Pre-mRNA Trans-Splicing Molecules for Genetic Correction
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Current gene therapy methods for treating genetic disorders like epidermolysis bullosa, cystic fibrosis, and autoimmune diseases face challenges such as low efficiency, size limitations for gene delivery, insertional mutagenesis, immune responses, and toxicity, particularly in addressing autosomal dominant disorders and aberrant splicing issues.
Innovation Solution
Development of pre-mRNA trans-splicing molecules (RTMs) with binding, splicing, and coding domains that target specific genes to correct defective mRNA transcripts through spliceosome-mediated RNA trans-splicing, allowing for precise correction of genetic defects in skin and epithelial disorders by forming chimeric RNA molecules.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If conventional gene therapy methods are used to treat genetic disorders, then gene delivery can be achieved, but efficiency is low and insertional mutagenesis occurs
Solution Approach 1:
The RTM molecule is divided into distinct functional domains: a binding domain (complementary to target pre-mRNA), a splicing domain (containing spliceosome recognition motifs), and a coding domain (carrying wild-type exon sequences). This segmentation allows the therapy to act locally on the target transcript without requiring genomic integration, thereby improving efficiency while avoiding insertional mutagenesis.
Solution Approach 2:
The RTM acts as an intermediary molecule that bridges the defectively spliced pre-mRNA and the spliceosome machinery. By providing exonic splicing enhancer sequences and wild-type coding sequences as intermediaries, the RTM redirects splicing to produce corrected mRNA without altering the genomic DNA, thus avoiding mutagenesis while restoring function.
2Reliability
If standard gene replacement therapy is applied, then functional gene can be introduced, but immune responses and toxicity occur
Solution Approach 1:
The RTM molecule is designed to be self-regulating through endogenous control mechanisms. Once delivered, the RTM uses the cell's own spliceosome machinery and regulatory networks to produce corrected mRNA. The binding domain ensures specificity to the target pre-mRNA, and the splicing domain leverages natural spliceosome recognition sequences, allowing the system to self-regulate expression levels and minimize immune recognition.
Solution Approach 2:
Instead of introducing a complete foreign gene that may trigger immune responses, the RTM copies only the specific wild-type exon sequences needed for correction. The binding domain is complementary to (copies the sequence of) the target pre-mRNA region, and this copied sequence is then spliced into the transcript, producing a corrected version that is immunologically more acceptable than full gene replacement.
3Manufacturing precision
If RNA trans-splicing is used to correct defective transcripts, then precision in correcting genetic defects is improved, but the complexity of the molecule increases
Solution Approach 1:
The RTM molecule exhibits local quality by concentrating correction functionality in specific domains: the binding domain provides sequence-specific targeting precision, the splicing domain contains optimized spliceosome recognition motifs for precise splicing control, and the coding domain carries only the necessary wild-type exon sequences. This localized functional organization achieves high correction precision while managing molecular complexity through domain specialization.
Solution Approach 2:
The RTM molecule is designed with multi-functionality: the binding domain provides target recognition, the splicing domain enables spliceosome recruitment and trans-splicing catalysis, and the coding domain supplies corrected amino acid sequences. Additionally, the splicing domain motifs are universal spliceosome recognition sequences that work across different target genes, making the RTM platform universally applicable to various splicing defects while maintaining controlled complexity through modular design.
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
RTMs effectively correct genetic defects by forming functional chimeric RNA molecules, reducing mutant transcript levels, and can be used to treat a range of skin disorders and cancers by targeting specific genes, offering improved efficiency and reduced adverse effects compared to conventional gene therapy.
Implementation Method 1
a) sequence(s) that is/are complementary to an exon sequence of a mammalian gene
Implementation Method 2
spliceosome mediated RNA trans-splicing... a portion of the RTM is trans-spliced to a portion of the target pre-mRNA to form a chimeric RNA molecule
Data Source
Figure 1
Figure 2A~2C
Figure 3~4
AI summary
The present invention relates to specific and markedly improved pre-mRNA trans-splicing molecule (RTM) molecules which are designed to correct specific genes expressed within cells to be targeted, and which are associated with epidermolysis bullosa, cystic fibrosis, pachyonychia congenital, and psoriasis or neurodermitis, as well as cancers of the skin. In particular, the RTMs of the present invention are genetically engineered to interact with a specific target pre-mRNA expressed in cells to be targeted so as to result in correction of genetic defects or reprogramming of gene expression responsible for a variety of different skin disorders.