Breast cancer is the most frequently diagnosed cancer in women worldwide. Although dramatically increased survival rates of early diagnosed cases have been observed, late diagnosed patients and metastatic cancer may still be considered fatal. The present study's main focus was on cancer‑associated fibroblasts (CAFs) which is an active component of the tumor microenvironment (TME) regulating the breast cancer ecosystem. Transcriptomic profiling and analysis of CAFs isolated from breast cancer skin metastasis, cutaneous basal cell carcinoma, and squamous cell carcinoma unravelled major gene candidates such as IL6, VEGFA and MFGE8 that induced co‑expression of keratins‑8/‑14 in the EM‑G3 cell line derived from infiltrating ductal breast carcinoma. Western blot analysis of selected keratins (keratin‑8, ‑14, ‑18, ‑19) and epithelial‑mesenchymal transition‑associated markers (SLUG, SNAIL, ZEB1, E‑/N‑cadherin, vimentin) revealed specific responses pointing to certain heterogeneity of the studied CAF populations. Experimental in vitro treatment using neutralizing antibodies against IL-6, VEGF‑A and MFGE8 attenuated the modulatory effect of CAFs on EM‑G3 cells. The present study provided novel data in characterizing and understanding the interactions between CAFs and EM‑G3 cells in vitro. CAFs of different origins support the pro‑inflammatory microenvironment and influence the biology of breast cancer cells. This observation potentially holds significant interest for the development of novel, clinically relevant approaches targeting the TME in breast cancer. Furthermore, its implications extend beyond breast cancer and have the potential to impact a wide range of other cancer types.

• Klíčová slova
• breast cancer, cell differentiation, epithelial‑mesenchymal interaction, neutralizing antibody, tumor microenvironment,
• MeSH
• antigeny povrchové MeSH
• fibroblasty asociované s nádorem * metabolismus   MeSH
• fibroblasty metabolismus   MeSH
• keratiny genetika   metabolismus   MeSH
• lidé MeSH
• maligní melanom kůže MeSH
• MFC-7 buňky MeSH
• mléčné bílkoviny genetika   metabolismus   MeSH
• nádorové buněčné linie MeSH
• nádorové mikroprostředí genetika   MeSH
• nádory prsu * farmakoterapie   genetika   metabolismus   MeSH
• prognóza MeSH
• transkriptom MeSH
• Check Tag
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• antigeny povrchové MeSH
• keratiny MeSH
• MFGE8 protein, human MeSH Prohlížeč
• mléčné bílkoviny MeSH

Protein phosphatase magnesium-dependent 1 delta (PPM1D) terminates cell response to genotoxic stress by negatively regulating the tumor suppressor p53 and other targets at chromatin. Mutations in the exon 6 of the PPM1D result in production of a highly stable, C-terminally truncated PPM1D. These gain-of-function PPM1D mutations are present in various human cancers but their role in tumorigenesis remains unresolved. Here we show that truncated PPM1D impairs activation of the cell cycle checkpoints in human non-transformed RPE cells and allows proliferation in the presence of DNA damage. Next, we developed a mouse model by introducing a truncating mutation in the PPM1D locus and tested contribution of the oncogenic PPM1DT allele to colon tumorigenesis. We found that p53 pathway was suppressed in colon stem cells harboring PPM1DT resulting in proliferation advantage under genotoxic stress condition. In addition, truncated PPM1D promoted tumor growth in the colon in Apcmin mice and diminished survival. Moreover, tumor organoids derived from colon of the ApcminPpm1dT/+ mice were less sensitive to 5-fluorouracil when compared to ApcminPpm1d+/+and the sensitivity to 5-fluorouracil was restored by inhibition of PPM1D. Finally, we screened colorectal cancer patients and identified recurrent somatic PPM1D mutations in a fraction of colon adenocarcinomas that are p53 proficient and show defects in mismatch DNA repair. In summary, we provide the first in vivo evidence that truncated PPM1D can promote tumor growth and modulate sensitivity to chemotherapy.

• MeSH
• chromatin účinky léků   MeSH
• exony genetika   MeSH
• fluoruracil farmakologie   MeSH
• karcinogeneze účinky léků   MeSH
• kontrolní body buněčného cyklu genetika   MeSH
• lidé MeSH
• mutace genetika   MeSH
• myši MeSH
• nádorový supresorový protein p53 genetika   MeSH
• nádory tračníku farmakoterapie   genetika   patologie   MeSH
• oprava DNA účinky léků   MeSH
• poškození DNA účinky léků   MeSH
• proliferace buněk účinky léků   MeSH
• protein familiární adenomatózní polypózy genetika   MeSH
• proteinfosfatasa 2C genetika   MeSH
• regulace genové exprese u nádorů účinky léků   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• adenomatous polyposis coli protein, mouse MeSH Prohlížeč
• chromatin MeSH
• fluoruracil MeSH
• nádorový supresorový protein p53 MeSH
• PPM1D protein, human MeSH Prohlížeč
• protein familiární adenomatózní polypózy MeSH
• proteinfosfatasa 2C MeSH
• Trp53 protein, mouse MeSH Prohlížeč

Head and neck squamous cell carcinoma is one of the most aggressive tumours and is typically diagnosed too late. Late diagnosis requires an urgent decision on an effective therapy. An individualized test of chemosensitivity should quickly indicate the suitability of chemotherapy and radiotherapy. No ex vivo chemosensitivity assessment developed thus far has become a part of general clinical practice. Therefore, we attempted to explore the new technique of coherence-controlled holographic microscopy to investigate the motility and growth of live cells from a head and neck squamous cell carcinoma biopsy. We expected to reveal behavioural patterns characteristic for malignant cells that can be used to imrove future predictive evaluation of chemotherapy. We managed to cultivate primary SACR2 carcinoma cells from head and neck squamous cell carcinoma biopsy verified through histopathology. The cells grew as a cohesive sheet of suspected carcinoma origin, and western blots showed positivity for the tumour marker p63 confirming cancerous origin. Unlike the roundish colonies of the established FaDu carcinoma cell line, the SACR2 cells formed irregularly shaped colonies, eliciting the impression of the collective invasion of carcinoma cells. Time-lapse recordings of the cohesive sheet activity revealed the rapid migration and high plasticity of these epithelial-like cells. Individual cells frequently abandoned the swiftly migrating crowd by moving aside and crawling faster. The increasing mass of fast migrating epithelial-like cells before and after mitosis confirmed the continuation of the cell cycle. In immunofluorescence, analogously shaped cells expressed the p63 tumour marker, considered proof of their origin from a carcinoma. These behavioural traits indicate the feasible identification of carcinoma cells in culture according to the proposed concept of the carcinoma cell dynamic phenotype. If further developed, this approach could later serve in a new functional online analysis of reactions of carcinoma cells to therapy. Such efforts conform to current trends in precision medicine.

• MeSH
• biopsie MeSH
• buněčný cyklus fyziologie   MeSH
• holografie metody   MeSH
• imunohistochemie MeSH
• lidé středního věku MeSH
• lidé MeSH
• mikroskopie metody   MeSH
• nádorové biomarkery metabolismus   MeSH
• nádorové buňky kultivované MeSH
• nádorové supresorové proteiny metabolismus   MeSH
• nádory hlavy a krku metabolismus   patologie   MeSH
• pohyb buněk fyziologie   MeSH
• senioři MeSH
• spinocelulární karcinom metabolismus   patologie   MeSH
• transkripční faktory metabolismus   MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• nádorové biomarkery MeSH
• nádorové supresorové proteiny MeSH
• TP63 protein, human MeSH Prohlížeč
• transkripční faktory MeSH

It is widely recognized that stromal fibroblasts significantly influence biological properties of multiple tumors including breast cancer. However, these epithelial-mesenchymal interactions seem to be essential in tumor biology and it is not fully clear whether this interaction is tumor type-specific or has a more general non-specific character. To elucidate this question, we tested the effect of cancer-associated fibroblasts (CAFs) isolated from different types of tumors (breast cancer skin metastasis, cutaneous basal cell carcinoma and melanoma, squamous cell carcinoma arising from oral cavity mucous membrane) on the EM-G3 breast cancer cell line. The results were compared with control experiments using normal human dermal fibroblasts, 3T3 mouse fibroblasts, and 3T3 fibroblasts influenced by the fibroblasts prepared from the basal cell carcinoma. Our results demonstrated that expression of luminal marker keratin 8 was influenced only by CAFs prepared from any tested tumors. In contrast, all tested types of fibroblasts showed a strong stimulatory effect on the expression of basal/myoepithelial marker keratin 14. The CAFs also elevated the number of cells with positivity for both keratins 8 and 14 that are similar to ductal originated precursor cells. The expression of proliferation marker Ki67 was not influenced by any of the tested fibroblasts. In conclusion, our data indicate that CAFs are able to influence the phenotype of a breast cancer cell line and this effect is based on a tumor type-unspecific mechanism. Finally, a clear functional difference between normal and CAFs was demonstrated.

• MeSH
• bazocelulární karcinom metabolismus   patologie   MeSH
• buňky 3T3 MeSH
• fibroblasty metabolismus   MeSH
• keratin-8 metabolismus   MeSH
• kokultivační techniky MeSH
• lidé MeSH
• melanom metabolismus   patologie   MeSH
• myši MeSH
• nádorové biomarkery metabolismus   MeSH
• nádorové buněčné linie MeSH
• nádory kůže metabolismus   patologie   sekundární   MeSH
• nádory prsu metabolismus   patologie   MeSH
• spinocelulární karcinom metabolismus   patologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• keratin-8 MeSH
• KRT8 protein, human MeSH Prohlížeč
• nádorové biomarkery MeSH

BACKGROUND: Transformed phenotypes are common to cell lines derived from various cancers. Proteome profiling is a valuable tool that may reveal uncharacteristic cell phenotypes in transformed cells. Changes in expression of glutathione S-transferases (GSTs) and other proteins interacting with glutathione (GSH) in model cell lines could be of particular interest. METHODS: We compared the phenotypes of breast cell lines EM-G3, HCC1937, MCF7 and MDA-MB-231 using 2-D electrophoresis (2-DE). We further separated GSH-binding proteins from the cell lines using affinity chromatography with GSH-Sepharose 4B, performed 2-DE analysis and identified the main protein spots. RESULTS: Correlation coefficients among 2-DE gels from the cell lines were lower than 0.65, pointing to dissimilarity among the cell lines. Differences in primary constituents of the cytoskeleton were shown by the 2-D protein maps and western blots. The spot patterns in gels of GSH-binding fractions from primary carcinoma-derived cell lines HCC1937 and EM-G3 were similar to each other, and they differed from the spot patterns of cell lines MCF7 and MDA-MB-231 that were derived from pleural effusions of metastatic mammary carcinoma patients. Major differences in the expression of GST P1-1 and carbonyl reductase [NADPH] 1 were observed among the cell lines, indicating differential abilities of the cell lines to metabolize xenobiotics. CONCLUSIONS: Our results confirmed the applicability of targeted affinity chromatography to proteome profiling and allowed us to characterize the phenotypes of four breast cancer cell lines.

• MeSH
• 2D gelová elektroforéza MeSH
• alkoholoxidoreduktasy metabolismus   MeSH
• capecitabinum MeSH
• chromatografie afinitní MeSH
• cisplatina aplikace a dávkování   MeSH
• deoxycytidin aplikace a dávkování   analogy a deriváty   MeSH
• dospělí MeSH
• fenotyp MeSH
• fluoruracil aplikace a dávkování   analogy a deriváty   MeSH
• glutathion-S-transferasa fí metabolismus   MeSH
• karboplatina aplikace a dávkování   MeSH
• kolorektální nádory farmakoterapie   metabolismus   patologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• míra přežití MeSH
• nádory hlavy a krku farmakoterapie   metabolismus   patologie   MeSH
• nádory plic farmakoterapie   metabolismus   patologie   MeSH
• nádory prsu farmakoterapie   metabolismus   patologie   MeSH
• neurofyziologie MeSH
• organoplatinové sloučeniny aplikace a dávkování   MeSH
• oxaliplatin MeSH
• paclitaxel aplikace a dávkování   MeSH
• protokoly protinádorové kombinované chemoterapie terapeutické užití   MeSH
• senioři MeSH
• studie případů a kontrol MeSH
• výsledek terapie MeSH
• western blotting MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Názvy látek
• alkoholoxidoreduktasy MeSH
• capecitabinum MeSH
• CBR1 protein, human MeSH Prohlížeč
• cisplatina MeSH
• deoxycytidin MeSH
• fluoruracil MeSH
• glutathion-S-transferasa fí MeSH
• GSTP1 protein, human MeSH Prohlížeč
• karboplatina MeSH
• organoplatinové sloučeniny MeSH
• oxaliplatin MeSH
• paclitaxel MeSH

BACKGROUND: Breast carcinomas represent a heterogeneous group of tumors diverse in behavior, outcome, and response to therapy. Identification of proteins resembling the tumor biology can improve the diagnosis, prediction, treatment selection, and targeting of therapy. Since the beginning of the post-genomic era, the focus of molecular biology gradually moved from genomes to proteins and proteomes and to their functionality. Proteomics can potentially capture dynamic changes in protein expression integrating both genetic and epigenetic influences. METHODS: We prepared primary cultures of epithelial cells from 23 breast cancer tissue samples and performed comparative proteomic analysis. Seven patients developed distant metastases within three-year follow-up. These samples were included into a metastase-positive group, the others formed a metastase-negative group. Two-dimensional electrophoretical (2-DE) gels in pH range 4-7 were prepared. Spot densities in 2-DE protein maps were subjected to statistical analyses (R/maanova package) and data-mining analysis (GUHA). For identification of proteins in selected spots, liquid chromatography-tandem mass spectrometry (LC-MS/MS) was employed. RESULTS: Three protein spots were significantly altered between the metastatic and non-metastatic groups. The correlations were proven at the 0.05 significance level. Nucleophosmin was increased in the group with metastases. The levels of 2,3-trans-enoyl-CoA isomerase and glutathione peroxidase 1 were decreased. CONCLUSION: We have performed an extensive proteomic study of mammary epithelial cells from breast cancer patients. We have found differentially expressed proteins between the samples from metastase-positive and metastase-negative patient groups.

• MeSH
• 2D gelová elektroforéza * MeSH
• dospělí MeSH
• epitelové buňky metabolismus   MeSH
• lidé středního věku MeSH
• lidé MeSH
• metastázy nádorů patologie   patofyziologie   MeSH
• nádorové buňky kultivované MeSH
• nádorové proteiny metabolismus   MeSH
• nádory prsu metabolismus   MeSH
• peptidové mapování MeSH
• proteomika MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Názvy látek
• nádorové proteiny MeSH