Can you detect early dementia from an email? A proof of principle study of daily computer use to detect cognitive and functional decline.

OBJECTIVE: To determine whether multiple computer use behaviours can distinguish between cognitively healthy older adults and those in the early stages of cognitive decline, and to investigate whether these behaviours are associated with cognitive and functional ability. METHODS: Older adults with cognitive impairment (n = 20) and healthy controls (n = 24) completed assessments of cognitive and functional abilities and a series of semi-directed computer tasks. Computer use behaviours were captured passively using bespoke software. RESULTS: The profile of computer use behaviours was significantly different in cognitively impaired compared with cognitively healthy control participants including more frequent pauses, slower typing, and a higher proportion of mouse clicks. These behaviours were significantly associated with performance on cognitive and functional assessments, in particular, those related to memory. CONCLUSION: Unobtrusively capturing computer use behaviours offers the potential for early detection of neurodegeneration in non-clinical settings, which could enable timely interventions to ultimately improve long-term outcomes.

ICORG 10-14: NEOadjuvant trial in Adenocarcinoma of the oEsophagus and oesophagoGastric junction International Study (Neo-AEGIS).

BACKGROUND: Neoadjuvant therapy is increasingly the standard of care in the management of locally advanced adenocarcinoma of the oesophagus and junction (AEG). In randomised controlled trials (RCTs), the MAGIC regimen of pre- and postoperative chemotherapy, and the CROSS regimen of preoperative chemotherapy combined with radiation, were superior to surgery only in RCTs that included AEG but were not powered on this cohort. No completed RCT has directly compared neoadjuvant or perioperative chemotherapy and neoadjuvant chemoradiation. The Neo-AEGIS trial, uniquely powered on AEG, and including comprehensive modern staging, compares both these regimens. METHODS: This open label, multicentre, phase III RCT randomises patients (cT2-3, N0-3, M0) in a 1:1 fashion to receive CROSS protocol (Carboplatin and Paclitaxel with concurrent radiotherapy, 41.4Gy/23Fr, over 5 weeks). The power calculation is a 10% difference in favour of CROSS, powered at 80%, two-sided alpha level of 0.05, requiring 540 patients to be evaluable, 594 to be recruited if a 10% dropout is included (297 in each group). The primary endpoint is overall survival, with a minimum 3-year follow up. Secondary endpoints include: disease free survival, recurrence rates, clinical and pathological response rates, toxicities of induction regimens, post-operative pathology and tumour regression grade, operative in-hospital complications, and health-related quality of life. The trial also affords opportunities for establishing a bio-resource of pre-treatment and resected tumour, and translational research. DISCUSSION: This RCT directly compares two established treatment regimens, and addresses whether radiation therapy positively impacts on overall survival compared with a standard perioperative chemotherapy regimen Sponsor: Irish Clinical Research Group (ICORG). TRIAL REGISTRATION: NCT01726452 . Protocol 10-14. Date of registration 06/11/2012.

Australian consensus guidelines for the safe handling of monoclonal antibodies for cancer treatment by healthcare personnel.

These consensus guidelines provide recommendations for the safe handling of monoclonal antibodies. Definitive recommendations are given for the minimum safe handling requirements to protect healthcare personnel. The seven recommendations cover: (i) appropriate determinants for evaluating occupational exposure risk; (ii) occupational risk level compared with other hazardous and non-hazardous drugs; (iii) stratification of risk based on healthcare personnel factors; (iv) waste products; (v) interventions and safeguards; (vi) operational and clinical factors and (vii) handling recommendations. The seventh recommendation includes a risk assessment model and flow chart for institutions to consider and evaluate clinical and operational factors unique to individual healthcare services. These guidelines specifically evaluated monoclonal antibodies used in the Australian cancer clinical practice setting; however, the principles may be applicable to monoclonal antibodies used in non-cancer settings. The guidelines are only applicable to parenterally administered agents.

Health-related quality of life across disease stages in patients with amyotrophic lateral sclerosis: results from a real-world survey.

BACKGROUND: Amyotrophic lateral sclerosis (ALS) is characterized by a rapid disease course, with disease severity being associated with declining health-related quality of life (HRQoL) in persons living with ALS (pALS). The main objective of this study was to assess the impact of disease progression on HRQoL across King's, Milano-Torino Staging (MiToS), and physician-judgement clinical staging. Additionally, we evaluated the impact of the disease on the HRQoL of care partners (cALS). METHODS: Data were sourced from the Adelphi ALS Disease Specific Programme (DSP)™, a cross-sectional survey of neurologists, pALS and cALS presenting in a real-world clinical setting between July 2020 and March 2021 in Europe and the United States. RESULTS: Neurologists (n = 142) provided data for 880 pALS. There were significant negative correlations between all three clinical staging systems and EuroQol (European Quality of Life) Five Dimension Five Level Scale (EQ-5D-5L) utility scores and visual analogue scale (VAS) ratings. Although not all differences were significant, 5-item Amyotrophic Lateral Sclerosis Assessment Questionnaire (ALSAQ-5) scores showed a stepwise increase in HRQoL impairment at each stage of the disease regardless of the staging system. At later stages, high levels of fatigue and substantial activity impairment were reported. As pALS disease states progressed, cALS also experienced a decline in HRQoL and increased burden. CONCLUSIONS: Across outcomes, pALS and cALS generally reported worse outcomes at later stages of the disease, highlighting an unmet need in this population for strategies to maximise QoL despite disease progression. Recognition and treatment of symptoms such as pain and fatigue may lead to improved outcomes for pALS and cALS.

Access to anticancer drugs: many evidence-based treatments are off-label and unfunded by the Pharmaceutical Benefits Scheme.

BACKGROUND: The off-label use of a drug refers to a use outside the terms of its approval by the Therapeutic Goods Administration (TGA). It is also possible to prescribe unlicensed drugs under the TGA's special access scheme. A high rate of off-label prescribing has previously been reported in cancer. Our study aimed to document the disparity between evidence-based clinical guidelines for anticancer therapy, product approval and funding status of these agents within an academic tertiary/quaternary cancer centre. METHODS: All chemotherapy protocols approved for use in our specialist oncology centre were assessed to determine if the drugs were off-label or unlicensed for that indication based on review of their current product information. The Pharmaceutical Benefits Scheme (PBS) funding status for each protocol was subsequently assessed. RESULTS: A total of 448 protocols containing 82 different drugs across 15 tumour groups was identified. Overall, 189 (42.2%) of protocols were off-label, and three (0.7%) were unlicensed. This resulted in all 192 protocols being unfunded by the PBS. Of the 189 off-label protocols, 132 (69.9%) were based on established evidence-based treatment guidelines, and a further 39 (20.6%) was based on phase II or III clinical trial data. CONCLUSION: Over 90% of off-label protocols are supported by established treatment guidelines or published peer-reviewed research even though the medications are not approved for that particular use by the TGA. However, these off-label protocols are unfunded by the PBS; this results in a marked inequality of access to appropriate medications for cancer patients across Australia.

Famine: causes, prevention, and relief.

Famines are generally caused by decline in food production in successive years brought about by poor weather, war, or both. The consequent complex interactions between prices, employment, and assets impoverish victims and lead to sharply increased mortality. Government policy is a key determinant as to whether or not these conditions mature into wisespread famine. India and Bangladesh have succeeded in controlling famines in recent years, but problems in most of Africa remain intractable due to civil unrest and a paucity of resources, including trained people, institutions, and infrastructure. General economic development and political consensus is needed to reduce Africa's vulnerability to famine. In this context, judiciously provided foreign aid can be of immense help.

Presynaptic kainate receptor mediation of frequency facilitation at hippocampal mossy fiber synapses.

Inhibition of transmitter release by presynaptic receptors is widespread in the central nervous system and is typically mediated via metabotropic receptors. In contrast, very little is known about facilitatory receptors, and synaptic activation of a facilitatory autoreceptor has not been established. Here we show that activation of presynaptic kainate receptors can facilitate transmitter release from hippocampal mossy fiber synapses. Synaptic activation of these presumed ionotropic kainate receptors is very fast (<10 ms) and lasts for seconds. Thus, these presynaptic kainate receptors contribute to the short-term plasticity characteristics of mossy fiber synapses, which were previously thought to be an intrinsic property of the synapse.

The actions of synaptically released zinc at hippocampal mossy fiber synapses.

Zn2+ is present at high concentrations in the synaptic vesicles of hippocampal mossy fibers. We have used Zn2+ chelators and the mocha mutant mouse to address the physiological role of Zn2+ in this pathway. Zn2+ is not involved in the unique presynaptic plasticities observed at mossy fiber synapses but is coreleased with glutamate from these synapses, both spontaneously and with electrical stimulation, where it exerts a strong modulatory effect on the NMDA receptors. Zn2+ tonically occupies the high-affinity binding site of NMDA receptors at mossy fiber synapses, whereas the lower affinity voltage-dependent Zn2+ binding site is occupied during action potential driven-release. We conclude that Zn2+ is a modulatory neurotransmitter released from mossy fiber synapses and plays an important role in shaping the NMDA receptor response at these synapses.

Roles for Gcn5p and Ada2p in transcription and nucleotide excision repair at the Saccharomyces cerevisiae MET16 gene.

Chromatin structure, transcription and repair of cyclobutane pyrimidine dimers at the MET16 gene of wild type, gcn5Delta and ada2Delta Saccharomyces cerevisiae cells were studied under repressing or derepressing conditions. These two components of the SAGA/ADA chromatin remodelling complexes are expendable for the basal transcription of MET16 but are mandatory for its full transcription induction. Despite their influence on transcription neither protein induces major changes in MET16 chromatin structure, but some minor ones occur. Repair at the coding region of the transcribed strand is faster than repair at non-transcribed regions in all strains and either growth condition. Moreover, the more MET16 is transcribed the faster the repair. The data show that by changing the transcription extent the rate of repair at each DNA strand is altered in a different way, confirming that repair at this locus is strongly modulated by its chromatin structure and transcription level. Deletion of GCN5 or ADA2 reduces repair at MET16. The results are discussed in light of the current understanding of Gcn5p and Ada2p functions, and they are the first to report a role for Ada2p in the nucleotide excision repair of the regulatory and transcribed regions of a gene.

Multiple control elements in the TRP1 promoter of Saccharomyces cerevisiae.

The TRP1 promoter generates two groups of mRNAs, transcript I and transcript II. The difference in size between the largest and smallest mRNAs is about 200 base pairs. A series of one-sided and internal deletions were constructed in vitro throughout the TRP1 promoter, and the effect of each deletion on transcription was assessed by Northern blotting. We showed that 395 base pairs of the TRP1 promoter were sufficient for the normal transcription of all RNAs and that the promoter contained two control domains. The control domain for transcript I consisted of one positive element and one negative element, while the control domain for transcript II contained two positive elements. The negative element, mapped between -293 and -318, expression of transcript I. Two regions of transcript I. Two regions (-280 to -236 and -235 to -209) were required for accurate initiation of transcript I. Each region contained sequences homologous to known consensus sequences of the TATA box.

Chromatin structure modulation in Saccharomyces cerevisiae by centromere and promoter factor 1.

CPF1 is an abundant basic-helix-loop-helix-ZIP protein that binds to the CDEI motif in Saccharomyces cerevisiae centromeres and in the promoters of numerous genes, including those encoding enzymes of the methionine biosynthetic pathway. Strains lacking CPF1 are methionine auxotrophs, and it has been proposed that CPF1 might positively influence transcription at the MET25 and MET16 genes by modulating promoter chromatin structure. We test this hypothesis and show that the regions surrounding the CDEI motifs in the MET25 and MET16 promoters are maintained in a nucleosome-free state and that this requires the entire CPF1 protein. However, the chromatin structure around the CDEI motifs does not change on derepression of transcription and does not correlate with the methionine phenotype of the cell. An intact CDEI motif but not CPF1 is required for transcriptional activation from a region of the MET25 upstream activation sequence. Our results suggest that CPF1 functions to modulate chromatin structure around the CDEI motif but that these changes at the MET25 and MET16 promoters do not explain how CPF1 functions to maintain methionine-independent growth. The presence of CPF1-dependent chromatin structures at these promoters leads to a weak repression of transcription.

Efficient expression of the Saccharomyces cerevisiae PGK gene depends on an upstream activation sequence but does not require TATA sequences.

The Saccharomyces cerevisiae PGK (phosphoglycerate kinase) gene encodes one of the most abundant mRNA and protein species in the cell. To identify the promoter sequences required for the efficient expression of PGK, we undertook a detailed internal deletion analysis of the 5' noncoding region of the gene. Our analysis revealed that PGK has an upstream activation sequence (UASPGK) located between 402 and 479 nucleotides upstream from the initiating ATG sequence which is required for full transcriptional activity. Deletion of this sequence caused a marked reduction in the levels of PGK transcription. We showed that PGK has no requirement for TATA sequences; deletion of one or both potential TATA sequences had no effect on either the levels of PGK expression or the accuracy of transcription initiation. We also showed that the UASPGK functions as efficiently when in the inverted orientation and that it can enhance transcription when placed upstream of a TRP1-IFN fusion gene comprising the promoter of TRP1 fused to the coding region of human interferon alpha-2.

Duplex Scorpion primers in SNP analysis and FRET applications.

Scorpions are fluorogenic PCR primers with a probe element attached at the 5'-end via a PCR stopper. They are used in real-time amplicon-specific detection of PCR products in homogeneous solution. Two different formats are possible, the 'stem-loop' format and the 'duplex' format. In both cases the probing mechanism is intramolecular. We have shown that duplex Scorpions are efficient probes in real-time PCR. They give a greater fluorescent signal than stem-loop Scorpions due to the vastly increased separation between fluorophore and quencher in the active form. We have demonstrated their use in allelic discrimination at the W1282X locus of the ABCC7 gene and shown that they can be used in assays where fluorescence resonance energy transfer is required.

Cbf1p modulates chromatin structure, transcription and repair at the Saccharomyces cerevisiae MET16 locus.

The presence of damage in the transcribed strand (TS) of active genes and its position in relation to nucleosomes influence nucleotide excision repair (NER) efficiency. We examined chromatin structure, transcription and repair at the MET16 gene of wild-type and cbf1Delta Saccharomyces cerevisiae cells under repressing or derepressing conditions. Cbf1p is a sequence-specific DNA binding protein required for MET16 chromatin remodelling. Irrespective of the level of transcription, repair at the MspI restriction fragment of MET16 exhibits periodicity in line with nucleosome positions in both strands of the regulatory region and the non-transcribed strand of the coding region. However, repair in the coding region of the TS is always faster, but exhibits periodicity only when MET16 is repressed. In general, absence of Cbf1p decreased repair in the sequences examined, although the effects were more dramatic in the Cbf1p remodelled area, with repair being reduced to the lowest levels within the nucleosome cores of this region. Our results indicate that repair at the promoter and coding regions of this lowly transcribed gene are dependent on both chromatin structure and the level of transcription. The data are discussed in light of current models relating NER and chromatin structure.

Imitation switch complexes.

The imitation switch (ISWI) family of chromatin remodelling ATPases is found in organisms ranging from yeast to mammals. ISWI ATPases assemble chromatin and slide and space nucleosomes, making the chromatin template fluid and allowing appropriate regulation of events such as transcription, DNA replication, recombination and repair. The site of action of the ATPases is determined, in part by the tissue type in which the enzyme is expressed and in part by the nature of the proteins associated with the enzyme. The ISWI complexes are generally conserved in composition and function across species. Roles in gene expression and DNA replication in heterochromatin, gene activation and repression in euchromatin, and functions related to maintaining chromosome architecture are associated with different complexes. Defects in ISWI-associated proteins may be associated with neurodegenerative disease, anencephaly, William's syndrome and melanotic tumours. Finally, the mechanism by which yeast Isw Ib influences gene transcription is discussed.

Fc-γ Receptor Polymorphisms, Cetuximab Therapy, and Survival in the NCIC CTG CO.17 Trial of Colorectal Cancer.

PURPOSE: Two germline Fc-γ receptor (FCGR) polymorphisms, rs1801274 [FCGR2A;His(H)131Arg(R)] and rs396991 [FCGR3A;Phe(F)158Val(V)] produce altered proteins through amino acid substitutions; both are reported to be associated with cetuximab-related outcomes. We performed a validation of these polymorphisms in NCIC CTG CO.17, a randomized trial of cetuximab monotherapy in refractory, metastatic colorectal cancer expressing EGFR. EXPERIMENTAL DESIGN: DNA extracted from formalin-fixed paraffin-embedded tissue was genotyped. In addition to log-rank tests, Cox proportional hazard models assessed their relationships with overall (OS) and progression-free survival (PFS), adjusting for clinically important prognostic factors, along with a polymorphism-treatment arm interaction term. RESULTS: Somatic KRAS status was wild-type for exon 2 in 153 (52%) of 293 patients, from whom tumor DNA was available. For FCGR2A H/H, a genotype-treatment interaction for KRAS wild-type patients was observed for OS (P = 0.03). In KRAS wild-type patients carrying FCGR2A H/H, cetuximab (vs. no cetuximab) improved survival substantially, with adjusted HRs (aHR) of 0.36 (OS) and 0.19 (PFS) and absolute benefits of 5.5 months (OS; P = 0.003) and 3.7 months (PFS; P = 0.02). In contrast, patients carrying FCGR2A R alleles (H/R or R/R) had aHRs of only 0.78 (OS; 2.8-month benefit) and 0.53 (PFS; 1.6-month benefit). No relationships were found for rs396991 (FCGR3A). CONCLUSIONS: In the CO.17 trial, cetuximab worked best for patients with KRAS wild-type colorectal cancers carrying FCGR2A H/H genotypes. Significantly lower benefits were observed in patients carrying germline FCGR2A R alleles. Clin Cancer Res; 22(10); 2435-44. ©2016 AACR.

Chromosome rearrangements induced by recombinant coliphage lambda placMu.

Operon fusions to lacZ, commonly used to study bacterial gene expression in vivo, are normally constructed using phage derivatives such as lambda placMu53 or Mud-1. These derivatives contain a part of trp operon, and we have found that, when integrated into the chromosome, recombination can occur at high frequency between this trp DNA and the chromosomal trp operon leading to chromosomal inversions which fuse lacZ to the trp promoter. Large segments of the chromosome can be inverted by such rearrangements and their occurrence can seriously complicate the isolation of regulatory mutations and other studies unless appropriate precautions are taken. This phenomenon provides a simple means of isolating inversions of defined chromosomal segments and determining the direction of transcription of some lacZ operon fusions.

Factors affecting heterologous gene expression in Saccharomyces cerevisiae.

The 'promoter' fragment from the yeast phosphoglycerate kinase (PGK) gene has been used to direct the expression of human interferon-alpha-2 (IFN alpha 2) on a high-copy-number plasmid in yeast. The yields of IFN alpha 2 are only 1-3% of yeast total protein, whereas the maximum yield of PGK produced by the PGK gene on a high-copy-number plasmid is at least 50%. IFN alpha 2 is turned over more rapidly than PGK but in addition a major reason for the relatively low level of IFN alpha 2 is that IFN-specific RNA levels are much lower. This does not reflect differences in plasmid copy number or integrity, or differences in the 5' and 3' untranslated regions of the transcripts or DNA flanking regions. It appears that the presence of heterologous coding sequences, or the absence of specific yeast sequences causes a reduction in heterologous RNA levels in yeast.

Efficient synthesis of enzymatically active calf chymosin in Saccharomyces cerevisiae.

We have constructed a high-efficiency expression vector to direct the synthesis of heterologous polypeptides in yeast. The vector is termed a sandwich expression vector as the heterologous gene is inserted between the 5' and 3' control regions of the efficiently expressed yeast PGK gene. We have used this vector to direct the expression of three derivatives of the calf chymosin cDNA gene; preprochymosin, prochymosin and chymosin. Prochymosin is synthesised to at least 5% of total yeast-cell protein and furthermore, it can be readily activated to produce an enzyme which has milk-clotting activity.

Somato-synaptic variation of GABA(A) receptors in cultured murine cerebellar granule cells: investigation of the role of the alpha6 subunit.

Electrophysiological investigation of cultured cerebellar murine granule cells revealed differences between the GABA(A) receptors at inhibitory synapses and those on the cell body. Specifically, mIPSCs decayed more rapidly than cell body receptors deactivated, the mean single channel conductance at the synapse (32 pS) was greater than that at cell body (21 pS) and only cell body receptors were sensitive to Zn(2+) (150 microM), which depressed response amplitude by 82+/-5% and almost doubled the rate of channel deactivation. The GABA(A) receptor alpha6 subunit is selectively expressed in cerebellar granule cells. Although concentrated at synapses, it is also found on extrasynaptic membranes. Using a mouse line (Deltaalpha6lacZ) lacking this subunit, we investigated its role in the somato-synaptic differences in GABA(A) receptor function. All differences between cell body and synaptic GABA(A) receptors observed in wild-type (WT) granule cells persisted in Deltaalpha6lacZ cells, thus demonstrating that they are not specifically due to the cellular distribution of the alpha6 subunit. However, mIPSCs from WT and Deltaalpha6lacZ cells differed in both their kinetics (faster decay in WT cells) and underlying single channel conductance (32 pS WT, 25 pS Deltaalpha6lacZ). This provides good evidence for a functional contribution of the alpha6 subunit to postsynaptic GABA(A) receptors in these cells. Despite this, deactivation kinetics of mIPSCs in WT and Deltaalpha6lacZ granule cells exhibited similar benzodiazepene (BDZ) sensitivity. This suggests that the enhanced BDZ-induced ataxia seen in Deltaalpha6lacZ mice may reflect physiological activity at extrasynaptic receptors which, unlike those at synapses, display differential BDZ-sensitivity in WT and Deltaalpha6lacZ granule cells (Jones, A.M., Korpi, E.R., McKernan, R.M., Nusser, Z., Pelz, R., Makela, R., Mellor, J.R., Pollard, S., Bahn, S., Stephenson, F.A., Randall, A.D., Sieghart, W., Somogyi, P., Smith, A.J.H., Wisden, W., 1997. Ligand-gated ion channel partnerships: GABA(A) receptor alpha(6) subunit inactivation inhibits delta subunit expression. Journal of Neuroscience 17, 1350-1362).