Long-Term Preservation of Cones and Improvement in Visual Function Following Gene Therapy in a Mouse Model of Leber Congenital Amaurosis Caused by Guanylate Cyclase-1 Deficiency

Leber congenital amaurosis (LCA) is a severe retinal dystrophy manifesting from early infancy as poor vision or blindness. Loss-of-function mutations in GUCY2D cause LCA1 and are one of the most common causes of LCA, accounting for 20% of all cases. Human GUCY2D and mouse Gucy2e genes encode guanylate cyclase-1 (GC), which is responsible for restoring the dark state in photoreceptors after light exposure. The Glicy2e(-/-) mouse shows partially diminished rod function, but an absence of cone function before degeneration. Although the cones appear morphologically normal, they exhibit mislocalization of proteins involved in phototransduction. In this study we tested the efficacy of an rAAV2/8 vector containing the human rhodopsin kinase promoter and the human GUCY2D gene. Following subretinal delivery of the vector in Glicy2e(-/-) mice, GC1 protein was detected in the rod and cone outer segments, and in transduced areas of retina cone transducin was appropriately localized to cone outer segments. Moreover, we observed a dose-dependent restoration of rod and cone function and an improvement in visual behavior of the treated mice. Most importantly, cone preservation was observed in transduced areas up to 6 months post injection. To date, this is the most effective rescue of the Glicy2e(-/-) mouse model of LCA and we propose that a vector, similar to the one used in this study, could be suitable for use in a clinical trial of gene therapy for LCA1

Development of an optimized AAV2/5 gene therapy vector for Leber congenital amaurosis owing to defects in RPE65

Leber congenital amaurosis is a group of inherited retinal dystrophies that cause severe sight impairment in childhood; RPE65-deficiency causes impaired rod photoreceptor function from birth and progressive impairment of cone photoreceptor function associated with retinal degeneration. In animal models of RPE65 deficiency, subretinal injection of recombinant adeno-associated virus (AAV) 2/2 vectors carrying RPE65 cDNA improves rod photoreceptor function, and intervention at an early stage of disease provides sustained benefit by protecting cone photoreceptors against retinal degeneration. In affected humans, administration of these vectors has resulted to date in relatively modest improvements in photoreceptor function, even when retinal degeneration is comparatively mild, and the duration of benefit is limited by progressive retinal degeneration. We conclude that the demand for RPE65 in humans is not fully met by current vectors, and predict that a more powerful vector will provide more durable benefit. With this aim we have modified the original AAV2/2 vector to generate AAV2/5-OPTIRPE65. The new configuration consists of an AAV vector serotype 5 carrying an optimized hRPE65 promoter and a codon-optimized hRPE65 gene. In mice, AAV2/5-OPTIRPE65 is at least 300-fold more potent than our original AAV2/2 vector

Differential Modulation of Retinal Degeneration by Ccl2 and Cx3cr1 Chemokine Signalling

Microglia and macrophages are recruited to sites of retinal degeneration where local cytokines and chemokines determine protective or neurotoxic microglia responses. Defining the role of Ccl2-Ccr2 and Cx3cl1-Cx3cr1 signalling for retinal pathology is of particular interest because of its potential role in age-related macular degeneration (AMD). Ccl2, Ccr2, and Cx3cr1 signalling defects impair macrophage trafficking, but have, in several conflicting studies, been reported to show different degrees of age-related retinal degeneration. Ccl2/Cx3cr1 double knockout (CCDKO) mice show an early onset retinal degeneration and have been suggested as a model for AMD. In order to understand phenotypic discrepancies in different chemokine knockout lines and to study how defects in Ccl2 and/or Cx3cr1 signalling contribute to the described early onset retinal degeneration, we defined primary and secondary pathological events in CCDKO mice. To control for genetic background variability, we compared the original phenotype with that of single Ccl2, Cx3cr1 and Ccl2/Cx3cr1 double knockout mice obtained from backcrosses of CCDKO with C57Bl/6 mice. We found that the primary pathological event in CCDKO mice develops in the inferior outer nuclear layer independently of light around postnatal day P14. RPE and vascular lesions develop secondarily with increasing penetrance with age and are clinically similar to retinal telangiectasia not to choroidal neovascularisation. Furthermore, we provide evidence that a third autosomal recessive gene causes the degeneration in CCDKO mice and in all affected re-derived lines and subsequently demonstrated co-segregation of the naturally occurring RD8 mutation in the Crb1 gene. By comparing CCDKO mice with re-derived CCl2−/−/Crb1Rd8/RD8, Cx3cr1−/−/Crb1Rd8/RD8 and CCl2−/−/Cx3cr1−/−/Crb1Rd8/RD8 mice, we observed a differential modulation of the retinal phenotype by genetic background and both chemokine signalling pathways. These findings indicate that CCDKO mice are not a model of AMD, but a model for an inherited retinal degeneration that is differentially modulated by Ccl2-Ccr2 and Cx3cl1-Cx3cr1 chemokine signalling

Generative Large Language Models are autonomous practitioners of evidence-based medicine

Background: Evidence-based medicine (EBM) is fundamental to modern clinical practice, requiring clinicians to continually update their knowledge and apply the best clinical evidence in patient care. The practice of EBM faces challenges due to rapid advancements in medical research, leading to information overload for clinicians. The integration of artificial intelligence (AI), specifically Generative Large Language Models (LLMs), offers a promising solution towards managing this complexity. Methods: This study involved the curation of real-world clinical cases across various specialties, converting them into .json files for analysis. LLMs, including proprietary models like ChatGPT 3.5 and 4, Gemini Pro, and open-source models like LLaMA v2 and Mixtral-8x7B, were employed. These models were equipped with tools to retrieve information from case files and make clinical decisions similar to how clinicians must operate in the real world. Model performance was evaluated based on correctness of final answer, judicious use of tools, conformity to guidelines, and resistance to hallucinations. Results: GPT-4 was most capable of autonomous operation in a clinical setting, being generally more effective in ordering relevant investigations and conforming to clinical guidelines. Limitations were observed in terms of model ability to handle complex guidelines and diagnostic nuances. Retrieval Augmented Generation made recommendations more tailored to patients and healthcare systems. Conclusions: LLMs can be made to function as autonomous practitioners of evidence-based medicine. Their ability to utilize tooling can be harnessed to interact with the infrastructure of a real-world healthcare system and perform the tasks of patient management in a guideline directed manner. Prompt engineering may help to further enhance this potential and transform healthcare for the clinician and the patient.Comment: Word count: 4548 words, Figures: 4, Tables:

Antibody predictors of mortality and lung function trends in myositis spectrum interstitial lung disease

Objectives: The impact of autoantibody profiles on the prognosis for idiopathic inflammatory myositis-associated interstitial lung disease (IIM-ILD) and myositis spectrum ILD with myositis-specific antibodies (MSAs) remains unclear. This retrospective cohort study examined whether serological profiles were associated with mortality or longitudinal lung function change. Methods: The baseline clinical/demographic characteristics and follow-up lung function data of consecutive adult patients with IIM-ILD or interstitial pneumonia with autoimmune features (IPAF) positive for MSAs (IPAF-MSA) were extracted from three hospitals. Univariate and multivariate Cox proportional hazards analyses were used to compare mortality between groups of patients with different autoantibodies. Regression models were used to analyse their lung function trends. Results: Of the 430 included patients, 81% met the IIM criteria, and the remaining 19% were diagnosed with IPAF-MSA. On univariate analysis, the risk factors associated with mortality included higher age, Charlson Comorbidity Index, and CRP; and lower BMI, baseline TLCO% and FEV1%. Compared with anti-MDA5 negativity, anti-MDA5 positivity (MDA5+) was associated with higher mortality in the first 3 months [hazard ratio (HR) 65.2, 95% CI 14.1, 302.0], while no significant difference was seen thereafter (HR 0.55, 95% CI 0.14, 2.28). On multivariate analysis, combined anti-synthetase antibodies were associated with a reduced risk of mortality (HR 0.63), although individually, mortality was reduced in patients with anti-Jo1+ (HR 0.61, 95% CI 0.4-0.87) and increased in patients with anti-PL7+ (HR 2.07, 95% CI 1.44-2.99). Anti-MDA5+ was associated with slow improvement in %FVC over the first 3 years, while anti-PL7+ was linked with a slow decline from 12 months onwards. Conclusion: Among the autoantibody profiles in myositis spectrum disorders, anti-MDA5+ and anti-PL7+ conferred higher mortality risks in patients with IIM-ILD. Survivors of an early peak of mortality in anti-MDA5+ disease appeared to have a favourable prognosis.Funding: No specific funding was received from any bodies in the public, commercial or not-for-profit sectors to carry out the work described in this article. Disclosure statement: J.H.: none declared, A.L.: none declared, J.M.: none declared, M.N.: none declared, S.S.A.: none declared, C.S.: none declared, C.O.: none declared, A.D.: consultant of: Boehringer Ingelhime, Brainomix, L.P.: none declared, S.A.: none declared, B.A.-M.: none declared, M.A.G.-G.: none declared, A.P.: none declared, A.W.: none declared, K.T.: none declared, H.R.: none declared, F.C.: none declared, V.K.: none declared, B.L.: none declared, A.V.W.: speakers bureau from Boehringer Ingelheim, Roche, and Veracyte, and consultant for: Boehringer Ingelheim, Roche, and Veracyte, S.N.: none declared, J.G.: none declared, E.A.R.: none declared, P.A.G.: speakers bureau from UCB, consultant for Eli Lilly, and Galapagos, and grant/research support from Corbus Pharmaceuticals. Acknowledgements: This work has previously been presented in abstract form at EULAR 2023. J.H. received personal support through grants from the King’s College Hospital Charity

Herschel -ATLAS: Extragalactic number counts from 250 to 500 microns

Aims. The Herschel-ATLAS survey (H-ATLAS) will be the largest area survey to be undertaken by the Herschel Space Observatory. It will cover 550 sq. deg. of extragalactic sky at wavelengths of 100, 160, 250, 350 and 500 μm when completed, reaching flux limits (5σ) from 32 to 145 mJy. We here present galaxy number counts obtained for SPIRE observations of the first ~14 sq. deg. observed at 250, 350 and 500 μm. Methods. Number counts are a fundamental tool in constraining models of galaxy evolution. We use source catalogs extracted from the H-ATLAS maps as the basis for such an analysis. Correction factors for completeness and flux boosting are derived by applying our extraction method to model catalogs and then applied to the raw observational counts. Results. We find a steep rise in the number counts at flux levels of 100–200 mJy in all three SPIRE bands, consistent with results from BLAST. The counts are compared to a range of galaxy evolution models. None of the current models is an ideal fit to the data but all ascribe the steep rise to a population of luminous, rapidly evolving dusty galaxies at moderate to high redshift

Effect of Gene Therapy on Visual Function in Leber's Congenital Amaurosis

Early-onset, severe retinal dystrophy caused by mutations in the gene encoding retinal pigment epithelium–specific 65-kD protein (RPE65) is associated with poor vision at birth and complete loss of vision in early adulthood. We administered to three young adult patients subretinal injections of recombinant adeno-associated virus vector 2/2 expressing RPE65 complementary DNA (cDNA) under the control of a human RPE65 promoter. There were no serious adverse events. There was no clinically significant change in visual acuity or in peripheral visual fields on Goldmann perimetry in any of the three patients. We detected no change in retinal responses on electroretinography. One patient had significant improvement in visual function on microperimetry and on dark-adapted perimetry. This patient also showed improvement in a subjective test of visual mobility. These findings provide support for further clinical studies of this experimental approach in other patients with mutant RPE65. (ClinicalTrials.gov number, NCT00643747.)Supported by grants from the U.K. Department of Health, the British Retinitis Pigmentosa Society, and the Special Trustees of Moorfields Eye Hospital, and by the Sir Jules Thorn Charitable Trust, the Wellcome Trust, the European Union (EVI-Genoret and Clinigene programs), the Medical Research Council, Foundation Fighting Blindness, Fight for Sight, the Ulverscroft Foundation, Fighting Blindness (Ireland), Moorfields Eye Hospital, and Institute of Ophthalmology Biomedical Research Centre for Ophthalmology, University College London

Herschel -ATLAS: The dust energy balance in the edge-on spiral galaxy UGC 4754

We use Herschel PACS and SPIRE observations of the edge-on spiral galaxy UGC 4754, taken as part of the H-ATLAS SDP observations, to investigate the dust energy balance in this galaxy. We build detailed SKIRT radiative models based on SDSS and UKIDSS maps and use these models to predict the far-infrared emission. We find that our radiative transfer model underestimates the observed FIR emission by a factor of two to three. Similar discrepancies have been found for other edge-on spiral galaxies based on IRAS, ISO, and SCUBA data. Thanks to the good sampling of the SED at FIR wavelengths, we can rule out an underestimation of the FIR emissivity as the cause for this discrepancy. Instead we support highly obscured star formation that contributes little to the optical extinction as a more probable explanation.This work used data from the UKIDSS DR5 and SDSS DR7. The UKIDSS project is defined in Lawrence et al. (2007) and uses the UKIRT Wide Field Camera (WFCAM; Casali et al. 2007). Funding for the SDSS and SDSS-II has been provided by the Alfred P. Sloan Foundation, the Participating Institutions, the National Science Foundation, the US Department of Energy, the National Aeronautics and Space Administration, the Japanese Monbukagakusho, the Max Planck Society, and the Higher Education Funding Council for England