Research Plan · Computational Genomics · 2025–2037

Calpainopathy / LGMD R1:
from mechanism to intervention

Limb-girdle muscular dystrophy type R1 (LGMD R1 / calpainopathy) is caused by mutations in the CAPN3 gene. It is progressive, currently without disease-modifying treatment, and affects tens of thousands worldwide. This research programme develops a four-pillar strategy from metabolic stabilisation to precision base editing.

Active · Strategy phase · Clinical horizon 2034–2037
The Disease

LGMD R1 — what we know and what we don't

Calpainopathy is the most common form of limb-girdle muscular dystrophy. It is caused by loss-of-function mutations in CAPN3, which encodes calpain-3 — a muscle-specific calcium-dependent protease essential for sarcomere remodelling and muscle homeostasis.

Without functional calpain-3, muscles undergo progressive fibrosis and atrophy. The disease typically presents in the second or third decade of life and leads to significant disability. There is currently no approved disease-modifying therapy.

The primary research target: The c.2242C>T mutation — one of the most common CAPN3 pathogenic variants — is a point mutation that is in principle correctable by adenine base editing (ABE). The research programme works toward a clinically viable ABE correction strategy for this mutation.

Why this target: c.2242C>T is among the most frequently reported pathogenic CAPN3 variants, it sits in an editing window reachable by current adenine base editors, and correcting it addresses a substantial share of the patient population rather than a single case. Adding c.550delA as a prime-editing target widens that share further. The programme's target selection follows from population coverage and editability, and it is stated here so that a reader can judge the rationale independently.
Research Strategy

Four-pillar framework

1
Precision gene editing
Adenine base editing (ABE) targeting the c.2242C>T CAPN3 mutation. Primary focus: efficiency, specificity, and off-target profile of available ABE systems (ABE8eY149V, compact ABE). Guide RNA design and in silico off-target prediction. iPSC-derived skeletal muscle cells as the primary test system — not cardiomyocytes, as CAPN3 is muscle-specific and cardiac expression is toxic (see cardiac safety section).
2
Anti-fibrosis strategies
Progressive fibrosis is the primary driver of disability in LGMD R1. Anti-fibrotic approaches — pirfenidone, losartan, TGF-β pathway inhibitors — are evaluated as potential adjunct therapies to slow disease progression independent of gene correction. Literature review and AI-augmented target identification.
3
Metabolic stabilisation
CAPN3 deficiency disrupts the autophagic-lysosomal pathway and mitochondrial function. Metabolic interventions — creatine supplementation, NAD+ precursors, mTORC1 modulation — are evaluated for their potential to maintain muscle mass and function as bridging therapies while gene editing approaches mature.
4
In vivo delivery
AAV-mediated delivery of base editors to skeletal muscle. Key challenges: AAV serotype selection for muscle tropism (AAV9, AAVrh74), cargo size constraints (split-intein approaches), immune response management. Monitoring developments in lipid nanoparticle delivery as an alternative.
iPSC Strategy

Cell banking and correction

A central component of the research strategy is the establishment of an iPSC bank from LGMD R1 patients carrying the c.2242C>T mutation. This serves two purposes:

Disease modelling
Patient-derived iPSC lines differentiated into skeletal muscle cells provide the primary in vitro model for testing base editing efficiency, specificity, and functional correction. Corrected lines serve as isogenic controls — the gold standard for demonstrating that the editing, not incidental genetic background, is responsible for functional improvement.
Therapeutic banking
Corrected iPSC lines (UCB-ABE, iPSC-ABE) serve as a potential therapeutic resource — autologous cell therapy using gene-corrected cells derived from the patient's own tissue. This strategy avoids immune rejection and is compatible with the clinical horizon of the programme (2034–2037).
Methodology

AI-augmented research approach

Literature synthesis
Continuous AI-augmented synthesis of the calpainopathy and base editing literature. The field moves fast — new ABE systems, new AAV serotypes, new anti-fibrotic candidates. AI enables continuous integration of new findings into the research strategy rather than periodic manual literature reviews.
In silico guide RNA design
Computational guide RNA design and off-target prediction for ABE correction of c.2242C>T. Tools: BE-Designer, CRISPick, CasOFFinder. Prioritisation of guides by editing window position, PAM availability, and predicted off-target profile. In silico results guide experimental prioritisation.
Network analysis
Protein interaction network analysis of CAPN3 and its interactors — identifying secondary targets for metabolic and anti-fibrotic intervention. AI-augmented pathway analysis (STRING, Reactome) to map downstream consequences of CAPN3 loss and identify druggable nodes.
Collaborative model
The EQUORA Institute does not operate a wet laboratory. The computational and strategic research outputs of this programme are designed to be directly usable by academic or clinical collaborators with experimental capacity. The Institute's contribution is the research strategy, the literature synthesis, and the computational groundwork.
Field Context — 2026

Why the timeline is shorter than expected

The field has accelerated on multiple fronts. Four developments directly affect the CAPN3 programme:

NCH / Sarepta
AAVrh74.tMCK.hCAPN3
Nationwide Children's Hospital demonstrated robust preclinical efficacy with AAVrh74-mediated CAPN3 gene replacement in the mouse model. Sarepta Therapeutics has licensed the programme. A Clinical Trial Readiness study (GRASP Consortium, Ann Clin Transl Neurol 2025) defined the outcome measures. IND submission is the next expected step — likely 2026–2027.
Genethon / Atamyo
AAV platform validation
Genethon has CAPN3 preclinical work underway. Their spin-off Atamyo Therapeutics has demonstrated in Phase 1b/2 trials for adjacent LGMD subtypes (ATA-200 for R5, ATA-100 for R9) that AAVrh74 gene replacement achieves 90%+ protein expression in muscle fibers with encouraging safety — validating the delivery platform directly applicable to CAPN3.
UCLA
Muscle-specific vector
A third independent track: UCLA is developing a muscle-specific AAV vector for CAPN3 gene replacement. Three parallel programmes (NCH/Sarepta, Genethon, UCLA) targeting the same gene significantly increases the probability and accelerates the timeline to first-in-human trials. Each uses distinct vector and promoter strategies, providing redundancy and comparative data.
CRISPR
c.550delA — second editing target
Beyond the c.2242C>T point mutation targeted by ABE, the c.550delA frameshift mutation is a second high-priority CAPN3 target — amenable to CRISPR-based exon skipping or prime editing correction. A dual-mutation strategy (ABE for c.2242C>T + prime editing for c.550delA) would cover a substantially larger fraction of the calpainopathy patient population.
bioRxiv 2026
Compact ABE — single vector AAV
New compact adenine base editors enabling single-vector AAV delivery were reported in May 2026 for DMD. The same approach is directly applicable to CAPN3 c.2242C>T correction — removing the split-intein size constraint that was the primary barrier to in vivo ABE delivery. ABE8eY149V (Protein & Cell 2026) adds a high-efficiency, low off-target variant.
WSiMD pilot
Weekly prednisolone — human data
NCT04054375 (McNally/Northwestern) provides the first human pilot data on weekly-pulsed glucocorticoids in a LGMD cohort including calpainopathy patients. Mechanism: GR–KLF15–MEF2C transcriptional switching producing anabolic rather than catabolic effects via H3K27ac remodeling. Investigated as a possible bridging approach while gene correction matures. NCT04054375 is a pilot study in a small cohort, and glucocorticoid regimens carry substantial known risks; any decision about them belongs to a treating neuromuscular clinician.
JOURNEY
Natural history study — trial access
The JOURNEY natural history study is a key pathway to accessing future interventional trials. Participation establishes the baseline functional measurements, biomarker profiles, and disease trajectory data required for gene therapy trial enrolment. Enrolment in JOURNEY (if available) is a near-term practical priority independent of the gene correction timeline.
Non-Classical Dimension

Calpainopathy as metabolic myopathy

An emerging research direction treats calpainopathy not as a classical muscular dystrophy — progressive fibrosis and atrophy driven by membrane fragility — but as a metabolic myopathy with a sarcomere remodelling arrest at its core. This has direct therapeutic implications.

PDGFRβ/STAT1 signalling as regenerative brake
A 2026 JCI study identified the PDGFRβ/STAT1/TGF-β signalling axis as a regulatory checkpoint in myocyte fusion and muscle regeneration. In CAPN3-deficient muscle, this axis may be chronically activated — functioning as a "regenerative brake" that prevents satellite cell-mediated repair. PDGFRβ inhibitors (SU16f) or STAT1 inhibitors (fludarabine) are candidate adjunct strategies to enhance regeneration independent of gene correction. This connects to CAPN3's known role in actin-cytoskeleton remodelling and reduced myonuclear accumulation in calpainopathy.
SOCE — store-operated calcium entry dysregulation
University of Florida (Barton / Wei-LaPierre, C3 grant 2022–2025): CAPN3 is essential for proper calcium handling in skeletal muscle. In LGMD2A/R1 mouse models, store-operated calcium entry (SOCE) is abnormally elevated at rest and fails to respond appropriately to exercise. A 2025 C3-funded follow-up grant is now testing SOCE chemical inhibitors in CAPN3-null mouse models and patient-derived muscle cells — making calcium handling modulation an active near-term therapeutic candidate independent of gene correction.
GSK-3β / Wnt-mTOR dysregulation
LGMD2A/R1 patient muscle shows severe reduction in Wnt pathway and mTOR signalling proteins. GSK-3β inhibitors — tideglusib and VP0.7 — restore Wnt and mTOR pathway activity in patient-derived myotubes (Biodonostia / CSIC, 2021). Tideglusib has completed phase 2 trials in Alzheimer's disease, where development was discontinued for lack of efficacy, and remains in clinical development for congenital myotonic dystrophy; it holds no marketing authorisation in the EU or UK for any indication. What makes repurposing evaluation feasible is the existing human safety dataset, rather than any approved status. This is a second metabolic intervention target alongside the NAD+/creatine/mTORC1 axis.
Mitochondrial energy deficit and IκBα/NF-κB dysregulation
Converging evidence from PPARδ agonist and pyruvate experiments in CAPN3-null models validates mitochondrial energy deficit as a therapeutically real node. CAPN3 normally cleaves IκBα, releasing NF-κB for muscle adaptation signalling — in CAPN3 deficiency, NF-κB is suppressed, leading to myonuclear apoptosis and transcriptional adaptation failure. The weekly prednisolone GR–KLF15 mechanism may partially compensate for this lost NF-κB regulation, explaining its anabolic profile at intermittent dosing.
Critical Safety Context

The cardiac toxicity problem — and its resolution

Early AAV-mediated CAPN3 gene transfer experiments in mice produced dose-dependent mortality from cardiac fibrosis — CAPN3 ectopic expression in the heart causes unregulated proteolysis. This was the primary obstacle to clinical translation.

The Genethon team (Isabelle Richard) solved this through two parallel strategies: (1) muscle-specific promoters including the CAPN3 promoter itself, combined with cardiac miRNA-208a target sequences in the 3′ UTR to suppress cardiac expression; (2) identification that the cardiac toxicity is species-specific — caused by titin splice isoform differences between mice and primates. In NHPs (and humans), titin splicing provides a buffering capacity for CAPN3 activity that mice lack. AAV9-desmin-hCAPN3 in NHPs showed no cardiac toxicity and therapeutic skeletal muscle expression.

This means the mouse cardiac toxicity data does not translate to humans — but the delivery construct must still use cardiac-suppressing design elements as a safety requirement. This is why muscle-specific promoters (tMCK, desmin, CAPN3 promoter + miR-208a target) are a non-negotiable design feature of any CAPN3 gene therapy construct.

Biomarkers & Registry

Monitoring tools and patient data

Urinary N-terminal titin fragment
A 2026 publication identified urinary N-terminal titin fragment as a biomarker in a LGMD2A/R1 cohort. Titin fragments released by abnormal CAPN3-mediated cleavage are detectable in urine — providing a non-invasive disease activity marker. Relevant for: monitoring disease progression, measuring response to metabolic or anti-fibrotic interventions, and tracking SOCE modulator efficacy without repeated muscle biopsy.
C3 Patient Registry — lgmd2a.iamrare.org
Coalition to Cure Calpain 3 (C3) operates a patient-powered natural history registry with 200+ participants from 41 countries in its first year. The registry has already supported recruitment for three clinical research studies. This is a parallel and complementary resource to the JOURNEY natural history study — and a direct route to connecting with the global calpainopathy research community.
Biological Resources

Patient-derived material

The programme has access to banked patient-derived biological material suitable for iPSC derivation, held for research use. The nature of the material and the genotype it carries are discussed directly with prospective academic collaborators rather than set out here. Access to patient-matched material is an unusual starting position for a computational programme, and it enables a tiered experimental approach once collaboration is established:

Primary line → mechanism assay
Differentiation to skeletal muscle allows direct testing of KLF15 transcriptional response, mitochondrial biogenesis, IκBα/NF-κB status, and sarcolemmal repair in patient-derived cells. ABE correction of the same line produces an isogenic control — the gold standard for demonstrating that the editing, rather than genetic background, is responsible for functional recovery.
Second line → validation and fibrosis model
An independently derived second line supports validation of the primary result, and serves as a direct model for the fibrosis and niche pillar. Mesenchymal-to-myofibroblast transition in the CAPN3-deficient context can be studied directly, and anti-fibrotic candidates (pirfenidone, losartan, TGF-β inhibitors) tested on patient-matched material.
Note: The EQUORA Institute does not operate a wet laboratory. The Institute's contribution is the research strategy and the computational groundwork; experimental execution would be conducted by a collaborating institution. Any transfer of biological material would proceed under the receiving institution's ethics approval and the terms of the applicable research consent, and is not discussed further on this page.
Broader Field Context

What the wider computational genomics field is producing

The CAPN3 programme sits within a rapidly accelerating field. Five platform-level developments are directly relevant — not as CAPN3-specific research, but as enabling technologies that reshape what is possible for any rare muscle disease.

1 — Prime editing enters clinical validation (2025)
Prime Medicine's PM359 produced the first human prime editing data: 82% of colony-forming cells with at least one repaired allele, under 2% unintended edits, in a Phase 1/2 trial for chronic granulomatous disease. Prime editing can make all 12 types of point mutation corrections plus small insertions and deletions — expanding the addressable mutation space beyond ABE's A→G conversions. For CAPN3, this means the c.550delA frameshift — not reachable by ABE — becomes a tractable prime editing target. David Liu won the 2025 Breakthrough Prize for base and prime editing.
2 — N-of-1 bespoke CRISPR: regulatory paradigm shift (2025–2026)
Baby KJ (CHOP/Penn Medicine, February 2025): first personalized in vivo CRISPR base editing therapy, designed and manufactured in 6 months for a unique CPS1 deficiency mutation. LNP-delivered ABE8e, 42% liver cell correction in patient-specific mouse model, no serious adverse effects. The FDA subsequently opened a draft guidance pathway for bespoke genome editing therapies — a regulatory precedent that could accelerate N-of-1 approaches for ultra-rare CAPN3 mutations. Proof that individualized gene editing is now a realistic clinical strategy, not a distant aspiration.
3 — LNP delivery to skeletal muscle: a paradigm shift (2025–2026)
Until 2024, LNPs were considered liver-tropism delivery vehicles — unsuitable for skeletal muscle. This is changing rapidly. A Myomerger-derived peptide (Laporte lab, IGBMC, Nov 2025) enhances skeletal muscle tropism of LNPs while reducing liver transduction. World Muscle Society Congress 2025 featured two keynotes on LNP delivery to muscle — a first. LNPs that target satellite cells (not just mature fibres) offer a fundamental advantage over AAV: sustained editing after muscle regeneration. This directly affects the CAPN3 delivery strategy: LNP-delivered compact ABE targeting muscle may become viable before AAV-based approaches reach the clinic.
4 — Engineered VLPs: non-viral, size-unlimited delivery (2022–2026)
Engineered virus-like particles (eVLPs, David Liu lab / Banskota et al. 2022; extended to prime editing RNPs 2024) deliver base editors and prime editors as ribonucleoproteins — not DNA. Key advantages: (1) no AAV cargo size limit — prime editors and large ABEs fit without split-intein; (2) transient expression reduces off-target exposure; (3) no viral genome integration risk; (4) glycoprotein swapping changes tissue tropism. In vivo base editing at therapeutic levels in liver, retina, and inner ear. Muscle tropism VLPs are the logical next development — directly relevant to CAPN3 ABE delivery without the cardiac-safety constraints of AAV-CAPN3 vectors.
5 — AlphaFold3 + AI for drug target identification (2024–2026)
AlphaFold3 (Nobel Prize in Chemistry 2024, Hassabis/Jumper/Baker) predicts structures of protein complexes — protein-protein, protein-DNA, protein-RNA, protein-ligand — with 76% accuracy in protein-ligand docking, 1.8× improvement over prior methods. Open-sourced November 2024. Directly applicable to the CAPN3 programme: (1) modelling the CAPN3-titin interaction at atomic resolution to understand why cardiac toxicity is species-specific; (2) identifying druggable pockets in the PDGFRβ/STAT1 and GSK-3β pathways for adjunct therapy design; (3) predicting guide RNA / target DNA structure for ABE off-target assessment. The AI-augmented research methodology of the EQUORA Institute is now running on the same tools the best structural biology labs in the world use.

Research roadmap — revised 2026

2025–2026
Strategy and computational phase. Four-pillar framework developed. In silico guide RNA design for c.2242C>T using ABE8eY149V and compact ABE systems. Metabolic and anti-fibrotic candidate identification. Field monitoring: NCH/Sarepta IND progress.
2026–2027
Collaboration initiation + parallel track monitoring. Engagement with academic partners for in vitro ABE validation. iPSC banking strategy finalized. NCH/Sarepta clinical trial expected to initiate — observing and learning from the gene replacement track. First working paper submitted on computational strategy.
2027–2029
In vitro ABE validation. c.2242C>T correction efficiency in patient-derived iPSC lines using compact ABE + ABE8eY149V. Off-target profiling. Anti-fibrotic and metabolic intervention testing in parallel. Gene replacement trial (NCH/Sarepta track) generating first human safety data — informing delivery strategy for ABE track.
2029–2031
In vivo ABE studies. Animal model ABE correction. Delivery strategy: single-vector compact ABE via AAVrh74 or AAV9. IND-enabling studies. Gene replacement trial may have early efficacy data by this point — establishing the clinical benchmark for ABE to exceed.
2031–2033
Clinical horizon for ABE. First-in-human ABE studies for c.2242C>T, contingent on preclinical success. Gene replacement may be approved or in late-stage trials by this point — ABE offers the advantage of permanent, precise correction vs. gene addition.
Method & Scope

How this page was produced

Trilith Method™ · Research Card
Programme status
Active · Strategy phase · Clinical horizon 2034–2037
Machine contributionI
Literature discovery and synthesis, candidate identification, computational modelling, and first drafts of this page. Volume and speed are the machine's contribution; none of it is treated as verified on its own.
VerificationII
Claims traced to primary sources rather than to summaries of them. Contested claims run through assert–refute–adjudicate across independent model families. Errors found after publication are corrected on this page with their date.
Human governanceIII
Problem selection, the evidentiary bar, and the decision to publish rest with Pölö (László Papp), EQUORA Institute, who holds editorial responsibility for this page.
Evidence basis
Stated in the Methodology section, per component. Where a source is a preprint, a pilot study, a single trial or a modelled estimate, the page says so at the point of use.
Version
v1.0 · 2026-06-27
Method
Trilith Method™ framework · how the method is being tested
Important · This page is not medical advice

Nothing on this page is a treatment recommendation. It describes a research strategy at the computational and planning stage. LGMD R1 / calpainopathy currently has no approved disease-modifying therapy, and none of the four pillars described here is an available treatment.

The compounds named on this page are research candidates, not options to act on. Several are unapproved for any indication, some have had development discontinued for lack of efficacy, and others are approved only for unrelated conditions. Their appearance here reflects a hypothesis worth testing, not evidence of benefit in calpainopathy.

Do not start, stop or change any medication on the basis of this page. This applies with particular force to glucocorticoid regimens, including the weekly-pulsed protocol discussed above: the human data is a small pilot study, and glucocorticoids carry substantial known risks that require clinical supervision. Decisions about treatment belong to a patient's own neuromuscular clinician.

The gene editing approaches described here are preclinical. Base editing and prime editing for CAPN3 are not clinically available for this condition anywhere. The timeline on this page is a research projection, and research projections of this kind frequently slip.

Editing strategies are variant-specific and do not generalise. An approach applicable to one CAPN3 variant is usually not applicable to another. Nothing here supports an inference about any individual's own genotype.

The EQUORA Institute is not a clinical provider. It does not diagnose, does not treat, does not supply materials to patients, and does not enrol anyone in trials. Families looking for a route into research should speak with a neuromuscular specialist centre and consider the disease registries and natural history studies named above, which are the established paths to trial access.

Declaration of interest: the lead investigator has a personal connection to this disease area. It is declared here as a matter of research practice. The scientific rationale on this page stands on the published literature and is stated so that it can be assessed without reference to that connection.

Research Lead

Principal investigator

Pölö (László Papp) — Founder, EQUORA Institute. Academic collaborators, clinical researchers, or patient organisations working in LGMD R1 / calpainopathy: lpapp@equora.institute