{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,2,28]],"date-time":"2026-02-28T04:28:45Z","timestamp":1772252925235,"version":"3.50.1"},"reference-count":54,"publisher":"MDPI AG","issue":"11","license":[{"start":{"date-parts":[[2021,11,22]],"date-time":"2021-11-22T00:00:00Z","timestamp":1637539200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"the Japanese Ministry of Education, Culture, Sports, Science, and Technology","award":["23500428, 23659458, 24390223, 26461272, and 26430054"],"award-info":[{"award-number":["23500428, 23659458, 24390223, 26461272, and 26430054"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Toxins"],"abstract":"<jats:p>All the botulinum type A neurotoxins available for clinical use are of the A1 subtype. We developed a subtype A2 low-molecular-weight (150 kD (kilo Dalton)) neurotoxin (A2NTX) with less spread and faster entry into the motor nerve terminal than A1 in vitro and in vivo. Preliminary clinical studies showed that its efficacy is superior to A1 toxins. We conducted an open study exploring its safety and tolerability profile in comparison with A1LL (LL type A1 toxin, or onabotulinumtoxinA) and a low-molecular-weight (150 kD) A1 neurotoxin (A1NTX). Those who had been using A1LL (n = 90; 50\u2013360 mouse LD50 units) or A1NTX (n = 30; 50\u2013580 units) were switched to A2NTX (n = 120; 25\u2013600 units) from 2010 to 2018 (number of sessions ~27, cumulative doses ~11,640 units per patient). The adverse events for A2NTX included weakness (n = 1, ascribed to alcoholic polyneuropathy), dysphagia (1), local weakness (4), and spread to other muscles (1), whereas those for A1LL or A1NTX comprised weakness (n = 2, A1NTX), dysphagia (8), ptosis (6), local weakness (7), and spread to other muscles (15). After injections, 89 out of 120 patients preferred A2NTX to A1 for the successive sessions. The present study demonstrated that A2NTX had clinical safety up to the dose of 500 units and was well tolerated compared to A1 toxins.<\/jats:p>","DOI":"10.3390\/toxins13110824","type":"journal-article","created":{"date-parts":[[2021,12,1]],"date-time":"2021-12-01T03:12:40Z","timestamp":1638328360000},"page":"824","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Clinical Safety and Tolerability of A2NTX, a Novel Low-Molecular-Weight Neurotoxin Derived from Botulinum Neurotoxin Subtype A2, in Comparison with Subtype A1 Toxins"],"prefix":"10.3390","volume":"13","author":[{"given":"Toshiaki","family":"Takeuchi","sequence":"first","affiliation":[{"name":"Department of Clinical Neuroscience, Graduate School of Medicine, Tokushima University, Tokushima 770-8503, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Tsuyoshi","family":"Okuno","sequence":"additional","affiliation":[{"name":"Department of Clinical Neuroscience, Graduate School of Medicine, Tokushima University, Tokushima 770-8503, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ai","family":"Miyashiro","sequence":"additional","affiliation":[{"name":"Department of Clinical Neuroscience, Graduate School of Medicine, Tokushima University, Tokushima 770-8503, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Tomoko","family":"Kohda","sequence":"additional","affiliation":[{"name":"Department of Veterinary Sciences, School of Life and Environmental Sciences, Osaka Prefecture University, Osaka 598-8531, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6434-9541","authenticated-orcid":false,"given":"Ryosuke","family":"Miyamoto","sequence":"additional","affiliation":[{"name":"Department of Clinical Neuroscience, Graduate School of Medicine, Tokushima University, Tokushima 770-8503, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yuishin","family":"Izumi","sequence":"additional","affiliation":[{"name":"Department of Clinical Neuroscience, Graduate School of Medicine, Tokushima University, Tokushima 770-8503, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Shunji","family":"Kozaki","sequence":"additional","affiliation":[{"name":"Department of Veterinary Sciences, School of Life and Environmental Sciences, Osaka Prefecture University, Osaka 598-8531, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6207-1080","authenticated-orcid":false,"given":"Ryuji","family":"Kaji","sequence":"additional","affiliation":[{"name":"Department of Clinical Neuroscience, Graduate School of Medicine, Tokushima University, Tokushima 770-8503, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2021,11,22]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"282","DOI":"10.1016\/j.pmrj.2010.02.006","article-title":"Safety considerations in the use of botulinum toxins in children with cerebral palsy","volume":"2","author":"Apkon","year":"2010","journal-title":"PM&R"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"446","DOI":"10.1016\/j.tins.2005.06.001","article-title":"Beyond BOTOX: Advantages and limitations of individual botulinum neurotoxins","volume":"28","author":"Davletov","year":"2005","journal-title":"Trends Neurosci."},{"key":"ref_3","first-page":"236","article-title":"Botulin\u2013the \u201cweapon of noiseless warfare\u201d","volume":"37","author":"Dolder","year":"1960","journal-title":"Vierteljahrsschr. Schweiz. Sanitatsoff."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"213","DOI":"10.1128\/jb.53.2.213-230.1947","article-title":"Practical media and control measures for producing highly toxic cultures of Clostridium botulinum, Type A","volume":"53","author":"Lewis","year":"1947","journal-title":"J. Bacteriol."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"502","DOI":"10.7861\/clinmedicine.1-6-502","article-title":"Biological weapons: The facts not the fiction","volume":"1","author":"Lamb","year":"2001","journal-title":"Clin. Med."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"D33","DOI":"10.1016\/j.vaccine.2009.08.059","article-title":"Botulism and vaccines for its prevention","volume":"27","author":"Smith","year":"2009","journal-title":"Vaccine"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1128\/jb.52.1.1-13.1946","article-title":"Botulinum toxin (type A); including a study of shaking with chloroform as a step in the isolation procedure","volume":"52","author":"Lamanna","year":"1946","journal-title":"J. Bacteriol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"613","DOI":"10.1126\/science.103.2681.613","article-title":"The purification and crystallization of Clostridium botulinum type A toxin","volume":"103","author":"Lamanna","year":"1946","journal-title":"Science"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"11","DOI":"10.1080\/10408410590912952","article-title":"Clostridium botulinum: A bug with beauty and weapon","volume":"31","author":"Shukla","year":"2005","journal-title":"Crit. Rev. Microbiol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"49","DOI":"10.1007\/164_2019_311","article-title":"Manufacturing and Clinical Formulations of Botulinum Neurotoxins","volume":"263","author":"Hasan","year":"2020","journal-title":"Handb. Exp. Pharmacol."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"469","DOI":"10.1016\/j.clindermatol.2003.11.003","article-title":"History of the clinical use of botulinum toxin A and B","volume":"21","author":"Carruthers","year":"2003","journal-title":"Clin. Dermatol."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"S2","DOI":"10.1002\/mds.20003","article-title":"Historical notes on botulism, Clostridium botulinum, botulinum toxin, and the idea of the therapeutic use of the toxin","volume":"19","author":"Erbguth","year":"2004","journal-title":"Mov. Disord."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"435","DOI":"10.1097\/01.icu.0000243018.97627.4c","article-title":"Use of botulinum toxin in strabismus","volume":"17","author":"Crouch","year":"2006","journal-title":"Curr. Opin. Ophthalmol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"559","DOI":"10.1007\/s00702-007-0728-2","article-title":"From poison to remedy: The chequered history of botulinum toxin","volume":"115","author":"Erbguth","year":"2008","journal-title":"J. Neural Transm."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"258","DOI":"10.7861\/clinmedicine.4-3-258","article-title":"The story of Clostridium botulinum: From food poisoning to Botox","volume":"4","author":"Ting","year":"2004","journal-title":"Clin. Med."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"1353","DOI":"10.1002\/mds.22157","article-title":"Long-term treatment with botulinum toxin type A in cervical dystonia has low immunogenicity by mouse protection assay","volume":"23","author":"Brin","year":"2008","journal-title":"Mov. Disord."},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Yin, L., Masuyer, G., Zhang, S., Zhang, J., Miyashita, S.I., Burgin, D., Lovelock, L., Coker, S.F., Fu, T.M., and Stenmark, P. (2020). Characterization of a membrane binding loop leads to engineering botulinum neurotoxin B with improved therapeutic efficacy. PLoS Biol., 18.","DOI":"10.1371\/journal.pbio.3000618"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"628","DOI":"10.1002\/mus.20033","article-title":"Botulinum toxins in neurological disease","volume":"29","author":"Comella","year":"2004","journal-title":"Muscle Nerve"},{"key":"ref_19","first-page":"19","article-title":"Structure and function of botulinum toxin","volume":"70","author":"Fujii","year":"1995","journal-title":"Hokkaido Igaku Zasshi"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"S77","DOI":"10.1016\/S0151-9638(09)74532-6","article-title":"Different botulinum toxins and their specifications","volume":"136","author":"Beylot","year":"2009","journal-title":"Ann. Dermatol. Venereol."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"145","DOI":"10.1007\/BF03033933","article-title":"Botulinum toxin therapy for cervical dystonia","volume":"9","author":"Jankovic","year":"2006","journal-title":"Neurotox. Res."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"767","DOI":"10.1111\/j.1348-0421.1995.tb03269.x","article-title":"Immunological characterization of the neurotoxin produced by Clostridium botulinum type A associated with infant botulism in Japan","volume":"39","author":"Kozaki","year":"1995","journal-title":"Microbiol. Immunol."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"753","DOI":"10.1023\/A:1020677417356","article-title":"Molecular composition of progenitor toxin produced by Clostridium botulinum type C strain 6813","volume":"18","author":"Watanabe","year":"1999","journal-title":"J. Protein Chem."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"733","DOI":"10.1016\/j.jmb.2006.07.040","article-title":"A structural perspective of the sequence variability within botulinum neurotoxin subtypes A1\u2013A4","volume":"362","author":"Arndt","year":"2006","journal-title":"J. Mol. Biol."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"5450","DOI":"10.1128\/IAI.73.9.5450-5457.2005","article-title":"Sequence variation within botulinum neurotoxin serotypes impacts antibody binding and neutralization","volume":"73","author":"Smith","year":"2005","journal-title":"Infect. Immun."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"199","DOI":"10.1016\/j.febslet.2010.11.045","article-title":"Botulinum neurotoxin subtype A2 enters neuronal cells faster than subtype A1","volume":"585","author":"Pier","year":"2011","journal-title":"FEBS Lett."},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Peck, M.W., Smith, T.J., Anniballi, F., Austin, J.W., Bano, L., Bradshaw, M., Cuervo, P., Cheng, L.W., Derman, Y., and Dorner, B.G. (2017). Historical perspectives and guidelines for botulinum neurotoxin subtype nomenclature. Toxins, 9.","DOI":"10.3390\/toxins9010038"},{"key":"ref_28","doi-asserted-by":"crossref","unstructured":"Davies, J.R., Masuyer, G., and Stenmark, P. (2020). Structural and biochemical characterization of botulinum neurotoxin subtype B2 binding to its receptors. Toxins, 12.","DOI":"10.3390\/toxins12090603"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"215","DOI":"10.1007\/s00216-012-5767-3","article-title":"De novo subtype and strain identification of botulinum neurotoxin type B through toxin proteomics","volume":"403","author":"Kalb","year":"2012","journal-title":"Anal. Bioanal. Chem."},{"key":"ref_30","doi-asserted-by":"crossref","unstructured":"Fonfria, E., Maignel, J., Lezmi, S., Martin, V., Splevins, A., Shubber, S., Kalinichev, M., Foster, K., Picaut, P., and Krupp, J. (2018). The expanding therapeutic utility of botulinum neurotoxins. Toxins, 10.","DOI":"10.3390\/toxins10050208"},{"key":"ref_31","doi-asserted-by":"crossref","unstructured":"Kull, S., Schulz, K.M., Weisemann, J., Kirchner, S., Schreiber, T., Bollenbach, A., Dabrowski, P.W., Nitsche, A., Kalb, S.R., and Dorner, M.B. (2015). Isolation and functional characterization of the novel Clostridium botulinum neurotoxin A8 subtype. PLoS ONE, 10.","DOI":"10.1371\/journal.pone.0116381"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"506","DOI":"10.1212\/WNL.45.3.506","article-title":"Comparison of therapeutic efficacies of type A and F botulinum toxins for blepharospasm: A double-blind, controlled study","volume":"45","author":"Mezaki","year":"1995","journal-title":"Neurology"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"1017","DOI":"10.1002\/1531-8257(199911)14:6<1017::AID-MDS1018>3.0.CO;2-3","article-title":"Combined use of type A and F botulinum toxins for blepharospasm: A double-blind controlled trial","volume":"14","author":"Mezaki","year":"1999","journal-title":"Mov. Disord."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"3149","DOI":"10.1001\/jama.1992.03480230041009","article-title":"The lethal botulinum toxic shot syndrome","volume":"267","author":"Walker","year":"1992","journal-title":"JAMA"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"231","DOI":"10.1016\/0168-1605(90)90016-X","article-title":"Distinct characters of Clostridium botulinum type A strains and their toxin associated with infant botulism in Japan","volume":"11","author":"Sakaguchi","year":"1990","journal-title":"Int. J. Food Microbiol."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"547","DOI":"10.1016\/0923-2508(93)90004-L","article-title":"Sequence of the gene coding for the neurotoxin of Clostridium botulinum type A associated with infant botulism: Comparison with other clostridial neurotoxins","volume":"144","author":"Willems","year":"1993","journal-title":"Res. Microbiol."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"275","DOI":"10.1254\/jphs.11121FP","article-title":"Type A1 but not type A2 botulinum toxin decreases the grip strength of the contralateral foreleg through axonal transport from the toxin-treated foreleg of rats","volume":"117","author":"Torii","year":"2011","journal-title":"J. Pharmacol. Sci."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"3894","DOI":"10.1128\/IAI.00536-13","article-title":"Characterization of botulinum neurotoxin A subtypes 1 through 5 by investigation of activities in mice, in neuronal cell cultures, and in vitro","volume":"81","author":"Whitemarsh","year":"2013","journal-title":"Infect. Immun."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"502","DOI":"10.1111\/1348-0421.12796","article-title":"Translocation domain of botulinum neurotoxin A subtype 2 potently induces entry into neuronal cells","volume":"64","author":"Kohda","year":"2020","journal-title":"Microbiol. Immunol."},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"32","DOI":"10.1016\/j.toxicon.2013.12.012","article-title":"Comparison between botulinum neurotoxin type A2 and type A1 by electrophysiological study in healthy individuals","volume":"81","author":"Mukai","year":"2014","journal-title":"Toxicon"},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"85","DOI":"10.1016\/j.toxicon.2015.09.025","article-title":"Clinical differences between A1 and A2 botulinum toxin subtypes","volume":"107","author":"Kaji","year":"2015","journal-title":"Toxicon"},{"key":"ref_42","first-page":"510","article-title":"Comparative study of spread of A1 and A2 subtypes of botulinum toxin preparations for blepharospasm: Proof-ofconcept randomized controlled trial","volume":"30","author":"Kaji","year":"2015","journal-title":"Mov. Disord."},{"key":"ref_43","first-page":"84","article-title":"Randomised double-blind clinical trial of botulinum toxin subtype A2 (A2NTX) in comparison with subtype A1 (onabotulinumtoxinA) (ClinicalTrials.gov NCT01910363) (P4.333)","volume":"14","author":"Kaji","year":"2015","journal-title":"Neurology"},{"key":"ref_44","first-page":"225","article-title":"Survey of practices employed by neurologists for the definition and management of secondary non-response to botulinum toxin in cervical dystonia","volume":"27","author":"Ferreira","year":"2012","journal-title":"Funct. Neurol."},{"key":"ref_45","first-page":"5","article-title":"Classification and mechanism of adverse drug reaction","volume":"65","author":"Shimizu","year":"2007","journal-title":"Nihon Rinsho"},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"93","DOI":"10.1016\/j.toxicon.2010.10.009","article-title":"Comparison of effects of botulinum toxin subtype A1 and A2 using twitch tension assay and rat grip strength test","volume":"57","author":"Torii","year":"2011","journal-title":"Toxicon"},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"524","DOI":"10.1111\/bcpt.12351","article-title":"Comparison of systemic toxicity between botulinum toxin subtypes A1 and A2 in mice and rats","volume":"116","author":"Torii","year":"2015","journal-title":"Basic Clin. Pharmacol. Toxicol."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"114","DOI":"10.1016\/j.toxicon.2018.08.017","article-title":"Comparison of efficacy and toxicity between botulinum toxin subtypes A1 and A2 in cynomolgus macaques","volume":"153","author":"Torii","year":"2018","journal-title":"Toxicon"},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"98","DOI":"10.3389\/fneur.2014.00098","article-title":"Spinal central effects of peripherally applied botulinum neurotoxin A in comparison between its subtypes A1 and A2","volume":"5","author":"Koizumi","year":"2014","journal-title":"Front. Neurol."},{"key":"ref_50","first-page":"216","article-title":"Falls in stroke patients: Risk factors and risk management","volume":"12","author":"Tsur","year":"2010","journal-title":"Isr. Med. Assoc. J."},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"368","DOI":"10.1136\/jnnp.2004.038166","article-title":"Ten year recurrence after first ever stroke in a Japanese community: The Hisayama study","volume":"76","author":"Hata","year":"2005","journal-title":"J. Neurol. Neurosurg. Psychiatry"},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1007\/s40268-014-0077-1","article-title":"Pharmaceutical, biological, and clinical properties of botulinum neurotoxin type A products","volume":"15","author":"Frevert","year":"2015","journal-title":"Drugs R&D"},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"e00795-16","DOI":"10.1128\/IAI.00795-16","article-title":"Entry of botulinum neurotoxin subtypes A1 and A2 into neurons","volume":"85","author":"Kroken","year":"2017","journal-title":"Infect. Immun."},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"9","DOI":"10.1016\/j.toxicon.2013.06.020","article-title":"Differences in immunological responses of polyclonal botulinum A1 and A2 antitoxin against A1 and A2 toxin","volume":"73","author":"Torii","year":"2013","journal-title":"Toxicon"}],"container-title":["Toxins"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2072-6651\/13\/11\/824\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T07:34:06Z","timestamp":1760168046000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2072-6651\/13\/11\/824"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,11,22]]},"references-count":54,"journal-issue":{"issue":"11","published-online":{"date-parts":[[2021,11]]}},"alternative-id":["toxins13110824"],"URL":"https:\/\/doi.org\/10.3390\/toxins13110824","relation":{"has-preprint":[{"id-type":"doi","id":"10.20944\/preprints202110.0121.v1","asserted-by":"object"}]},"ISSN":["2072-6651"],"issn-type":[{"value":"2072-6651","type":"electronic"}],"subject":[],"published":{"date-parts":[[2021,11,22]]}}}