Phosphorylation-Mediated Molecular Pathway Changes in Human Pituitary Neuroendocrine Tumors Identified by Quantitative Phosphoproteomics.
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SUBMITTER: Li J
PROVIDER: S-EPMC8471408 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Acetylation at lysine residue in a protein mediates multiple cellular biological processes, including tumorigenesis. This study aimed to investigate the acetylated protein profile alterations and acetylation-mediated molecular pathway changes in human nonfunctional pituitary neuroendocrine tumors (NF-PitNETs). The anti-acetyl antibody-based label-free quantitative proteomics was used to analyze the acetylomes between NF-PitNETs (n = 4) and control pituitaries (n = 4). A total of 296 acetylated proteins with 517 acetylation sites was identified, and the majority of which were significantly down-acetylated in NF-PitNETs (p<0.05 or only be quantified in NF-PitNETs/controls). These acetylated proteins widely functioned in cellular biological processes and signaling pathways, including metabolism, translation, cell adhesion, and oxidative stress. The randomly selected acetylated phosphoglycerate kinase 1 (PGK1), which is involved in glycolysis and amino acid biosynthesis, was further confirmed with immunoprecipitation and western blot in NF-PitNETs and control pituitaries. Among these acetylated proteins, 15 lysine residues within 14 proteins were down-acetylated and simultaneously up-ubiquitinated in NF-PitNETs to demonstrate a direct competition relationship between acetylation and ubiquitination. Moreover, the potential effect of protein acetylation alterations on NF-PitNETs invasiveness was investigated. Overlapping analysis between acetylomics data in NF-PitNETs and transcriptomics data in invasive NF-PitNETs identified 26 overlapped molecules. These overlapped molecules were mainly involved in metabolism-associated pathways, which means that acetylation-mediated metabolic reprogramming might be the molecular mechanism to affect NF-PitNET invasiveness. This study provided the first acetylomic profiling and acetylation-mediated molecular pathways in human NF-PitNETs, and offered new clues to elucidate the biological functions of protein acetylation in NF-PitNETs and discover novel biomarkers for early diagnosis and targeted therapy of NF-PitNETs.
| S-EPMC8546192 | biostudies-literature
Project description:Background and aimAbout a third of Pituitary Neuroendocrine Tumors (PitNETs) may show aggressive behavior. Many efforts have been performed for identifying possible predictive factors to early determine the future behavior of PitNETs. Programmed cell death ligand 1 (PD-L1) expression was associated with a more aggressive biology in different solid tumors, but its role in PitNET is not well-established yet. Our study aims to analyze PD-L1 expression in a surgical cohort of PitNETs to determine its association with radiological invasion and pathology findings, as well as with long-term recurrence rates.MethodsWe performed a retrospective analysis in a series of 86 PitNETs. Clinical presentation and radiological features of the preoperative period were collected, as well as pathological data and follow-up data. The rate of PD-L1 expression was immunohistochemically evaluated and expressed as a tumor proportion score (TPS). We assessed its relationship with cavernous sinus invasion and Trouillas' classification as primary outcomes. Secondary outcomes included the TPS' relationship with histopathological markers of proliferation, hormonal expression, tumor size and long-term recurrence rates. We calculated the optimal cut-point for the primary outcomes while maximizing the product of the sensitivity and specificity and then we evaluated the significance of secondary outcomes with logistic regression analysis.ResultsEighty-six patients were included in the analysis; 50 cases were non-functional PitNETs. The TPS for PD-L1 showed a highly right-skewed distribution in our sample, as 30.2% of patients scored 0. Using Trouillas' classification, we found that "proliferative" cases have a significantly higher probability to express PD-L1 in more than 30% of tumor cells (OR: 5.78; CI 95%: 1.80-18.4). This same cut-point was also associated with p53 expression. A positive association was found between PD-L1 expression and GH expression (p = 0.001; OR: 5.44; CI 95%: 1.98-14.98), while an inverse relationship was found with FSH/LH expression (p = 0.014; OR = 0.27, CI 95%: 0.10-0.76). No association was found with CS invasion, tumor size, bone erosion or dura invasion. We could not find any association between PD-L1 expression and recurrence.ConclusionsPD-L1 expression was associated with proliferative grades of Trouillas' classification and p53 expression. We also confirmed a higher expression of PD-L1 in somatotroph tumors. Larger studies are necessary to investigate the relationship between PD-L1 expression and aggressive behaviors.
| S-EPMC10526513 | biostudies-literature
Project description:The modification of intracellular proteins by monosaccharides of O-linked ?-N-acetylglucosamine (O-GlcNAc) is an essential and dynamic PTM of metazoans. The addition and removal of O-GlcNAc is catalyzed by the O-GlcNAc transferase (OGT) and O-GlcNAcase, respectively. One mechanism by which O-GlcNAc is thought to mediate proteins is by regulating phosphorylation. To provide insight into the pathways regulated by O-GlcNAc, we have utilized SILAC-based quantitative proteomics to carry out comparisons of site-specific phosphorylation in OGT wild-type and Null cells. Quantitation of the phosphoproteome demonstrated that of 5529 phosphoserine, phosphothreonine, and phosphotyrosine sites, 232 phosphosites were upregulated and 133 downregulated in the absence of O-GlcNAc. Collectively, these data suggest that deletion of OGT has a profound effect on the phosphorylation of cell cycle and DNA damage response proteins. Key events were confirmed by biochemical analyses and demonstrate an increase in the activating autophosphorylation event on ATM (Ser1987) and on ATM's downstream targets p53, H2AX, and Chk2. Together, these data support widespread changes in the phosphoproteome upon removal of O-GlcNAc, suggesting that O-GlcNAc regulates processes such as the cell cycle, genomic stability, and lysosomal biogenesis. All MS data have been deposited in the ProteomeXchange with identifier PXD001153 (http://proteomecentral.proteomexchange.org/dataset/PXD001153).
| S-EPMC4564869 | biostudies-literature
Project description:The most common genetic drivers of pituitary neuroendocrine tumors (PitNETs) lie within mutational hotspots, which are genomic regions where variants tend to cluster. Some of these hotspot defects are unique to PitNETs, while others are associated with additional neoplasms. Hotspot variants in GNAS and USP8 are the most common genetic causes of acromegaly and Cushing's disease, respectively. Although it has been proposed that these genetic defects could define specific clinical phenotypes, results are highly variable among studies. In contrast, DICER1 hotspot variants are associated with a familial syndrome of cancer predisposition, and only exceptionally occur as somatic changes. A small number of non-USP8-driven corticotropinomas are due to somatic hotspot variants in USP48 or BRAF; the latter is a well-known mutational hotspot in cancer. Finally, somatic variants affecting a hotspot in SF3B1 have been associated with multiple cancers and, more recently, with prolactinomas. Since the associations of BRAF, USP48, and SF3B1 hotspot variants with PitNETs are very recent, their effects on clinical phenotypes are still unknown. Further research is required to fully define the role of these genetic defects as disease biomarkers and therapeutic targets.
| S-EPMC10705109 | biostudies-literature
Project description:BackgroundInvasiveness 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.MethodsThe 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.ResultsIn 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.ConclusionsThis 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.
| S-EPMC7429628 | biostudies-literature
Project description:The clinical characteristics of growth hormone (GH)-producing pituitary adenomas/somatotroph pituitary neuroendocrine tumors (GHomas/somatotroph PitNETs) vary across patients. In this study, we aimed to integrate the genetic alterations, protein expression profiles, transcriptomes, and clinical characteristics of GHomas/somatotroph PitNETs to identify molecules associated with acromegaly characteristics. Targeted capture sequencing and copy number analysis of 36 genes and nontargeted proteomics analysis were performed on fresh-frozen samples from 121 sporadic GHomas/somatotroph PitNETs. Targeted capture sequencing revealed GNAS as the only driver gene, as previously reported. Classification by consensus clustering using both RNA sequencing and proteomics revealed many similarities between the proteome and the transcriptome. Gene ontology analysis was performed for differentially expressed proteins between wild-type and mutant GNAS samples identified by nontargeted proteomics and involved in G protein-coupled receptor (GPCR) pathways. The results suggested that GNAS mutations impact endocrinological features in acromegaly through GPCR pathway induction. ATP2A2 and ARID5B correlated with the GH change rate in the octreotide loading test, and WWC3, SERINC1, and ZFAND3 correlated with the tumor volume change rate after somatostatin analog treatment. These results identified a biological connection between GNAS mutations and the clinical and biochemical characteristics of acromegaly, revealing molecules associated with acromegaly that may affect medical treatment efficacy.
| S-EPMC9701206 | biostudies-literature
Project description:IntroductionAngiotensin-converting enzyme 2 (ACE2) is the receptor of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The effects of SARS-CoV-2 on normal pituitary glands function or pituitary neuroendocrine tumors (PitNETs) have not yet been elucidated. Thus, the present study aimed to investigate the potential risks of SARS-CoV-2 infection on the impairment of pituitary glands and the development of PitNETs.MethodsPitNETs tissues were obtained from 114 patients, and normal pituitary gland tissues were obtained from the autopsy. The mRNA levels of ACE2 and angiotensin II receptor type 1 (AGTR1) were examined by quantitative real-time PCR. Immunohistochemical staining was performed for ACE2 in 69 PitNETs and 3 normal pituitary glands. The primary tumor cells and pituitary cell lines (MMQ, GH3 and AtT-20/D16v-F2) were treated with diminazene aceturate (DIZE), an ACE2 agonist, with various dose regimens. The pituitary hormones between 43 patients with SARS-CoV-2 infection were compared with 45 healthy controls.ResultsPituitary glands and the majority of PitNET tissues showed low/negative ACE2 expression at both the mRNA and protein levels, while AGTR1 showed high expression in normal pituitary and corticotroph adenomas. ACE2 agonist increased the secretion of ACTH in AtT-20/D16v-F2 cells through downregulating AGTR1. The level of serum adrenocorticotropic hormone (ACTH) was significantly increased in COVID-19 patients compared to normal controls (p < 0.001), but was dramatically decreased in critical cases compared to non-critical patients (p = 0.003).ConclusionsThis study revealed a potential impact of SARS-CoV-2 infection on corticotroph cells and adenomas.
| S-EPMC8009460 | biostudies-literature
Project description:Pituitary neuroendocrine tumors (PitNETs) are common, generally benign tumors with complex clinical characteristics related to hormone hypersecretion and/or growing sellar tumor mass. PitNETs can be classified based on the expression pattern of anterior pituitary hormones and three main transcriptions factors (TF), SF1, PIT1 and TPIT that regulate differentiation of adenohypophysial cells. Here, we have extended this classification based on the global transcriptomics landscape using tumor tissue from a well-defined cohort comprising 51 PitNETs of different clinical and histological types. The molecular profiles were compared with current classification schemes based on immunohistochemistry. Our results identified three main clusters of PitNETs that were aligned with the main pituitary TFs expression patterns. Our analyses enabled further identification of specific genes and expression patterns, including both known and unknown genes, that could distinguish the three different classes of PitNETs. We conclude that the current classification of PitNETs based on the expression of SF1, PIT1 and TPIT reflects three distinct subtypes of PitNETs with different underlying biology and partly independent from the expression of corresponding hormones. The transcriptomic analysis reveals several potentially targetable tumor-driving genes with previously unknown role in pituitary tumorigenesis.
| S-EPMC8579660 | biostudies-literature
Project description:BackgroundPituitary neuroendocrine tumors (PitNETs) are the second most common intracranial tumor. We lacked a comprehensive understanding of the pathogenesis and heterogeneity of these tumors.MethodsWe performed high-precision single-cell RNA sequencing for 2679 individual cells obtained from 23 surgically resected samples of the major subtypes of PitNETs from 21 patients. We also performed single-cell multi-omics sequencing for 238 cells from 5 patients.ResultsUnsupervised clustering analysis distinguished all tumor subtypes, which was in accordance with the classification based on immunohistochemistry and provided additional information. We identified 3 normal endocrine cell types: somatotrophs, lactotrophs, and gonadotrophs. Comparisons of tumor and matched normal cells showed that differentially expressed genes of gonadotroph tumors were predominantly downregulated, while those of somatotroph and lactotroph tumors were mainly upregulated. We identified novel tumor-related genes, such as AMIGO2, ZFP36, BTG1, and DLG5. Tumors expressing multiple hormone genes showed little transcriptomic heterogeneity. Furthermore, single-cell multi-omics analysis demonstrated that the tumor had a relatively uniform pattern of genome with slight heterogeneity in copy number variations.ConclusionsOur single-cell transcriptome and single-cell multi-omics analyses provide novel insights into the characteristics and heterogeneity of these complex neoplasms for the identification of biomarkers and therapeutic targets.
| S-EPMC8563320 | biostudies-literature
Project description:The purpose of this study was to analyze the impact of copy number variations (CNV) on sporadic pituitary neuroendocrine tumors (PitNETs) prognosis, to identify specific prognosis markers according to the known clinico-pathological classification. CGH array analysis was performed on 195 fresh-frozen PitNETs (56 gonadotroph, 11 immunonegative, 56 somatotroph, 39 lactotroph and 33 corticotroph), with 5 years post-surgery follow-up (124 recurrences), classified according to the five-tiered grading classification (invasion, Ki-67, mitotic index and p53 positivity). Effect of alterations on recurrence was studied using logistic regression models. Transcriptomic analysis of 32 lactotroph tumors was performed. The quantity of CNV was dependent on tumor type: higher in lactotroph (median(min-max) = 38% (0-97) of probes) compared to corticotroph (11% (0-77)), somatotroph (5% (0-99)), gonadotroph (0% (0-10)) and immunonegative tumors (0% (0-17). It was not predictive of recurrence in the whole cohort. In lactotroph tumors, genome instability, especially quantity of gains, significantly predicted recurrence independently of invasion and proliferation (p-value = 0.02, OR = 1.2). However, no specific CNV was found as a prognostic marker. Transcriptomic analysis of the genes included in the CNV and associated with prognosis didn't show significantly overrepresented pathway. In somatotroph and corticotroph tumors, USP8 and GNAS mutations were not associated with genome disruption or recurrence respectively. To conclude, CGH array analysis showed genome instability was dependent on PitNET type. Lactotroph tumors were highly altered and the quantity of altered genome was associated with poorer prognosis though the mechanism is unclear, whereas gonadotroph and immunonegative tumors showed the same 'quiet' profile, leaving the mechanism underlying tumorigenesis open to question.
| S-EPMC7653703 | biostudies-literature