SMN2 C840T Base Editing for Stable SMN Protein
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Solution Overview
Problem
Current gene therapies for spinal muscular atrophy (SMA), such as antisense oligonucleotides, provide only transient improvements and do not address the underlying issue of SMN protein instability, leading to a lack of a lasting cure.
Innovation Solution
Employing base editing strategies to target the SMN2 genomic locus, specifically editing the C840T mutation and removing degrons like the EMLA sequence to stabilize and increase the production of functional SMN protein.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If antisense oligonucleotides are used to treat SMA, then SMN2 exon 7 splicing is improved, but the treatment effect is only transient and not curative
Solution Approach 1:
The patent performs preliminary genetic modification by converting the C840T mutation in the SMN2 gene before the therapeutic effect is needed. This preliminary base editing creates a stable, heritable change in the genome that will continuously produce functional SMN protein throughout the patient's life, rather than requiring repeated administrations of antisense oligonucleotides.
Solution Approach 2:
The patent creates a corrected copy of the defective SMN2 gene by using base editing to convert the mutant C840T sequence back to the functional C6 sequence. This creates a genomic copy that behaves like the wild-type SMN1 gene, continuously producing stable, full-length SMN protein without the degradation issues of the original SMN2 transcript.
2Quantity of substance
If SMN2 gene is used to compensate for SMN1 loss, then some functional SMN protein is produced, but the protein is truncated and unstable
Solution Approach 1:
The patent changes the critical parameter at position 840 in the SMN2 gene from T (thymine) to C (cytosine) through base editing. This single nucleotide parameter change fundamentally alters the splicing outcome, preventing exon 7 skipping and ensuring production of the full-length, stable SMN protein isoform instead of the truncated, unstable variant.
Solution Approach 2:
The patent converts the harmful C840T mutation into a benefit by using the same genomic location to perform a reverse conversion (T to C). This transforms the defective SMN2 gene, which normally produces unstable truncated protein, into a functional gene that produces stable full-length SMN protein, effectively turning a disease-causing element into a therapeutic asset.
3Duration of action of stationary object
If base editing is used to correct SMN2 gene, then lasting cure is achieved, but the editing precision and efficiency must be optimized
Solution Approach 1:
The patent uses a carefully designed single-guide RNA (sgRNA) as an intermediary to direct the base editor to the precise C840T location in the SMN2 gene. The sgRNA provides sequence-specific targeting, ensuring that editing occurs only at the intended genomic locus and not at off-target sites, thereby achieving high precision while maintaining efficient correction of the mutation.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The method results in a significant increase in SMN protein levels and stability, potentially offering a lasting cure for SMA by enhancing the functional activity of the SMN2 protein.
Implementation Method 1
employing base editing strategies to target the SMN2 genomic locus, specifically editing the C840T mutation
Data Source
Figure 1A~1B
Figure 2A~2B
Figure 2C
AI summary
The disclosure provides methods, base editors, vectors encoding base editors and cognate gRNAs, and compositions and kits comprise said components, for installing nucleobase edits to the SMN2 locus to increase the activity and/or amount and/or stability of SMN2 protein in a cell, thereby treating Spinal Muscular Atrophy. In certain aspect, the disclosure provides compositions and methods to edit C840T of exon 7 of the SMN2 gene, or installing another one or more nucleobase edits which have the effect of removing or inactivating a degron, such as the C -terminal portion of the region encoded by exon 6 or the 4-amino acid region encoded by exon 8 (i.e., the EMLA (SEQ ID NO: 466) -tail) so as to remove or limit their degron activity to reduce, mitigate, or eliminate the intracellular degradation of the SMN2 protein.