The overexpression of MYC genes is frequently found in many human cancers, including adult and pediatric malignant brain tumors. Targeting MYC genes continues to be challenging due to their undruggable nature. Using our prediction algorithm, the nine-amino-acid activation domain (9aaTAD) has been identified in all four Yamanaka factors, including c-Myc. The predicted activation function was experimentally demonstrated for all these short peptides in transactivation assay. We generated a set of c-Myc constructs (1-108, 69-108 and 98-108) in the N-terminal regions and tested their ability to initiate transcription in one hybrid assay. The presence and absence of 9aaTAD (region 100-108) in the constructs strongly correlated with their activation functions (5-, 3- and 67-times respectively). Surprisingly, we observed co-activation function of the myc region 69-103, called here acetyl-TAD, previously described by Faiola et al. (Mol Cell Biol 25:10220-10234, 2005) and characterized in this study as a new domain collaborating with the 9aaTAD. We discovered strong interactions on a nanomolar scale between the Myc-9aaTAD activation domains and the KIX domain of CBP coactivator. We showed conservation of the 9aaTADs in the MYC family. In summary for the c-Myc oncogene, the acetyl-TAD and the 9aaTAD domains jointly mediated activation function. The c-Myc protein is largely intrinsically disordered and therefore difficult to target with small-molecule inhibitors. For the c-Myc driven tumors, the strong c-Myc interaction with the KIX domain represents a promising druggable target.
- MeSH
- aktivace transkripce MeSH
- lidé MeSH
- proteinové domény MeSH
- protoonkogenní proteiny c-myc * metabolismus genetika MeSH
- sekvence aminokyselin MeSH
- vazba proteinů * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Závěrečná zpráva o řešení grantu Agentury pro zdravotnický výzkum MZ ČR
nestr.
Multiformní glioblastom (GBM) je běžný maligní nádor mozku u dospělých a navzdory agresivní léčbě zůstává prakticky nevyléčitelný se špatnou prognózou a mediánem přežití 12 měsíců po resekci, radioterapii a chemoterapii. Terapeutické přístupy k imunoterapii GBM mají omezenou úspěšnost, zejména v důsledku chráněné bariéry mozku a imunosupresivního mikroprostředí nádoru. Nový výzkum naznačuje možnost použití imunitních efektorových buněk, které účinně zničí maligní GBM buňky. Naše předchozí studie ukázala, že vrozené gamma-delta (gd) T buňky byly vysoce reaktivní na nádor GBM. Cílem navrhovaného projektu je buněčná a molekulární analýza gd T buněk u pacientů před a během léčby včetně podrobné charakterizace jejich interakcí s GBM buňkami a identifikace významných markerů v mikroprostředí nádorů, které jsou odpovědné za rezistenci na terapii a relapsu onemocnění. Současné výzkumné záměry zaměřené na odstranění reziduálních nádorových buněk pomocí gd T buněk poskytují platformu pro imunoterapii GBM.; Glioblastoma multiforme (GBM) is the common malignant brain tumour in adults and despite aggressive treatment it remains virtually incurable with a poor prognosis and median survival of 12 months following resection, radiotherapy and chemotherapy. Therapeutic approaches to GBM immunotherapy have had limited success, principally due to the protected brain barrier and the immunosuppressive tumour microenvironment. New research indicates the possibility of using immune effector cells that effectively destroy malignant GBM cells. Our previous study showed innate gamma-delta (gd) T cells were highly reactive to GBM tumour. The aim of the proposed project is cellular and molecular analyses of gd T cells in patients before and during treatment, including the detailed characterization of their interactions with GBM cells and identification of significant markers within the tumour microenvironment responsible for therapy resistance and relapse of the disease. The current research interests aiming to eliminate residual tumour cells by gd T cells provide a platform for GBM immunotherapy.
- Klíčová slova
- tumour microenvironment, nádorové mikroprostředí, glioblastoma, glioblastom, Gammadelta T buňky, Gammadelta T cells,
- NLK Publikační typ
- závěrečné zprávy o řešení grantu AZV MZ ČR
Recent studies have underscored the importance of gamma-delta (γδ) T cells in mediating potent MHC-unrestricted cytotoxicity in numerous malignancies. Here, we analyzed Vδ1 and Vδ2 γδ T cell subsets in newly diagnosed chronic myeloid leukemia (CML) patients (n = 40) who had initiated tyrosine kinase inhibitor (TKI) therapy including imatinib (n = 22), nilotinib (n = 14) and dasatinib (n = 4). Patient peripheral blood samples were analyzed at diagnosis and monitored prospectively at 3, 6, 12 and 18 months post-TKI. γδ T cells isolated from healthy donors and CML patients were used against K562, LAMA-84 and KYO-1 cell lines and against primary CML cells in cytotoxicity assays. We found large expansions of Vδ1 and Vδ2 T cells in patients at diagnosis compared to age-matched healthy donors (n = 40) (p < 0.0001). The γδ T cell reconstitution in patients on imatinib and also on nilotinib showed significant reductions of Vδ1 T cell and Vδ2 T cell absolute counts at 3 months compared to diagnosis. Importantly, Vδ1 and Vδ2 T absolute cell counts remained at normal levels from 3 months throughout the follow-up. Next, we observed susceptibility to specific lysis of primary CML tumor cells by Vδ1 T cells from healthy donors. Furthermore, we determined inherent cytotoxic reactivity by autologous patients' Vδ1 T lymphocytes against primary CML tumor cells. Finally, the TCR clonality profiles showed in CML patients mostly polyclonal repertoires regardless of the TKI. Our results provide further evidence into γδ T cell antileukemia immunity in CML that might be beneficial for long-term disease control and treatment outcome.
- MeSH
- buněčné linie MeSH
- chronická myeloidní leukemie * farmakoterapie metabolismus MeSH
- imatinib mesylát farmakologie terapeutické užití MeSH
- lidé MeSH
- myeloidní leukemie * metabolismus MeSH
- receptory antigenů T-buněk gama-delta metabolismus MeSH
- T-lymfocyty - podskupiny MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Previously, the Nine amino acid TransActivation Domain (9aaTAD) was identified in the Gal4 region 862-870 (DDVYNYLFD). Here, we identified 9aaTADs in the distal Gal4 orthologs by our prediction algorithm and found their conservation in the family. The 9aaTAD function as strong activators was demonstrated. We identified adjacent Gal4 region 871-811 (DEDTPPNPKKE) as a natural 9aaTAD inhibitory domain located at the extreme Gal4 terminus. Moreover, we identified conserved Gal4 region 172-185 (FDWSEEDDMSDGLP), which was capable to reverse the 9aaTAD inhibition. In conclusion, our results uncover the existence of the cryptic inhibitory domains, which need to be carefully implemented in all functional studies with transcription factors to avoid incorrect conclusions.
BACKGROUND: Plasmacytoid dendritic cells (pDCs) play prominent roles in mediating innate and adaptive immune responses. However, it is unclear how pDCs contribute to the immunosuppressive tumor microenvironment described in multiple myeloma (MM). METHODS: Newly diagnosed myeloma patients (MM, n = 37) were analyzed to determine the pDC counts in comparison to peripheral blood (PB, n = 53) and bone marrow (BM, n = 10) samples of age-matched healthy donors (HD) using flow cytometry. Second, proliferation of myeloma tumor cells in the presence of freshly isolated pDCs was examined. Third, production of IFNα by pDCs co-cultured with MM cells was determined by intracellular staining. RESULTS: We found a highly significant reduction of circulating pDCs (p < 0.0001) and in bone marrow (p < 0.0001) of MM patients compared to HD. We also observed a significant decrease of pDCs (p = 0.004) in BM in patients with monoclonal gammopathy of undetermined significance (MGUS, n = 12). Importantly, we determined that pDCs promote proliferation specifically of MM cells and not the stromal cells and that pDCs secrete IFNα upon co-culture with MM tumor cells. CONCLUSIONS: Our results show altered pDC frequencies in the BM microenvironment in MGUS and MM patients at diagnosis. We showed the tumor-promoting function of pDCs that may mediate immune deficiencies affecting long-term disease control and treatment outcome.
- Publikační typ
- časopisecké články MeSH
Závěrečná zpráva o řešení grantu Agentury pro zdravotnický výzkum MZ ČR
Nestr.
Cílem tohoto projektu je analýza V?1, V?2 a V?3 gamma-delta (??) populací T-lymfocytů u pacientů s chronickou lymfocytární leukémií (CLL). Lidské ?? T buňky jsou účinnými efektorovými buňkami přirozené imunity podílící se na protinádorovému imunitnímu dozoru. Náše pilotní data ukazují významnou reaktivitu ?? T buněk proti primárním CLL buňkám. Naším cílem je detailní analýza populací ?? T buněk včetně stanovení expanzí, klonality, celogenomového profilování, protinádorové cytotoxicity a interakcí s CLL buňkami. Dále bude provedena analýza unikátní kohorty dárců s monoklonální B-buněčnou lymfocytózou (MBL), která přispěje k objasnění příčin a mechanismů patogeneze CLL onemocnění. MBL je biologicky podobné s CLL a je přijímáno jako její prekurzor. Je velmi důležité, abychom dosáhli lepšího porozumění nejen o ?? cytotoxicitě proti maligním CLL buňkám, ale také o reaktivitě ?? T-buněk proti CLL i přes signalizaci od stromatu kostní dřeně. Tato analýza přinese nové výsledky důležité pro budoucí imunoterapeutické aplikace určené k odstranění residuálních CLL buněk.; This project aims to investigate the V?1, V?2 and V?3 subsets of gamma-delta (??) T cells in patients with chronic lymphocytic leukemia (CLL). Human ?? T cells are potent effector cells of innate immunity involved in anti-tumour immune surveillance. Our pilot in vitro data support the observations of prominent reactivity of ?? T cells against primary CLL cells but the detailed analysis of ?? expansions, clonality and anti-CLL reactivity are currently missing. In addition, a unique cohort of monoclonal B-cell lymphocytosis (MBL) donors will be investigated and will lead to important insights into mechanisms of CLL disease pathogenesis. MBL has been shown to display biological similarities to CLL and is accepted as its precursor state. It is crucial that we achieve a better understanding of not only ?? cytotoxicity against malignant CLL cells, but also the efficacy of ?? T cells on CLL cells despite of signaling from bone marrow stroma that will generate novel results highly relevant for future immunotherapeutic applications designed to eliminate the residual CLL cells.
- MeSH
- biologická terapie MeSH
- chronická lymfatická leukemie terapie MeSH
- imunoterapie MeSH
- intraepiteliální lymfocyty MeSH
- mezibuněčná komunikace MeSH
- protinádorové látky imunologicky aktivní MeSH
- Konspekt
- Patologie. Klinická medicína
- NLK Obory
- onkologie
- cytologie, klinická cytologie
- alergologie a imunologie
- NLK Publikační typ
- závěrečné zprávy o řešení grantu AZV MZ ČR
Broad changes in human innate and adaptive immunity are associated with advanced age. The age-related alteration of gene expression was reported for both T and B lymphocytes. We analysed the genome-wide expression profiles (n=20) of naive and whole B cell populations from young and early aged healthy donors under 60 years. We revealed large homogeneity of all analysed genome-wide expression profiles but did not identified any significant gene deregulation between young (30-45 years) and early aged healthy donors (50-60 years). We argue that B cells avoid the aging program on molecular level until 60 years of age. Our results demonstrate the potential of hematopoietic stem cells to generate uncompromised lymphocytes in early elderly. These are very encouraging findings for the general health and the immunity maintenance would not need any intervention to naive B cells. Rather, a suitable immune stimulation in healthy body environment warrants further research into aging of older elderly.
- MeSH
- adaptivní imunita genetika imunologie MeSH
- B-lymfocyty imunologie metabolismus MeSH
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- přirozená imunita genetika imunologie MeSH
- receptor interleukinu-7 - alfa-podjednotka genetika imunologie metabolismus MeSH
- stanovení celkové genové exprese metody MeSH
- stárnutí genetika imunologie MeSH
- transkriptom genetika imunologie MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
