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Safety and efficacy of glofitamab for relapsed/refractory large B-cell lymphoma in a multinational real-world study
Get PDFGlofitamab, a bispecific antibody targeting CD20 and CD3, is approved for relapsed/refractory diffuse large B-cell lymphoma (r/r DLBCL) after at least 2 prior treatment lines, but real-world data are scarce. In this retrospective, multicenter, multinational study, we evaluated the outcomes of 70 patients with r/r DLBCL treated with glofitamab as part of the compassionate use patient program in Germany, Austria, and Switzerland. The median number of prior treatment lines was 4, with 71% of patients refractory to their last treatment. Cytokine release syndrome was observed in 40% of patients (grade 3-4 in 2%), immune effector cell–associated neurotoxicity syndrome in 10% (grade 3 in 1%), and infections in 31% (grade 5 in 3%). The overall response rate was 46%, with 27% achieving complete responses (CR) and 19% partial responses. The median progression-free survival (PFS) was 3.6 months, whereas the median overall survival was 5.7 months. Notably, 13 patients (19%) were in CR 6 months after initiating glofitamab and exhibited durable responses. Elevated lactate dehydrogenase is the most robust predictor of inferior outcome. Patients pretreated with bendamustine within 6 months prior to glofitamab initiation exhibited significantly reduced PFS, suggesting that bendamustine may impair T-cell fitness and hence glofitamab efficacy. In summary, glofitamab demonstrates promising efficacy and a manageable safety profile in heavily pretreated patients with r/r DLBCL in a real-world scenario and the optimal sequence of treatments should use T-cell–depleting agents before glofitamab with caution
Neuropsychiatric changes following striatal stroke - results from the observational PostPsyDis study
Get PDFBACKGROUND: Ischemic stroke can lead to neuropsychiatric sequelae such as depression and post-traumatic stress disorder (PTSD), resulting in poorer functional outcomes. The POST-stroke PSYchological DIStress PostPsyDis; NCT01187342) study aimed to investigate whether ischemic lesions in the striatum increase the risk of depression and PTSD after stroke. METHODS: This monocenter, observational, case-control study included 84 ischemic stroke patients with striatal (n = 54) and non-striatal ischemic brain lesions (n = 30). Primary study endpoints included symptoms of depression (assessed via the Geriatric Depression Scale; GDS-30) and PTSD (assessed via the Posttraumatic Symptom Scale; PTSS-10) 90 days post-stroke. A normative functional connectome was used to obtain a measure of striatal connectivity to the rest of the brain ("striatal network"). Network damage scores were used to estimate damage of each lesion to the striatal network. RESULTS: Patients with striatal lesions had higher GDS-30 scores at 90 days post-stroke (median 5.6 vs. 3.0; Cohen's d = 0.39; p = 0.057), indicating a small to moderate effect. However, no meaningful group differences were observed in the incidence of depression or PTSD. In multivariable regression analyses, striatal infarction had an adjusted beta coefficient (β) of 1.9 (95%CI 0.19-3.7; p = 0.076) for GDS-10 and 1.8 (95%CI -1.9-5.5; p = 0.25) for PTSS-10 scores after 90 days. Only female sex was independently associated with PTSD severity (adjusted β = 5.1, 95% CI 1.3-8.8; p = 0.008). Analyzing lesion connectivity to the striatal network did not change these findings. CONCLUSIONS: Taken together, the PostPsyDis study suggests a high rate of psychiatric morbidity in stroke patients. Moreover, the study suggests increased neuropsychiatric symptoms in patients with striatal lesions. There is a clear need for larger studies to investigate the role of the striatum in post-stroke neuropsychiatric disorders. TRIAL REGISTRATION: ClinicalTrials.gov (NCT01187342) Registered 23 August 2009, https://clinicaltrials.gov/study/NCT01187342
Nicotinamide modulates gut microbial metabolic potential and accelerates recovery in mild-to-moderate COVID-19
Get PDFCellular NAD(+) depletion, altered tryptophan metabolism and gut microbiome dysbiosis are associated with disease progression and unfavourable clinical outcomes in COVID-19. Here, we show that supplementing tryptophan metabolism with nicotinamide alleviates COVID-19 symptoms. We evaluate a 4-week intervention with a novel nicotinamide formulation (1,000 mg) in a prospective, double-blind, randomized, placebo-controlled trial in 900 symptomatic outpatients with PCR-proven COVID-19. In the primary analysis population of participants at risk for severe COVID-19, 57.6% of those receiving nicotinamide and 42.6% receiving placebo recover from their performance drop at week 2 (P = 0.004). Nicotinamide is also beneficial for returning to normal activities (P = 0.009). Effects on gut metagenomic signatures parallel clinical efficacy, suggesting that nicotinamide influences COVID-19-associated faecal microbiome changes. After 6 months, responders to nicotinamide in acute COVID-19 show fewer post-COVID symptoms than placebo responders (P = 0.010). No relevant safety signals are observed. Overall, our results show that nicotinamide leads to faster recovery of physical performance and modulates COVID-19-associated faecal microbiome changes
BEscreen: a versatile toolkit to design base editing libraries
Get PDFBase editing enables the high-throughput screening of genetic variants for phenotypic effects. Base editing screens require the design of single guide RNA (sgRNA) libraries to enable either gene- or variant-centric approaches. While computational tools supporting the design of sgRNAs exist, no solution offers versatile and scalable library design enabling all major use cases. Here, we introduce BEscreen, a comprehensive base editing guide design tool provided as a web server (bescreen.ostendorflab.org) and as a command line tool. BEscreen provides variant-, gene-, and region-centric modes to accommodate various screening approaches. The variant mode accepts genomic coordinates, amino acid changes, or rsIDs as input. The gene mode designs near-saturation libraries covering the entire coding sequence of given genes or transcripts, and the region mode designs all possible guides for given genomic regions. BEscreen enables selection of guides by biological consequence, it features comprehensive customization of base editor characteristics, and it offers optional annotation using Ensembl’s Variant Effect Predictor. In sum, BEscreen is a highly versatile tool to design base editing screens for a wide range of use cases with seamless scalability from individual variants to large, near-saturation libraries
Unravelling heart failure cellular signalling heterogeneity with spatial transcriptomics
No full textSuper-resolution imaging of native fluorescent photoreceptors in chytrid fungal eyes
Get PDFPhotoorientation in motile fungal zoospores is mediated by rhodopsin guanylyl cyclases (RGCs). In certain chytrids, these photoreceptors form heterodimers consisting of a visible-light-absorbing RGC paired with neorhodopsin (NeoR), a rhodopsin distinguished by its unique spectral properties: far-red absorption and high fluorescence. Leveraging the native fluorescence of NeoR, we detected RGCs in living zoospores of the fungus Rhizoclosmatium globosum. The reversible photoswitching of bistable NeoR enabled super-resolution microscopy, facilitating single-molecule detection and quantification of NeoR proteins within individual zoospores. This approach also revealed the precise localization of RGCs within the rumposome, a chytrid-specific organelle hypothesized to mediate photoreception. Fluorescence tracking across different stages of the chytrid life cycle and the analysis of transcriptomic data confirmed that RGCs are predominantly present during the zoospore stage. Functional assays of recombinantly expressed RGC heterodimers with modified substrate specificity revealed that only one of the two pseudo-symmetric nucleotide-binding sites is catalytically active. Strikingly, disrupting nucleotide binding in the non-catalytic site enhanced light-triggered cyclase activity by up to ninefold, indicating an allosteric regulatory mechanism in heterodimeric RGCs
Single-stranded HDR templates with truncated Cas12a-binding sequences improve knock-in efficiencies in primary human T cells
Get PDFCRISPR-Cas12a gene editing offers an alternative to Cas9-based methods, providing better targeting of AT-rich regions, simplified guide RNA manufacturing, and high specificity. However, the efficacy of donor-based editing is subject to various factors, with template format playing a crucial role. Currently, the predominant non-viral template format for homology-directed repair (HDR) after nuclease-induced DNA breaks is double-stranded DNA, which is toxic when transfected at high doses. Others have demonstrated that using single-stranded DNA (ssDNA) with flanking double-stranded Cas-target-sequences (CTS) as a template for Cas9-mediated gene editing can mitigate this toxicity and increase knock-in efficiency. Here, we investigate CTS design for AsCas12a Ultra by exploring PAM orientation and binding requirements. Additionally, we rule out ssDNase activity of AsCas12a under cell-physiological Mg2(+) conditions. Finally, we showcase the advantage of ssDNA donors with CTS (ssCTS) at high doses for delivering clinically relevant transgenes of varying sizes into three TCR-CD3 complex genes (TRAC, CD3ζ, CD3ε), achieving up to 90% knock-in rates for a 0.8kb-insert at the CD3ε locus. Long-read sequencing confirmed higher HDR rates and revealed that CTS reduced partial integration events compared to unmodified ssDNA. Overall, AsCas12a and ssCTS represent a platform for highly efficient knock-in in primary human T cells with minimal toxicity
Butyrylcholinesterase and 24 h-urinary copper excretion as compliance assessment in long-term treated Wilson's disease
Get PDFBACKGROUND AND AIM OF THE STUDY: Compliance is the most challenging aspect of long-term therapy in Wilson's disease (WD). Evidence is presented that butyrylcholinesterase (CHE) can be used as a sensitive biomarker to detect compliance problems in long-term treated WD-patients. METHODS: For the present retrospective, monocentric study demographical and treatment related data of 108 WD-patients (who had been treated at the Clinic of Neurology of the university hospital in Düsseldorf (Germany) between 2/2005 and 1/2021) were extracted from the charts. These patients underwent 2003 therapy control visits. The present study focuses on the analysis of three parameters of copper metabolism (serum levels of ceruloplasmin (CER-S), copper (CU-S) and the 24 h-urinary copper excretion (24 h-UCU)) and the serum levels of CHE (CHE-S). A patient was classified to be non-compliant when in his charts at least 8 24 h-UCU-values were found, and all his 24 h-UCU-values were larger than 60 μg/d (N-COM8-group). A patient was classified to be compliant when at least one of at least 8 24 h-UCU-values was lower than or equal to 60 μg/d (COM8-group). RESULTS: CHE-S was significantly (p < 0.05) different between thus defined compliant or non-compliant patients. Neither CU-S nor CER-S nor calculated free (non-ceruloplasmin-bound) serum copper levels (NCC-S) were significantly different between the NCOM8- and COM8-group. Analysis of the area under the curve and sensitivity and specificity by means of ROC-curves underlined the sensitivity of CHE-S in contrast to the insensitivity of CU-S and CER-S to detect patients who had been classified as compliant/non-compliant on the basis of their 24 h-UCU-values. CONCLUSION: When compliance of WD-patients is classified on the basis of their 24 h-urinary copper excretion CHE-S is more sensitive to detect problems of non-compliance than serum levels of copper, of non-ceruloplasmin bound free copper or ceruloplasmin. Therefore, CHE-S may be used as an easy to determine further biomarker for compliance assessment in long-term treatment of WD in addition to 24 h-UCU
Endogenous SNAP-tagging of Munc13-1 for monitoring synapse nanoarchitecture
Get PDFSynaptic function is governed by highly regulated protein machineries, whose abundance and spatial localization change continually. Studies to determine dynamic changes in synaptic protein nanoarchitecture typically rely on immunolabeling or on the expression of fluorescent proteins. The former employs chemical fluorophores and signal amplification but requires fixation. The latter enables monitoring of proteins by live microscopy but uses suboptimal fluorophores. Self-labeling tags have been introduced to combine the advantages of these two approaches, and here we introduce a knock-in mouse line where the essential presynaptic protein Munc13–1 is endogenously fused to the self-labeling SNAP tag. We demonstrate efficient Munc13–1-SNAP labeling in fixed cultured neurons and in brain sections by various SNAP dyes, as well as by a novel far-red and cell impermeable compound, SBG-SiR-d12. We introduce and characterize SBG-SiR-d12 as a highly efficient dye for SNAP-tag labeling of extracellular epitopes and of intracellular proteins such as Munc13–1 in fixed and permeabilized tissue. Finally, we show that Munc13–1-SNAP can be labeled in living neurons and monitored through live-cell imaging using confocal and super resolution microscopy. We conclude that the Unc13a(SNAP) mouse line is a useful tool for labeling the presynaptic compartment and for the analysis of presynaptic nanoarchitectural dynamics, with potential for wide adoption
Hypertensive Erkrankungen in der Schwangerschaft (HES): Diagnostik und Therapie. Leitlinie der DGGG, OEGGG und SGGG (S2k-Level, AWMF-Registernummer 015/018, Juni 2024) [Hypertensive Disorders in Pregnancy (HDP): Diagnostics and Therapy. Guideline of the DGGG, OEGGG and SGGG (S2k-Level, AWMF Registry No. 015/018, June 2024)]
No full textAIM: This S2k guideline of the German Society of Gynecology and Obstetrics (DGGG) contains consensus-based recommendations for the care and treatment of women with hypertension in pregnancy. It aims to serve as a guide for all professions involved in the care of pregnant women and to improve interprofessional and interdisciplinary cooperation. A new focus was placed on patientsʼ long-term health beyond the postpartum period. METHODS: The existing S2k guideline was revised and the relevant literature reviewed. Where new questions arose, they were formulated and developed in PICO format. A targeted systematic literature search was carried out using PubMed. Other international guidelines were also consulted. After summarizing and presenting the available data, recommendations and statements were developed, discussed, and agreed on by the guideline group. RECOMMENDATIONS: The recommendations cover prediction, prevention, diagnosis, and treatment from the moment hypertensive disease is detected in pregnancy as well as postpartum, in the puerperium, and during breastfeeding. A major change from the previous version of the guideline is the reduction in blood pressure levels that should be achieved during pregnancy. Suggestions are made on how to proceed with regards to the long-term health of mother and child, which the guideline group believes is currently regulated inadequately in the German healthcare system