide
phosphatidylinositol 3 kinase
protein kinase G
pituitary tumor-transforming gene
ribosomal S6 kinase
- Ser or S:
solute carrier family 2 facilitated glucose transporter member 4
soluble N-ethylmaleimide-sensitive fusion attachment protein
soluble N-ethylmaleimide-sensitive factor attachment protein receptor
SPARC-like protein 1
- Thr or T:
tandem mass tag
- Tyr or Y:
vascular endothelial growth factor
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The authors also acknowledge Professor Xuejun Li from Xiangya Hospital and Professor Dominic M. Desiderio from University of Tennessee Health Science Center to assist in obtaining human tissues.
This work was supported by the Shandong First Medical University Talent Introduction Funds (to X.Z.), the Hunan Provincial Hundred Talent Plan (to X.Z.), the SCIBP supported project (No. SCIBP2019090006), and China “863” Plan Project (Grant No. 2014AA020610-1 to XZ).
The authors declare that they have no conflict of interest.
Four pituitary adenoma tissue samples, obtained from the Department of Neurosurgery of Xiangya Hospital, Central South University, were approved by the Xiangya Hospital Medical Ethics Committee of Central South University. Post-mortem control pituitary tissue samples, obtained from the Memphis Regional Medical Center (n = 5), were approved by the University of Tennessee Health Science Center Internal Review Board.
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Liu, D., Li, J., Li, N. et al. Integration of quantitative phosphoproteomics and transcriptomics revealed phosphorylation-mediated molecular events as useful tools for a potential patient stratification and personalized treatment of human nonfunctional pituitary adenomas. EPMA Journal 11, 419–467 (2020). https://doi.org/10.1007/s13167-020-00215-0
- Nonfunctional pituitary adenomas
- Tandem mass tag (TMT) labeling
- TiO2 enrichment
- Quantitative phosphoproteomics
- Differentially expressed genes
- Overlapped molecule (phosphoprotein
- invasive DEG)
- Signaling pathway
- Patient stratification
- Prognostic/predictive assessment
- Personalized treatment