Ocular Gene Therapy with Adeno-associated Virus Vectors: Current Outlook for Patients and Researchers

In this “Perspective”, we discuss ocular gene therapy – the patient’s perspective, the various strategies of gene replacement and gene editing, the place of adenoassociated virus vectors, routes of delivery to the eye and the remaining question - “why does immunity continue to limit efficacy?” Through the coordinated efforts of patients, researchers, granting agencies and industry, and after many years of pre-clinical studies, biochemical, cellular, and animal models, we are seeing clinical trials emerge for many previously untreatable heritable ocular disorders. The pathway to therapies has been led by the successful treatment of the RPE65 form of Leber congenital amaurosis with LUXTURNATM. In some cases, immune reactions to the vectors continue to occur, limiting efficacy. The underlying mechanisms of inflammation require further study, and new vectors need to be designed that limit the triggers of immunity. Researchers studying ocular gene therapies and clinicians enrolling patients in clinical trials must recognize the current limitations of these therapies to properly manage expectations and avoid disappointment, but we believe that gene therapies are well on their way to successful, widespread utilization to treat heritable ocular disorders

The Intrinsic Two-Dimensional Size of Sagittarius A*

We report the detection of the two-dimensional structure of the radio source associated with the Galactic Center black hole, Sagittarius A*, obtained from Very Long Baseline Array (VLBA) observations at a wavelength of 7mm. The intrinsic source is modeled as an elliptical Gaussian with major axis size 35.4 x 12.6 R_S in position angle 95 deg East of North. This morphology can be interpreted in the context of both jet and accretion disk models for the radio emission. There is supporting evidence in large angular-scale multi-wavelength observations for both source models for a preferred axis near 95 deg. We also place a maximum peak-to-peak change of 15% in the intrinsic major axis size over five different epochs. Three observations were triggered by detection of near infrared (NIR) flares and one was simultaneous with a large X-ray flare detected by NuSTAR. The absence of simultaneous and quasi-simultaneous flares indicates that not all high energy events produce variability at radio wavelengths. This supports the conclusion that NIR and X-ray flares are primarily due to electron excitation and not to an enhanced accretion rate onto the black hole.Comment: accepted for publication in Ap

A Millisecond Interferometric Search for Fast Radio Bursts with the Very Large Array

We report on the first millisecond timescale radio interferometric search for the new class of transient known as fast radio bursts (FRBs). We used the Very Large Array (VLA) for a 166-hour, millisecond imaging campaign to detect and precisely localize an FRB. We observed at 1.4 GHz and produced visibilities with 5 ms time resolution over 256 MHz of bandwidth. Dedispersed images were searched for transients with dispersion measures from 0 to 3000 pc/cm3. No transients were detected in observations of high Galactic latitude fields taken from September 2013 though October 2014. Observations of a known pulsar show that images typically had a thermal-noise limited sensitivity of 120 mJy/beam (8 sigma; Stokes I) in 5 ms and could detect and localize transients over a wide field of view. Our nondetection limits the FRB rate to less than 7e4/sky/day (95% confidence) above a fluence limit of 1.2 Jy-ms. Assuming a Euclidean flux distribution, the VLA rate limit is inconsistent with the published rate of Thornton et al. We recalculate previously published rates with a homogeneous consideration of the effects of primary beam attenuation, dispersion, pulse width, and sky brightness. This revises the FRB rate downward and shows that the VLA observations had a roughly 60% chance of detecting a typical FRB and that a 95% confidence constraint would require roughly 500 hours of similar VLA observing. Our survey also limits the repetition rate of an FRB to 2 times less than any known repeating millisecond radio transient.Comment: Submitted to ApJ. 13 pages, 9 figure

Techno-Economic Feasibility Analysis of a Fully Mobile Radiation Oncology System using Monte Carlo Simulation

PURPOSEDisparities in radiation oncology (RO) can be attributed to geographic location, socioeconomic status, race, sex, and other societal factors. One potential solution is to implement a fully mobile (FM) RO system to bring radiotherapy to rural areas and reduce barriers to access. We use Monte Carlo simulation to quantify techno-economic feasibility with uncertainty, using two rural Missouri scenarios.METHODSRecently, a semimobile RO system has been developed by building an o-ring linear accelerator (linac) into a mobile coach that is used for temporary care, months at a time. Transitioning to a more FM-RO system, which changes location within a given day, presents technical challenges including logistics and quality assurance. This simulation includes cancer census in both northern and southeastern Missouri, multiple treatment locations within a given day, and associated expenditures and revenues. A subset of patients with lung, breast, and rectal diseases, treated with five fractions, was simulated in the FM-RO system.RESULTSThe FM-RO can perform all necessary quality assurance tests as suggested in national medical physics guidelines within 1.5 hours, thus demonstrating technological feasibility. In northern and southeastern Missouri, five-fraction simulations\u27 net incomes were, in US dollars (USD), $1.55 ± 0.17 million (approximately 74 patients/year) and$3.65 USD ± 0.25 million (approximately 98 patients/year), respectively. The number of patients seen had the highest correlation with net income as well as the ability to break-even within the simulation. The model does not account for disruptions in care or other commonly used treatment paradigms, which may lead to differences in estimated economic return. Overall, the mobile system achieved a net benefit, even for the most negative simulation scenarios.CONCLUSIONOur simulations suggest technologic success and economic viability for a FM-RO system within rural Missouri and present an interesting solution to address other geographic disparities in access to radiotherapy

Core handling, transportation and processing for the South Pole ice core (SPICEcore) project

An intermediate-depth (1751 m) ice core was drilled at the South Pole between 2014 and 2016 using the newly designed US Intermediate Depth Drill. The South Pole ice core is the highest-resolution interior East Antarctic ice core record that extends into the glacial period. The methods used at the South Pole to handle and log the drilled ice, the procedures used to safely retrograde the ice back to the National Science Foundation Ice Core Facility (NSF-ICF), and the methods used to process and sample the ice at the NSF-ICF are described. The South Pole ice core exhibited minimal brittle ice, which was likely due to site characteristics and, to a lesser extent, to drill technology and core handling procedures

VAST: An ASKAP Survey for Variables and Slow Transients

The Australian Square Kilometre Array Pathfinder (ASKAP) will give us an unprecedented opportunity to investigate the transient sky at radio wavelengths. In this paper we present VAST, an ASKAP survey for Variables and Slow Transients. VAST will exploit the wide-field survey capabilities of ASKAP to enable the discovery and investigation of variable and transient phenomena from the local to the cosmological, including flare stars, intermittent pulsars, X-ray binaries, magnetars, extreme scattering events, interstellar scintillation, radio supernovae and orphan afterglows of gamma ray bursts. In addition, it will allow us to probe unexplored regions of parameter space where new classes of transient sources may be detected. In this paper we review the known radio transient and variable populations and the current results from blind radio surveys. We outline a comprehensive program based on a multi-tiered survey strategy to characterise the radio transient sky through detection and monitoring of transient and variable sources on the ASKAP imaging timescales of five seconds and greater. We also present an analysis of the expected source populations that we will be able to detect with VAST.Comment: 29 pages, 8 figures. Submitted for publication in Pub. Astron. Soc. Australi

NLRP3 inflammasome blockade reduces liver inflammation and fibrosis in experimental NASH in mice

Background & Aims: NOD-like receptor protein 3 (NLRP3) inflammasome activation occurs in Non-alcoholic fatty liver disease (NAFLD). We used the first small molecule NLRP3 inhibitor, MCC950, to test whether inflammasome blockade alters inflammatory recruitment and liver fibrosis in two murine models of steatohepatitis. Methods: We fed foz/foz and wild-type mice an atherogenic diet for 16 weeks, gavaged MCC950 or vehicle until 24 weeks, then determined NAFLD phenotype. In mice fed an methionine/choline deficient (MCD) diet, we gavaged MCC950 or vehicle for 6 weeks and determined the effects on liver fibrosis. Results: In vehicle-treated foz/foz mice, hepatic expression of NLRP3, pro-IL-1β, active caspase-1 and IL-1β increased at 24 weeks, in association with cholesterol crystal formation and NASH pathology; plasma IL-1β, IL-6, MCP-1, ALT/AST all increased. MCC950 treatment normalized hepatic caspase 1 and IL-1β expression, plasma IL-1β, MCP-1 and IL-6, lowered ALT/AST, and reduced the severity of liver inflammation including designation as NASH pathology, and liver fibrosis. In vitro, cholesterol crystals activated Kupffer cells and macrophages to release IL-1β; MCC950 abolished this, and the associated neutrophil migration. MCD diet-fed mice developed fibrotic steatohepatitis; MCC950 suppressed the increase in hepatic caspase 1 and IL-1β, lowered numbers of macrophages and neutrophils in the liver, and improved liver fibrosis. Conclusion: MCC950, an NLRP3 selective inhibitor, improved NAFLD pathology and fibrosis in obese diabetic mice. This is potentially attributable to the blockade of cholesterol crystal-mediated NLRP3 activation in myeloid cells. MCC950 reduced liver fibrosis in MCD-fed mice. Targeting NLRP3 is a logical direction in pharmacotherapy of NASH.Supported by NIH Project grants R2 AA023574 and U01 AA022489 (to AEF), Australian NHMRC project grants 1084136 and 1044288 (to GCF), and 1086786 (to MAC and AABR), and Deutsche Forschungsgemeinschaft WR 173/3-1 (to AW). Matthew Cooper is an NHMRC Principle Research Fellow (1059354

The inner junction protein CFAP20 functions in motile and non-motile cilia and is critical for vision

Motile and non-motile cilia are associated with mutually-exclusive genetic disorders. Motile cilia propel sperm or extracellular fluids, and their dysfunction causes primary ciliary dyskinesia. Non-motile cilia serve as sensory/signalling antennae on most cell types, and their disruption causes single-organ ciliopathies such as retinopathies or multi-system syndromes. CFAP20 is a ciliopathy candidate known to modulate motile cilia in unicellular eukaryotes. We demonstrate that in zebrafish, cfap20 is required for motile cilia function, and in C. elegans, CFAP-20 maintains the structural integrity of non-motile cilia inner junctions, influencing sensory-dependent signalling and development. Human patients and zebrafish with CFAP20 mutations both exhibit retinal dystrophy. Hence, CFAP20 functions within a structural/functional hub centered on the inner junction that is shared between motile and non-motile cilia, and is distinct from other ciliopathy-associated domains or macromolecular complexes. Our findings suggest an uncharacterised pathomechanism for retinal dystrophy, and potentially for motile and non-motile ciliopathies in general