PTS Gene Exon 3 Skipping Correction in Monocytes
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Solution Overview
Problem
Human monocytes and monocyte-derived cells have reduced activity of the 6-pyruvoyltetrahydropterin synthase (PTS) protein due to exon 3 skipping, leading to a weakened immune system and increased susceptibility to infectious diseases and cancer, as they produce an inactive protein due to cell-type specific alternative splicing.
Innovation Solution
Editing the genomic sequence of the PTS gene in monocytes, macrophages, dendritic cells, or their precursor cells to inhibit exon 3 skipping by altering splicing factor recognition sites or inhibiting the expression/activity of splicing factors such as SRSF1 and SRSF3, using CRISPR/Cas systems, meganucleases, zinc finger nucleases, or antisense oligonucleotides to restore PTPS protein activity.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Adaptability or versatility
If alternative splicing occurs in monocytes and monocyte-derived cells, then cell-type specific gene expression is achieved, but PTPS protein activity is lost due to exon 3 skipping
Solution Approach 1:
The patent extracts and removes the intronic sequences (intron 2 and/or intron 3) that contain splicing factor recognition sites responsible for exon 3 skipping. By taking out these harmful intronic elements, the patent eliminates the aberrant splicing mechanism while preserving the desired PTPS protein expression in monocytes and monocyte-derived cells.
Solution Approach 2:
The patent changes the genomic sequence parameters by introducing point mutations in the intronic recognition sites. This alters the binding affinity of splicing factors (such as SRSF1 and SRSF3) to their recognition sites, thereby changing the splicing outcome from exon skipping to exon inclusion, and restoring PTPS protein activity.
2Adaptability or versatility
If splicing factor recognition sites are present in intron 2 or intron 3, then alternative splicing is regulated, but exon 3 skipping occurs leading to inactive protein production
Solution Approach 1:
The patent converts the harmful effect of splicing factor recognition sites (which cause exon 3 skipping) into a beneficial outcome by strategically modifying these sites. The modified recognition sites now promote exon 3 inclusion instead of skipping, transforming the originally harmful splicing regulation mechanism into a beneficial one that produces active PTPS protein.
Solution Approach 2:
The patent changes the nucleotide sequence parameters of the splicing factor recognition sites in intron 2 and/or intron 3. By introducing specific point mutations, the patent alters the binding characteristics of splicing factors, thereby changing the splicing outcome from harmful (exon skipping) to beneficial (exon inclusion).
3Reliability
If PTPS protein activity is low in monocytes, then immune system function is weakened, but genetic modification to increase activity may have off-target effects
Solution Approach 1:
The patent applies local quality modification by targeting only specific intronic sequences (intron 2 and/or intron 3) containing splicing factor recognition sites. The genetic modification is localized to these specific regions, leaving the rest of the PTS gene and other genomic regions unchanged, thereby minimizing off-target effects while achieving the desired increase in PTPS protein activity.
Solution Approach 2:
The patent takes out and removes the harmful intronic sequences that cause exon 3 skipping, rather than introducing foreign genetic material. This extraction approach reduces the risk of off-target effects compared to methods that require introducing exogenous genes or widespread genomic modifications.
Data Source
AI summary
The present disclosure relates generally to compositions and methods for inhibiting exon 3 skipping in the 6-pyruvoyltetrahydropterin synthase (PTS) gene in monocytes, monocyte-derived cells such as macrophages and dendritic cells of the precursor cells thereof. The inhibition can be achieved with genome editing of the genomic sequence to remove certain splicing factor recognition sites or inhibiting the expression or activity of the splicing factors that contribute to the cell-specific exon skipping. Monocytes and monocyte-derived cells that have reduced exon skipping in the PTS gene can generate more potent immune response and thus are useful in preventing or treating diseases such as infectious diseases and cancer.