At the end of the year, we are providing a BIND Project 2022 overview on the research and developments made in understanding the role of dystrophin in the brain. Below, you can read a summary of the work performed by the consortium, and a list of papers that were published. In short, the past year the BIND Project focused on the following activities:
- Behavioural and neuropsychological phenotyping of dystrophic mouse models lacking different brain isoforms & Impact of brain dystrophins restoration on dystrophic mice phenotype
- Localisation and interactome analysis of dystrophin isoforms in the mouse and human brain
- Impact of brain dystrophins restoration on dystrophic mice phenotype
- Deep functional phenotyping of Duchenne muscular dystrophy and Becker muscular dystrophy patients
- Structural integrity in the DMD and BMD brain and in dystrophic mdx mice
- Data integration and FAIRification
BIND Project 2022 overview
1. Behavioural and neuropsychological phenotyping of dystrophic mouse models lacking different brain isoforms & Impact of brain dystrophins restoration on dystrophic mice phenotype
WP3 and WP4 published a paper where is mentioned that exon 51 skipping with a PMO antisense oligonucleotide restores in-frame dystrophin, leading to improved muscle pathology and function in mdx52. In addition to this, they focused on the differences in dystrophin isoforms in mdx and mdx52 mice. Secondly, the group found out that fear and anxiety behaviours, measured using freezing response, are similar between mdx and mdx52, suggesting they are caused by absence of Dp427. Social behaviour testing showed increased social interaction in male mdx52 compared to wt and mdx23. These results implicate Dp140 in abnormal social behaviour.
The groups performed restoration of Dp140 to try and rescue abnormal mdx52 phenotypes. Exon 53 skipping and production of in frame DMD transcript was confirmed. The Dp140 overexpression resulted in normal glutamatergic transmission and normal social behaviour in mdx52 mice. The results of this study were published very recently.
2. Localisation and interactome analysis of dystrophin isoforms in the mouse and human brain
WP2 focused on the localisation of dystrophin isoforms in mouse and human brain. BaseScope probes targeting Dp427b, Dp427m, Dp427p2, Dp140, Dp71 and Dp40 have been used in quantitative and qualitative analysis of human brain, showing that there is different isoform expression in different areas. Progress on the mouse brain work included the optimisation of immunohistochemistry (IHC) detection of dystrophin using the antibody NCL-DYS1, and optimisation of the fixation methodology.
Characterisation of dystrophin protein complexes in the mouse brain are being performed. First experiments were done using the MANDYS1 antibody for immunoprecipitation and mass spec optimisation is ongoing.
3. Impact of brain dystrophins restoration on dystrophic mice phenotype
The WP4 group focused on optimizing the tools and delivery methods. PMO and tcDNA antisense oligonucleotides delivery were compared using different techniques, as well as alternative exon skipping tools. Regarding dystrophin protein restoration, these levels are lower in the mdx52 compared to the more commonly used mdx23. This is similar to discrepancy observed in the muscle of mdx52 using AAV-U7.
Behavioural outcomes validated in WP3 were used to assess impact of partial Dp427 rescue. The conclusion is that these deficits could be reversible in patients. The reason for only partial rescue is not related to the absence of dystrophin in muscle, but could be linked to absence of Dp140 or to insufficient protein restoration, or to postnatal treatment being too late.
4. Deep functional phenotyping of Duchenne muscular dystrophy and Becker muscular dystrophy patients
The Part 1 study involves online questionnaires: in REDCap and DAWBA. The total target number for all sites in the protocol is 800; over 200 participants have now completed Part 1. Sites have been implementing measures to improve recruitment. The more successful route has been to approach families in clinic. The genotype distribution is close to expectations, so no adjustments are needed at this stage.
Part 2 includes the in-clinic assessments and optional additional MRI (WP6). The total target number for all sites in the protocol is 300; 105 participants (35%) have now completed Part 2 and 193 (64%) have consented. Genotype breakdown of cohort that has completed Part 2 assessments is as expected and does not need adjustment at this stage. WP5 should be liaising closely with WP6 on recruiting patients for scanning.
5. Structural integrity in the DMD and BMD brain and in dystrophic mdx mice
UCL provided an update on the status of clinical MRI. UCL, UNEW and LUMC have started scanning patients. At UCL, 6-7 patients have been scanned, although two were moving so data cannot be used. LUMC have scanned 4-5 patients. UNEW have scanned at least 3 patients. RegionH (Copenhagen) are about to start scanning. Paris and Rome are not yet ready to scan. Regarding the preclinical MRI, all mice have been scanned.
6. Data integration and FAIRification
The main focus of WP7 is data integration through FAIRification of data. To achieve this goal and build on top of previous findings, the group has built knowledge graphs based on databases and text mining to represent previous knowledge related to the comorbidities that are observed in Duchenne and Becker patients. This step enabled to bring in previous knowledge on these comorbidities in the context of Duchenne and Becker and beyond. In 2022 we have assessed the value of the knowledge graphs using competency questions and by comparing the results of queries to lists compiled by domain experts. One example is the review of dystrophin interacting partners. Papers reporting on dystrophin interactors were shortlisted and compared to queries on knowledge graphs performed in. WP7. This allowed to identify genes related to comorbidities that are directly interacting with dystrophin.
Among the activities performed in 2022, Bring Your Own Data (BYOD) workshops were held for all WPs with the purpose of FAIRifying the BIND datasets,. In preparation for the WP5 patient phenotyping data being transferred to the Duchenne Data Repository, the questionnaire was also FAIRified. Data will be findable, accessible and reusable for users external to BIND. Data will be interoperable both within and outside BIND. The FAIRification of BIND datasets will facilitate data integration and analytics in WP7. Multiple partners were involved in the FAIRification process with LUMC leading the data FAIRification, while DDF and EBRAINS focused on the metadata.
List of BIND publications
- Partial Restoration of Brain Dystrophin and Behavioral Deficits by Exon Skipping in the Muscular Dystrophy X-Linked (mdx) Mouse. Zarrouki F, Relizani K, Bizot F, Tensorer T, Garcia L, Vaillend C, Goyenvalle A. Ann Neurol. 2022 Aug;92(2):213-229. doi: 10.1002/ana.26409.
- Brain Dp140 alters glutamatergic transmission and social behaviour in the mdx52 mouse model of Duchenne muscular dystrophy. Hashimoto Y, Kuniishi H, Sakai K, Fukushima Y, Du X, Yamashiro K, Hori K, Imamura M, Hoshino M, Yamada M, Araki T, Sakagami H, Takeda S, Itaka K, Ichinohe N, Muntoni F, Sekiguchi M, Aoki Y. Prog Neurobiol. 2022 Sep;216:102288. doi: 10.1016/j.pneurobio.2022.102288.
- Investigating the role of dystrophin isoform deficiency in motor function in Duchenne muscular dystrophy. Chesshyre M, Ridout D, Hashimoto Y, Ookubo Y, Torelli S, Maresh K, Ricotti V, Abbott L, Gupta VA, Main M, Ferrari G, Kowala A, Lin YY, Tedesco FS, Scoto M, Baranello G, Manzur A, Aoki Y, Muntoni F. J Cachexia Sarcopenia Muscle. 2022 Apr;13(2):1360-1372. doi: 10.1002/jcsm.12914.
- Startle responses in Duchenne muscular dystrophy: a novel biomarker of brain dystrophin deficiency. Maresh K, Papageorgiou A, Ridout D, Harrison NA, Mandy W, Skuse D, Muntoni F. Brain. 2022 Feb 8:awac048. doi: 10.1093/brain/awac048.