85 research outputs found
Dual LSD1 and HDAC6 Inhibition Induces Doxorubicin Sensitivity in Acute Myeloid Leukemia Cells
Defects in epigenetic pathways are key drivers of oncogenic cell proliferation. We developed a LSD1/HDAC6 multitargeting inhibitor (iDual), a hydroxamic acid analogue of the clinical candidate LSD1 inhibitor GSK2879552. iDual inhibits both targets with IC50 values of 540, 110, and 290 nM, respectively, against LSD1, HDAC6, and HDAC8. We compared its activity to structurally similar control probes that act by HDAC or LSD1 inhibition alone, as well as an inactive null compound. iDual inhibited the growth of leukemia cell lines at a higher level than GSK2879552 with micromolar IC50 values. Dual engagement with LSD1 and HDAC6 was supported by dose dependent increases in substrate levels, biomarkers, and cellular thermal shift assay. Both histone methylation and acetylation of tubulin were increased, while acetylated histone levels were only mildly affected, indicating selectivity for HDAC6. Downstream gene expression (CD11b, CD86, p21) was also elevated in response to iDual treatment. Remarkably, iDual synergized with doxorubicin, triggering significant levels of apoptosis with a sublethal concentration of the drug. While mechanistic studies did not reveal changes in DNA repair or drug efflux pathways, the expression of AGPAT9, ALOX5, BTG1, HIPK2, IFI44L, and LRP1, previously implicated in doxorubicin sensitivity, was significantly elevated
Focused screening identifies different sensitivities of human TET oxygenases to the oncometabolite 2-hydroxyglutarate
Ten-eleven translocation enzymes (TETs) are Fe(II)/2-oxoglutarate (2OG) oxygenases that catalyze the sequential oxidation of 5-methylcytosine to 5-hydroxymethylcytosine, 5-formylcytosine, and 5-carboxylcytosine in eukaryotic DNA. Despite their roles in epigenetic regulation, there is a lack of reported TET inhibitors. The extent to which 2OG oxygenase inhibitors, including clinically used inhibitors and oncometabolites, modulate DNA modifications via TETs has been unclear. Here, we report studies on human TET1–3 inhibition by a set of 2OG oxygenase-focused inhibitors, employing both enzyme-based and cellular assays. Most inhibitors manifested similar potencies for TET1–3 and caused increases in cellular 5hmC levels. (R)-2-Hydroxyglutarate, an oncometabolite elevated in isocitrate dehydrogenase mutant cancer cells, showed different degrees of inhibition, with TET1 being less potently inhibited than TET3 and TET2, potentially reflecting the proposed role of TET2 mutations in tumorigenesis. The results highlight the tractability of TETs as drug targets and provide starting points for selective inhibitor design
Vertical Field Effect Transistor based on Graphene-WS2 Heterostructures for flexible and transparent electronics
The celebrated electronic properties of graphene have opened way for
materials just one-atom-thick to be used in the post-silicon electronic era. An
important milestone was the creation of heterostructures based on graphene and
other two-dimensional (2D) crystals, which can be assembled in 3D stacks with
atomic layer precision. These layered structures have already led to a range of
fascinating physical phenomena, and also have been used in demonstrating a
prototype field effect tunnelling transistor - a candidate for post-CMOS
technology. The range of possible materials which could be incorporated into
such stacks is very large. Indeed, there are many other materials where layers
are linked by weak van der Waals forces, which can be exfoliated and combined
together to create novel highly-tailored heterostructures. Here we describe a
new generation of field effect vertical tunnelling transistors where 2D
tungsten disulphide serves as an atomically thin barrier between two layers of
either mechanically exfoliated or CVD-grown graphene. Our devices have
unprecedented current modulation exceeding one million at room temperature and
can also operate on transparent and flexible substrates
Atomically thin boron nitride: a tunnelling barrier for graphene devices
We investigate the electronic properties of heterostructures based on
ultrathin hexagonal boron nitride (h-BN) crystalline layers sandwiched between
two layers of graphene as well as other conducting materials (graphite, gold).
The tunnel conductance depends exponentially on the number of h-BN atomic
layers, down to a monolayer thickness. Exponential behaviour of I-V
characteristics for graphene/BN/graphene and graphite/BN/graphite devices is
determined mainly by the changes in the density of states with bias voltage in
the electrodes. Conductive atomic force microscopy scans across h-BN terraces
of different thickness reveal a high level of uniformity in the tunnel current.
Our results demonstrate that atomically thin h-BN acts as a defect-free
dielectric with a high breakdown field; it offers great potential for
applications in tunnel devices and in field-effect transistors with a high
carrier density in the conducting channel.Comment: 7 pages, 5 figure
ECMO for COVID-19 patients in Europe and Israel
Since March 15th, 2020, 177 centres from Europe and Israel have joined the study, routinely reporting on the ECMO support they provide to COVID-19 patients. The mean annual number of cases treated with ECMO in the participating centres before the pandemic (2019) was 55. The number of COVID-19 patients has increased rapidly each week reaching 1531 treated patients as of September 14th. The greatest number of cases has been reported from France (n = 385), UK (n = 193), Germany (n = 176), Spain (n = 166), and Italy (n = 136) .The mean age of treated patients was 52.6 years (range 16–80), 79% were male. The ECMO configuration used was VV in 91% of cases, VA in 5% and other in 4%. The mean PaO2 before ECMO implantation was 65 mmHg. The mean duration of ECMO support thus far has been 18 days and the mean ICU length of stay of these patients was 33 days. As of the 14th September, overall 841 patients have been weaned from ECMO
support, 601 died during ECMO support, 71 died after withdrawal of ECMO, 79 are still receiving ECMO support and for 10 patients status n.a. . Our preliminary data suggest that patients placed
on ECMO with severe refractory respiratory or cardiac failure secondary to COVID-19 have a reasonable (55%) chance of survival. Further extensive data analysis is expected to provide invaluable information on the demographics, severity of illness, indications and different ECMO management strategies in these patients
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Effect of Hydrocortisone on Mortality and Organ Support in Patients With Severe COVID-19: The REMAP-CAP COVID-19 Corticosteroid Domain Randomized Clinical Trial.
Importance: Evidence regarding corticosteroid use for severe coronavirus disease 2019 (COVID-19) is limited. Objective: To determine whether hydrocortisone improves outcome for patients with severe COVID-19. Design, Setting, and Participants: An ongoing adaptive platform trial testing multiple interventions within multiple therapeutic domains, for example, antiviral agents, corticosteroids, or immunoglobulin. Between March 9 and June 17, 2020, 614 adult patients with suspected or confirmed COVID-19 were enrolled and randomized within at least 1 domain following admission to an intensive care unit (ICU) for respiratory or cardiovascular organ support at 121 sites in 8 countries. Of these, 403 were randomized to open-label interventions within the corticosteroid domain. The domain was halted after results from another trial were released. Follow-up ended August 12, 2020. Interventions: The corticosteroid domain randomized participants to a fixed 7-day course of intravenous hydrocortisone (50 mg or 100 mg every 6 hours) (n = 143), a shock-dependent course (50 mg every 6 hours when shock was clinically evident) (n = 152), or no hydrocortisone (n = 108). Main Outcomes and Measures: The primary end point was organ support-free days (days alive and free of ICU-based respiratory or cardiovascular support) within 21 days, where patients who died were assigned -1 day. The primary analysis was a bayesian cumulative logistic model that included all patients enrolled with severe COVID-19, adjusting for age, sex, site, region, time, assignment to interventions within other domains, and domain and intervention eligibility. Superiority was defined as the posterior probability of an odds ratio greater than 1 (threshold for trial conclusion of superiority >99%). Results: After excluding 19 participants who withdrew consent, there were 384 patients (mean age, 60 years; 29% female) randomized to the fixed-dose (n = 137), shock-dependent (n = 146), and no (n = 101) hydrocortisone groups; 379 (99%) completed the study and were included in the analysis. The mean age for the 3 groups ranged between 59.5 and 60.4 years; most patients were male (range, 70.6%-71.5%); mean body mass index ranged between 29.7 and 30.9; and patients receiving mechanical ventilation ranged between 50.0% and 63.5%. For the fixed-dose, shock-dependent, and no hydrocortisone groups, respectively, the median organ support-free days were 0 (IQR, -1 to 15), 0 (IQR, -1 to 13), and 0 (-1 to 11) days (composed of 30%, 26%, and 33% mortality rates and 11.5, 9.5, and 6 median organ support-free days among survivors). The median adjusted odds ratio and bayesian probability of superiority were 1.43 (95% credible interval, 0.91-2.27) and 93% for fixed-dose hydrocortisone, respectively, and were 1.22 (95% credible interval, 0.76-1.94) and 80% for shock-dependent hydrocortisone compared with no hydrocortisone. Serious adverse events were reported in 4 (3%), 5 (3%), and 1 (1%) patients in the fixed-dose, shock-dependent, and no hydrocortisone groups, respectively. Conclusions and Relevance: Among patients with severe COVID-19, treatment with a 7-day fixed-dose course of hydrocortisone or shock-dependent dosing of hydrocortisone, compared with no hydrocortisone, resulted in 93% and 80% probabilities of superiority with regard to the odds of improvement in organ support-free days within 21 days. However, the trial was stopped early and no treatment strategy met prespecified criteria for statistical superiority, precluding definitive conclusions. Trial Registration: ClinicalTrials.gov Identifier: NCT02735707
Effect of angiotensin-converting enzyme inhibitor and angiotensin receptor blocker initiation on organ support-free days in patients hospitalized with COVID-19
IMPORTANCE Overactivation of the renin-angiotensin system (RAS) may contribute to poor clinical outcomes in patients with COVID-19.
Objective To determine whether angiotensin-converting enzyme (ACE) inhibitor or angiotensin receptor blocker (ARB) initiation improves outcomes in patients hospitalized for COVID-19.
DESIGN, SETTING, AND PARTICIPANTS In an ongoing, adaptive platform randomized clinical trial, 721 critically ill and 58 non–critically ill hospitalized adults were randomized to receive an RAS inhibitor or control between March 16, 2021, and February 25, 2022, at 69 sites in 7 countries (final follow-up on June 1, 2022).
INTERVENTIONS Patients were randomized to receive open-label initiation of an ACE inhibitor (n = 257), ARB (n = 248), ARB in combination with DMX-200 (a chemokine receptor-2 inhibitor; n = 10), or no RAS inhibitor (control; n = 264) for up to 10 days.
MAIN OUTCOMES AND MEASURES The primary outcome was organ support–free days, a composite of hospital survival and days alive without cardiovascular or respiratory organ support through 21 days. The primary analysis was a bayesian cumulative logistic model. Odds ratios (ORs) greater than 1 represent improved outcomes.
RESULTS On February 25, 2022, enrollment was discontinued due to safety concerns. Among 679 critically ill patients with available primary outcome data, the median age was 56 years and 239 participants (35.2%) were women. Median (IQR) organ support–free days among critically ill patients was 10 (–1 to 16) in the ACE inhibitor group (n = 231), 8 (–1 to 17) in the ARB group (n = 217), and 12 (0 to 17) in the control group (n = 231) (median adjusted odds ratios of 0.77 [95% bayesian credible interval, 0.58-1.06] for improvement for ACE inhibitor and 0.76 [95% credible interval, 0.56-1.05] for ARB compared with control). The posterior probabilities that ACE inhibitors and ARBs worsened organ support–free days compared with control were 94.9% and 95.4%, respectively. Hospital survival occurred in 166 of 231 critically ill participants (71.9%) in the ACE inhibitor group, 152 of 217 (70.0%) in the ARB group, and 182 of 231 (78.8%) in the control group (posterior probabilities that ACE inhibitor and ARB worsened hospital survival compared with control were 95.3% and 98.1%, respectively).
CONCLUSIONS AND RELEVANCE In this trial, among critically ill adults with COVID-19, initiation of an ACE inhibitor or ARB did not improve, and likely worsened, clinical outcomes.
TRIAL REGISTRATION ClinicalTrials.gov Identifier: NCT0273570
Histone lysine and DNA methylation: dynamic marks in the chromatin
Ten-eleven translocation (TET) enzymes are epigenetic enzymes that are responsible for the iterative oxidation of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxycytosine (5caC) on genomic DNA using a catalytic 2-oxoglutarate (2OG) dependent oxygenase domain. These epigenetic marks are associated with transcriptional activation and silencing depending on the modification and context. Furthermore, 5fC and 5caC participate in active demethylation pathways to revert back to cytosine (C) creating a dynamic cycle. TET enzymes play an essential role in shaping the epigenome and loss of TET2’s function has frequently been described in haematopoietic malignancies with notable examples being myelodysplastic syndrome (MDS) and acute myeloid leukaemia (AML). To investigate the functional role of TET enzymes in a cellular context in greater detail, chemical probes can aid in understanding the effect of TET inhibition and can elicit a biological response. Here, two robust orthogonal high-throughput screening technologies were developed for kinetic analysis of TET enzymes and inhibitor profiling. Examination of a focused small-molecule library revealed differences in potency amongst TET isoforms and 2OG-dependent oxygenases. These results provide the initial scaffolds as a starting point for further inhibitor development.
A myriad of epigenetic modifications is found on histones in nucleosome complexes. A common post translational modification is Nε-lysine methylation which is linked to transcriptional silencing or activation depending on the location within the nucleosome. The methylation levels are under dynamic control of histone lysine methyltransferases (KMTs) and histone lysine demethylases (KDMs) while readout is performed by chromatin binding modules. In order to study the biological role of lysine methylation and participation in orchestrating the epigenetic landscape, chemical probes can be used. However, the relative high number of structurally related proteins and corresponding domains found in humans consisting of KMTs (SET: ~25), KDMs (JumonjiC domain: ~25) and chromatin binding modules (tudor domain (TD): ~25; plant homeodomain (PHD): >100) makes it challenging to produce both potent and specific inhibitors. The development of inhibitors for Nε-trimethyllysine chromatin binding modules to date, have been plagued with relatively weak inhibition but also with off-target effects. To aid in the development of inhibitors, epigenetic tools were developed, and their effectiveness was explored against a panel of chromatin binding modules. The tools were then applied in probing the PHD of multiple KDMs including KDM5s (JARID1s) and KDM7B (PHF8). The collective results show that these new tools can be utilized for the development of potent and selective inhibitors of chromatin binding modules that may in the future be used for targeted therapies
Expanding the Chemical Repertoire of DNA Nanomaterials Generated by Rolling Circle Amplification
Rolling circle amplification (RCA) is a powerful tool for the construction of DNA nanomaterials such as hydrogels, high-performance scaffolds and DNA nanoflowers (DNFs), hybrid materials formed of DNA and magnesium pyrophosphate. Such DNA nanomaterials have great potential in therapeutics, imaging, protein immobilisation, and drug delivery, yet limited chemistry is available to expand their functionality. Here, we present an orthogonal strategy to produce densely modified RCA products and DNFs. We show that it is possible to selectively functionalise the DNA component of these materials, their protein cargo, or both, thereby greatly expanding the chemical repertoire available to these systems. We then use this methodology to construct DNFs bearing multiple surface aptamers capable of binding to cancer cells that overexpress the HER2 oncobiomarker, demonstrating the therapeutic and diagnostic potential of this chemistry.</p
Expanding the chemical repertoire of DNA nanomaterials generated by rolling circle amplification.
Rolling circle amplification (RCA) is a powerful tool for the construction of DNA nanomaterials such as hydrogels, high-performance scaffolds and DNA nanoflowers (DNFs), hybrid materials formed of DNA and magnesium pyrophosphate. Such DNA nanomaterials have great potential in therapeutics, imaging, protein immobilisation, and drug delivery, yet limited chemistry is available to expand their functionality. Here, we present orthogonal strategies to produce densely modified RCA products and DNFs. We provide methods to selectively modify the DNA component and/or the protein cargo of these materials, thereby greatly expanding the range of chemical functionalities available to these systems. We have used our methodology to construct DNFs bearing multiple surface aptamers and peptides capable of binding to cancer cells that overexpress the HER2 oncobiomarker, demonstrating their potential for diagnostic and therapeutic applications
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