Oncogenic Viral Protein Interactions with p53 Family Proteins.

BACKGROUND: Cellular transformation induced by oncogenic viruses is a complex process including viral molecules, host cells and environmental factors. Viruses alone are unable to reproduce and thus they need a host to use their signalling, proteosynthetic and metabolic pathways. One target host molecule is the p53 tumour suppressor. Viral proteins functionally inactivate p53 and deregulate the expression of proteins active during apoptosis, cell proliferation and DNA damage response. Hepatitis virus B HbX protein and hepatitis virus C proteins NS2 and NS5A interact with p53 and prevent its localisation to the nucleus and thus reduce its transcriptional activity. Another mechanism lies in elevated p53 degradation caused by the BZLF1 protein of the Epstein-Barr virus, the LANA protein of the Kaposi sarcoma virus and human papilloma virus E6. The Merkel cell polyomavirus large T antigen does not interact directly with p53, however it acts through downregulation of p53 mediated transcription. The tax protein of human T cell lymphotropic virus type 1 modifies p53 posttranslationally and thus blocks its interaction with other factors of transcription machinery. Due to its tumour suppressor function and role in the maintenance of the genome integrity, the p53 protein is one of the best studied proteins. Following this, evolutionary homologues with important developmental functions p63 and p73 are intensively studied as well. Their roles in oncogenesis have not been clarified yet. PURPOSE: This review describes some of their known interactions with oncogenic viral proteins.

[Molecular Pathology of Colorectal Cancer, Microsatellite Instability - the Detection, the Relationship to the Pathophysiology and Prognosis]. / Molekulární patologie kolorektálního karcinomu, mikrosatelitová nestabilita - zpusob detekce, vztah k patofyziologii a prognóze.

BACKGROUND: Colorectal carcinoma (CRC) is third most common cancer worldwide with very heterogenous character. In most cases, it is caused by sporadic events leading to disruption of epithelial cells of the colon. The minority evolves from germline mutations associated with hereditary cancer syndromes. Mechanisms leading to mutations of oncogenes, tumour suppressors and genes of DNA repair mechanisms include: 1. chromosomal instability, 2. microsatellite instability and 3. CpG island methylator phenotype. Microsatellite instability (MSI) usually arises from a germline mutation of the component of mismatch repair machinery (MMR) or somatic hypermethylation of the MLH1 promoter. The diagnostic approaches include PCR methods and immunohistochemistry for the detection of the loss of MMR part. The aim of our study was to characterise the cohort of ongoing study of gut microbiome in CRC patients considering MSI. MATERIAL AND METHODS: The consecutive study group consisted of 103 patients diagnosed with CRC. The cohort consisted of 45 women (43.7%) and 58 men (56.3%). Patient age at the time of diagnosis was within the range of 31-83 years (median 66 years). The expression of MLH1, MSH2, MSH6 and PMS2 proteins was detected by immunohistochemical method and the positivity was correlated with the stage and the localization of the primary tumour. RESULTS: The MMR status was determined by immunohistochemical method in 43 (41.7%) from the existing total of 103 patients. MSI was detected in 11 (25.6%) cases while 32 (74.4%) were microsatellite stabile. With the respect to cancer clasification the most cases of MSI was detected in stage II (8 cases; 22.2%). In regard to localization of primary tumour, MSI rather correlates to right site CRC, while microsatellite stable tumours do not show any site preferences. CONCLUSION: Considering low number of MMR status determination in study group, statistic evaluation is inaccurate so far. However there is a trend in our cohort in relation to determination of the portion of MSI in CRC population and also in localization of primary tumour according to literature.Key words: colorectal carcinoma - microsatellite instability - Lynch syndrome The work was supported by the project MEYS - NPS I - LO1413 and AZV 16-31966A. The authors declare they have no potential conflicts of interest concerning drugs, products, or services used in the study. The Editorial Board declares that the manuscript met the ICMJE recommendation for biomedical papers.Submitted: 13. 3. 2017Accepted: 26. 3. 2017.

[Circulating Myeloid Suppressor Cells and Their Role in Tumour Immunology]. / Cirkulující myeloidní supresorové bunky a jejich úloha v nádorové imunologii.

BACKGROUND: Myeloid-derived suppressor cells (MDSCs) are heterogenic population of multipotent progenitors of myeloid lineage. For their immunosuppressive effect, MDSC are responsible for tumour escape from the host immune surveillance. Furthermore, MDSCs support tumour by promotion of angiogenesis and metastasis. Membrane markers of human MDSCs are myeloid markers CD11b and CD13, these cells are HLA-Drlow/- and expression of CD15 or CD14 differentiate them into granulocytic (Gr-MDSCs) and monocytic (Mo-MDSCs), resp. PATIENTS AND METHODS: Using flow cytometry, we investigated Mo-MDSC counts in peripheral blood of non-cancer individuals - control group (n = 61), breast (n = 39) and colorectal (n = 52) cancer patients. These cells were detected as CD45+CD11b+CD33+CD14+HLA-Drlow/- and quantified as percentage of total white blood cells and as absolute count. RESULTS: In control group, circulating Mo-MDSCs was gender-and age-independent and the average value was 1.09% and 0.073 × 109/l. Breast cancer patients had higher circulating Mo-MDSCs compared to control group with average values: 3.57% and 0.229 × 109/l (p < 0.001) and we also observed increase in Mo-MDSC number after granulopoietic growth factors administration (p = 0.043). Colorectal cancer patients had higher average number of circulating Mo-MDSCs compared to control group: 1.71% a 0.125 × 109/l (p = 0.003) and its number did not correlate with tumour clinicopathological stage, localization of primary tumour (colon vs. rectum), site (left vs. right) and microsatellite instability. CONCLUSION: Increased number of MDSCs in circulation and within tumour microenvironment has been associated with immune suppression and tumour progression. Colorectal cancer patients at diagnosis showed higher circulating Mo-MDSCs possibly reflecting immunosuppressive effect of tumour microenvironment. Change of Mo-MDSC number from baseline level need to be evaluated in the context of CRC patients outcome. Recombinant granulopoietic growth factors increase number of circulating Mo-MDSCs and the effect of this phenomenon on cancer prognosis remains to be elucidated.Key words: myeloid-derived suppressor cells - colorectal cancer - breast cancer - immunology - immunosuppression - G-CSF This work was supported by MEYS by NPU I (LO1413), grant AZV 16-31966A and MH DRO 00209805. The authors declare they have no potential conflicts of interest concerning drugs, products, or services used in the study. The Editorial Board declares that the manuscript met the ICMJE recommendation for biomedical papers.Submitted: 11. 3. 2017Accepted: 26. 3. 2017.

[Endoplasmic Reticulum Chaperones at the Tumor Cell Surface and in the Extracellular Space]. / Chaperony endoplazmatického retikula na povrchu nádorové bunky a v extracelulárním prostredí.

BACKGROUND: Endoplasmic reticulum chaperones are stress induced proteins capable of translocation into cytosol, cell membrane or extracellular space. The chaperones are transported from the endoplasmic reticulum particularly under endoplasmic reticulum stress conditions, while their constitutive extracellular expression was found in many cancers. Cell surface or extracellular endoplasmic reticulum chaperones take up distinct functions compared to their endoplasmic reticulum resident variants because they act like multifunctional receptors and thus affect cell signaling and proliferation. AIM: The presented review focuses primarily on endoplasmic reticulum chaperones expression on the cell surface of cancer cells and into extracellular space. The work describes possible mechanisms of chaperones translocation to the cancer cell surface, including KDEL transport mechanism and retrotranslocation and the influence of chaperone posttranslation modifications on their localization within the cell. Well described cancer cell surface endoplasmic reticulum chaperones include GRP78, GRP94, calreticulin and calnexin that are involved in cancer cell signaling in different ways. The attention is also paid to immunogenic properties of membrane-localized chaperones for their ability to participate in immune reactions. They can take part in innate and adaptive immune response through their interaction with toll-like receptors or during the antigen presentation as well as in tumor-specific immunity. The expression of endoplasmic reticulum chaperones on the cancer cells surface is potentially exploitable in specific antitumor therapy as well as vaccine therapy, thus the final part of this review is dedicated to this topic.Key words: endoplasmic reticulum - glucose-regulated proteins - molecular chaperones - KDEL sequence - immunobiologyThis work was supported by the project MEYS - NPS I - LO1413.The authors declare they have no potential conflicts of interest concerning drugs, products, or services used in the study.The Editorial Board declares that the manuscript met the ICMJE recommendation for biomedical papers.Submitted: 17. 5. 2016Accepted: 20. 7. 2016.

[In situ proximity ligation assay for detection of proteins, their interactions and modifications]. / Detekce proteinu, proteinových interakcí a modifikací s vyuzitím "proximity ligation assay" in situ.

To understand cellular processes and events responsible for their perturbations, proteomic analyses are needed in bio-medical research and clinical diagnostics. Several techniques based on specifically binding reagents (antibodies) or recombinant proteins (GFP fusion protein, methods of fluorescence/ bio-luminescence resonance energy transfer) are generally used to study protein location and activity resulting from secondary modifications and interactions. The in situ proximity ligation assay represents a novel technique of in situ protein imaging using DNA as a reporter molecule and DNA amplification processes. This method enables direct visualization of single molecules, their levels, modifications and pattern of interactions in individual fixed cells and tissues. Proximity probes consist of specific antibody with attached oligonucleotides that are used as reporter molecules for identification of such events. Proximity probes guide the formation of a circular DNA strand when bound in close proximity. The DNA circle after that serves as a template for rolling  circle amplification allowing the interaction to be visualized. Compared to available proteomic techniques benefiting from genetic engineering, in situ PLA enables study of endogenous proteins in their natural environment and thus can be used for clinical specimens. The areas of applicability where proximity ligation procedure can be used include any research field where protein interaction measurements are important, such as signal-ing pathway studies, monitoring of pharmacological treatment targets and oncological diagnostics.