Proteomic analysis of brain metastatic lung adenocarcinoma reveals intertumoral heterogeneity and specific alterations associated with the timing of brain metastases.

BACKGROUND: Brain metastases are associated with considerable negative effects on patients' outcome in lung adenocarcinoma (LADC). Here, we investigated the proteomic landscape of primary LADCs and their corresponding brain metastases. MATERIALS AND METHODS: Proteomic profiling was conducted on 20 surgically resected primary and brain metastatic LADC samples via label-free shotgun proteomics. After sample processing, peptides were analyzed using an Ultimate 3000 pump coupled to a QExactive HF-X mass spectrometer. Raw data were searched using PD 2.4. Further data analyses were carried out using Perseus, RStudio and GraphPad Prism. Proteomic data were correlated with clinical and histopathological parameters and the timing of brain metastases. Mass spectrometry-based proteomic data are available via ProteomeXchange with identifier PXD027259. RESULTS: Out of the 6821 proteins identified and quantified, 1496 proteins were differentially expressed between primary LADCs and corresponding brain metastases. Pathways associated with the immune system, cell-cell/matrix interactions and migration were predominantly activated in the primary tumors, whereas pathways related to metabolism, translation or vesicle formation were overrepresented in the metastatic tumors. When comparing fast- versus slow-progressing patients, we found 454 and 298 differentially expressed proteins in the primary tumors and brain metastases, respectively. Metabolic reprogramming and ribosomal activity were prominently up-regulated in the fast-progressing patients (versus slow-progressing individuals), whereas expression of cell-cell interaction- and immune system-related pathways was reduced in these patients and in those with multiple brain metastases. CONCLUSIONS: This is the first comprehensive proteomic analysis of paired primary tumors and brain metastases of LADC patients. Our data suggest a malfunction of cellular attachment and an increase in ribosomal activity in LADC tissue, promoting brain metastasis. The current study provides insights into the biology of LADC brain metastases and, moreover, might contribute to the development of personalized follow-up strategies in LADC.

Short infusion of paclitaxel imbalances plasmatic lipid metabolism and correlates with cardiac markers of acute damage in patients with breast cancer.

PURPOSE: Although paclitaxel-based chemotherapy is widely used for treating breast cancer, paclitaxel therapy has been associated with several adverse effects. Such adverse effects have primarily been associated with long-term regimens, but some acute effects are being increasingly reported in the literature. In this context, the present study analyzed the systemic proteomic profiles of women diagnosed with breast cancer at the first cycle of short paclitaxel infusion (n = 30). Proteomic profiles thus obtained were compared with those of breast cancer patients without chemotherapy (n = 50), as well as with those of healthy controls (n = 40). METHODS: Plasma samples were evaluated by label-free LC-MS to obtain systemic proteomic profiles. Putative dysregulated pathways were identified and validated by in silico analysis of proteomic profiles. RESULTS: Our results identified 188 proteins that were differentially expressed in patients who received a single short paclitaxel infusion when compared to patients who did not receive the infusion. Gene ontology analysis indicated that the cholesterol pathway may be dysregulated by paclitaxel in these patients. Validation analysis showed that paclitaxel treatment significantly reduced plasma high-density lipoprotein levels and increased plasma hydroperoxide levels when compared to breast cancer patients without chemotherapy. Furthermore, augmented C-reactive protein and creatine kinase fraction MB were found to be significantly higher in paclitaxel-treated patients in comparison with healthy controls. CONCLUSIONS: Taken together, these data suggest that a single dose of short paclitaxel infusion is sufficient to trigger significant alterations in lipid metabolism, which puts breast cancer patients at risk for increased incidence of cardiovascular disease.

Discovering the infectome of human endothelial cells challenged with Aspergillus fumigatus applying a mass spectrometry label-free approach.

Blood vessel invasion is a key feature of invasive aspergillosis. This angioinvasion process contributes to tissue thrombosis, which can impair the access of leukocytes and antifungal drugs to the site of infection. It has been demonstrated that human umbilical vein endothelial cells (HUVECs) are activated and assume a prothrombotic phenotype following contact with Aspergillus fumigatus hyphae or germlings, a process that is independent of fungus viability. However, the molecular mechanisms by which this pathogen can activate endothelial cells, together with the endothelial pathways that are involved in this process, remain unknown. Using a label-free approach by High Definition Mass Spectrometry (HDMS(E)), differentially expressed proteins were identified during HUVEC-A. fumigatus interaction. Among these, 89 proteins were determined to be up- or down-regulated, and another 409 proteins were exclusive to one experimental condition: the HUVEC control or HUVEC:AF interaction. The in silico predictions provided a general view of which biological processes and/or pathways were regulated during HUVEC:AF interaction, and they mainly included cell signaling, immune response and hemostasis pathways. This work describes the first global proteomic analysis of HUVECs following interaction with A. fumigatus germlings, the fungus morphotype that represents the first step of invasion and dissemination within the host. BIOLOGICAL SIGNIFICANCE: A. fumigatus causes the main opportunistic invasive fungal infection related to neutropenic hematologic patients. One of the key steps during the establishment of invasive aspergillosis is angioinvasion but the mechanism associated with the interaction of A. fumigatus with the vascular endothelium remains unknown. The identification of up- and down-regulated proteins expressed by human endothelial cells in response to the fungus infection can contribute to reveal the mechanism of endothelial response and, to understand the physiopathology of this high mortality disease. This article is part of a Special Issue entitled: Trends in Microbial Proteomics.

Stability of human mesenchymal stem cells during in vitro culture: considerations for cell therapy.

Ex vivo expansion and manipulation of human mesenchymal stem cells are important approaches to immunoregulatory and regenerative cell therapies. Although these cells show great potential for use, issues relating to their overall nature emerge as problems in the field. The need for extensive cell quantity amplification in vitro to obtain sufficient cell numbers for use, poses a risk of accumulating genetic and epigenetic abnormalities that could lead to sporadic malignant cell transformation. In this study, we have examined human mesenchymal stem cells derived from bone marrow, over extended culture time, using cytogenetic analyses, mixed lymphocyte reactions, proteomics and gene expression assays to determine whether the cultures would retain their potential for use in subsequent passages. Results indicate that in vitro cultures of these cells demonstrated chromosome variability after passage 4, but their immunomodulatory functions and differentiation capacity were maintained. At the molecular level, changes were observed from passage 5 on, indicating initiation of differentiation. Together, these results lead to the hypothesis that human mesenchymal stem cells cultures can be used successfully in cell therapy up to passage 4. However, use of cells from higher passages would have to be analysed case by case.