Click chemistry enables comprehensive preclinical evaluation of targeted epigenetic therapies
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE88751 | GEO | 2017/07/13
SECONDARY ACCESSION(S): PRJNA348447
REPOSITORIES: GEO
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Project description:The success of targeted therapies hinges on our ability to understand the molecular and cellular mechanism of action of these agents. Here we modify various BET bromodomain inhibitors, an exemplar novel targeted therapy, to create functionally conserved compounds that are amenable to click-chemistry and can be used as molecular probes in vitro and in vivo. Using click-proteomics and click-sequencing we provide new mechanistic insights to explain the gene regulatory function of BRD4 and the transcriptional changes invoked by BET inhibitors. In mouse models of acute leukaemia, we use high resolution microscopy and flow cytometry to highlight the underappreciated heterogeneity of drug activity within tumour cells located in different tissue compartments. We also demonstrate the differential distribution and effects of the drug in normal and malignant cells in vivo. These data provide critical insights that reveal the cellular and molecular details for the efficacy and limitations of these agents. This study provides a framework for the pre-clinical assessment of other conventional and targeted therapies.
2017-07-13 | GSE88749 | GEO
Project description:The success of targeted therapies hinges on our ability to understand the molecular and cellular mechanism of action of these agents. Here we modify various BET bromodomain inhibitors, an exemplar novel targeted therapy, to create functionally conserved compounds that are amenable to click-chemistry and can be used as molecular probes in vitro and in vivo. Using click-proteomics and click-sequencing we provide new mechanistic insights to explain the gene regulatory function of BRD4 and the transcriptional changes invoked by BET inhibitors. In mouse models of acute leukaemia, we use high resolution microscopy and flow cytometry to highlight the underappreciated heterogeneity of drug activity within tumour cells located in different tissue compartments. We also demonstrate the differential distribution and effects of the drug in normal and malignant cells in vivo. These data provide critical insights that reveal the cellular and molecular details for the efficacy and limitations of these agents. This study provides a framework for the pre-clinical assessment of other conventional and targeted therapies.
2017-07-13 | GSE88748 | GEO
Project description:The success of targeted therapies hinges on our ability to understand the molecular and cellular mechanism of action of these agents. Here we modify various BET bromodomain inhibitors, an exemplar novel targeted therapy, to create functionally conserved compounds that are amenable to click-chemistry and can be used as molecular probes in vitro and in vivo. Using click-proteomics and click-sequencing we provide new mechanistic insights to explain the gene regulatory function of BRD4 and the transcriptional changes invoked by BET inhibitors. In mouse models of acute leukaemia, we use high resolution microscopy and flow cytometry to highlight the underappreciated heterogeneity of drug activity within tumour cells located in different tissue compartments. We also demonstrate the differential distribution and effects of the drug in normal and malignant cells in vivo. These data provide critical insights that reveal the cellular and molecular details for the efficacy and limitations of these agents. This study provides a framework for the pre-clinical assessment of other conventional and targeted therapies.
2017-07-13 | GSE88750 | GEO
Project description:The success of targeted therapies hinges on our ability to understand the molecular and cellular mechanism of action of these agents. Here we modify various BET bromodomain inhibitors, an exemplar novel targeted therapy, to create functionally conserved compounds that are amenable to click-chemistry and can be used as molecular probes in vitro and in vivo. Using click-proteomics and click-sequencing we provide new mechanistic insights to explain the gene regulatory function of BRD4 and the transcriptional changes invoked by BET inhibitors. In mouse models of acute leukaemia, we use high resolution microscopy and flow cytometry to highlight the underappreciated heterogeneity of drug activity within tumour cells located in different tissue compartments. We also demonstrate the differential distribution and effects of the drug in normal and malignant cells in vivo. These data provide critical insights that reveal the cellular and molecular details for the efficacy and limitations of these agents. This study provides a framework for the pre-clinical assessment of other conventional and targeted therapies.
2017-07-13 | GSE88747 | GEO
Project description:The success of targeted therapies hinges on our ability to understand the molecular and cellular mechanism of action of these agents. Here we modify various BET bromodomain inhibitors, an exemplar novel targeted therapy, to create functionally conserved compounds that are amenable to click-chemistry and can be used as molecular probes in vitro and in vivo. Using click-proteomics and click-sequencing we provide new mechanistic insights to explain the gene regulatory function of BRD4 and the transcriptional changes invoked by BET inhibitors. In mouse models of acute leukaemia, we use high resolution microscopy and flow cytometry to highlight the underappreciated heterogeneity of drug activity within tumour cells located in different tissue compartments. We also demonstrate the differential distribution and effects of the drug in normal and malignant cells in vivo. These data provide critical insights that reveal the cellular and molecular details for the efficacy and limitations of these agents. This study provides a framework for the pre-clinical assessment of other conventional and targeted therapies.
2017-07-13 | GSE88746 | GEO
Project description:The success of new therapies hinges on our ability to understand their molecular and cellular mechanisms of action. We modified BET bromodomain inhibitors, an epigenetic-based therapy, to create functionally conserved compounds that are amenable to click chemistry and can be used as molecular probes in vitro and in vivo. We used click proteomics and click sequencing to explore the gene regulatory function of BRD4 (bromodomain containing protein 4) and the transcriptional changes induced by BET inhibitors. In our studies of mouse models of acute leukemia, we used high-resolution microscopy and flow cytometry to highlight the heterogeneity of drug activity within tumor cells located in different tissue compartments. We also demonstrate the differential distribution and effects of BET inhibitors in normal and malignant cells in vivo. This study provides a potential framework for the preclinical assessment of a wide range of drugs.
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Project description:Click chemistry enables comprehensive preclinical evaluation of targeted epigenetic therapies
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Project description:The ability to observe cells in live organisms is essential for understanding their function in complex in vivo milieus. A major challenge today has been the limited ability to perform higher multiplexing beyond four to six colors to define cell subtypes in vivo. Here, a click chemistry-based strategy is presented for higher multiplexed in vivo imaging in mouse models. The method uses a scission-accelerated fluorophore exchange (SAFE), which exploits a highly efficient bioorthogonal mechanism to completely remove fluorescent signal from antibody-labeled cells in vivo. It is shown that the SAFE-intravital microscopy imaging method allows 1) in vivo staining of specific cell types in dorsal and cranial window chambers of mice, 2) complete un-staining in minutes, 3) in vivo click chemistries at lower (µm) and thus non-toxic concentrations, and 4) the ability to perform in vivo cyclic imaging. The potential utility of the method is demonstrated by 12 color imaging of immune cells in live mice.
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Project description:Biphenyl-4,4'-dicarboxylic acid derivatives containing either azide or acetylene functional groups were inserted into UiO-67 metal organic frameworks (MOFs) via post synthetic linker exchange. Sequential and orthogonal click reactions could be performed on these modified MOFs by incubating the crystals with small molecule substrates bearing azide or acetylene groups in the presence of a copper catalyst. 1H NMR of digested MOF samples showed that up to 50% of the incorporated linkers could be converted to their "clicked" triazole products. Powder X-ray diffraction confirmed that the UiO-67 structure was maintained throughout all transformations. The click reaction efficiency is discussed in context of MOF crystallite size and pore size. As the incorporation of clicked linkers could be controlled by post synthetic exchange, this work introduces a powerful method of quickly introducing orthogonal modifications into known MOF architectures.
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Project description:The level of cancer biomarkers in cells, tissues or body fluids can be used for the prediction of the presence of cancer or can even indicate the stage of the disease. Alpha-fetoprotein (AFP) is the most commonly used biomarker for early screening and diagnosis of hepatocellular carcinoma (HCC). Here, a combination of three techniques (click chemistry, the biotin-streptavidin-biotin sandwich strategy and the use of antigen-antibody interactions) were combined to implement a sensitive fluorescent immunosensor for AFP detection. Three types of functionalized glasses (dibenzocyclooctyne- (DBCO-), thiol- and epoxy-terminated surfaces) were biotinylated by employing the respective adequate click chemistry counterparts (biotin-thiol or biotin-azide for the first class, biotin-maleimide or biotin-DBCO for the second class and biotin-amine or biotin-thiol for the third class). The anti-AFP antibody was immobilized on the surfaces via a biotin-streptavidin-biotin sandwich technique. To evaluate the sensing performance of the differently prepared surfaces, fluorescently labeled AFP was spotted onto them via microchannel cantilever spotting (µCS). Based on the fluorescence measurements, the optimal microarray design was found and its sensitivity was determined.
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