The incidence of cutaneous malignant melanoma is increasing worldwide. While the treatment of initial stages of the disease is simple, the advanced disease frequently remains fatal despite novel therapeutic options . This requires identification of novel therapeutic targets in melanoma. Similarly to other types of tumours, the cancer microenvironment plays a prominent role and determines the biological properties of melanoma. Importantly, melanoma cell-produced exosomes represent an important tool of intercellular communication within this cancer ecosystem. We have focused on potential differences in the activity of exosomes produced by melanoma cells towards melanoma-associated fibroblasts and normal dermal fibroblasts. Cancer-associated fibroblasts were activated by the melanoma cell-produced exosomes significantly more than their normal counterparts, as assessed by increased transcription of genes for inflammation-supporting cytokines and chemokines, namely IL-6 or IL-8. We have observed that the response is dependent on the duration of the stimulus via exosomes and also on the quantity of exosomes. Our study demonstrates that melanoma-produced exosomes significantly stimulate the tumour-promoting proinflammatory activity of cancer-associated fibroblasts. This may represent a potential new target of oncologic therapy .

• Klíčová slova
• Cancer-associated fibroblasts, Exosomes, IL-6, IL-8, Melanoma, Proinflammatory cytokine,
• MeSH
• exozómy metabolismus   MeSH
• fibroblasty metabolismus   patologie   MeSH
• lidé MeSH
• melanom experimentální metabolismus   patologie   MeSH
• nádorové buňky kultivované MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Profilin 1 is a crucial actin regulator, interacting with monomeric actin and several actin-binding proteins controlling actin polymerization. Recently, it has become evident that this profilin isoform associates with microtubules via formins and interferes with microtubule elongation at the cell periphery. Recruitment of microtubule-associated profilin upon extensive actin polymerizations, for example, at the cell edge, enhances microtubule growth, indicating that profilin contributes to the coordination of actin and microtubule organization. Here, we provide further evidence for the profilin-microtubule connection by demonstrating that it also functions in centrosomes where it impacts on microtubule nucleation.

• MeSH
• aktiny metabolismus   MeSH
• Caco-2 buňky MeSH
• centrozom metabolismus   MeSH
• forminy metabolismus   MeSH
• genový knockout MeSH
• lidé MeSH
• melanom experimentální metabolismus   patologie   MeSH
• mikrofilamentové proteiny metabolismus   MeSH
• mikrotubuly metabolismus   MeSH
• myši MeSH
• nádory kůže metabolismus   patologie   MeSH
• polymerizace MeSH
• profiliny genetika   metabolismus   MeSH
• signální transdukce genetika   MeSH
• transfekce MeSH
• tubulin metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• aktiny MeSH
• forminy MeSH
• mikrofilamentové proteiny MeSH
• PFN1 protein, human MeSH Prohlížeč
• Pfn1 protein, mouse MeSH Prohlížeč
• profiliny MeSH
• tubulin MeSH

The globally increasing incidence of cancer, including melanoma, requires novel therapeutic strategies. Development of successful novel drugs is based on clear identification of the target mechanisms responsible for the disease progression. The specific cancer microenvironment represents a critically important aspect of cancer biology, which cannot be properly studied in simplistic cell culture conditions. Among other traditional options, the study of melanoma cell growth on the chicken chorioallantoic membrane offers several significant advantages. This model offers increased complexity compared to usual in silico culture models and still remains financially affordable. Using this model, we studied the growth of three established human melanoma cell lines: A2058, BLM, G361. The combination of histology, immunohistochemistry with the application of human-specific antibodies, intravascular injection of contrast material such as filtered Indian ink, Mercox solution and phosphotungstic acid, and X-ray micro-CT and live-cell monitoring was employed. Melanoma cells spread well on the chicken chorioallantoic membrane. However, invasion into the stroma of the chorioallantoic membrane and the limb primordium graft was rare. The melanoma cells also significantly influenced the architecture of the blood vessel network, resulting in the orientation of the vessels to the site of the tumour cell inoculation. The system of melanoma cell culture on the chorioallantoic membrane is suitable for the study of melanoma cell growth, particularly of rearrangement of the host vascular pattern after cancer cell implantation. The system also has promising potential for further development.

• Klíčová slova
• Cancer microenvironment, Cancer-associated fibroblasts, Chorioallantoic membrane, Embryo, Melanoma, Melanoma invasiveness,
• MeSH
• biologické modely * MeSH
• chorioalantoická membrána metabolismus   patologie   MeSH
• imunohistochemie MeSH
• kur domácí MeSH
• kuřecí embryo MeSH
• lidé MeSH
• melanom experimentální metabolismus   patologie   MeSH
• nádorové buňky kultivované MeSH
• zvířata MeSH
• Check Tag
• kuřecí embryo MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Immunotherapy emerges as a fundamental approach in cancer treatment. Up to date, the efficacy of numerous different immunotherapies has been evaluated. The use of microorganisms or their parts for immune cell activation, referred to as Pathogen-Associated Molecular Patterns (PAMPs), represents highly promising concept. The therapeutic effect of PAMPs can be further amplified by suitable combination of different types of PAMPs such as Toll like receptor (TLR) agonists and phagocytosis activating ligands. Previously, we used the combination of phagocytosis activating ligand (mannan) and mixture of TLR agonists (resiquimod (R-848), poly(I:C), inactivated Listeria monocytogenes) for successful treatment of melanoma in murine B16-F10 model. In the present study, we optimized the composition and timing of previously used mixture. Therapeutic mixture based on well-defined chemical compounds consisted of mannan anchoring to tumor cell surface by biocompatible anchor for membranes (BAM) and TLR agonists resiquimod, poly(I:C), and lipoteichoic acid (LTA). The optimization resulted in (1) eradication of advanced stage progressive melanoma in 83% of mice, (2) acquisition of resistance to tumor re-transplantation, and (3) potential anti-metastatic effect. After further investigation of mechanisms, underlying anti-tumor responses, we concluded that both innate and adaptive immunity are activated and involved in these processes. We tested the efficacy of our treatment in Panc02 murine model of aggressive pancreatic tumor as well. Simultaneous application of agonistic anti-CD40 antibody was necessary to achieve effective therapeutic response (80% recovery) in this model. Our results suggest that herein presented immunotherapeutic approach is a promising cancer treatment strategy with the ability to eradicate not only primary tumors but also metastases.

• Klíčová slova
• Cancer immunotherapy, Melanoma B16-F10, Metastasis, Panc02, Phagocytosis, TLR agonists,
• MeSH
• adenokarcinom imunologie   patologie   terapie   MeSH
• fagocytóza * MeSH
• imidazoly terapeutické užití   MeSH
• imunoterapie MeSH
• kyseliny teichoové terapeutické užití   MeSH
• lipopolysacharidy terapeutické užití   MeSH
• mannany terapeutické užití   MeSH
• melanom experimentální imunologie   patologie   terapie   MeSH
• myši inbrední C57BL MeSH
• nádorové buněčné linie MeSH
• nádory slinivky břišní imunologie   patologie   terapie   MeSH
• neutrofily imunologie   MeSH
• poly I-C terapeutické užití   MeSH
• toll-like receptory agonisté   MeSH
• tumor burden účinky léků   MeSH
• tumor infiltrující lymfocyty imunologie   MeSH
• zvířata MeSH
• Check Tag
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• imidazoly MeSH
• kyseliny teichoové MeSH
• lipopolysacharidy MeSH
• lipoteichoic acid MeSH Prohlížeč
• mannany MeSH
• poly I-C MeSH
• resiquimod MeSH Prohlížeč
• toll-like receptory MeSH

We report that tumor cells without mitochondrial DNA (mtDNA) show delayed tumor growth, and that tumor formation is associated with acquisition of mtDNA from host cells. This leads to partial recovery of mitochondrial function in cells derived from primary tumors grown from cells without mtDNA and a shorter lag in tumor growth. Cell lines from circulating tumor cells showed further recovery of mitochondrial respiration and an intermediate lag to tumor growth, while cells from lung metastases exhibited full restoration of respiratory function and no lag in tumor growth. Stepwise assembly of mitochondrial respiratory (super)complexes was correlated with acquisition of respiratory function. Our findings indicate horizontal transfer of mtDNA from host cells in the tumor microenvironment to tumor cells with compromised respiratory function to re-establish respiration and tumor-initiating efficacy. These results suggest pathophysiological processes for overcoming mtDNA damage and support the notion of high plasticity of malignant cells.

• MeSH
• citrátsynthasa metabolismus   MeSH
• elektronový transportní řetězec metabolismus   MeSH
• energetický metabolismus MeSH
• homologní transplantace MeSH
• melanom experimentální patologie   MeSH
• messenger RNA metabolismus   MeSH
• mitochondriální DNA metabolismus   MeSH
• mitochondrie genetika   metabolismus   ultrastruktura   MeSH
• myši inbrední BALB C MeSH
• myši inbrední C57BL MeSH
• myši inbrední NOD MeSH
• myši SCID MeSH
• myši MeSH
• NADH-dehydrogenasa genetika   metabolismus   MeSH
• nádorové buněčné linie MeSH
• nádory plic patologie   sekundární   MeSH
• proliferace buněk MeSH
• reaktivní formy kyslíku metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• citrátsynthasa MeSH
• elektronový transportní řetězec MeSH
• messenger RNA MeSH
• mitochondriální DNA MeSH
• NADH-dehydrogenasa MeSH
• reaktivní formy kyslíku MeSH

AIM: We investigated differences of metastatic spread of normal proteinase-activated receptor-2 (Par2+/+) melanoma B16 in Par2-/- (knock-out) animals compared to C57Bl6 mice. MATERIALS AND METHODS: Nine knock-out mice B6.Cg-F2rl1tm1Mslb/J (Par2-/-) and nine C57Bl6/J controls were subcutaneously inoculated with B16 melanoma tissue cells. Twelve days after inoculation, all primary tumors were removed. Survival and metastatic spread was followed for up to 100 days after primary tumor extirpation. RESULTS: Excised primary tumors were on average larger in Par2-/- mice (360 mm3 vs. 221 mm3 in C57Bl6/J). Distant spontaneous metastases developed in only 3 of 9 of Par2-/- mice in comparison to 6 of 9 controls. The average survival time was 84 days in Par2-/- animals compared to 63 days in C57Bl6/J mice. CONCLUSION: Host Par2 melanoma model contributes to the limitation of local cancer progression in one area, while on the other hand is important for enhancing distant metastatic spread.

• Klíčová slova
• PAR2, melanoma, metastasis, murine model,
• MeSH
• imunohistochemie MeSH
• melanom experimentální genetika   metabolismus   patologie   MeSH
• myši inbrední C57BL MeSH
• myši knockoutované MeSH
• myši MeSH
• receptor PAR-2 genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• receptor PAR-2 MeSH

The application of the phagocytic receptor agonists in cancer immunotherapy was studied. Agonists (laminarin, molecules with terminal mannose, N-Formyl-methioninyl-leucyl-phenylalanine) were firmly anchored to the tumor cell surface. When particular agonists of phagocytic receptors were used together with LPS (Toll-like receptor agonist), high synergy causing tumour shrinkage and a temporary or permanent disappearance was observed. Methods of anchoring phagocytic receptor agonists (charge interactions, anchoring based on hydrophobic chains, covalent bonds) and various regimes of phagocytic agonist/LPS mixture applications were tested to achieve maximum therapeutic effect. Combinations of mannan/LPS and f-MLF/LPS (hydrophobic anchors) in appropriate (pulse) regimes resulted in an 80% and 60% recovery for mice, respectively. We propose that substantial synergy between agonists of phagocytic and Toll-like receptors (TLR) is based on two events. The TLR ligand induces early and massive inflammatory infiltration of tumors. The effect of this cell infiltrate is directed towards tumor cells, bearing agonists of phagocytic receptors on their surface. The result of these processes was effective killing of tumor cells. This novel approach represents exploitation of innate immunity mechanisms for treating cancer.

• MeSH
• aktivace makrofágů účinky léků   MeSH
• analýza přežití MeSH
• cytokiny metabolismus   MeSH
• glukany MeSH
• imunoterapie * MeSH
• ligandy MeSH
• lipopolysacharidy farmakologie   MeSH
• mannosa chemie   MeSH
• melanom experimentální farmakoterapie   imunologie   patologie   MeSH
• modely nemocí na zvířatech MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• polysacharidy farmakologie   terapeutické užití   MeSH
• proliferace buněk účinky léků   MeSH
• průtoková cytometrie MeSH
• receptory imunologické agonisté   metabolismus   MeSH
• signální transdukce účinky léků   MeSH
• toll-like receptory metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• cytokiny MeSH
• glukany MeSH
• laminaran MeSH Prohlížeč
• ligandy MeSH
• lipopolysacharidy MeSH
• mannosa MeSH
• phagocytosis receptor MeSH Prohlížeč
• polysacharidy MeSH
• receptory imunologické MeSH
• toll-like receptory MeSH

Studies over the past decade have clearly shown that s.c. implant of primary and cultured tumor cells rarely leads to the occurrence of metastatic disease. Orthotopic transplantation of cell suspensions, surgical orthotopic implantation (SOI) of cancer tissue fragments resulted in metastases in many cancer types reaching 100% successful rate. We compared two metastatic models - heterotopic model of Lewis lung cancer and orthotopic B16 mouse melanoma. Both models were syngeneic with high metastatic ratio in C57BL/6 mice after transplantation of cancer cells, by injection into subcutaneous region of mice tail and without surgical intervention. The conclusion is that the localisation of cancer cell injection is a crucial condition for metastatic potential. The site with 100% haematogenous and lymph metastasis rate, after simple injection of cancer cells only, has been defined in mice, without dependence on the genetically predisposition and tumor cell line.

• MeSH
• červený fluorescenční protein MeSH
• karcinom plic Lewisové metabolismus   patologie   MeSH
• luminescentní proteiny metabolismus   MeSH
• melanom experimentální metabolismus   patologie   MeSH
• metastázy nádorů * MeSH
• modely nemocí na zvířatech MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• zelené fluorescenční proteiny metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Názvy látek
• luminescentní proteiny MeSH
• zelené fluorescenční proteiny MeSH

Nuclease from tomato (TBN1) was produced by in planta biotechnology purified and tested for its anticarcinogenic properties. The nuclease was cytostatic after its intratumoral administration to nude mice bearing human melanoma or prostate carcinoma or after tumor targeting by TBN1 administration intravenously as conjugate with polyethylene glycol (PEG). Inhibitory effects of TBN1 on tumor growth were comparable to effects of bovine seminal RNase (BS-RNase), but the inhibition was reached at about ten times lower protein concentration. Simultaneously, TBN1 exhibited a lower degree of embryotoxicity compared to BS-RNAse and other nucleases. TBN1 showed significant stability in vivo, because it was readily detected after its administration intratumorally or intravenously by the fluorescence methods. Intravenous administration of TBN1-PEG caused significant inhibition of tumor proliferation without obvious degenerative changes, while direct administration of TBN1 into melanoma tumors led to rapid tumor tissue degeneration. The fact can be essential for the mode of TBN1 biological action that mature nuclease is a small (36 kDa) thermostable glycoprotein that has ability to destroy human 28S, 18S, 7S and 5.8S RNA, circular RNAs, double-stranded RNA in vitro and shows DNase and 3'nucleotidase activities.

• MeSH
• antitumorózní látky farmakologie   MeSH
• apoptóza účinky léků   MeSH
• dvouvláknová RNA genetika   metabolismus   MeSH
• embryo savčí enzymologie   patologie   MeSH
• endodeoxyribonukleasy farmakologie   MeSH
• fluorescenční protilátková technika MeSH
• glykosylace MeSH
• lidé MeSH
• melanom experimentální patologie   MeSH
• míra přežití MeSH
• myši nahé MeSH
• myši MeSH
• nádorové buňky kultivované MeSH
• nádory prostaty patologie   MeSH
• rekombinantní proteiny terapeutické užití   MeSH
• skot MeSH
• Solanum lycopersicum enzymologie   MeSH
• testikulární nádory patologie   MeSH
• western blotting MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• myši MeSH
• skot MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antitumorózní látky MeSH
• dvouvláknová RNA MeSH
• endodeoxyribonukleasy MeSH
• rekombinantní proteiny MeSH

We report a syngeneic model of spontaneous metastatic B16-F10 mouse melanoma in C57/BL6 mice with a very high metastatic frequency that mimics clinical metastatic melanoma. The B16 melanoma cells were injected between the skin and cartilage on the dorsal side of the ear. The model generated lymphatic and visceral metastases in all of the tested animals. In mice with large primary tumors, tumor weight correlated with the tumor growth time and also with the number of metastases in lymph nodes and organs. The dorsal ear space between the skin and cartilage enables both lymphatic and hematogenous metastatic spread. The model should be useful to study the mechanism of melanoma metastasis and to develop therapy for this currently untreatable disease.

• MeSH
• lymfatické metastázy MeSH
• melanom experimentální patologie   sekundární   MeSH
• modely nemocí na zvířatech MeSH
• myši inbrední C57BL MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• transplantace nádorů MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH