Interferon-induced transmembrane proteins IFITM1 and IFITM3 (IFITM1/3) play a role in both RNA viral restriction and in human cancer progression. Using immunohistochemical staining of FFPE tissue, we identified subgroups of cervical cancer patients where IFITM1/3 protein expression is inversely related to metastasis. Guide RNA-CAS9 methods were used to develop an isogenic IFITM1/IFITM3 double null cervical cancer model in order to define dominant pathways triggered by presence or absence of IFITM1/3 signalling. A pulse SILAC methodology identified IRF1, HLA-B, and ISG15 as the most dominating IFNγ inducible proteins whose synthesis was attenuated in the IFITM1/IFITM3 double-null cells. Conversely, SWATH-IP mass spectrometry of ectopically expressed SBP-tagged IFITM1 identified ISG15 and HLA-B as dominant co-associated proteins. ISG15ylation was attenuated in IFNγ treated IFITM1/IFITM3 double-null cells. Proximity ligation assays indicated that HLA-B can interact with IFITM1/3 proteins in parental SiHa cells. Cell surface expression of HLA-B was attenuated in IFNγ treated IFITM1/IFITM3 double-null cells. SWATH-MS proteomic screens in cells treated with IFITM1-targeted siRNA cells resulted in the attenuation of an interferon regulated protein subpopulation including MHC Class I molecules as well as IFITM3, STAT1, B2M, and ISG15. These data have implications for the function of IFITM1/3 in mediating IFNγ stimulated protein synthesis including ISG15ylation and MHC Class I production in cancer cells. The data together suggest that pro-metastatic growth associated with IFITM1/3 negative cervical cancers relates to attenuated expression of MHC Class I molecules that would support tumor immune escape.

• Klíčová slova
• CAS9 gene editing, Cervical cancer, IFITM1, Interferon, MHC Class I molecule, SILAC mass spectrometry,
• MeSH
• buněčné linie MeSH
• diferenciační antigeny fyziologie   MeSH
• lidé MeSH
• membránové proteiny fyziologie   MeSH
• MHC antigeny I. třídy metabolismus   MeSH
• nádory děložního čípku metabolismus   MeSH
• proteiny vázající RNA fyziologie   MeSH
• proteosyntéza fyziologie   MeSH
• Check Tag
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• diferenciační antigeny MeSH
• IFITM3 protein, human MeSH Prohlížeč
• leu-13 antigen MeSH Prohlížeč
• membránové proteiny MeSH
• MHC antigeny I. třídy MeSH
• proteiny vázající RNA MeSH

The incidence of cutaneous squamous cell carcinoma (cSCC) is rising. Whilst the majority are cured surgically, aggressive metastatic cSCC carry a poor prognosis. Inactivating mutations in transforming growth factor beta (TGF-β) receptors have been identified amongst genetic drivers of sporadic tumours and murine models of cSCC, suggesting a tumour suppressor function for TGF-β in normal skin. However, paradoxically, TGF-β acts as a tumour promoter in some murine model systems. Few studies have analysed the role of TGF-β/activin signalling in human normal skin, hyper-proliferative skin disorders and cSCC. Antibodies recognising phospho-SMAD proteins which are activated during canonical TGF-β/activin signalling were validated for use in immunohistochemistry. A tissue microarray comprising FFPE lesional and perilesional tissue from human primary invasive cSCC (n=238), cSCC in-situ (n=2) and keratocanthoma (n=9) were analysed in comparison with tissues from normal human scalp (n=10). Phosphorylated SMAD2 and SMAD3 were detected in normal interfollicular epidermal keratinocytes and were also highly localised to inner root sheath, matrix cells and Keratin 15 positive cells. Lesional cSCC tissue had significantly reduced activated SMAD2/3 compared to perilesional tissue, consistent with a tumour suppressor role for SMAD2/3 activators in cSCC. Increased cSCC tumour thickness inversely correlated with the presence of phospho-SMADs in tumour tissue suggesting that a reduction in canonical TGF-β/activin signalling may be associated with disease progression.

• Klíčová slova
• FFPE, IHC, SMAD, TGF-β, carcinoma,
• Publikační typ
• časopisecké články MeSH

The TP63 gene encodes two major protein variants that differ in their N-terminal sequences and have opposing effects. In breast, ΔNp63 is expressed by immature stem/progenitor cells and mature myoepithelial/basal cells and is a characteristic feature of basal-like triple-negative breast cancers (TNBCs). The expression and potential role of TAp63 in the mammary gland and breast cancers is less clear, partly due to the lack of studies that employ p63 isoform-specific antibodies. We used immunohistochemistry with ΔNp63-specific or TAp63-specific monoclonal antibodies to investigate p63 isoforms in 236 TNBCs. TAp63, but not ΔNp63, was seen in tumour-associated lymphocytes and other stromal cells. Tumour cells showed nuclear staining for ΔNp63 in 17% of TNBCs compared to 7.3% that were positive for TAp63. Whilst most TAp63+ tumours also contained ΔNp63+ cells, the levels of the two isoforms were independent of each other. ΔNp63 associated with metaplastic and medullary cancers, and with a basal phenotype, whereas TAp63 associated with androgen receptor, BRCA1/2 wild-type status and PTEN positivity. Despite the proposed effects of p63 on proliferation, Ki67 did not correlate with either p63 isoform, nor did they associate with p53 mutation status. ΔNp63 showed no association with patient outcomes, whereas TAp63+ patients showed fewer recurrences and improved overall survival. These findings indicate that both major p63 protein isoforms are expressed in TNBCs with different tumour characteristics, indicating distinct functional activities of p63 variants in breast cancer. Analysis of individual p63 isoforms provides additional information into TNBC biology, with TAp63 expression indicating improved prognosis.

• Klíčová slova
• Androgen receptor, BRCA1, BRCA2, PTEN, TAp63, Triple-negative breast cancer, ΔNp63,
• MeSH
• fenotyp MeSH
• fosfohydroláza PTEN genetika   MeSH
• lidé MeSH
• lokální recidiva nádoru genetika   MeSH
• mutace genetika   MeSH
• nádorové supresorové proteiny genetika   MeSH
• protein - isoformy genetika   MeSH
• protein BRCA1 genetika   MeSH
• protein BRCA2 genetika   MeSH
• regulace genové exprese u nádorů genetika   MeSH
• transkripční faktory genetika   MeSH
• triple-negativní karcinom prsu genetika   metabolismus   mortalita   MeSH
• Check Tag
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• BRCA1 protein, human MeSH Prohlížeč
• BRCA2 protein, human MeSH Prohlížeč
• fosfohydroláza PTEN MeSH
• nádorové supresorové proteiny MeSH
• protein - isoformy MeSH
• protein BRCA1 MeSH
• protein BRCA2 MeSH
• PTEN protein, human MeSH Prohlížeč
• TP63 protein, human MeSH Prohlížeč
• transkripční faktory MeSH

Plasticity of cancer cells, manifested by transitions between epithelial and mesenchymal phenotypes, represents a challenging issue in the treatment of neoplasias. Both epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET) are implicated in the processes of metastasis formation and acquisition of stem cell-like properties. Mouse double minute (MDM) 2 and MDMX are important players in cancer progression, as they act as regulators of p53, but their function in EMT and metastasis may be contradictory. Here, we show that the EMT phenotype in multiple cellular models and in clinical prostate and breast cancer samples is associated with a decrease in MDM2 and increase in MDMX expression. Modulation of EMT-accompanying changes in MDM2 expression in benign and transformed prostate epithelial cells influences their migration capacity and sensitivity to docetaxel. Analysis of putative mechanisms of MDM2 expression control demonstrates that in the context of defective p53 function, MDM2 expression is regulated by EMT-inducing transcription factors Slug and Twist. These results provide an alternative context-specific role of MDM2 in EMT, cell migration, metastasis, and therapy resistance.

• Klíčová slova
• MDM2/MDMX, SNAI2/SLUG, TWIST, epithelial-mesenchymal transition, prostate/breast cancer,
• MeSH
• epitelo-mezenchymální tranzice fyziologie   MeSH
• fenotyp MeSH
• heterografty MeSH
• jaderné proteiny biosyntéza   MeSH
• lidé MeSH
• myši nahé MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• nádory prostaty genetika   metabolismus   patologie   MeSH
• nádory prsu genetika   metabolismus   patologie   MeSH
• proteiny buněčného cyklu MeSH
• protoonkogenní proteiny c-mdm2 biosyntéza   MeSH
• protoonkogenní proteiny biosyntéza   MeSH
• transfekce MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• jaderné proteiny MeSH
• MDM2 protein, human MeSH Prohlížeč
• MDM4 protein, human MeSH Prohlížeč
• proteiny buněčného cyklu MeSH
• protoonkogenní proteiny c-mdm2 MeSH
• protoonkogenní proteiny MeSH

Glioblastoma is the most common and the most aggressive type of brain cancer. Aberrations of the RTK/RAS/PI3K-, p53-, and RB cell signaling pathways were recognized as a core requirement for pathogenesis of glioblastoma. The p53 tumor suppressor functions as a transcription factor transactivating expression of its target genes in response to various stress stimuli. We determined the p53 status in 36 samples of glioblastoma by functional analyses FASAY and split assay. Seventeen p53 mutations were detected and further analyzed by cDNA and gDNA sequencing in 17 patients (47.2 %). Fifteen (88.2 %) of the mutations were missense mutations causing amino acid substitutions, seven of them exhibited temperature-sensitivity. Two mutations were determined as short deletions, one of them causing formation of premature termination codon in position 247. Fluorescent in situ hybridization revealed the loss of the p53-specific 17p13.3 locus in four of 33 analyzed samples (12 %). In 12 out of 30 samples (40 %), the p53 protein accumulation was shown by immunoblotting. There was high (80 %) concordance between the presence of the clonal p53 mutation and the p53 protein accumulation.

• MeSH
• delece genu MeSH
• geny p53 * MeSH
• glioblastom genetika   metabolismus   MeSH
• hybridizace in situ fluorescenční MeSH
• kvasinky MeSH
• lidé středního věku MeSH
• lidé MeSH
• mutace * MeSH
• mutační analýza DNA MeSH
• nádorové buněčné linie MeSH
• nádorový supresorový protein p53 genetika   metabolismus   MeSH
• nádory mozku genetika   metabolismus   MeSH
• orgánová specificita MeSH
• sekvenční analýza DNA MeSH
• senioři MeSH
• teplota MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• nádorový supresorový protein p53 MeSH
• TP53 protein, human MeSH Prohlížeč