bims-prodis 2018-10-07 papers

Issue of 2018–10–07
three papers selected by
Nancy Gough, Bioserendipity

J Proteomics. 2018 Sep 26. pii: S1874-3919(18)30351-8. [Epub ahead of print]

Proteomic profiling of the proteolytic events in the secretome of the transformed phenotype of melanocyte-derived cells using Terminal Amine Isotopic Labeling of Substrates.

Tarcísio Liberato, Isabella Fukushima, Eduardo S Kitano, Solange M T Serrano, Roger Chammas, André Zelanis.

The comprehensive profiling of the repertoire of secreted proteins from cancer cells samples provides information on the signaling events in oncogenesis as well as on the cross-talk between normal and tumoral cells. Moreover, the analysis of post-translational modifications in secreted proteins may unravel biological circuits regulated by irreversible modifications such as proteolytic processing. In this context, we used Terminal Amine Isotopic Labeling of Substrates (TAILS) to perform a system-wide investigation on the N-terminome of the secretomes derived from a paired set of mouse cell lines: Melan-a (a normal melanocyte) and Tm1 (its transformed phenotype). Evaluation of the amino acid identities at the scissile bond in internal peptides revealed significant differences, suggesting distinct proteolytic processes acting in the normal and tumoral secretomes. The mapping and annotation of cleavage sites in the tumoral secretome suggested functional roles of active proteases in central biological processes related to oncogenesis, such as the processing of growth factors, cleavage of extracellular matrix proteins and the shedding of ectopic domains from the cell surface, some of which may represent novel processed forms of the corresponding proteins. In the context of the tumor microenvironment, these results suggest important biological roles of proteolytic processing in murine melanoma secreted proteins.

Keywords: Cancer; Melanoma; N-terminomics; Proteomics; Secretome; TAILS

DOI: https://doi.org/10.1016/j.jprot.2018.09.010

Lung Cancer. 2018 Oct;pii: S0169-5002(18)30498-7. [Epub ahead of print]124 90-94

Automated chromogenic multiplexed immunohistochemistry assay for diagnosis and predictive biomarker testing in non-small cell lung cancer.

Marius Ilie, Mélanie Beaulande, Marame Hamila, Gilles Erb, Véronique Hofman, Paul Hofman.

OBJECTIVES: The current challenge in the management of non-small cell lung cancer (NSCLC) in pathology laboratories is to combine immunohistochemistry (IHC) and molecular approaches on increasingly smaller biopsies and the need to reserve a fair amount of tumor material for molecular analyses with increasingly larger panels. The latest lung cancer classification, especially in the setting of poorly differentiated tumors, requires an IHC workup to allow for accurate diagnosis and also to preserve as much tissue as possible for molecular testing. Thus, it is recommended to reduce use of the term NSCLC not otherwise specified as much as possible and classify tumors according to their specific histologic subtype. This implies limiting the number of tissue slides despite the existence of specific and sensitive biomarkers (ALK, ROS1, BRAF V600E, PD-L1) and the obligation to distinguish lung adenocarcinoma (TTF-1 positive) from squamous cell carcinoma (p40 positive).
MATERIALS AND METHODS: Samples from 18 patients with NSCLC, previously characterized for histologic and genomic/immune features, were included. Two multiplexed IHC assays were developed, for diagnosis and immunophenotyping including TTF1, p40, PD-L1, and pan-Keratin antibodies, and for molecular profiling panel including ALK, ROS1 and BRAF V600E antibodies.
RESULTS: We developed two sensitive multiplexed IHC assays to comprehensively characterize major NSCLC histotypes and FDA-cleared predictive biomarkers, without antigenicity loss, steric interference or increased cross-reactivity. The assays rely on standard antigen retrieval and automated staining protocols, limiting the need for validation strategies.
CONCLUSION: Our multiplexed IHC approach provides a unique sample-sparing tool to characterize limited tissue samples in lung oncology and making it an alternative method in the clinical setting for therapeutic decision making of advanced NSCLC, provided that validation in a larger population is performed.

Keywords: Chromogenic; Diagnosis; Multiplexed immunohistochemistry; Non-small cell lung cancer; Predictive biomarkers

DOI: https://doi.org/10.1016/j.lungcan.2018.07.037

J Pharm Sci. 2018 Sep 26. pii: S0022-3549(18)30552-5. [Epub ahead of print]

Dysregulation of mucosal membrane transporters and drug metabolizing enzymes in ulcerative colitis.

Pascal Erdmann, Henrike Bruckmueller, Paul Martin, Diana Busch, Sierk Haenisch, Janett Müller, Anna Wiechowska-Kozlowska, Lars Ivo Partecke, Claus-Dieter Heidecke, Ingolf Cascorbi, Marek Drozdzik, Stefan Oswald.

Intestinal transporters and metabolizing enzymes are important factors of the intestinal absorption barrier. Because there is evidence that their expression and function may be affected during inflammatory conditions, we investigated gene expression, protein abundance and regulation of relevant intestinal transporters and metabolizing enzymes in the intestinal mucosa of ulcerative colitis (UC) patients. Specimens from inflamed and non-inflamed tissues of 10 UC patients as well as colonic control tissues of 10 patients without inflammation were subjected to gene (9 enzymes, 15 transporters, 9 cytokines) and microRNA (N=54) expression analysis. Protein abundance was quantified by LC-MS/MS-based targeted proteomics. Gene expression of several metabolizing enzymes (e.g. CYP2C9, UGT1A1) and transporters as ABCB1 (ABCB1), ABCG2 (ABCG2) and monocarboxylate transporter 1 (MCT1, SLC16A1) were significantly decreased during inflammation and negatively correlated to microRNAs. Contrary, multidrug resistance-protein 4 (MRP4, ABCC4), organic anion transporting-polypeptide 2B1 (OATP2B1, SLCO2B1) and organic cation transporter-like 2 (ORCTL2, SLC22A18) were significantly elevated in inflamed tissue. However, on protein level these findings could only be confirmed for MCT1. UC is associated with complex changes in the intestinal expression of enzymes, transporters, cytokines and microRNAs, which may affect efficacy of anti-inflammatory drug therapy or the disease state itself.

Keywords: cytokines; inflammatory bowel disease; intestinal transporters; metabolizing enzymes; microRNA

DOI: https://doi.org/10.1016/j.xphs.2018.09.024