An Allosteric Inhibitor of KRas Identified Using a Barcoded Rapid Assay Microchip Platform

Protein catalyzed capture agents (PCCs) are synthetic antibody surrogates that can target a wide variety of biologically relevant proteins. As a step toward developing a high-throughput PCC pipeline, we report on the preparation of a barcoded rapid assay platform for the analysis of hits from PCC library screens. The platform is constructed by first surface patterning a micrometer scale barcode composed of orthogonal ssDNA strands onto a glass slide. The slide is then partitioned into microwells, each of which contains multiple copies of the full barcode. Biotinylated candidate PCCs from a click screen are assembled onto the barcode stripes using a complementary ssDNA-encoded cysteine-modified streptavidin library. This platform was employed to evaluate candidate PCC ligands identified from an epitope targeted in situ click screen against the two conserved allosteric switch regions of the Kirsten rat sarcoma (KRas) protein. A single microchip was utilized for the simultaneous evaluation of 15 PCC candidate fractions under more than a dozen different assay conditions. The platform also permitted more than a 10-fold savings in time and a more than 100-fold reduction in biological and chemical reagents relative to traditional multiwell plate assays. The best ligand was shown to exhibit an in vitro inhibition constant (IC_(50)) of ∼24 μM

An Allosteric Inhibitor of KRas Identified Using a Barcoded Rapid Assay Microchip Platform

Protein catalyzed capture agents (PCCs) are synthetic antibody surrogates that can target a wide variety of biologically relevant proteins. As a step toward developing a high-throughput PCC pipeline, we report on the preparation of a barcoded rapid assay platform for the analysis of hits from PCC library screens. The platform is constructed by first surface patterning a micrometer scale barcode composed of orthogonal ssDNA strands onto a glass slide. The slide is then partitioned into microwells, each of which contains multiple copies of the full barcode. Biotinylated candidate PCCs from a click screen are assembled onto the barcode stripes using a complementary ssDNA-encoded cysteine-modified streptavidin library. This platform was employed to evaluate candidate PCC ligands identified from an epitope targeted in situ click screen against the two conserved allosteric switch regions of the Kirsten rat sarcoma (KRas) protein. A single microchip was utilized for the simultaneous evaluation of 15 PCC candidate fractions under more than a dozen different assay conditions. The platform also permitted more than a 10-fold savings in time and a more than 100-fold reduction in biological and chemical reagents relative to traditional multiwell plate assays. The best ligand was shown to exhibit an in vitro inhibition constant (IC_(50)) of ∼24 μM

Allosteric Inhibitor of KRas Identified Using a Barcoded Assay Microchip Platform

Protein catalyzed capture agents (PCCs) are synthetic antibody surrogates that can target a wide variety of biologically relevant proteins. As a step toward developing a high-throughput PCC pipeline, we report on the preparation of a barcoded rapid assay platform for the analysis of hits from PCC library screens. The platform is constructed by first surface patterning a micrometer scale barcode composed of orthogonal ssDNA strands onto a glass slide. The slide is then partitioned into microwells, each of which contains multiple copies of the full barcode. Biotinylated candidate PCCs from a click screen are assembled onto the barcode stripes using a complementary ssDNA-encoded cysteine-modified streptavidin library. This platform was employed to evaluate candidate PCC ligands identified from an epitope targeted in situ click screen against the two conserved allosteric switch regions of the Kirsten rat sarcoma (KRas) protein. A single microchip was utilized for the simultaneous evaluation of 15 PCC candidate fractions under more than a dozen different assay conditions. The platform also permitted more than a 10-fold savings in time and a more than 100-fold reduction in biological and chemical reagents relative to traditional multiwell plate assays. The best ligand was shown to exhibit an in vitro inhibition constant (IC₅₀) of ∼24 μM

The impact of assistive technology on burden and psychological well-being in informal caregivers of people with dementia (ATTILA Study).

INTRODUCTION: Assistive technology and telecare (ATT) may alleviate psychological burden in informal caregivers of people with dementia. This study assessed the impact of ATT on informal caregivers' burden and psychological well-being. METHODS: Individuals with dementia and their informal caregivers were recruited to a randomized-controlled trial assessing effectiveness of ATT. Caregivers were allocated to two groups according to their cared-for person's randomization to a full or basic package of ATT and were assessed on caregiver burden, state anxiety, and depression. Caregivers' data from three assessments over 6 months of the trial were analyzed. RESULTS: No significant between- or within-group differences at any time point on caregivers' burden, anxiety, and depression levels were found. DISCUSSION: Full ATT for people with dementia did not impact caregivers' psychological outcomes compared to basic ATT. The length of follow up was restricted to 6 months

Assessment of need and practice for assistive technology and telecare for people with dementia-The ATTILA (Assistive Technology and Telecare to maintain Independent Living At home for people with dementia) trial.

INTRODUCTION: The objective of this study was to define current assistive technology and telecare (ATT) practice for people with dementia living at home. METHODS: This is a randomized controlled trial (N = 495) of ATT assessment and ATT installation intervention, compared with control (restricted ATT package). ATT assessment and installation data were collected. Qualitative work identified value networks delivering ATT, established an ATT assessment standard. RESULTS: ATT was delivered by public and not-for-profit telecare networks. ATT assessments showed 52% fidelity to the ATT assessment standard. Areas of assessment most frequently leading to identifying ATT need were daily activities (93%), memory (89%), and problem-solving (83%). ATT needs and recommendations were weakly correlated (τ = 0.242; P < .000), with ATT recommendations and installations moderately correlated (τ = -0.470; P < .000). Half (53%) of recommended technology was not installed. Safety concerns motivated 38% of installations. DISCUSSION: Assessment recommendations were routinely disregarded at the point of installation. ATT was commonly recommended for safety and seldom for supporting leisure.Funding for the trial came from the NIHR Health Technology Assessment program (HTA 10/50/02). Neither the funders nor the sponsor have been involved in the preparation or publication of this article. P.B. received financial support from TauRx Therapeutics. J.O.B. received financial support from TauRx, GE Healthcare, Avid/Lilly, and Eisai