Photodecomposition pathways for photoactivatable platinum(IV) diazido anticancer complexes

Photo-activatable platinum(IV) diazido complexes with the general formula of trans,trans,trans-[Pt(N3)2(OH)(X)(py)(am1)] (X is a hydroxide or carboxylate and am1 is an aliphatic/aromatic amine) show dark-stability under physiological conditions but can induce a photo-cytotoxic effect in cancer cells after irradiation with UVA, blue and/or green light. These platinum(IV) diazido complexes can platinate DNA and induce different lesions that are distinctly different from those generated by the anticancer drug cis-platin. Through the use of EPR, multinuclear NMR, and UV-visible absorption spectroscopy, as well as mass spectrometry and some cell studies, this thesis aims to investigate the pathways of photochemical decomposition and in particular the release of azidyl ligands and their subsequent involvement in photo-cytotoxicity. Firstly, the irradiation of trans,trans,trans-[Pt(N3)2(OH)2(py)2] (40, py = pyridine) with blue light in the presence of the spin trap 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) led to the detection of a characteristic quartet-of-triplets EPR spectrum, assigned to the azidyl radical adduct, DMPO-14N3. Irradiation of 15N-40, led to the detection of a quartet-of-doublets EPR spectrum as assigned to the DMPO-15N3 spin adduct. This confirmed that the ●N3 radicals arose from the platinum(IV)- bound azide. The DMPO-N3 spin adduct was also detected from the photoirradiation of trans,trans,trans-[Pt(N3)2(OH)2(MA)(py)] (44, MA = methylamine) with blue light. A greater yield in the DMPO-N3 spin adduct was formed in PBS/D2O. This effect was attributed to the Brownian motion of the ●N3 radicals. Interestingly, photoirradiation in the cell culture medium-, RPMI-1640 led to a reduction in the DMPO-N3 spin adduct. This reduction was accredited to the variety of components present in the cell culture medium which could behave as radical quenchers. Alternative nitrone spin traps, -4-pyridyl-1-oxide-N-tert-butylnitrone (4-POBN) and 5-diethoxyphosphoryl-5-methyl-1-pyrroline-N-oxide (DEPMPO) also led to the trapping of the azidyl radicals from irradiated 40, forming the 4-POBN-N3 (triplet-of-quartets) and DEPMPO-N3 (octet-of-triplets) spin adducts EPR spectra, respectively. DEPMPO was the most efficient azidyl radical trap; with the DEPMPO-N3 spin adduct possessing the longest lifetime in aqueous media. Extending the wavelength of activation to green light (517 nm), also led to the detection of DMPO-N3 from the photo-irradiation of 40 in RPMI-1640, trans,trans,trans-[Pt(N3)2(OH)(SAD)(py)2] (56,SAD=succinate), trans,trans,trans-[Pt(N3)2(OH)(ethyl-methyl-SAD)(py)2] (57) and trans,trans,trans-[Pt(N3)2(OH)(N-MI)(py)2] (58, N-MI = N-methylisatoate) in H2O/DMF. The DEPMPO-N3 was also detected from the gamma-ray irradiation of 40, which appears to be the first report of the activation of a platinum(IV) diazido anticancer complex with gamma-rays, a procedure which might be useful of clinical use. The azidyl radicals generated from irradiated 40 were unreactive towards both glycine and L-tyrosine. However, in the presence of L-tryptophan the azidyl radicals were quenched. Detection of free azide by 14N NMR spectroscopy confirmed that this quenching mechanism proceeded through a one-electron transfer pathway. The photo-cytotoxicity of 40 was supressed in the presence of low doses of L-tryptophan in A2780 ovarian cancer cells. From this study, it was deduced that the photo-cytotoxicity of 40 is comprised of both an acute (radical) and chronic (DNA platination) based mechanisms. Additionally, certain cancers have reported on the depleted serum levels of L-tryptophan, in particular ovarian cancer cells. This suggests the extent of the photo-cytotoxicity of 40 can be controlled in the presence of L-tryptophan. Photo-irradiation of 40 in the presence of melatonin, an analogue of L-tryptophan was also performed. Despite, the structural similarity between L-tryptophan and melatonin, the presence of the 5-methoxy substituent present in melatonin induced an alternative photo-decomposition pathway of 40. Photo-irradiation of 40 in the presence of melatonin with blue light also led to the detection of the quartet EPR spectrum assigned to the hydroxyl radical adduct, DMPO-OH. Through multinuclear NMR and mass spectrometry the quenching of both the azidyl and hydroxyl radicals by melatonin was determined. Additionally, mass spectrometry also detected a mass adduct attributed to a platinum(II)-melatonin complex. This dual antioxidant and metal-binding ability of melatonin was attributed to the observed photo-protective effect in A2780 ovarian cancer cells. Melatonin regulates circadian rhythms with a maximum concentration during dark hours. Consequently, treatment of antineoplastic tissue with 40 during the hours of melatonin production may be ineffective. Platinum accumulation and absorption has been suggested to be mediated by organic cation transporters (OCTs), in particular OCT2. Irradiation of 40 was monitored in the presence of cimetidine, an OCT2 inhibitor. A new strong absorption band at ca. 354 nm was assigned to an SPtII LMCT band. High resolution mass spectroscopy identified a mass adduct assigned to a novel platinum(II)-cimetidine species. The loss of coordinated pyridine from irradiated 40 as detected by 1H NMR spectroscopy and the quenching of the azidyl radical by cimetidine were correlated with the observed photo-protective effect in HaCaT keratinocytes cells

Evolution of high pathogenicity of H5 avian influenza virus: haemagglutinin cleavage site selection of reverse-genetics mutants during passage in chickens

Low pathogenicity avian influenza viruses (LPAIVs) are generally asymptomatic in their natural avian hosts. LPAIVs can evolve into highly pathogenic forms, which can affect avian and human populations with devastating consequences. The switch to highly pathogenic avian influenza virus (HPAIV) from LPAIV precursors requires the acquisition of multiple basic amino acids in the haemagglutinin cleavage site (HACS) motif. Through reverse genetics of an H5N1 HPAIV, and experimental infection of chickens, we determined that viruses containing five or more basic amino acids in the HACS motif were preferentially selected over those with three to four basic amino acids, leading to rapid replacement with virus types containing extended HACS motifs. Conversely, viruses harbouring low pathogenicity motifs containing two basic amino acids did not readily evolve to extended forms, suggesting that a single insertion of a basic amino acid into the cleavage site motif of low-pathogenic viruses may lead to escalating selection for extended motifs. Our results may explain why mid-length forms are rarely detected in nature. The stability of the short motif suggests that pathogenicity switching may require specific conditions of intense selection pressure (such as with high host density) to boost selection of the initial mid-length HACS forms

Effects of antiplatelet therapy on stroke risk by brain imaging features of intracerebral haemorrhage and cerebral small vessel diseases: subgroup analyses of the RESTART randomised, open-label trial

Background Findings from the RESTART trial suggest that starting antiplatelet therapy might reduce the risk of recurrent symptomatic intracerebral haemorrhage compared with avoiding antiplatelet therapy. Brain imaging features of intracerebral haemorrhage and cerebral small vessel diseases (such as cerebral microbleeds) are associated with greater risks of recurrent intracerebral haemorrhage. We did subgroup analyses of the RESTART trial to explore whether these brain imaging features modify the effects of antiplatelet therapy

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

Large expert-curated database for benchmarking document similarity detection in biomedical literature search

Document recommendation systems for locating relevant literature have mostly relied on methods developed a decade ago. This is largely due to the lack of a large offline gold-standard benchmark of relevant documents that cover a variety of research fields such that newly developed literature search techniques can be compared, improved and translated into practice. To overcome this bottleneck, we have established the RElevant LIterature SearcH consortium consisting of more than 1500 scientists from 84 countries, who have collectively annotated the relevance of over 180 000 PubMed-listed articles with regard to their respective seed (input) article/s. The majority of annotations were contributed by highly experienced, original authors of the seed articles. The collected data cover 76% of all unique PubMed Medical Subject Headings descriptors. No systematic biases were observed across different experience levels, research fields or time spent on annotations. More importantly, annotations of the same document pairs contributed by different scientists were highly concordant. We further show that the three representative baseline methods used to generate recommended articles for evaluation (Okapi Best Matching 25, Term Frequency-Inverse Document Frequency and PubMed Related Articles) had similar overall performances. Additionally, we found that these methods each tend to produce distinct collections of recommended articles, suggesting that a hybrid method may be required to completely capture all relevant articles. The established database server located at https://relishdb.ict.griffith.edu.au is freely available for the downloading of annotation data and the blind testing of new methods. We expect that this benchmark will be useful for stimulating the development of new powerful techniques for title and title/abstract-based search engines for relevant articles in biomedical research.Peer reviewe

Search for new particles in events with energetic jets and large missing transverse momentum in proton-proton collisions at root s=13 TeV

A search is presented for new particles produced at the LHC in proton-proton collisions at root s = 13 TeV, using events with energetic jets and large missing transverse momentum. The analysis is based on a data sample corresponding to an integrated luminosity of 101 fb(-1), collected in 2017-2018 with the CMS detector. Machine learning techniques are used to define separate categories for events with narrow jets from initial-state radiation and events with large-radius jets consistent with a hadronic decay of a W or Z boson. A statistical combination is made with an earlier search based on a data sample of 36 fb(-1), collected in 2016. No significant excess of events is observed with respect to the standard model background expectation determined from control samples in data. The results are interpreted in terms of limits on the branching fraction of an invisible decay of the Higgs boson, as well as constraints on simplified models of dark matter, on first-generation scalar leptoquarks decaying to quarks and neutrinos, and on models with large extra dimensions. Several of the new limits, specifically for spin-1 dark matter mediators, pseudoscalar mediators, colored mediators, and leptoquarks, are the most restrictive to date.Peer reviewe

Combined searches for the production of supersymmetric top quark partners in proton-proton collisions at root s=13 TeV

A combination of searches for top squark pair production using proton-proton collision data at a center-of-mass energy of 13 TeV at the CERN LHC, corresponding to an integrated luminosity of 137 fb(-1) collected by the CMS experiment, is presented. Signatures with at least 2 jets and large missing transverse momentum are categorized into events with 0, 1, or 2 leptons. New results for regions of parameter space where the kinematical properties of top squark pair production and top quark pair production are very similar are presented. Depending on themodel, the combined result excludes a top squarkmass up to 1325 GeV for amassless neutralino, and a neutralinomass up to 700 GeV for a top squarkmass of 1150 GeV. Top squarks with masses from 145 to 295 GeV, for neutralino masses from 0 to 100 GeV, with a mass difference between the top squark and the neutralino in a window of 30 GeV around the mass of the top quark, are excluded for the first time with CMS data. The results of theses searches are also interpreted in an alternative signal model of dark matter production via a spin-0 mediator in association with a top quark pair. Upper limits are set on the cross section for mediator particle masses of up to 420 GeV

Measurements of Higgs boson production cross sections and couplings in the diphoton decay channel at root s=13 TeV

Measurements of Higgs boson production cross sections and couplings in events where the Higgs boson decays into a pair of photons are reported. Events are selected from a sample of proton-proton collisions at root s = 13TeV collected by the CMS detector at the LHC from 2016 to 2018, corresponding to an integrated luminosity of 137 fb(-1). Analysis categories enriched in Higgs boson events produced via gluon fusion, vector boson fusion, vector boson associated production, and production associated with top quarks are constructed. The total Higgs boson signal strength, relative to the standard model (SM) prediction, is measured to be 1.12 +/- 0.09. Other properties of the Higgs boson are measured, including SM signal strength modifiers, production cross sections, and its couplings to other particles. These include the most precise measurements of gluon fusion and vector boson fusion Higgs boson production in several different kinematic regions, the first measurement of Higgs boson production in association with a top quark pair in five regions of the Higgs boson transverse momentum, and an upper limit on the rate of Higgs boson production in association with a single top quark. All results are found to be in agreement with the SM expectations.Peer reviewe

Probing effective field theory operators in the associated production of top quarks with a Z boson in multilepton final states at root s=13 TeV

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Search for a heavy Higgs boson decaying into two lighter Higgs bosons in the tau tau bb final state at 13 TeV

A search for a heavy Higgs boson H decaying into the observed Higgs boson h with a mass of 125 GeV and another Higgs boson h(S) is presented. The h and h(S) bosons are required to decay into a pair of tau leptons and a pair of b quarks, respectively. The search uses a sample of proton-proton collisions collected with the CMS detector at a center-of-mass energy of 13TeV, corresponding to an integrated luminosity of 137 fb(-1). Mass ranges of 240-3000 GeV for m(H) and 60-2800 GeV for m(hS) are explored in the search. No signal has been observed. Model independent 95% confidence level upper limits on the product of the production cross section and the branching fractions of the signal process are set with a sensitivity ranging from 125 fb (for m(H) = 240 GeV) to 2.7 fb (for m(H) = 1000 GeV). These limits are compared to maximally allowed products of the production cross section and the branching fractions of the signal process in the next-to-minimal supersymmetric extension of the standard model.Peer reviewe