The preclinical development of microRNA-based gene therapies for inherited neurodegenerative diseases is accompanied by translational challenges. Due to the inaccessibility of the brain to... Show moreThe preclinical development of microRNA-based gene therapies for inherited neurodegenerative diseases is accompanied by translational challenges. Due to the inaccessibility of the brain to periodically evaluate therapy effects, accessible and reliable biomarkers indicative of dosing, durability and therapeutic efficacy in the central nervous system are very much needed. This is particularly important for viral vector-based gene therapies, in which a one-time administration results in long-term expression of active therapeutic molecules in the brain. Recently, extracellular vesides have been identified as carriers of RNA species, including microRNAs, and proteins in all biological fluids, whilst becoming potential sources of biomarkers for diagnosis. In this study, we investigated the secretion and potential use of circulating miRNAs associated with extracellular vesicles as suitable sources to monitor the expression and durability of gene therapies in the brain. Neuronal cells derived from induced pluripotent stern cells were treated with adeno-associated viral vector serotype 5 carrying an engineered microRNA targeting huntingtin or ataxin3 gene sequences, the diseases-causing genes of Huntington disease and spinocerebellar ataxia type 3, respectively. After treatment, the secretion of mature engineered microRNA molecules was confirmed, with extracellular microRNA levels correlating with viral dose and cellular microRNA expression in neurons. We further investigated the detection of engineered microRNAs over time in the CSF of non-human primates after a single intrastriatal injection of adeno-associated viral vector serotype 5 carrying a huntingtin-targeting engineered microRNA. Quantifiable engineered microRNA levels enriched in extracellular vesicles were detected in the CSF up to 2 years after brain infusion. Altogether, these results confirm the long-term expression of adeno-associated viral vector serotype 5-delivered microRNAs and support the use of extracellular vesicle-associated microRNAs as novel translational pharmacokinetic markers in ongoing clinical trials of gene therapies for neurodegenerative diseases. Show less
Spronck, E.A.; Valles, A.; Lampen, M.H.; Montenegro-Miranda, P.S.; Keskin, S.; Heijink, L.; ... ; Haan, M. de 2021
Huntington disease (HD) is a fatal, neurodegenerative genetic disorder with aggregation of mutant Huntingtin protein (mutHTT) in the brain as a key pathological mechanism. There are currently no... Show moreHuntington disease (HD) is a fatal, neurodegenerative genetic disorder with aggregation of mutant Huntingtin protein (mutHTT) in the brain as a key pathological mechanism. There are currently no disease modifying therapies for HD; however, HTT-lowering therapies hold promise. Recombinant adeno-associated virus serotype 5 expressing a microRNA that targets HTT mRNA (AAV5-miHTT) is in development for the treatment of HD with promising results in rodent and minipig HD models. To support a clinical trial, toxicity studies were performed in non-human primates (NHP, Macaca fascicularis) and Sprague-Dawley rats to evaluate the safety of AAV5-miHTT, the neurosurgical administration procedure, vector delivery and expression of the miHTT transgene during a 6-month observation period. For accurate delivery of AAV5-miHTT to the striatum, real-time magnetic resonance imaging (MRI) with convection-enhanced delivery (CED) was used in NHP. Catheters were successfully implanted in 24 NHP, without neurological symptoms, and resulted in tracer signal in the target areas. Widespread vector DNA and miHTT transgene distribution in the brain was found, particularly in areas associated with HD pathology. Intrastriatal administration of AAV5-miHTT was well tolerated with no clinically relevant changes in either species. These studies demonstrate the excellent safety profile of AAV5-miHTT, the reproducibility and tolerability of intrastriatal administration, and the delivery of AAV5-miHTT to the brain, which support the transition of AAV5-miHTT into clinical studies. Show less
Main recommendations1. ESGE/ESGAR recommend computed tomographic colonography (CTC) as the radiological examination of choice for the diagnosis of colorectal neoplasia. Strong recommendation, high... Show moreMain recommendations1. ESGE/ESGAR recommend computed tomographic colonography (CTC) as the radiological examination of choice for the diagnosis of colorectal neoplasia. Strong recommendation, high quality evidence. ESGE/ESGAR do not recommend barium enema in this setting. Strong recommendation, high quality evidence.2. ESGE/ESGAR recommend CTC, preferably the same or next day, if colonoscopy is incomplete. The timing depends on an interdisciplinary decision including endoscopic and radiological factors. Strong recommendation, low quality evidence. ESGE/ESGAR suggests that, in centers with expertise in and availability of colon capsule endoscopy (CCE), CCE preferably the same or the next day may be considered if colonoscopy is incomplete. Weak recommendation, low quality evidence.3. When colonoscopy is contraindicated or not possible, ESGE/ESGAR recommend CTC as an acceptable and equally sensitive alternative for patients with alarm symptoms. Strong recommendation, high quality evidence. Because of lack of direct evidence, ESGE/ESGAR do not recommend CCE in this situation. Very low quality evidence. ESGE/ESGAR recommend CTC as an acceptable alternative to colonoscopy for patients with non-alarm symptoms. Strong recommendation, high quality evidence. In centers with availability, ESGE/ESGAR suggests that CCE may be considered in patients with non-alarm symptoms. Weak recommendation, low quality evidence.4. Where there is no organized fecal immunochemical test (FIT)-based population colorectal screening program, ESGE/ESGAR recommend CTC as an option for colorectal cancer screening, providing the screenee is adequately informed about test characteristics, benefits, and risks, and depending on local service- and patient-related factors. Strong recommendation, high quality evidence. ESGE/ESGAR do not suggest CCE as a first-line screening test for colorectal cancer. Weak recommendation, low quality evidence.5. ESGE/ESGAR recommend CTC in the case of a positive fecal occult blood test (FOBT) or FIT with incomplete or unfeasible colonoscopy, within organized population screening programs. Strong recommendation, moderate quality evidence. ESGE/ESGAR also suggest the use of CCE in this setting based on availability. Weak recommendation, moderate quality evidence.6. ESGE/ESGAR suggest CTC with intravenous contrast medium injection for surveillance after curative-intent resection of colorectal cancer only in patients in whom colonoscopy is contraindicated or unfeasible. Weak recommendation, low quality evidence. There is insufficient evidence to recommend CCE in this setting. Very low quality evidence.7. ESGE/ESGAR suggest CTC in patients with high risk polyps undergoing surveillance after polypectomy only when colonoscopy is unfeasible. Weak recommendation, low quality evidence. There is insufficient evidence to recommend CCE in post-polypectomy surveillance. Very low quality evidence.8. ESGE/ESGAR recommend against CTC in patients with acute colonic inflammation and in those who have recently undergone colorectal surgery, pending a multidisciplinary evaluation. Strong recommendation, low quality evidence. 9. ESGE/ESGAR recommend referral for endoscopic polypectomy in patients with at least one polyp >= 6 mm detected at CTC or CCE. Follow-up CTC may be clinically considered for 6-9-mm CTC-detected lesions if patients do not undergo polypectomy because of patient choice, comorbidity, and/or low risk profile for advanced neoplasia. Strong recommendation, moderate quality evidence.Source and scopeThis is an update of the 2014-15 Guideline of the European Society of Gastrointestinal Endoscopy (ESGE) and the European Society of Gastrointestinal and Abdominal Radiology (ESGAR). It addresses the clinical indications for the use of imaging alternatives to standard colonoscopy. A targeted literature search was performed to evaluate the evidence supporting the use of computed tomographic colonography (CTC) or colon capsule endoscopy (CCE). The Grading of Recommendations Assessment, Development and Evaluation (GRADE) system was adopted to define the strength of recommendations and the quality of evidence. Show less
Main recommendations1 ESGE/ESGAR recommend computed tomographic colonography (CTC) as the radiological examination of choice for the diagnosis of colorectal neoplasia.Strong recommendation, high... Show moreMain recommendations1 ESGE/ESGAR recommend computed tomographic colonography (CTC) as the radiological examination of choice for the diagnosis of colorectal neoplasia.Strong recommendation, high quality evidence.ESGE/ESGAR do not recommend barium enema in this setting.Strong recommendation, high quality evidence.2 ESGE/ESGAR recommend CTC, preferably the same or next day, if colonoscopy is incomplete. The timing depends on an interdisciplinary decision including endoscopic and radiological factors.Strong recommendation, low quality evidence.ESGE/ESGAR suggests that, in centers with expertise in and availability of colon capsule endoscopy (CCE), CCE preferably the same or the next day may be considered if colonoscopy is incomplete.Weak recommendation, low quality evidence.3 When colonoscopy is contraindicated or not possible, ESGE/ESGAR recommend CTC as an acceptable and equally sensitive alternative for patients with alarm symptoms.Strong recommendation, high quality evidence.Because of lack of direct evidence, ESGE/ESGAR do not recommend CCE in this situation.Very low quality evidence.ESGE/ESGAR recommend CTC as an acceptable alternative to colonoscopy for patients with non-alarm symptoms.Strong recommendation, high quality evidence.In centers with availability, ESGE/ESGAR suggests that CCE may be considered in patients with non-alarm symptoms.Weak recommendation, low quality evidence.4 Where there is no organized fecal immunochemical test (FIT)-based population colorectal screening program, ESGE/ESGAR recommend CTC as an option for colorectal cancer screening, providing the screenee is adequately informed about test characteristics, benefits, and risks, and depending on local service- and patient-related factors.Strong recommendation, high quality evidence.ESGE/ESGAR do not suggest CCE as a first-line screening test for colorectal cancer.Weak recommendation, low quality evidence.5 ESGE/ESGAR recommend CTC in the case of a positive fecal occult blood test (FOBT) or FIT with incomplete or unfeasible colonoscopy, within organized population screening programs.Strong recommendation, moderate quality evidence.ESGE/ESGAR also suggest the use of CCE in this setting based on availability.Weak recommendation, moderate quality evidence.6 ESGE/ESGAR suggest CTC with intravenous contrast medium injection for surveillance after curative-intent resection of colorectal cancer only in patients in whom colonoscopy is contraindicated or unfeasibleWeak recommendation, low quality evidence.There is insufficient evidence to recommend CCE in this setting.Very low quality evidence.7 ESGE/ESGAR suggest CTC in patients with high risk polyps undergoing surveillance after polypectomy only when colonoscopy is unfeasible.Weak recommendation, low quality evidence.There is insufficient evidence to recommend CCE in post-polypectomy surveillance.Very low quality evidence.8 ESGE/ESGAR recommend against CTC in patients with acute colonic inflammation and in those who have recently undergone colorectal surgery, pending a multidisciplinary evaluation.Strong recommendation, low quality evidence.9 ESGE/ESGAR recommend referral for endoscopic polypectomy in patients with at least one polyp ≥ 6 mm detected at CTC or CCE.Follow-up CTC may be clinically considered for 6 – 9-mm CTC-detected lesions if patients do not undergo polypectomy because of patient choice, comorbidity, and/or low risk profile for advanced neoplasia.Strong recommendation, moderate quality evidence. Show less
Gene therapy for severe hemophilia B is advancing and offers sustained disease amelioration with a single treatment. We have reported the efficacy and safety of AMT-060, an investigational gene... Show moreGene therapy for severe hemophilia B is advancing and offers sustained disease amelioration with a single treatment. We have reported the efficacy and safety of AMT-060, an investigational gene therapy comprising an adeno-associated virus serotype 5 capsid encapsidating the codon-optimized wild-type human factor IX (WT hFIX) gene with a liver-specific promoter, in patients with severe hemophilia B. Treatment with 2 x 10(13) gc/kg AMT-060 showed sustained and durable FIX activity of 3%-13% and a substantial reduction in spontaneous bleeds without T cell-mediated hepatoxicity. To achieve higher FIX activity, we modified AMT-060 to encode the R338L "Padua" FIX variant that has increased specific activity (AMT-061). We report the safety and increased FIX activity ofAMT-061 in non-human primates. Animals (n = 3/group) received intravenous AMT-060 (5 x 10(12) gc/kg), AMT-061 (ranging from 5 x 10(11) to 9 x 10(13) gc/kg), or vehicle. Human FIX protein expression, FIX activity, and coagulation markers including D-dimer and thrombin-antithrombin complexes were measured. At equal doses, AMT-060 and AMT-061 resulted in similar human FIX protein expression, but FIX activity was 6.5-fold enhanced using AMT-061. Both vectors show similar safety and transduction profiles. Thus, AMT-061 holds great promise as a more potent FIX replacement gene therapy with a favorable safety profile. Show less
Zoest, R.A. van; Law, M.; Sabin, C.A.; Vaartjes, I.; Valk, M. van der; Arends, J.E.; ... ; Elst-Laurijssen, D.H. 2019
Huntington disease (HD) is a fatal neurodegenerative disorder caused by an autosomal dominant CAG repeat expansion in the huntingtin (HTT) gene. The translated expanded polyglutamine repeat in the... Show moreHuntington disease (HD) is a fatal neurodegenerative disorder caused by an autosomal dominant CAG repeat expansion in the huntingtin (HTT) gene. The translated expanded polyglutamine repeat in the HTT protein is known to cause toxic gain of function. We showed previously that strong HTT lowering prevented neuronal dysfunction in HD rodents and minipigs after single intracranial injection of adeno-associated viral vector serotype 5 expressing a microRNA targeting human HTT (AAV5-miHTT). To evaluate long-term efficacy, AAV5-miHTT was injected into the striatum of knockin Q175 HD mice, and the mice were sacrificed 12 months post-injection. AAV5-miHTT caused a dose-dependent and sustained HTT protein reduction with subsequent suppression of mutant HTT aggregate formation in the striatum and cortex. Functional proof of concept was shown in transgenic R6/2 HD mice. Eight weeks after AAV5-miHTT treatment, a significant improvement in motor coordination on the rotarod was observed. Survival analysis showed that a single AAV5-miHTT treatment resulted in a significant 4-week increase in median survival compared with vehicle-treated R6/2 HD mice. The combination of long-term HTT lowering, reduction in aggregation, prevention of neuronal dysfunction, alleviation of HD-like symptoms, and beneficial survival observed in HD rodents treated with AAV5-miHTT supports the continued development of HTT-lowering gene therapies for HD. Show less
