Inflammatory breast cancer (IBC) has a poor prognosis because of the lack of specific biomarkers and its late diagnosis. An accurate and rapid diagnosis implemented early enough can significantly improve the disease outcome. Vibrational spectroscopy has proven to be useful for cell and tissue characterization based on the intrinsic molecular information. Here, we have applied infrared and Raman microspectroscopy and imaging to differentiate between non-IBC and IBC at both cell and tissue levels. Two human breast cancer cell lines (MDA-MB-231 and SUM-149), 20 breast cancer patients (10 non-IBC and 10 IBC), and 4 healthy volunteer biopsies were investigated. Fixed cells and tissues were analyzed by FTIR microspectroscopy and imaging, while live cells were studied by Raman microspectroscopy. Spectra were analyzed by hierarchical cluster analysis (HCA) and images by common k-means clustering algorithms. For both cell suspensions and single cells, FTIR spectroscopy showed sufficient high inter-group variability to delineate MDA-MB-231 and SUM-149 cell lines. Most significant differences were observed in the spectral regions of 1096–1108 and 1672–1692 cm⁻¹. Analysis of live cells by Raman microspectroscopy gave also a good discrimination of these cell types. The most discriminant regions were 688–992, 1019–1114, 1217–1375 and 1516–1625 cm⁻¹. Finally, k-means cluster analysis of FTIR images allowed delineating non-IBC from IBC tissues. This study demonstrates the potential of vibrational spectroscopy and imaging to discriminate between non-IBC and IBC at both cell and tissue levels.

It was reported that lumican inhibits the activity of metalloproteinase MMP-14 and melanoma cell migration in vitro and in vivo. Moreover, Snail triggers epithelial-to-mesenchymal transition and the metastatic potential of cancer cells. Therefore, the aim of this study was to examine the effect of lumican on Mock and Snail overexpressing melanoma B16F1 cells in vivo. Lung metastasis was analyzed after intravenous injections of Mock-B16F1 and Snail-B16F1 cells in Lum^+/+ and Lum^−/− mice. At day 14, mice were sacrificed, and lungs were collected. The number of lung metastatic nodules was significantly higher in mice injected with Snail-B16F1 cells as compared to mice injected with Mock-B16F1 cells confirming the pro-metastatic effect of Snail. This effect was stronger in Lum^−/− mice as compared to Lum^+/+, suggesting that endogenous lumican of wild-type mice significantly inhibits metastasis to lungs. Scanning electron and confocal microscopy investigations demonstrated that lumican inhibits the development of elongated cancer cell phenotypes which are known to develop invadopodia releasing MMPs. Moreover, lumican was shown to affect the expression of cyclin D1, cortactin, vinculin, hyaluronan synthase 2, heparanase, MMP-14 and the phosphorylation of FAK, AKT, p130 Cas and GSK3α/β. Altogether, these data demonstrated that lumican significantly inhibits lung metastasis in vivo, as well as cell invasion in vitro, suggesting that a lumican-based strategy targeting Snail-induced metastasis could be useful for melanoma treatment.

Ovarian cancer remains one of the most fatal cancers due to a lack of robust screening methods of detection at early stages. Extracellular matrix (ECM) mediates interactions between cancer cells and their microenvironment via specific molecules. Lumican, a small leucine-rich proteoglycan (SLRP), maintains ECM integrity and inhibits both melanoma primary tumor development, as well as metastatic spread. The aim of this study was to analyze the effect of lumican on tumor growth of murine ovarian epithelial cancer. C57BL/6 wild type mice (n = 12) and lumican-deficient mice (n = 10) were subcutaneously injected with murine ovarian epithelial carcinoma ID8 cells, and then sacrificed after 18 days. Analysis of tumor volumes demonstrated an inhibitory effect of endogenous lumican on ovarian tumor growth. The ovarian primary tumors were subjected to histological and immunohistochemical staining using anti-lumican, anti-αv integrin, anti-CD31 and anti-cyclin D1 antibodies, and then further examined by label-free infrared spectral imaging (IRSI), second harmonic generation (SHG) and Picrosirius red staining. The IR tissue images allowed for the identification of different ECM tissue regions of the skin and the ovarian tumor. Moreover, IRSI showed a good correlation with αv integrin immunostaining and collagen organization within the tumor. Our results demonstrate that lumican inhibits ovarian cancer growth mainly by altering collagen fibrilogenesis.

Biochemical and FTIR approaches to discriminate ovarian cancer cell-derived EVs.

Sulfated glycosaminoglycans (GAGs), namely, chondroitin sulfate (CS), dermatan sulfate, keratan sulfate, heparin, and heparan sulfate, are linear complex polysaccharides that are covalently attached to core proteins to form proteoglycans. They are present at the cell surface and in the extracellular matrix and play a key role in the regulation of cellular microenvironmental effectors. To better understand the biological functions of GAGs and particularly of CS-E (4,6-disulfated) at the molecular level, structurally well-defined oligosaccharides are necessary. Chemically synthesised biotinylated conjugates are useful to study the interactions with proteins at the intra- and extracellular levels. Herein, FTIR spectroscopy was used to characterise nine chondroitin oligosaccharides, including biotinylated or reducing CS-E di-, tetra- and hexasaccharides as well as their non-sulfated analogs. Spectral features characteristic of the vibrational modes of oligosaccharides (1640, 1626, 1565, 1418, 1375, and 1160 cm⁻¹), CS-E (1280–1200, 1134, 1065, 1034, 1000, 927, and 866–860, 815 cm⁻¹), and biotin (1681, 1460, 1425, and 792 cm⁻¹) were identified. FTIR spectroscopy was sensitive enough to reveal structural microheterogeneity, allowing distinguishing C-4 from C-6 sulfated isoforms. CS-E- and biotin-specific signatures were obtained via difference spectra. PCA plots revealed three distinct groups: biotinylated oligosaccharides, CS-E biotinylated oligosaccharides and CS-E reducing oligosaccharides. Furthermore, the first component clearly distinguished sulfated from non-sulfated forms, while component two tended to discriminate according to the chain length, exclusively for non-sulfated oligosaccharides. Identifying the spectral signatures of these oligosaccharides is an important step for future research on the monitoring of the internalisation of oligosaccharide- and cell-penetrating peptide-bound forms in drug-delivery studies.