bims-prodis 2019-05-26 papers

bims-prodis

Biomed News

on Proteomics in disease

Issue of 2019–05–26
twenty-one papers selected by
Nancy Gough, Bioserendipity

Identification of Predictive Proteins and Biological Pathways for the Tumorigenicity of Vestibular Schwannoma by Proteomic Profiling.
In-depth proteomic analysis of human bronchoalveolar lavage fluid towards the biomarker discovery for lung cancers.
Serum Proteomics on the Basis of Discovery of Predictive Biomarkers of Response to Androgen Deprivation Therapy in Advanced Prostate Cancer.
Inflammaging Determines Health and Disease in Lumbar discs - Evidence from Differing Proteomic Signatures of Healthy, Ageing and Degenerating Discs.
Identification of proteomic signatures associated with COPD frequent exacerbators.
iTRAQ comparison of proteomic profiles of endometrial receptivity.
Modulation of phospho-proteins by interferon-alpha and valproic acid in acute myeloid leukemia.
Proteomic insight into the pathogenesis of CAPN5-vitreoretinopathy.
LABEL-FREE CERVICOVAGINAL FLUID PROTEOME PROFILING REFLECTS THE CERVIX NEOPLASTIC TRANSFORMATION.
Proteomic Profiling of Signaling Networks Modulated by G-CSF/Plerixafor/Busulfan-Fludarabine Conditioning in Acute Myeloid Leukemia Patients in Remission or with Active Disease prior to Allogeneic Stem Cell Transplantation.
Proteomic Bioprofiles and Mechanistic Pathways of Progression to Heart Failure.
2D-DIGE as a strategy to identify serum protein biomarkers to monitor pharmacological efficacy in dopamine-dictated states of Parkinson's disease and schizophrenia.
From clinic to mechanism: Proteomics-based assessment of angiogenesis in adrenal pheochromocytoma.
Differential proteome analysis in adolescent idiopathic scoliosis patients with thoracolumbar/lumbar curvatures.
LGALS3 is connected to CD74 in a previously unknown protein network that is associated with poor survival in patients with AML.
Pan-cancer analysis connects tumor matrisome to immune response.
The serum proteomics tracking of hepatocellular carcinoma early recurrence following radical resection.
Leukocyte proteomic profiling in first-episode schizophrenia patients: does oxidative stress play central roles in the pathophysiology network of schizophrenia?
Three kinds of corneal host cells contribute differently to corneal neovascularization.
Novel Biomarkers to Assess In-Utero Effects of Maternal Opioid Use: First Steps towards Understanding Short and Long Term Neurodevelopmental Sequelae.
Simultaneous Integration of Multi-omics Data Improves the Identification of Cancer Driver Modules.

Proteomics Clin Appl. 2019 May 23. e1800175

Identification of Predictive Proteins and Biological Pathways for the Tumorigenicity of Vestibular Schwannoma by Proteomic Profiling.

Jianhui Xu, Yang Zhang, Yuxuan Shi, Dongming Yin, Peidong Dai, Weidong Zhao, Tianyu Zhang.

   PURPOSE: Vestibular schwannomas (VSs) are benign tumors that account for 8%-10% of all intracranial tumors. So far, the tumorigenesis of VS has not been fully elucidated. This study is designed to identify differently expressed proteins involved in VS tumorigenesis.
EXPERIMENTAL DESIGN: We used an isobaric tag for relative and absolute quantification (iTRAQ) approach to characterize the protein expression profiles from pooled VS tissues(n = 12) and pooled matched normal vestibular tissues(n = 12).
RESULTS: A total of 933 differentially expressed proteins were identified between VS and the matched normal vestibular tissues, with 489 being upregulated and 444 being downregulated. Bioinformatics analyses were performed according to Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses. Several of the differentially expressed proteins were validated by western blotting analyses, and upregulation of LGALS1, ANXA1, GRB2, and STAT1 was validated in VS tissue by immunohistochemistry.
CONCLUSIONS AND CLINICAL RELEVANCE: Our study represents the successful application of iTRAQ technology to an investigation of VS. Many of the differentially expressed proteins identified here have not been linked to VS before, and these dysregulated proteins may provide potential biomarkers for human VS diagnosis. This article is protected by copyright. All rights reserved.

Keywords:  ANXA1; GRB2; LGALS1; STAT1 ; iTRAQ; proteomics; vestibular schwannoma

DOI:  https://doi.org/10.1002/prca.201800175
Proteomics Clin Appl. 2019 May 23. e1900028

In-depth proteomic analysis of human bronchoalveolar lavage fluid towards the biomarker discovery for lung cancers.

Seo Young Sim, Yu Ri Choi, Jun Hyung Lee, Jae Min Lim, Seung-Eun Lee, Kwang Pyo Kim, Jin Young Kim, Seung Hyeun Lee, Min-Sik Kim.

   PURPOSE: Lung cancer is among the most common cancers. Bronchoalveolar lavage fluid (BALF) is a fluid that can be easily obtained from patients with lung cancers. We aimed at developing a novel proteomic method of the molecule-based sensitive detection of biomarkers from BALF.
EXPERIMENTAL DESIGN: BALF samples were collected from segmental bronchus of 14 patients with lung cancers from Kyung Hee Hospital. First, BALF proteome was depleted using a depletion column, and then peptides were prepared from the enriched low abundant proteins and fractionated by high pH reverse phase liquid chromatography. Each fraction was analyzed by LC-MS/MS. Data are available via ProteomeXchange with identifier PXD012645.
RESULTS: We developed a novel method for in-depth proteomic analysis of BALF by combining antibody-based depletion of high abundant proteins from BALF with high pH peptide fractionation. Peptides were analyzed on a high resolution Orbitrap Fusion mass spectrometer. MaxQuant search resulted in the identification of 4,615 protein groups mapped to 4,534 genes.
CONCLUSIONS AND CLINICAL RELEVANCE: We found that our method outperformed conventional BALF proteomic methods and believe that this method would facilitate the biomarker research for the lung cancer. In addition, we showed BALF would be a great source of biomarkers of lung diseases. This article is protected by copyright. All rights reserved.

Keywords:  bronchoalveolar lavage fluid (BALF); depletion; high-pH fractionation; lung cancer

DOI:  https://doi.org/10.1002/prca.201900028
Clin Genitourin Cancer. 2019 Mar 15. pii: S1558-7673(18)30661-X. [Epub ahead of print]

Serum Proteomics on the Basis of Discovery of Predictive Biomarkers of Response to Androgen Deprivation Therapy in Advanced Prostate Cancer.

Manish Kohli, Ann L Oberg, Douglas W Mahoney, Shaun M Riska, Robert Sherwood, Yuzi Zhang, Roman M Zenka, Deepak Sahasrabudhe, Rui Qin, Sheng Zhang.

   BACKGROUND: We investigated the serum proteome of hormone-sensitive prostate cancer patients to determine candidate biomarkers associated with androgen deprivation therapy (ADT) efficacy.
PATIENTS AND METHODS: Serum proteomes generated using isobaric mass tags for relative and absolute quantitation were analyzed using reverse-phase liquid chromatography coupled to tandem mass spectrometry. The advanced hormone-sensitive prostate cancer cohorts studied were: (1) untreated "paired" pre-ADT and 4-month post-ADT hormone-sensitive patients (n = 15); (2) "early ADT failure" patients (n = 10) in whom ADT treatment failed within a short period of time; and (3) "late ADT failure" patients (n = 10) in whom ADT treatment failed after a prolonged response time. Differential abundance was assessed, and ingenuity pathway analysis (IPA) was used to identify interaction networks in selected candidates from these comparisons.
RESULTS: Between "post-ADT" and combined "early" and "late" ADT failure groups 149 differentially detected candidates were observed, and between "early" and "late" ADT failure groups 98 candidates were observed; 47 candidates were common in both comparisons. IPA network enrichment analysis of the 47 candidates identified 3 interaction networks (P < .01) including 17-β-estradiol, nuclear factor kappa-light-chain enhancer of activated B cells complex, and P38 mitogen-activated protein kinases as pathways with potential markers of response to ADT.
CONCLUSION: A global proteomic analysis identified pathways with markers of ADT response, which will need validation in independent data sets.

Keywords:  Hormonal therapy; Predictive factors; Prostate cancer; Proteomics; Serum

DOI:  https://doi.org/10.1016/j.clgc.2019.03.006
Spine J. 2019 May 21. pii: S1529-9430(19)30192-5. [Epub ahead of print]

Inflammaging Determines Health and Disease in Lumbar discs - Evidence from Differing Proteomic Signatures of Healthy, Ageing and Degenerating Discs.

S Rajasekaran, Chitraa Tangavel, Sri Vijay Anand, Dilip Chand Raja, Sharon Miracle Nayagam, Monica Steffi Matchado, M Raveendran, Ajoy Prasad Shetty, Rishi Mugesh Kanna, K Dharmalingam.

   BACKGROUND CONTEXT: The true understanding of aging and disc degeneration (DD) is still elusive. MRI has not helped our attempts to understand the health and disease status of the discs as it reflects mainly the end morphological changes and not the changes at a molecular level. Understanding degeneration at a molecular level through proteomics might allow differentiation from normal aging and also aid in the development of biomarkers for early diagnosis and preventive therapies.
PURPOSE: To utilize proteomics to understand the molecular basis of healthy, aging and degenerating discs and conclusively differentiate normal aging and degeneration.
STUDY DESIGN: Proteomic analysis of human intervertebral disc samples METHODS: L4-L5 disc samples from three groups were acquired and subjected to proteomic analysis. Samples from individuals aged in the 2nd, 3rd, and 4th decades were used to represent young healthy discs (Group A). Those from MRI normal donors aged in the 5th, 6th and 7th decades represented normal aging (Group B). Five degenerated discs obtained from patients at surgery represented degeneration (Group C). The entire proteome map and alteration in protein expressions were further analyzed using bioinformatics analysis. This was a self-funded project.
RESULTS: There were 84 common proteins. Specific proteins numbered 225 in A, 315 in B and 283 in C. By gene ontology biological process identification, Group A predominated with extracellular matrix organization, cytoskeletal structural and normal metabolic proteins. Group B differed in having additional basal expression of immune response, complement inhibitors and senescence proteins. Group C was different, with up regulation of proteins associated with oxidative stress response, positive regulators of apoptosis, innate immune response, complement activation and defense response to gram positive bacteria indicating ongoing Inflammaging.
CONCLUSIONS: Our study documented diverse proteome signatures between the young, aging and degenerating discs. Inflammaging was the main differentiator between normal biological aging and DD.
CLINICAL SIGNIFICANCE: Multiple inflammatory molecules unique to DD were identified, allowing the possibility of developing specific biomarkers for early diagnosis and thereby provide evidence-based metrics for preventive measures rather than surgical intervention and also to monitor progress of the disease.

Keywords:  Apoptosis; Disc degeneration; Inflammaging; Intervertebral disc; Low back pain; Proteomics; aging disc

DOI:  https://doi.org/10.1016/j.spinee.2019.04.023
Life Sci. 2019 May 20. pii: S0024-3205(19)30403-5. [Epub ahead of print]

Identification of proteomic signatures associated with COPD frequent exacerbators.

Pengbo Sun, Ruin Ye, Cuihong Wang, Shuang Bai, Li Zhao.

   AIMS: Acute exacerbation is a major event that alters the natural course of chronic obstructive pulmonary disease (COPD), and recurrent exacerbation results in worse clinical outcomes and greater economic consequences. While some patients suffer frequent exacerbations, others experience no exacerbations; this study was designed to detect proteins that were differentially abundant in COPD frequent exacerbators and assess whether those expression profiles are unique among COPD patients.
MAIN METHODS: Tandem mass tag labeled quantitative proteomics combined with two-dimensional liquid chromatography-tandem mass spectrometry was used to detect the changes in the lung proteome in COPD frequent exacerbators and infrequent exacerbators. A series of bioinformatics analyses were performed to screen potential signatures of COPD frequent exacerbations. The accuracy of proteomic results was further verified by western blot studies.
KEY FINDINGS: Compared with infrequent exacerbators, 23 proteins in the lung tissues from frequent exacerbators showed significant degrees of differential expression; combined bioinformatics analyses of proteome indicated that the immune network for IgA production and the phenylalanine metabolism pathway were associated with frequent exacerbations. The Western blot analysis confirmed the expression pattern of three significantly regulated proteins (HLA-DQA1, pIgR and biglycan).
SIGNIFICANCE: These findings indicate that immune response might play a key role in the pathophysiological mechanisms of COPD frequent exacerbations. Our results make a crucial contribution to the search for a comprehensive understanding of potential pathophysiological mechanisms associated with the frequent exacerbations of COPD, and might provide guidance for treating frequent exacerbations.

Keywords:  COPD; Frequent exacerbators; Human lung tissue; Immune response; Proteomic profile

DOI:  https://doi.org/10.1016/j.lfs.2019.05.047
J Proteomics. 2019 May 15. pii: S1874-3919(19)30153-8. [Epub ahead of print]203 103381

iTRAQ comparison of proteomic profiles of endometrial receptivity.

Silvia Pérez-Debén, José Bellver, Pilar Alamá, Stefania Salsano, Alicia Quiñonero, Patricia Sebastian-Leon, Patricia Díaz-Gimeno, Francisco Domínguez.

  Endometrial receptivity is a limiting step in human reproduction. A disruption in the development of endometrial receptivity is responsible for recurrent implantation failures (RIF) of endometrial origin. To understand the molecular mechanisms behind the endometrial receptivity process, we used the isobaric tag for relative and absolute quantitation (iTRAQ) method to compare three different endometrial statuses: fertile women, intrauterine device (IUD) carriers, and RIF patients. Overall, iTRAQ allowed identified 1889 non-redundant proteins. Of these, 188 were differentially expressed proteins (DEP) (p-value < .05). Pairwise comparisons revealed 133 significant DEP in fertile vs. IUD carriers and 158 DEP in RIF vs. IUD carriers. However, no DEP were identified between fertile and RIF patients. Western blot validation of three DEP involved in endometrial receptivity (plastin 2, lactotransferrin, and lysozyme) confirmed our iTRAQ results. Moreover, functional KEGG enrichment revealed that complement and coagulation cascades and peroxisome were the two most significant pathways for the RIF vs. IUD comparison and ribosome and spliceosome for the fertile vs. IUD comparison, as possible important pathways involved in the endometrial receptivity acquisition. The lack of DEP between fertile and RIF patient endometria suggest that idiopathic RIF may not have an endometrial origin, with other as-yet-unknown factors involved. SIGNIFICANCE: A pilot study where a comparison of the endometrial protein profile from women with different endometrial receptive grade (fertile women, IUD carriers and RIF patients) during the same period of time (overlapping with the window of implantation) of a hormone replacement therapy was performed using a high-throughput proteomic technique. This approach lead us to better understand the molecular mechanisms undergoing endometrial receptivity, a time-limiting step to achieve pregnancy in humans. Moreover, the number of samples per group (10 Fertile women, 10 IUD carriers and 8 RIF patients) according to the methodology here employed (8plex iTRAQ), give more robustness to our results. Our findings confirm that an IUD introduces numerous changes in the endometrial protein profile when compared to fertile and RIF endometria, revealing some key proteins involved in endometrial receptivity. Finding no significant differences between Fertile and RIF patient endometria could suggest that other as-yet-unknown factors could be involved in the etiology of idiopathic RIF.

Keywords:  Endometrial receptivity; Endometrium; Intrauterine device (IUD); Isobaric tags for relative and absolute quantification (iTRAQ); Recurrent implantation failure (RIF)

DOI:  https://doi.org/10.1016/j.jprot.2019.103381
J Cancer Res Clin Oncol. 2019 May 20.

Modulation of phospho-proteins by interferon-alpha and valproic acid in acute myeloid leukemia.

Rakel Brendsdal Forthun, Monica Hellesøy, André Sulen, Reidun Kristin Kopperud, Gry Sjøholt, Øystein Bruserud, Emmet McCormack, Bjørn Tore Gjertsen.

   PURPOSE: Valproic acid (VPA) is suggested to be therapeutically beneficial in combination with interferon-alpha (IFNα) in various cancers. Therefore, we examined IFNα and VPA alone and in combinations in selected AML models, examining immune regulators and intracellular signaling mechanisms involved in phospho-proteomics.
METHODS: The anti-leukemic effects of IFNα and VPA were examined in vitro and in vivo. We mapped the in vitro phosphoprotein modulation by IFNα-2b and human IFNα-Le in MOLM-13 cells by IMAC/2D DIGE/MS analysis and phospho-flow cytometry, and in primary healthy and AML patient-derived PBMCs by CyTOF. In vivo, IFNα-Le and VPA efficacy were investigated in the immunodeficient NOD/Scid IL2γ-/- MOLM-13Luc+ mouse model and the syngeneic immunocompetent BNML rat model.
RESULTS: IFNα-2b and IFNα-Le differed in the modulation of phospho-proteins involved in protein folding, cell stress, cell death and p-STAT6 Y641, whereas VPA and IFNα-Le shared signaling pathways involving phosphorylation of Akt (T308), ERK1/2 (T202/T204), p38 (T180/Y182), and p53 (S15). Both IFNα compounds induced apoptosis synergistically with VPA in vitro. However, in vivo, VPA monotherapy increased survival, but no benefit was observed by IFNα-Le treatment. CyTOF analysis of primary human PBMCs indicated that lack of immune-cell activation could be a reason for the absence of response to IFNα in the animal models investigated.
CONCLUSIONS: IFNα-2b and IFNα-Le showed potent and synergistic anti-leukemic effects with VPA in vitro but not in leukemic mouse and rat models in vivo. The absence of IFNα immune activation in lymphocyte subsets may potentially explain the limited in vivo anti-leukemic effect of IFNα-monotherapy in AML.

Keywords:  AML; CyTOF; IFNα; Phospho-flow; Phosphoproteome; VPA

DOI:  https://doi.org/10.1007/s00432-019-02931-1
Sci Rep. 2019 May 20. 9(1): 7608

Proteomic insight into the pathogenesis of CAPN5-vitreoretinopathy.

Gabriel Velez, Jing Yang, Angela S Li, Stephen H Tsang, Alexander G Bassuk, Vinit B Mahajan.

CAPN5 Neovascular Inflammatory Vitreoretinopathy (CAPN5-NIV; OMIM 193235) is a poorly-understood rare, progressive inflammatory intraocular disease with limited therapeutic options. To profile disease effector proteins in CAPN5-NIV patient vitreous, liquid vitreous biopsies were collected from two groups: eyes from control subjects (n = 4) with idiopathic macular holes (IMH) and eyes from test subjects (n = 12) with different stages of CAPN5-NIV. Samples were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Protein expression changes were evaluated by principal component analysis, 1-way ANOVA (significant p-value < 0.05), hierarchical clustering, gene ontology, and pathway representation. There were 216 differentially-expressed proteins (between CAPN5-NIV and control vitreous), including those unique to and abundant in each clinical stage. Gene ontology analysis revealed decreased synaptic signaling proteins in CAPN5-NIV vitreous compared to controls. Pathway analysis revealed that inflammatory mediators of the acute phase response and the complement cascade were highly-represented. The CAPN5-NIV vitreous proteome displayed characteristic enrichment of proteins and pathways previously-associated with non-infectious posterior uveitis, rhegmatogenous retinal detachment (RRD), age-related macular degeneration (AMD), proliferative diabetic retinopathy (PDR), and proliferative vitreoretinopathy (PVR). This study expands our knowledge of affected molecular pathways in CAPN5-NIV using unbiased, shotgun proteomic analysis rather than targeted detection platforms. The high-levels and representation of acute phase response proteins suggests a functional role for the innate immune system in CAPN5-NIV pathogenesis.

DOI: https://doi.org/10.1038/s41598-019-44031-7
J Mass Spectrom. 2019 May 22.

LABEL-FREE CERVICOVAGINAL FLUID PROTEOME PROFILING REFLECTS THE CERVIX NEOPLASTIC TRANSFORMATION.

Natalia L Starodubtseva, Alexander G Brzhozovzkiy, Anna E Bugrova, Alexey S Kononikhin, Maria I Indeykina, Kiril I Gusakov, Vitaliy V Chagovets, Niso M Nazarova, Vladimir E Frankevich, Gennady T Sukhikh, Eugene N Nikolaev.

  Cervicovaginal fluid (CVF) is a valuable source of clinical information about the female reproductive tract in both non-pregnant and pregnant women. The aim of this study is to specify the CVF proteome at different stages of cervix neoplastic transformation by label-free quantitation approach based on LC-MS/MS method. The proteome composition of CVF from 40 women of reproductive age with HPV-associated cervix neoplastic transformation (LSIL, HSIL, CANCER) was investigated. Hierarchical clustering and principal component analysis (PCA) of the proteomic data obtained by a label-free quantitation approach shows the distribution of the sample set between four major clusters (NILM, LSIL, HSIL and CANCER) depending on the form of cervical lesion. Multisample ANOVA with subsequent Welch's t-test resulted in 117 that changed significantly across the four clinical stages, including 27 proteins significantly changed in cervical cancer. Some of them were indicated as promising biomarkers previously (ACTN4, VTN, ANXA1, CAP1, ANXA2, MUC5B). CVF proteomic data from the discovery stage was analyzed by the PLS-DA method to build a statistical model, allowing to differentiate severe dysplasia (HSIL, CANCER) from the mild/normal stage (NILM and LSIL) and ROC AUC were obtained on an independent set of 33 samples. The sensitivity of the model was 77% and the specificity - 94%, AUC was equal to 0.87. CVF proteome proved to be reflect the stage of cervical epithelium neoplastic process.

Keywords:  HSIL; LSIL; Label-free proteomics; cervical cancer; cervicovaginal fluid; mass-spectrometry

DOI:  https://doi.org/10.1002/jms.4374
Acta Haematol. 2019 May 21. 1-9

Proteomic Profiling of Signaling Networks Modulated by G-CSF/Plerixafor/Busulfan-Fludarabine Conditioning in Acute Myeloid Leukemia Patients in Remission or with Active Disease prior to Allogeneic Stem Cell Transplantation.

Zhihong Zeng, Wenbin Liu, Christopher B Benton, Sergej Konoplev, Hongbo Lu, Rui-Yu Wang, Julianne Chen, Elizabeth Shpall, Keith A Baggerly, Richard Champlin, Marina Konopleva.

  To characterize intracellular signaling in peripheral blood (PB) cells of acute myeloid leukemia (AML) patients undergoing pretransplant conditioning with CXCR4 inhibitor plerixafor, granulocyte colony-stimulating factor (G-CSF), and busulfan plus fludarabine (Bu+Flu) chemotherapy, we profiled 153 proteins in 33 functional groups using reverse phase protein array. CXCR4 inhibition mobilized AML progenitors and clonal AML cells, and this was associated with molecular markers of cell cycle progression. G-CSF/plerixafor and G-CSF/plerixafor/Bu+Flu modulated distinct signaling networks in AML blasts of patients undergoing conditioning with active disease compared to nonleukemic PB cells of patients in remission. We identified AML-specific proteins that remained aberrantly expressed after chemotherapy, representing putative chemoresistance markers in AML.

Keywords:  Acute myeloid leukemia; Allogeneic stem cell transplantation; CXCR4; Plerixafor; Proteomic profiling of signaling

DOI:  https://doi.org/10.1159/000495456
Circ Heart Fail. 2019 May;12(5): e005897

Proteomic Bioprofiles and Mechanistic Pathways of Progression to Heart Failure.

João Pedro Ferreira, Job Verdonschot, Timothy Collier, Ping Wang, Anne Pizard, Christian Bär, Jens Björkman, Alessandro Boccanelli, Javed Butler, Andrew Clark, John G Cleland, Christian Delles, Javier Diez, Nicolas Girerd, Arantxa González, Mark Hazebroek, Anne-Cécile Huby, Wouter Jukema, Roberto Latini, Joost Leenders, Daniel Levy, Alexandre Mebazaa, Harald Mischak, Florence Pinet, Patrick Rossignol, Naveed Sattar, Peter Sever, Jan A Staessen, Thomas Thum, Nicolas Vodovar, Zhen-Yu Zhang, Stephane Heymans, Faiez Zannad.

  Background Identifying the mechanistic pathways potentially associated with incident heart failure (HF) may provide a basis for novel preventive strategies. Methods and Results To identify proteomic biomarkers and the potential underlying mechanistic pathways that may be associated with incident HF defined as the first hospitalization for HF, a nested-matched case-control design was used with cases (incident HF) and controls (without HF) selected from 3 cohorts (>20 000 individuals). Controls were matched on cohort, follow-up time, age, and sex. Two independent sample sets (a discovery set with 286 cases and 591 controls and a replication set with 276 cases and 280 controls) were used to discover and replicate the findings. Two hundred fifty-two circulating proteins in the plasma were studied. Adjusting for the matching variables age, sex, and follow-up time (and correcting for multiplicity of tests), 89 proteins were found to be associated with incident HF in the discovery phase, of which 38 were also associated with incident HF in the replication phase. These 38 proteins pointed to 4 main network clusters underlying incident HF: (1) inflammation and apoptosis, indicated by the expression of the TNF (tumor necrosis factor)-family members; (2) extracellular matrix remodeling, angiogenesis and growth, indicated by the expression of proteins associated with collagen metabolism, endothelial function, and vascular homeostasis; (3) blood pressure regulation, indicated by the expression of natriuretic peptides and proteins related to the renin-angiotensin-aldosterone system; and (4) metabolism, associated with cholesterol and atherosclerosis. Conclusions Clusters of biomarkers associated with mechanistic pathways leading to HF were identified linking inflammation, apoptosis, vascular function, matrix remodeling, blood pressure control, and metabolism. These findings provide important insight on the pathophysiological mechanisms leading to HF. Clinical Trial Registration: URL: https://www.clinicaltrials.gov . Unique identifier: NCT02556450.

Keywords:  apoptosis; atherosclerosis; blood pressure; heart failure; proteomics

DOI:  https://doi.org/10.1161/CIRCHEARTFAILURE.118.005897
Neuropsychiatr Dis Treat. 2019 ;15 1031-1044

2D-DIGE as a strategy to identify serum protein biomarkers to monitor pharmacological efficacy in dopamine-dictated states of Parkinson's disease and schizophrenia.

Ashish Kumar Gupta, Gaurav Khunger Kumar, Komal Rani, Ruchika Pokhriyal, Mohd Imran Khan, Domada Ratna Kumar, Vinay Goyal, Manjari Tripathi, Rishab Gupta, Rakesh Kumar Chadda, Perumal Vanamail, Ashok Kumar Mohanty, Gururao Hariprasad.

   Objectives: Parkinson's disease and schizophrenia are clinical scenarios that occur due to dopaminergic deficit and hyperactivity in the midbrain, respectively. Current pharmacological interventions for these two diseases therefore aim to restore normal dopamine levels in the midbrain. But during therapy, there is a overshooting of dopamine concentrations that result in hallucinations in Parkinson's disease patients and extra-pyramidal symptoms in schizophrenic patients. This causes a lot of inconvenience to the patents and the clinicians. There are no tests currently available to monitor drug efficacy in these two neuropsychiatric diseases.
Materials and methods: Parkinson's disease and schizophrenic naïve patients were recruited. Serum proteins isolated from these two clinical phenotypes were labeled with fluorescent cyanine dyes and analyzed by two-dimensional difference in gel electrophoresis proteomic experiment. Differentially expressed spots that had consistent expression pattern across five sets of biological replicate gels were trypsin digested and subjected to mass spectrometric analysis for protein identification. Validation experiments were done for the identified proteins using antibody-based assay on a patient cohort that included naïve, treated, and those who had side effects.
Results: Serum α- and β-globin chains were identified as differentially expressed proteins having threefold higher expressions in Parkinson's patients as compared to schizophrenia. Interestingly, concentrations of these two proteins had an inverse correlation across clinical phenotypes in the dopaminergic spectrum. RBC contamination as a source for these proteins was ruled out.
Conclusion: There is a clear association of free serum globin with dopaminergic clinical states. This lays a platform for protein biomarker-based monitoring of pharmacological efficacy in Parkinson's disease and schizophrenia.

Keywords:  Parkinson’s disease; biomarkers; difference gel electrophoresis; dopamine; gel-based proteomics; pharmacological efficacy; schizophrenia

DOI:  https://doi.org/10.2147/NDT.S198559
J Cell Physiol. 2019 May 20.

From clinic to mechanism: Proteomics-based assessment of angiogenesis in adrenal pheochromocytoma.

Fukang Sun, Ran Zhuo, Wenming Ma, Da Yang, Tingwei Su, Lei Ye, Danfeng Xu, Weiqing Wang.

  Adrenal pheochromocytoma (PCC) is a very rare tumor that stems from chromaffin cells, which can develop into malignant tumor. During the operation, abundant blood vessels were often observed in PCC than other adrenal tumors, which increases the difficulty and risk of the surgery. Therefore, it is important to investigate the mechanism of PCC angiogenesis. Twelve surgical specimens of PCC from Ruijin Hospital, Shanghai Jiaotong University were grouped into high and low microvessel density (MVD) group. They were also divided into rich blood supply and nonenriched blood supply group, according to computed tomography (CT) manifestation. Comparative proteomic analysis based on liquid chromatography-tandem mass spectrometry (LC-MS/MS) and bioinformatics analysis revealed that 206 proteins differentially regulated in the high MVD group compared with low MVD group (p < 0.05). Besides, 61 proteins were discovered to be significantly changed when the 12 samples were grouped according to CT manifestation. By intersecting the differentially changed protein from MVD and CT grouping, 25 proteins were filtered out, with pathological function. COX4I2 was verified to be increased gradually with angiogenesis with increasing severity, and PLAT was shown to be decreased with angiogenesis in PCC, by quantitative reverse-transcription polymerase chain reaction and immunohistochemistry. The quantitative proteomics result indicated that the tumor angiogenesis in PCC is associated with hypoxia. COX4I2 and PLAT were highly correlated with blood supply in PCC which contribute to angiogenesis in PCC, which could be used as biomarkers to better indicate tumor angiogenesis, or targets to regress tumor angiogenesis as treatment.

Keywords:  angiogenesis; computed tomography; microvessel density; pheochromocytoma; quantitative proteomics

DOI:  https://doi.org/10.1002/jcp.28769
BMC Musculoskelet Disord. 2019 May 24. 20(1): 247

Differential proteome analysis in adolescent idiopathic scoliosis patients with thoracolumbar/lumbar curvatures.

Hiroto Makino, Shoji Seki, Isao Kitajima, Hiraku Motomura, Makiko Nogami, Yasuhito Yahara, Naoko Ejiri, Tomoatsu Kimura.

   BACKGROUND: Although the pathogenesis of adolescent idiopathic scoliosis (AIS) remains unclear, there are little evidences of the pathogenesis in patients with thoracolumbar/lumbar AIS. The purpose of this study was to identify proteins or proteomes that may be causally related to the pathogenesis of AIS with structured thoracolumbar/lumbar curvature using two-dimensional fluorescence difference gel electrophoresis (2D-DIGE).
METHODS: A total of 20 control volunteers and 61 AIS in patients with thoracolumbar/lumbar curvature were included. First, the plasma samples of each five AIS with pure thoracolumbar/lumbar curvature and control samples were subjected to 2D-DIGE analysis. Protein spots that were expressed differently by the AIS and control groups were selected and identified by nanoscale liquid chromatography-tandem mass spectrometry (nanoLC-MS/MS) analysis. To characterize the differently-expressed proteins in AIS patients, we performed functional pathway analysis using the Protein ANalysis THrough Evolutionary Relationships (PANTHER) system. Additionally, the proteins were compared between control and AIS using western blotting. Lastly, prospectively collected 15 control and 41 AIS with thoracolumbar/lumbar curvature samples were compared to the differentially expressed proteins.
RESULTS: A total of 3862 ± 137 spots were detected, of which 11 spots met the criteria when compared with controls. Nine proteins were identified by nanoLC-MS/MS. Functional analysis showed the association of the proteins in AIS patients with blood coagulation using the PANTHER system. Of the proteins, vitamin D binding protein (DBP) significantly correlated with Cobb angle in thoracolumbar/lumbar curvatures. DBP expression of the prospectively collected AIS samples were significantly higher than those of controls (P < 0.05).
CONCLUSIONS: This study suggests that DBP and several coagulation-related proteins may play a role in the pathogenesis of AIS. DBP appears to be a marker of severity of AIS with thoracolumbar/lumbar curvature.

Keywords:  Adolescent idiopathic scoliosis; Thoracolumbar curvature; Two-dimensional fluorescence difference gel electrophoresis; Vitamin D binding protein

DOI:  https://doi.org/10.1186/s12891-019-2640-y
EBioMedicine. 2019 May 16. pii: S2352-3964(19)30326-3. [Epub ahead of print]

LGALS3 is connected to CD74 in a previously unknown protein network that is associated with poor survival in patients with AML.

Peter P Ruvolo, Chenyue W Hu, Yihua Qiu, Vivian R Ruvolo, Robin L Go, Stefan E Hubner, Kevin R Coombes, Michael Andreeff, Amina A Qutub, Steven M Kornblau.

   BACKGROUND: Galectin 3 (LGALS3) gene expression is associated with poor survival in acute myeloid leukemia (AML) but the prognostic impact of LGALS3 protein expression in AML is unknown. LGALS3 supports diverse survival pathways including RAS mediated cascades, protein expression and stability of anti-apoptotic BCL2 family members, and activation of proliferative pathways including those mediated by beta Catenin. CD74 is a positive regulator of CD44 and CXCR4 signaling and this molecule may be critical for AML stem cell function. At present, the role of LGALS3 and CD74 in AML is unclear. In this study, we examine protein expression of LGALS3 and CD74 by reverse phase protein analysis (RPPA) and identify new protein networks associated with these molecules. In addition, we determine prognostic potential of LGALS3, CD74, and their protein networks for clinical correlates in AML patients.
METHODS: RPPA was used to determine relative expression of LGALS3, CD74, and 229 other proteins in 231 fresh AML patient samples and 205 samples were from patients who were treated and evaluable for outcome. Pearson correlation analysis was performed to identify proteins associated with LGALS3 and CD74. Progeny clustering was performed to generate protein networks. String analysis was performed to determine protein:protein interactions in networks and to perform gene ontology analysis. Kaplan-Meir method was used to generate survival curves.
FINDINGS: LGALS3 is highest in monocytic AML patients and those with elevated LGALS3 had significantly shorter remission duration compared to patients with lower LGALS3 levels (median 21.9 vs 51.3 weeks, p = 0.016). Pearson correlation of LGALS3 with 230 other proteins identifies a distinct set of 37 proteins positively correlated with LGALS3 expression levels with a high representation of proteins involved in AKT and ERK signaling pathways. Thirty-one proteins were negatively correlated with LGALS3 including an AKT phosphatase. Pearson correlation of proteins associated with CD74 identified 12 proteins negatively correlated with CD74 and 16 proteins that are positively correlated with CD74. CD74 network revealed strong association with CD44 signaling and a high representation of apoptosis regulators. Progeny clustering was used to build protein networks based on LGALS3 and CD74 associated proteins. A strong relationship of the LGALS3 network with the CD74 network was identified. For AML patients with both the LGALS3 and CD74 protein cluster active, median overall survival was only 24.3 weeks, median remission duration was 17.8 weeks, and no patient survived beyond one year.
INTERPRETATION: The findings from this study identify for the first time protein networks associated with LGALS3 and CD74 in AML. Each network features unique pathway characteristics. The data also suggest that the LGALS3 network and the CD74 network each support AML cell survival and the two networks may cooperate in a novel high risk AML population. FUND: Leukemia Lymphoma Society provided funds to SMK for RPPA study of AML patient population. Texas Leukemia provided funds to PPR and SMK to study CD74 and LGALS3 expression in AML patients using RPPA. No payment was involved in the production of this manuscript.

Keywords:  Acute myeloid leukemia; CD74; LGALS3; Proteomics; RPPA

DOI:  https://doi.org/10.1016/j.ebiom.2019.05.025
NPJ Precis Oncol. 2019 ;3 15

Pan-cancer analysis connects tumor matrisome to immune response.

Su Bin Lim, Melvin Lee Kiang Chua, Joe Poh Sheng Yeong, Swee Jin Tan, Wan-Teck Lim, Chwee Teck Lim.

  Recent sequencing efforts unveil genomic landscapes of tumor microenvironment. A key compartment in this niche is the extracellular matrix (ECM) and its related components - matrisome. Yet, little is known about the extent to which matrisome pattern is conserved in progressive tumors across diverse cancer types. Using integrative genomic approaches, we conducted multi-platform assessment of a measure of deregulated matrisome associated with tumor progression, termed as tumor matrisome index (TMI), in over 30,000 patient-derived samples. Combined quantitative analyses of genomics and proteomics reveal that TMI is closely associated with mutational load, tumor pathology, and predicts survival across different malignancies. Interestingly, we observed an enrichment of specific tumor-infiltrating immune cell populations, along with signatures predictive of resistance to immune checkpoint blockade immunotherapy, and clinically targetable immune checkpoints in TMIhigh tumors. B7-H3 emerged as a particularly promising target for anti-tumor immunity in these tumors. Here, we show that matrisomal abnormalities could represent a potential clinically useful biomarker for prognostication and prediction of immunotherapy response.

Keywords:  Cancer genomics; Predictive markers

DOI:  https://doi.org/10.1038/s41698-019-0087-0
Cancer Manag Res. 2019 ;11 2935-2946

The serum proteomics tracking of hepatocellular carcinoma early recurrence following radical resection.

Hongzhi Liu, Hui Chen, Xiaomo Wu, Ying Sun, Yingchao Wang, Yongyi Zeng, Geng Chen, Xiaolong Liu, Xiaohua Xing, Bixing Zhao, Jingfeng Liu.

  Purpose: There is still lacking of highly sensitive and specific biomarkers for the prediction of hepatocellular carcinoma (HCC) early recurrence, which has hindered further improvement of the clinical outcomes. We aim to find highly sensitive and specific biomarkers for the prediction of HCC recurrence. Patients and methods: By using isobaric tags for relative and absolute quantitation (iTRAQ)-based multidimensional liquid chromatography-tandem mass spectrometry (2D LC-MS/MS) technique, we have quantitatively investigated and monitored the proteome alterations of a series of serum after radical resection during the follow-up of 4 HCC patients. Results: A total of 27 differentially abundant proteins (DAPs) in serum were identified to be closely associated with the early recurrence of HCC, and these DAPs were particularly concentrated within ERK1/2 and nuclear factor-κ beta signaling pathways, suggesting the dysregulation of these two pathways played an important role in the pathological process of HCC early recurrence. Further investigation of a cohort of patients confirmed that the high serum level of PGK1 was closely associated with HCC early recurrence and poor prognosis. In addition, the serum level of PGK1 could be complementary with AFP to further improve the sensitivity and specificity for predicting the relapse of HCC. Conclusion: PGK1 might be an independent factor for the recurrence of HCC. And the PGK1 could be complementary with AFP to further improve the sensitivity and specificity in prognostic prediction of HCC relapse.

Keywords:  PGK1; early recurrence; hepatocellular carcinoma; iTRAQ; serum proteomics

DOI:  https://doi.org/10.2147/CMAR.S190561
Antioxid Redox Signal. 2019 May 24.

Leukocyte proteomic profiling in first-episode schizophrenia patients: does oxidative stress play central roles in the pathophysiology network of schizophrenia?

Jie Jiang, Chao Peng, Liya Sun, Jing Li, Ying Qing, Xiaowen Hu, Xuhan Yang, Yan Li, Chuangye Xu, Juan Zhang, Jie Min, Xingwang Li, Shengying Qin, Mei Lin, Liwen Tan, Chunling Wan.

Although several underlying etiologic implications for schizophrenia (SZ) have been proposed, the crosstalk between them are rarely explored systematically. The aim of the present study was to illustrate the pathogenic mechanism of SZ through describing a systematical pathophysiology network using proteomic signatures in first-episode SZ patients. A total of 3152 proteins were identified in leukocytes, and 475 of these proteins were significantly altered in SZ. Functional analysis revealed that cell redox homeostasis was dramatically disrupted, demonstrated as upregulated glycolysis, mitochondrial oxidative phosphorylation, and thioredoxin-centered antioxidant system. We also identified activated complement system and caspase-independent apoptosis. In addition, increased pyruvate and lactate levels and decreased lactate-to-pyruvate ratios were observed in plasma of SZ patients. The results here lead to the hypothesis that increased oxidative stress is caused by metabolic upregulation and complement activation, which induces protein damage and cell apoptosis, thus contributing to the development of schizophrenia.

DOI: https://doi.org/10.1089/ars.2019.7805
EBioMedicine. 2019 May 21. pii: S2352-3964(19)30327-5. [Epub ahead of print]

Three kinds of corneal host cells contribute differently to corneal neovascularization.

Haiyang Yu, Liyao Sun, Jing Cui, Yan Li, Yu Yan, Xi Wei, Chao Wang, Fanqian Song, Wentong Jiang, Yifan Liu, Hongyan Ge, Hua Qian, Xiaoguang Li, Xianling Tang, Ping Liu.

   BACKGROUND: Corneal neovascularization (angiogenesis and lymphangiogenesis) compromises corneal transparency and transplant survival, however, the molecular mechanisms of corneal host epithelial and stromal cells in neovascularization have not yet been fully elucidated. Furthermore, the contribution and mechanism of corneal host endothelial cells involved in neovascularization are largely unexplored.
METHODS: Liquid chromatography-mass spectrometry, immunoblotting, and ELISA were used to screen and identify potential neovascularization-related factors in human full-thickness vascularized corneal tissues. Lipopolysaccharide was used to induce inflammation in three kinds of corneal host cells in vitro, including corneal epithelial, stromal, and endothelial cells. Fungus was used to establish an animal model of corneal neovascularization in vivo. Tube formation and spheroid sprouting assays were used to evaluate the contribution of three kinds of corneal host cells to the degree of neovascularization under various stimuli. Matrix metalloproteinase (MMP)-2, alpha-crystallin A chain (CRYAA), galectin-8, Bcl-2, neuropilin-2, MMP-9 plasmids, and recombinant human fibronectin were used to identify the key proteins of corneal host cells involved in corneal inflammatory neovascularization.
FINDINGS: All three kinds of corneal host cells influenced corneal neovascularization to varying degrees. MMP-9 in human corneal epithelial cells, MMP-2, and CRYAA in human corneal stromal cells, and MMP-2 and galectin-8 in human corneal endothelial cells are potential key proteins that participate in corneal inflammatory neovascularization.
INTERPRETATION: Our data indicated that both the effects of key proteins and corneal host cells involved should be considered for the treatment of corneal inflammatory neovascularization.

Keywords:  CRYAA; Corneal cells; Galectin-8; MMP-2; MMP-9; Neovascularization

DOI:  https://doi.org/10.1016/j.ebiom.2019.05.026
Genes Brain Behav. 2019 May 23.

Novel Biomarkers to Assess In-Utero Effects of Maternal Opioid Use: First Steps towards Understanding Short and Long Term Neurodevelopmental Sequelae.

Laura Goetzl, Tara Thompson-Felix, Nune Darbinian, Nana Merabova, Salim Merali, Carmen Merali, Kathryne Sanserino, Tamara Tatevosian, Bruno Fant, Mathieu E Wimmer.

  Maternal opioid use disorder is common, resulting in significant neonatal morbidity and cost. Currently, it is not possible to predict which opioid-exposed newborns will require pharmacotherapy for neonatal abstinence syndrome. Further, little is known regarding the effects of maternal opioid use disorder on the developing human brain. We hypothesized that novel methodologies utilizing fetal central nervous system-derived extracellular vesicles isolated from maternal blood can address these gaps in knowledge. Plasma from opioid users and controls between 9 and 21 weeks was precipitated and extracellular vesicles were isolated. Mu opioid and cannabinoid receptor levels were quantified. Label free proteomics studies and unbiased small RNA next generation sequencing was performed in paired fetal brain tissue. Maternal opioid use disorder increased mu opioid receptor protein levels in extracellular vesicles independent of opioid equivalent dose. Moreover, cannabinoid receptor levels in extracellular vesicles were upregulated with opioid exposure indicating cross talk with endocannabinoids. Maternal opioid use disorder was associated with significant changes in extracellular vesicle protein cargo and fetal brain micro RNA expression, especially in male fetuses. Many of the altered cargo molecules and micro RNAs identified are associated with adverse clinical neurodevelopmental outcomes. Our data suggest that assays relying on extracellular vesicles isolated from maternal blood extracellular vesicles may provide information regarding fetal response to opioids in the setting of maternal opioid use disorder. Prospective clinical studies are needed to evaluate the association between extracellular vesicle biomarkers, risk of neonatal abstinence syndrome, and neurodevelopmental outcomes.

Keywords:  cannabinoid; extracellular vesicles; fetal; maternal opioid use disorder; micro RNA

DOI:  https://doi.org/10.1111/gbb.12583
Cell Syst. 2019 May 22. pii: S2405-4712(19)30147-4. [Epub ahead of print]8(5): 456-466.e5

Simultaneous Integration of Multi-omics Data Improves the Identification of Cancer Driver Modules.

Dana Silverbush, Simona Cristea, Gali Yanovich-Arad, Tamar Geiger, Niko Beerenwinkel, Roded Sharan.

  The identification of molecular pathways driving cancer progression is a fundamental challenge in cancer research. Most approaches to address it are limited in the number of data types they employ and perform data integration in a sequential manner. Here, we describe ModulOmics, a method to de novo identify cancer driver pathways, or modules, by integrating protein-protein interactions, mutual exclusivity of mutations and copy number alterations, transcriptional coregulation, and RNA coexpression into a single probabilistic model. To efficiently search and score the large space of candidate modules, ModulOmics employs a two-step optimization procedure that combines integer linear programming with stochastic search. Applied across several cancer types, ModulOmics identifies highly functionally connected modules enriched with cancer driver genes, outperforming state-of-the-art methods and demonstrating the power of using multiple omics data types simultaneously. On breast cancer subtypes, ModulOmics proposes unexplored connections supported by an independent patient cohort and independent proteomic and phosphoproteomic datasets.

Keywords:  cancer; cancer drivers; cancer pathways; data integration; driver modules; integer linear programming; mutual exclusivity; simultaneous optimization

DOI:  https://doi.org/10.1016/j.cels.2019.04.005