Reducing huntingtin by immunotherapy delays disease progression in a mouse model of Huntington disease.

In Huntington disease (HD), the mutant huntingtin (mtHTT) protein is the principal cause of pathological changes that initiate primarily along the cortico-striatal axis. mtHTT is ubiquitously expressed and there is, accordingly, growing recognition that HD is a systemic disorder with functional interplay between the brain and the periphery. We have developed a monoclonal antibody, C6-17, targeting an exposed region of HTT near the aa586 Caspase 6 cleavage site. As recently published, mAB C6-17 can block cell-to-cell propagation of mtHTT in vitro. In order to reduce the burden of the mutant protein in vivo, we queried whether extracellular mtHTT could be therapeutically targeted in YAC128 HD mice. In a series of proof of concept experiments, we found that systemic mAB C6-17 treatment resulted in the distribution of the mAB C6-17 to peripheral and CNS tissues and led to the reduction of HTT protein levels. Compared to CTRL mAB or vehicle treated mice, the mAB C6-17 treated YAC128 animals showed improved body weight and motor behaviors, a delayed progression in motor deficits and reduced striatal EM48 immunoreactivity. These results provide the first proof of concept for the feasibility and therapeutic efficacy of an antibody-based anti-HTT passive immunization approach and suggest this modality as a potential new HD treatment strategy.

Outcome of Budd-Chiari Syndrome Patients Treated With Direct Oral Anticoagulants: An Austrian Multicenter Study.

BACKGROUND AND AIMS: Direct oral anticoagulants (DOACs) may simplify management of Budd-Chiari syndrome (BCS). Here, we report our experience with off-label use of DOACs for anticoagulation in BCS. METHODS: The safety of DOAC vs vitamin K antagonist treatment as well as associated clinical outcomes were retrospectively assessed in 47 BCS patients treated at 6 Austrian centers. RESULTS: Mean age at study inclusion was 37.9 ± 14.0 years and mean Model for End-Stage Liver Disease was 13.1 ± 5.1. Overall, 63.8% (n = 30) of patients had decompensated liver disease, and 87.2% (n = 41) showed clinical signs of portal hypertension. During a median follow-up of 82.5 (interquartile range, 43.1-121.8) months, 43 (91.5%) patients received anticoagulation alone or following interventional treatment, including 22 (46.8%) patients treated with DOACs (edoxaban: 10, apixaban: 4, rivaroxaban: 3, dabigatran: 3, more than one DOAC sequentially: 2) for a median of 24.4 (interquartile range, 5.7-35.1) months. While 72.7% (n = 16 of 22) of patients were switched from low-molecular-weight heparin (n = 12) or vitamin K antagonist (n = 4) to DOAC after disease stabilization or improvement, 27.3% (n = 6 of 22) of BCS patients were initially treated with DOAC. Complete response (European Association for the Study of the Liver criteria) was achieved or maintained in 14 (63.6%) of 22 patients, with ongoing response in 2 patients, while disease progressed in 6 patients (including 2 patients with hepatocellular carcinoma). Four major spontaneous bleedings (18.2%; incidence rate 8.8 per 100 patient-years; n = 2 upper gastrointestinal bleeding, n = 1 lower gastrointestinal bleeding, n = 1 hepatocellular carcinoma rupture), 7 minor bleedings, and 1 major procedure-related bleeding (4.5%; 2.2 per 100 patient-years) occurred during DOAC therapy. Overall transplant-free survival was 91.6% at 5 years. CONCLUSIONS: DOACs seem to be effective and safe for long-term anticoagulation in patients with BCS, but confirmation by larger prospective studies is needed.

Inhibiting cellular uptake of mutant huntingtin using a monoclonal antibody: Implications for the treatment of Huntington's disease.

Huntington's disease (HD) is caused by a highly polymorphic CAG trinucleotide expansion in the gene encoding for the huntingtin protein (HTT). The resulting mutant huntingtin protein (mutHTT) is ubiquitously expressed but also exhibits the ability to propagate from cell-to-cell to disseminate pathology; a property which may serve as a new therapeutic focus. Accordingly, we set out to develop a monoclonal antibody (mAB) targeting a particularly exposed region close to the aa586 caspase-6 cleavage site of the HTT protein. This monoclonal antibody, designated C6-17, effectively binds mutHTT and is able to deplete the protein from cell culture supernatants. Using cell-based assays, we demonstrate that extracellular secretion of mutHTT into cell culture media and its subsequent uptake in recipient HeLa cells can be almost entirely blocked by mAB C6-17. Immunohistochemical stainings of post-mortem HD brain tissue confirmed the specificity of mAB C6-17 to human mutHTT aggregates. These findings demonstrate that mAB C6-17 not only successfully engages with its target, mutHTT, but also inhibits cell uptake suggesting that this antibody could interfere with the pathological processes of mutHTT spreading in vivo.

Quantitative in vitro and in vivo models to assess human IgE B cell receptor crosslinking by IgE and EMPD IgE targeting antibodies.

Targeting plasma IgE by therapeutic mABs like Omalizumab (Xolair®) is current clinical practice for severe allergic conditions or other IgE related diseases like chronic urticaria. As an alternative to soluble IgE targeting, IgE supply can be lowered by targeting the Extracellular Membrane Proximal Domain (EMPD) of the IgE B cell receptor (BCR) present on IgE switched B cells. This ultimately leads to apoptosis of these cells upon IgE BCR crosslinking. Since tools to selectively assess the efficacy of IgE BCR crosslinking by IgE targeting antibodies are limited, a readily quantifiable cell model was developed that allows to specifically address IgE BCR crosslinking activity in vitro. The new cell model allowed for a direct quantitative comparison of anti-EMPD IgE therapeutic prototype antibody 47H4 with anti-IgE(Ce3) directed therapeutic antibody Omalizumab and with a newly selected anti-human EMPD IgE monoclonal antibody, designated mAB 15cl12. Furthermore, a complementing mouse model was developed that allows for in vivo validation of antibodies addressing human EMPD IgE. It carries a targetable humanized EMPD IgE sequence that has been introduced by seamless genomic replacement of the endogenous EMPD encoding sequence. The model allowed to directly compare IgE lowering activity of two anti-human EMPD IgE therapeutic antibodies in vivo. Our tools provide the means for quantitative assessment of IgE BCR crosslinking activity which is increasingly gaining attention with respect to forthcoming second generation anti-IgE clinical candidates such as Ligelizumab or other clinical candidates featuring additional effector functions such as IgE BCR crosslinking activity.

Activation of the mouse histone deacetylase 1 gene by cooperative histone phosphorylation and acetylation.

Histone deacetylase 1 (HDAC1) is a major regulator of chromatin structure and gene expression. Tight control of HDAC1 expression is essential for normal cell cycle progression of mammalian cells. HDAC1 mRNA levels are regulated by growth factors and by changes in intracellular deacetylase activity levels. Stimulation of the mitogen-activated protein kinase cascade by anisomycin or growth factors, together with inhibition of deacetylases by trichostatin A (TSA), leads to stable histone H3 phosphoacetylation and strongly induced HDAC1 expression. In contrast, activation of the nucleosomal response by anisomycin alone results only in transient phosphoacetylation of histone H3 without affecting HDAC1 mRNA levels. The transcriptional induction of the HDAC1 gene by anisomycin and TSA is efficiently blocked by H89, an inhibitor of the nucleosomal response. Detailed studies of the kinetics of histone acetylation and phosphorylation show that the two modifications are synergistic and essential for induced HDAC1 transcription. Activation of the HDAC1 gene by anisomycin together with TSA or by growth factors is accompanied by phosphoacetylation of HDAC1 promoter-associated histone H3. Our results present evidence for a precise regulatory mechanism which allows induction of the HDAC1 gene in response to proliferation signals and modulation of HDAC1 expression dependent on intracellular deacetylase levels.