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Peptides have potential bioactive functions, and the peptidomics landscape has been broadly investigated for various diseases, including cancer. In this chapter, we reviewed the past four years of literature available and selected 16 peer-reviewed publications exploring peptidomics in diagnosis, prognosis, and treatment in cancer research. We highlighted their main aims, mass spectrometry-based peptidomics, multi-omics, data-driven and in silico strategies, functional assays, and clinical applications. Moreover, we underscored several levels of difficulties in translating the peptidomics findings to clinical practice, aiming to learn with the accumulated knowledge and guide upcoming studies. Finally, this review reinforces the peptidomics robustness in discovering potential candidates for monitoring the several stages of cancer disease and therapeutic treatment, leveraging the management of cancer patients in the future., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Objective: To integrate all the available data published in the English literature regarding the protein diagnostic and/or prognostic markers in salivary gland tumors identified by mass spectrometry (MS)-based discovery proteomics., Design: An extensive search was carried out using MEDLINE/PubMed, EMBASE, Web of Science, and Scopus databases. Manual searching in Google Scholar and assessment of the reference list of the included articles also was performed. The risk of bias was assessed by the Joanna Briggs Institute Critical Appraisal tool for the specific type of study., Results: A total of 1092 articles were initially retrieved within which 6 were used for data extraction, resulting in 145 cases of salivary gland tumors. The data was composted by eleven salivary gland tumor types. In total, 2136 proteins were detected by MS-based discovery proteomics in salivary gland tumors. Ninety-one proteins were proposed as potential diagnostic and/or prognostic markers. Of these, some have been identified in one or more studies, whereas fifteen were in common across studies and a total of seventy-six were non-repeat proteins., Conclusion: In summary, we compiled data about the proteomic profile of potential diagnostic and/or prognostic protein markers of the salivary gland tumors detected by MS-based discovery proteomics. The proteins ANXA1, ANXA5, CAPG, CRYAB, FGB, GNB2L1, IGHG1, PPIA, S100A9, and SOD1 were proposed as the most common potential diagnostic markers of salivary gland tumors., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Different regions of oral squamous cell carcinoma (OSCC) have particular histopathological and molecular characteristics limiting the standard tumor-node-metastasis prognosis classification. Therefore, defining biological signatures that allow assessing the prognostic outcomes for OSCC patients would be of great clinical significance. Using histopathology-guided discovery proteomics, we analyze neoplastic islands and stroma from the invasive tumor front (ITF) and inner tumor to identify differentially expressed proteins. Potential signature proteins are prioritized and further investigated by immunohistochemistry (IHC) and targeted proteomics. IHC indicates low expression of cystatin-B in neoplastic islands from the ITF as an independent marker for local recurrence. Targeted proteomics analysis of the prioritized proteins in saliva, combined with machine-learning methods, highlights a peptide-based signature as the most powerful predictor to distinguish patients with and without lymph node metastasis. In summary, we identify a robust signature, which may enhance prognostic decisions in OSCC and better guide treatment to reduce tumor recurrence or lymph node metastasis.

ADAM17 (a disintegrin and metalloproteinase 17) is a plasma membrane metalloprotease involved in proteolytic release of the extracellular domain of many cell surface molecules, a process known as ectodomain shedding. Through this process, ADAM17 is implicated in several aspects of tumor growth and metastasis in a broad range of tumors, including head and neck squamous cell carcinomas (HNSCC). In this study, mass spectrometry-based proteomics approaches revealed glypican-1 (GPC1) as a new substrate for ADAM17, and its shedding was confirmed to be metalloprotease-dependent, induced by a pleiotropic agent (PMA) and physiologic ligand (EGF), and inhibited by marimastat. In addition, immunoblotting analysis of GPC1 in the extracellular media from control and ADAM17shRNA pointed to a direct involvement of ADAM17 in the cleavage of GPC1. Moreover, mass spectrometry-based interactome analysis of GPC1 revealed biological functions and pathways related mainly to cellular movement, adhesion and proliferation, which were events also modulated by up regulation of full length and cleavage GPC1. Altogether, we showed that GPC1 is a novel ADAM17 substrate, thus the function of GPC1 may be modulated by proteolysis signaling., Biological Significance: Inhibition of metalloproteases as a therapeutic approach has failed because there is limited knowledge of the degradome of individual proteases as well as the cellular function of cleaved substrates. Using different proteomic techniques, this study uncovered novel substrates that can be modulated by ADAM17 in oral squamous cell carcinoma cell line. Glypican-1 was validated as a novel substrate for ADAM17, with important function in adhesion, proliferation and migration of carcinoma cells. Therefore, this study opens new avenues regarding the proteolysis-mediated function of GPC1 by ADAM17., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The mechanism of alternative splicing in the transcriptome may increase the proteome diversity in eukaryotes. In proteomics, several studies aim to use protein sequence repositories to annotate MS experiments or to detect differentially expressed proteins. However, the available protein sequence repositories are not designed to fully detect protein isoforms derived from mRNA splice variants. To foster knowledge for the field, here we introduce SpliceProt, a new protein sequence repository of transcriptome experimental data used to investigate for putative splice variants in human proteomes. Current version of SpliceProt contains 159 719 non-redundant putative polypeptide sequences. The assessment of the potential of SpliceProt in detecting new protein isoforms resulting from alternative splicing was performed by using publicly available proteomics data. We detected 173 peptides hypothetically derived from splice variants, which 54 of them are not present in UniprotKB/TrEMBL sequence repository. In comparison to other protein sequence repositories, SpliceProt contains a greater number of unique peptides and is able to detect more splice variants. Therefore, SpliceProt provides a solution for the annotation of proteomics experiments regarding splice isofoms. The repository files containing the translated sequences of the predicted splice variants and a visualization tool are freely available at http://lbbc.inca.gov.br/spliceprot., (© 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Class III peroxidases are present as large multigene families in all land plants. This large number of genes together with the diversity of processes catalyzed by peroxidases suggests possible functional specialization of each isoform. However, assigning a precise role for each individual peroxidase gene has continued to be a major bottleneck. Here we investigated the enzyme activity and translational profile of class III peroxidases during stem development of sugarcane as a first step in the estimation of physiological functions of individual isoenzymes. Internodes at three different developmental stages (young, developing and mature) were divided into pith (inner tissue) and rind (outer tissue) fractions. The rind of mature internodes presented the highest enzymatic activity and thus could be considered the ideal tissue for the discovery of peroxidase gene function. In addition, activity staining of 2DE gels revealed different isoperoxidase profiles and protein expression regulation among different tissue fractions. In-gel tryptic digestion of excised spots followed by peptide sequencing by LC-MS/MS positively matched uncharacterized peroxidases in the sugarcane database SUCEST. Multiple spots matching the same peroxidase gene were found, which reflects the generation of more than one isoform from a particular gene by post-translational modifications. The identified sugarcane peroxidases appear to be monocot-specific sequences with no clear ortholog in dicot model plant Arabidopsis thaliana., (Copyright © 2012 Elsevier Masson SAS. All rights reserved.)

Physical protein-protein interactions are fundamental to all biological processes and are organized in complex networks. One branch of the kinome network is the evolutionarily conserved NIMA-related serine/threonine kinases (Neks). Most of the 11 mammalian Neks studied so far are related to cell cycle regulation, and due to association with diverse human pathologies, Neks are promising chemotherapeutic targets. Human Nek6 was associated to carcinogenesis, but its interacting partners and signaling pathways remain elusive. Here we introduce hNek6 as a highly connected member in the human kinase interactome. In a more global context, we performed a broad data bank comparison based on degree distribution analysis and found that the human kinome is enriched in hubs. Our networks include a broad set of novel hNek6 interactors as identified by our yeast two-hybrid screens classified into 18 functional categories. All of the tested interactions were confirmed, and the majority of tested substrates were phosphorylated in vitro by hNek6. Notably, we found that hNek6 N-terminal is important to mediate the interactions with its partners. Some novel interactors also colocalized with hNek6 and γ-tubulin in human cells, pointing to a possible centrosomal interaction. The interacting proteins link hNek6 to novel pathways, for example, Notch signaling and actin cytoskeleton regulation, or give new insights on how hNek6 may regulate previously proposed pathways such as cell cycle regulation, DNA repair response, and NF-κB signaling. Our findings open new perspectives in the study of hNek6 role in cancer by analyzing its novel interactions in specific pathways in tumor cells, which may provide important implications for drug design and cancer therapy.

The article focuses on analyzing exclusive proteins associated with malignant phenotype acquisition in pleomorphic adenoma (PA), highlighting their structural and functional roles. Topics include the identification of exclusive proteins in PA, residual PA, and carcinoma ex pleomorphic adenoma (CXPA), their biological processes, and their potential implications in the pathogenesis of PA and CXPA.

Human papillomavirus (HPV) infection has recently been linked to a subset of cancers affecting the oral cavity. However, the molecular mechanisms underlying HPV-driven oral squamous cell carcinoma (OSCC) onset and progression are poorly understood. Methods: We performed MS-based proteomics profiling based on HPV status in OSCC in young patients, following biological characterization and cell assays to explore the proteome functional landscape. Results: Thirty-nine proteins are differentially abundant between HPV (+) and HPV (−) OSCC. Among them, COPS3, DYHC1, and S100A8 are unfavorable for tumor recurrence and survival, in contrast to A2M and Serpine1, low levels of which show an association with better DFS. Remarkably, S100A8 is considered an independent prognostic factor for lower survival rates, and at high levels, it alters tumor-associated immune profiling, showing a lower proportion of M1 macrophages and dendritic cells. HPV (+) OSCC also displayed the pathogen-associated patterns receptor that, when activated, triggered the S100A8 and NFκB inflammatory responses. Conclusion: HPV (+) OSCC has a peculiar microenvironment pattern distinctive from HPV (−), involving the expression of pathogen-associated pattern receptors, S100A8 overexpression, and NFκB activation and responses, which has important consequences in prognosis and may guide therapeutic decisions. [ABSTRACT FROM AUTHOR]

Objective: To analyze the proteomic profile of salivary pleomorphic adenoma (PA) and carcinoma ex pleomorphic adenoma (CXPA) samples and correlate them with the malignant transformation of the PA. Materials and methods: Thirty samples (10 PA, 16 CXPA, and 4 residual PA) were microdissected and submitted to liquid chromatography‐tandem mass spectrometry (LC‐MS/MS). The proteomic data and protein identification were analyzed through LC‐MS/MS spectra using the MaxQuant software. Results: The proteomic analysis identified and quantified a total of 240 proteins in which 135 were found in PA, residual PA, and CXPA. The shared proteins were divided into six subgroups, and the proteins that showed statistically significant differences (p > 0.05) and fold‐change > or <2.5 in one subgroup to another subgroup were included. Seven proteins (Apolipoprotein A‐I—APOA1, haptoglobin—HP, protein of the synaptonemal complex 1—SYCP1, anion transport protein of band 3—SLC4A1, subunit μ1 of AP‐1 complex—AP1M1, beta subunit of hemoglobin—HBB, and dermcidin—DCD) were classified as potential protein signatures, being HP, AP1M1, and HBB with higher abundance for PA to residual PA, APOA1 with higher abundance for PA to CXPA, SLC4A1 with lower abundance in the PA to CXPA, SYCP1with lower abundance for residual PA to CXPA, and DCD with higher abundance in the CXPA with epithelial differentiation to myoepithelial differentiation. Conclusions: In this work, we demonstrated the comparative proteomic profiling of PA, residual PA, and CXPA, and seven were proposed as protein signatures, some of which may be associated with the malignant phenotype acquisition. [ABSTRACT FROM AUTHOR]

Introduction: Oral squamous cell carcinoma (OSCC) ranks among the top 10 leading causes of cancer worldwide, with 5-year survival rate of about 50%, high lymph node metastasis, and relapse rates. The OSCC diagnosis, prognosis, and treatment are mostly based on the clinical TNM classification. There is an urgent need for the discovery of biomarkers and therapeutic targets to assist in the clinical decision-making process. Areas covered: We summarize proteomic studies of the OSCC tumor, immune microenvironment, potential liquid biopsy sites, and post-translational modifications trying to retrieve information in the discovery and verification or (pre)validation phases. The search strategy was based on the combination of MeSH terms and expert refinement. Expert opinion: Untargeted combined with targeted proteomics are strategies that provide reliable and reproducible quantitation of proteins and are the methods of choice of many groups worldwide. Undoubtedly, proteomics has been contributing to the understanding of OSCC progression and uncovers potential candidates as biomarker or therapeutic targets. Nevertheless, none of these targets are available in the clinical practice yet. The scientific community needs to overcome the limitations by investing in robust experimental designs to strengthen the value of the findings, leveraging the translation of knowledge, and further supporting clinical decisions. [ABSTRACT FROM AUTHOR]

Only few studies on snake venoms were dedicated to deeply characterize the toxin secretion of animals from the Colubridae family, despite the fact that they represent the majority of snake diversity. As a consequence, some evolutionary trends observed in venom proteins that underpinned the evolutionary histories of snake toxins were based on data from a minor parcel of the clade. Here, we investigated the proteins of the totally unknown venom from Phalotrismertensi (Dipsadinae subfamily), in order to obtain a detailed profile of its toxins and to appreciate evolutionary tendencies occurring in colubrid venoms. By means of integrated omics and functional approaches, including RNAseq, Sanger sequencing, high-resolution proteomics, recombinant protein production, and enzymatic tests, we verified an active toxic secretion containing up to 21 types of proteins. A high content of Kunitz-type proteins and C-type lectins were observed, although several enzymatic components such as metalloproteinases and an L-amino acid oxidase were also present in the venom. Interestingly, an arguable venom component of other species was demonstrated as a true venom protein and named svLIPA (snake venom acid lipase). This finding indicates the importance of checking the actual protein occurrence across species before rejecting genes suggested to code for toxins, which are relevant for the discussion about the early evolution of reptile venoms. Moreover, trends in the evolution of some toxin classes, such as simplification of metalloproteinases and rearrangements of Kunitz and Wap domains, parallel similar phenomena observed in other venomous snake families and provide a broader picture of toxin evolution. [ABSTRACT FROM AUTHOR]

Background: Snake Venom Metalloproteinases (SVMPs) are amongst the key enzymes that contribute to the high toxicity of snake venom. We have recently shown that snake venoms from the Bothrops genus activate the Complement system (C) by promoting direct cleavage of C-components and generating anaphylatoxins, thereby contributing to the pathology and spread of the venom. The aim of the present study was to isolate and characterize the C-activating protease from Bothrops pirajai venom. Results: Using two gel-filtration chromatography steps, a metalloproteinase of 23 kDa that activates Complement was isolated from Bothrops pirajai venom. The mass spectrometric identification of this protein, named here as C-SVMP, revealed peptides that matched sequences from the P-I class of SVMPs. C-SVMP activated the alternative, classical and lectin C-pathways by cleaving the α-chain of C3, C4 and C5, thereby generating anaphylatoxins C3a, C4a and C5a. In vivo, C-SVMP induced consumption of murine complement components, most likely by activation of the pathways and/or by direct cleavage of C3, leading to a reduction of serum lytic activity. Conclusion: We show here that a P-I metalloproteinase from Bothrops pirajai snake venom activated the Complement system by direct cleavage of the central C-components, i.e., C3, C4 and C5, thereby generating biologically active fragments, such as anaphylatoxins, and by cleaving the C1-Inhibitor, which may affect Complement activation control. These results suggest that direct complement activation by SVMPs may play a role in the progression of symptoms that follow envenomation. Author Summary: The genus Bothrops inflicts the vast majority of snakebites in Central and South America and is responsible for 90% of snake envenomations in Brazil. Envenomations are characterized by prominent local effects, including edema and necrosis, and by systemic manifestations such as hemorrhage, coagulopathy and acute renal failure. Several components have been isolated from Bothrops venoms, and the snake venom metalloproteinases (SVMPs) are key enzymes contributing to the high toxicity of the venoms. Previously, we analyzed the pro-inflammatory properties of snake venoms from the genus Bothrops and demonstrated that several of them were potent activators of the Complement (C) system. C3a, C4a and C5a were generated in venom-treated sera not only through C-activation but also by direct cleavage of C-components. In the present study, we have isolated and characterized a metalloproteinase from Bothrops pirajai snake venom, named here as C-SVMP, which interferes with all three complement pathways, generating potent pro-inflammatory fragments, such as C3a, C4a and C5a. Our data suggest that C-activation by Bothrops pirajai venom is due to activity of an SVMP, which may play a role in the progression of symptoms that follow envenomation. [ABSTRACT FROM AUTHOR]

In this study a label-free proteomic approach was used to investigate the composition of the layer of protein adsorbed to rough titanium (Ti) after exposure to human blood plasma. The influence of the protein layer on the surface free energy (SFE) of the Ti was evaluated by contact angle measurements. Ti discs were incubated with blood plasma for 180 min at 37 °C, and the proteins recovered were subjected to liquid chromatography coupled to tandem mass spectrometry analysis. A total of 129 different peptides were identified and assigned to 25 distinct plasma proteins. The most abundant proteins were fibronectin, serum albumin, apolipoprotein A-I, and fibrinogen, comprising 74.54% of the total spectral counts. Moreover, the protein layer increased the SFE of the Ti (p < 0.05). The layer adsorbed to the rough Ti surface was composed mainly of proteins related to cell adhesion, molecule transportation, and coagulation processes, creating a polar and hydrophilic interface for subsequent interactions with host cells. [ABSTRACT FROM PUBLISHER]

Dental cementum (DC) covers the tooth root and has important functions in tooth attachment and position. DC can be lost to disease, and regeneration is currently unpredictable due to limited understanding of DC formation. This study used a model of experimentally-induced apposition (EIA) in mice to identify proteins associated with new DC formation. Mandibular first molars were induced to super-erupt for 6 and 21 days after extracting opposing maxillary molars. Decalcified and formalin-fixed paraffin-embedded mandible sections were prepared for laser capture microdissection. Microdissected protein extracts were analyzed by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS), and the data submitted to repeated measure ANOVA test (RM-ANOVA, alpha = 5%). A total of 519 proteins were identified, with 97 (18.6%) proteins found exclusively in EIA sites and 50 (9.6%) proteins exclusively expressed in control sites. Fifty six (10.7%) proteins were differentially regulated by RM-ANOVA (p < 0.05), with 24 regulated by the exclusive effect of EIA (12 proteins) or the interaction between EIA and time (12 proteins), including serpin 1a, procollagen C-endopeptidase enhancer, tenascin X (TNX), and asporin (ASPN). In conclusion, proteomic analysis demonstrated significantly altered protein profile in DC under EIA, providing new insights on DC biology and potential candidates for tissue engineering applications. Significance Dental cementum (DC) is a mineralized tissue that covers the tooth root surface and has important functions in tooth attachment and position. DC and other periodontal tissues can be lost to disease, and regeneration is currently unpredictable due to lack of understanding of DC formation. This study used a model of experimentally-induced apposition (EIA) in mice to promote new cementum formation, followed by laser capture microdissection (LCM) and liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) proteomic analysis. This approach identified proteins associated with new cementum formation that may be targets for promoting cementum regeneration. [ABSTRACT FROM AUTHOR]

Abstract: Secreted class III peroxidases (EC 1.11.1.7) are implicated in a broad range of physiological processes throughout the plant life cycle. However, the unambiguous determination of the precise biological role of an individual class III peroxidase isoenzyme is still a difficult task due to genetic redundancy and broad substrate specificity in vitro. In addition, many difficulties are encountered during extraction and analysis of cell wall proteins. Since class III peroxidases are also secreted into the apoplast, the use of suspension cell cultures can facilitate isolation and functional characterization of individual isoforms. Here, we report on the characterization of class III peroxidases secreted in the spent medium of sugarcane suspension cell cultures. After treatment with specific inducers of cell wall lignification, peroxidases were isolated and activities assayed with guaiacol, syringaldazine and coniferyl alcohol. Enzymatic activity was not significantly different after treatments, regardless of the substrate, with the exception of methyl-jasmonate treatment, which led to a decreased guaiacol peroxidase activity. Remarkably, peroxidases isolated from the medium were capable of oxidizing syringaldazine, an analog to sinapyl alcohol, suggesting that sugarcane cultures can produce peroxidases putatively correlated to lignification. A proteomic approach using activity staining of 2-DE gels revealed a complex isoperoxidase profile, composed predominantly of cationic isoforms. Individual spots were excised and analyzed by LC-ESI-Q-TOF and homology-based search against the Sugarcane EST Database resulted in the identification of several proteins. Spatio-temporal expression pattern of selected genes was determined for validation of identified class III peroxidases that were preferentially expressed during sugarcane stem development. [Copyright &y& Elsevier]

Oligodendrocytes produce and maintain the myelin sheath of axons in the central nervous system. Because misassembled myelin sheaths have been associated with brain disorders such as multiple sclerosis and schizophrenia, recent advances have been made towards the description of the oligodendrocyte proteome. The identification of splice variants represented in the proteome is as important as determining the level of oligodendrocyte-associated proteins. Here, we used an oligodendrocyte proteome dataset deposited in ProteomeXchange to search against a customized protein sequence file containing computationally predicted splice variants. Our approach resulted in the identification of 39 splice variants, including one variant from the GTPase KRAS gene and another from the human glutaminase gene family. We also detected the mRNA expression of five selected splice variants and demonstrated that a fraction of these have their canonical proteins participating in direct protein-protein interactions. In conclusion, we believe our findings contribute to the molecular characterization of oligodendrocytes and may encourage other research groups working with central nervous system disorders to investigate the biological significance of these splice variants. The splice variants identified in this study may encode proteins that could be targeted in novel treatment strategies and diagnostic methods. Significance Several disorders of the central nervous system (CNS) are associated with misassembled myelin sheaths, which are produced and maintained by oligodendrocytes (OL). Recently, the OL proteome has been explored to identify key proteins and molecular functions associated with CNS disorders. We developed an innovative approach to select, with a higher level of confidence, a relevant list of splice variants from a proteome dataset and detected the mRNA expression of five selected variants: EEF1D , KRAS , MFF , SDR39U1 , and SUGT1 . We also described splice variants extracted from OL proteome data. Among the splice variants identified, some are from genes previously linked to CNS and related disorders. Our findings may contribute to oligodendrocyte characterization and encourage other research groups to investigate the biological role of splice variants and to improve current treatments and diagnostic methods for CNS disorders. [ABSTRACT FROM AUTHOR]

Abstract: Dental cementum (DC) is a bone-like tissue covering the tooth root and responsible for attaching the tooth to the alveolar bone (AB) via the periodontal ligament (PDL). Studies have unsuccessfully tried to identify factors specific to DC versus AB, in an effort to better understand DC development and regeneration. The present study aimed to use matched human DC and AB samples (n=7) to generate their proteomes for comparative analysis. Bone samples were harvested from tooth extraction sites, whereas DC samples were obtained from the apical root portion of extracted third molars. Samples were denatured, followed by protein extraction reduction, alkylation and digestion for analysis by nanoAcquity HPLC system and LTQ-FT Ultra. Data analysis demonstrated that a total of 318 proteins were identified in AB and DC. In addition to shared proteins between these tissues, 105 and 83 proteins exclusive to AB or DC were identified, respectively. This is the first report analyzing the proteomic composition of human DC matrix and identifying putative unique and enriched proteins in comparison to alveolar bone. These findings may provide novel insights into developmental differences between DC and AB, and identify candidate biomarkers that may lead to more efficient and predictable therapies for periodontal regeneration. Biological significance: Periodontal disease is a highly prevalent disease affecting the world population, which involves breakdown of the tooth supporting tissues, the periodontal ligament, alveolar bone, and dental cementum. The lack of knowledge on specific factors that differentiate alveolar bone and dental cementum limits the development of more efficient and predictable reconstructive therapies. In order to better understand cementum development and potentially identify factors to improve therapeutic outcomes, we took the unique approach of using matched patient samples of dental cementum and alveolar bone to generate and compare a proteome list for each tissue. A potential biomarker for dental cementum was identified, superoxide dismutase 3 (SOD3), which is found in cementum and cementum-associated cells in mouse, pig, and human tissues. These findings may provide novel insights into developmental differences between alveolar bone and dental cementum, and represent the basis for improved and more predictable therapies. [Copyright &y& Elsevier]