[New Technologies for In Vivo Cancer Diagnostics]. / Nové technologie pouzívané pro in vivo diagnostiku nádoru.

BACKGROUND: The treatment of oncological diseases is based on the combination of surgery, representing the key step for the removal of the tumor tissue, radiotherapy, chemotherapy, and hormone therapy. However, the surgery is often accompanied by issue of determining the boundaries of the tumor. Prior the operation, the surgeon has information on preoperative findings, which indicate the location and extent of the tumor, but does not specify a clear boundary between the tumor and healthy tissue. This area cannot be recognized visually or by touch in most cases and when the tumor is not removed completely the patient has to undergo reoperation. AIM: Therefore, a number of research centers began to deal with the development of technology that would provide information about the state of the tissue in real time directly during surgery and would not require the collection or storage of tissue samples. These include MarginProbe, Spectropen tissue and spectroscopic scanner devices. Another group consists of imaging techniques using mass spectrometry approaches to determine the tissue specificity. Recently, the intraoperative mass spectrometry (REIMS) technique has undergone tremendous development. It uses an electronic scalpel using by the surgeon for cutting the tissue, when the resulting aerosol is discharged into the mass spectrometer that in tenths of seconds measures mass spectra of phospholipids, which are specific to the operated tissue (tumor or healthy). In the Czech Republic this technology has been already used for research purposes for the detection of drug deposited in the tumor and healthy tissue of mice suffering from melanoma. The obtained results show that with this apparatus it would be possible fundamentally affect the treatment and its efficacy in oncology as well. We will inform you about these new technologies and elucidate their principles and utilization.Key words: surgery - cancer - tumors - molecular diagnostics - mass spectrometry - databaseThis 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: 6. 9. 2016.

[p SRM, SWATH and HRM -  targeted proteomics approaches on TripleTOF 5600+ mass spectrometer and their applications in oncology research]. / p SRM, SWATH a HRM -  cílené proteomické prístupy na hmotnostním spektrometru TripleTOF 5600+ a jejich aplikace v onkologickém výzkumu.

Development of novel diagnostic and therapeutic approaches in cancer research requires sensitive and quantitative assays for determination of cancer associated proteins in clinical samples. Novel quantitative targeted proteomic approaches are overviewed in this communication. A major advantage of selected reaction monitoring (SRM) and pseudo- SRM lies in the selective and sensitive quantification of selected proteins in large sample sets. As such, they represent an alternative to immunochemical approaches. On the other hand, the potential of HRM and SWATH lies in recording of digital fingerprints, which enable post acquisition quantitative proteomic data mining on a similar basis to SRM. This article shows applications of targeted proteomics in a number of cancer research studies where they were used for quantification and validation of current or potential protein bio-markers and to study their role in cancer development and progression.

[Methods for studying tumor cell migration and invasiveness]. / Metody studia bunecné migrace a invazivity nádorových bunek.

Migration and invasiveness are phenotypic characteristics of cells that contribute to physiological processes, such as wound healing or embryogenesis and they are involved in serious pathological processes, namely in tumor cell metastasis. Availability of methods for studying migration and invasiveness of the cells is important for understanding molecular basis of these processes. In the case of cancer, migration, invasiveness and metastatic potential of tumor cells are key factors that determine clinical prognosis of the patients. This communication provides an overview of in vitro and in vivo methods which are used to study cell migration, invasion and metastasis. In vitro meth-ods for studying cell migration include simple two dimensional assays (scratch -  wound assay and the assay based on the effect of hepatocyte growth factor) and methods based on chemotaxis (Dunns chamber, videomicroscopy of cells, the use of carriers with chemoattractants). Methods for studying both cell migration and invasiveness in vitro include more complex systems based on the principle of the Boyden chamber (transwell migration/ invasive test, analysis of cell migration and invasion in xCELLigence system, confocal microscopy based approaches) as well as analysis of cell migration in microchannels. Our overview of in vivo methods provides an introduction into model organisms and methods used in this field, with an emphasis on the study of cancer metastasis in mouse models. The methods described in this review are mainly involved in larger research projects aiming at developing new diagnostic and therapeutic approaches in oncology.

[Development of PCR methods and their applications in oncological research and practice]. / Vývoj metod zalozených na PCR a jejich aplikace v onkologickém výzkumu a praxi.

Since its discovery, PCR has become a conventional method of molecular biology research laboratories and an indispensable tool in diagnostic medicine. Multiple variants of the PCR technique were developed, which enable the analysis of different bio-logical materials at different amounts and reaction conditions. This article briefly summarizes the PCR approaches and points out their applications in oncological research and practice.

[Quantitative mass spectrometry and its utilization in oncology]. / Kvantitativní hmotnostní spektrometrie a její vyuzití v onkologii.

Cancers are genetically and clinically very heterogeneous diseases; therefore, various proteomic studies have been trying to find bio-markers which can facilitate prognosis, diagnosis or treatment of these oncological diseases. The mass spectrometry is an effective tool for identification, quantitation, and characterization of biomolecules in the complex bio-logical samples. The first step suitable for selection of bio-markers called discovery proteomics provides a detailed analysis of the samples contributing to the identification of proteins, comparison of their presence in the samples, and selection of the convenient candidates for the prospective bio-markers. The next step of proteomics analysis is directed towards verification of chosen bio-markers with the approach called targeted proteomics. This technique evaluates presence and quantity of the proteins (biomarkers) in clinically precisely defined samples. This article focuses on the description of various approaches suitable for the quantitative analysis of the proteins connected with mass spectrometry.

A combined immunoprecipitation and mass spectrometric approach to determine deltaNp63-interacting partners.

Expression of p63 is essential for the formation of epidermis and other stratifying epithelia. Moreover p63 is highly expressed in several epithelial cancers and is involved in tumourigenesis and controlling chemo-sensitivity. The identification of p63 interacting partners is essential for understanding the complex network of gene regulation managing epithelial development and could also help to reveal signalling pathways participating in UV-damage response in human skin. We used a proteomic approach to identify proteins that interact with deltaNp63. Proteins were isolated by immunoprecipitation with deltaNp63 specific antibody and analysed by mass spectrometry. We identified 23 proteins as potential deltaNp63 binding partners that were not present in negative control samples. These results will be evaluated using other methods.

[Chemosensitivity prediction in tumor cells ex vivo--difficulties and limitations of the method]. / Predikce citlivosti nádorových bunek k chemoterapeutikum ex vivo--úskali a limitace vlastní metody.

Certain hope is entertained in the prediction of chemosensitivity in vitro/ ex vivo for the purpose of selecting the most effective treatment of malignant diseases with minimal patient loading. The possible choice of an effective substance based on the results of a simple ex vivo test would increase the success of the treatment in case of standard chemotherapy failure or in the treatment of primary chemoresistant tumor. MTT test seems to be an easy process for the prediction of chemosensitivity of isolated malignant cells ex vivo, however each method represents a simple tool, which can provide false results if incorrectly preformed. Numerous limitations significantly reduce the successful evaluation and constituent aspects of the methodic press to further reflections about the proper application of the test.

[Actual state of ex vivo chemoresistance testing of malignant tumors in Masaryk Memorial Cancer Institute Brno]. / Soucasný stav testování chemorezistence nádoru ex vivo v Masarykove Onkologickém Ustavu v Brne.

Chemoresistance assay results may play a role in cancer management decision process. Since August 2006 testing chemoresistance has been tested according to a protocol that was designed for this reason in our institute (Masaryk Memorial Cancer Institute). Five groups of different types of cancer in particular clinical stages were defined for chemosensitivity testing with: (1) metastatic malignant melanoma, (2) soft tissue sarcoma (STS), either primary or recurrent/metastic, (3) primary or metastatic renal cancer, (4) recurrent ovarian cancer and (5) other diagnosis "on clinician's request". In the period from September 2006 to November 2007, 25 samples of malignant melanoma (reproducible results in 9 cases), 29 samples of STS (relevant data in 11 cases), 36 samples of renal cancer (relevant results in 20 samples) and 16 samples of ovarian cancer (reproducible results in 11 cases) were acquired. Sensitivity to certain chemotherapy agent observed ex vivo does not necessarily mean that the cancer would also be sensitive to the same agent in vivo, however, ex vivo resistance with following in vivo sensitivity of the tumour has not been observed to date. The cultivation of malignant cells is very uncertain in solid tumours, which consist of several malignant cell multiclones (benign/stromal cells may outgrow malignant cells). This cultivation uncertainty as well as the unique complexity of human metabolism makes clinical application of chemoresistance testing at least very questionable. The small number of successfully evaluated samples has not yet provided us to carry out proper statistical evaluation and clinical application.