Trigger Nucleic Acids and RNA-Binding Proteins for Gene Upregulation
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Solution Overview
Problem
Existing therapies for genetic disorders and cancer primarily focus on downregulating gene expression, while diseases like haploinsufficiency disorders and autosomal recessive disorders are characterized by a decrease in protein expression, and methods to increase gene expression are limited and face challenges in delivering large nucleic vectors efficiently.
Innovation Solution
The use of engineered nucleic acids and RNA-binding proteins, such as Interleukin Enhancer Binding Factor 3 (ILF3) or fragments thereof, to upregulate gene expression by targeting RNA decay and recruiting transcription factors and chromatin remodelers, with RNA-targeting Cas proteins that do not have nuclease activity, and delivery via lipid nanoparticles.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If existing overexpression systems are used to increase gene expression, then gene expression levels are increased, but cells are inundated with non-physiological levels of gene expression
Solution Approach 1:
The patent uses RNA-binding proteins with tunable binding affinities to achieve physiological levels of gene expression. By modifying the RNA-binding protein's affinity for the target mRNA, the system can precisely control expression levels to match physiological conditions rather than producing non-physiological overexpression.
Solution Approach 2:
The patent introduces RNA-binding proteins as intermediaries that bind to specific RNA sequences to regulate gene expression. These proteins act as mediators between the genetic material and the expression machinery, enabling controlled upregulation of gene expression at physiological levels through mechanisms such as stabilizing mRNA or enhancing translation efficiency.
2Quantity of substance
If large nucleic vectors are used to deliver gene expression machinery, then gene expression can be increased, but delivery to cells becomes difficult
Solution Approach 1:
The patent divides the gene expression system into separate functional components: small RNA molecules (such as guide RNAs or aptamers) and protein components (such as RNA-binding proteins with transcriptional activation domains). These segmented components can be delivered separately using smaller, more efficient vectors, avoiding the delivery problems associated with large nucleic vectors while maintaining the ability to increase gene expression.
3Quantity of substance
If CRISPR-based methods targeting DNA are used to upregulate gene expression, then gene expression can be increased, but the targeting window is very narrow around the transcription start site
Solution Approach 1:
The patent transitions from DNA-targeting (one-dimensional constraint at the transcription start site) to RNA-targeting (multi-dimensional freedom across the entire transcript). By targeting RNA molecules rather than DNA, the system gains access to the entire length of the transcript as a potential targeting window, dramatically increasing the number of possible guide sequence locations and design options.
4Quantity of substance
If DNA-targeting methods are used for gene upregulation, then gene expression can be increased, but antisense transcripts cannot be targeted
Solution Approach 1:
The patent uses RNA-binding proteins that can recognize and bind to RNA sequences, including antisense transcripts. By targeting the RNA copy rather than the DNA template, the system can specifically regulate antisense transcripts that are transcribed from the opposite strand, providing a capability that DNA-targeting methods inherently lack.
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
This approach effectively increases gene expression in a tissue-specific manner, overcoming limitations of existing methods by allowing broader targeting and more efficient delivery, particularly for diseases characterized by decreased gene expression.
Implementation Method 1
the RNA-binding protein, Interleukin Enhancer Binding Factor 3 (ILF3), or fragments thereof may be used to increase gene expression
Implementation Method 2
delivery via lipid nanoparticles
Data Source
AI summary
The present disclosure, at least in part, relates to compositions (e.g., engineered nucleic acids and engineered proteins) and methods for increasing gene expression. The engineered proteins include RNA-binding proteins (e.g., RNA-binding proteins that comprise a Interleukin Enhancer Binding Factor 3 (ILF3) sequence, a Cas sequence, or a combination thereof). In some aspects, the disclosure provides methods of identifying engineered nucleic acids that are shorter in length than a gene of interest to induce expression of the gene of interest and also provides RNA-binding proteins for inducing gene expression.