ALAS1 Gene Editing via Modified gRNA for Porphyria Therapy

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Solution Overview

Problem

There is a need for developing safe and effective therapies for treating and preventing ALAS1-related diseases and disorders, such as Porphyria, which are caused by the over-expression of the 5′-Aminolevulinate Synthase 1 (ALAS1) enzyme.

Innovation Solution

The use of guide RNAs (gRNAs) that target the ALAS1 genomic locus, specifically comprising spacer sequences from SEQ ID NOs: 25-48 and 83-112, in combination with a Cas9 endonuclease, to induce cutting efficiency and reduce ALAS1 expression.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Productivity

If CRISPR-Cas9 systems are used to target ALAS1 genomic locus, then cutting efficiency is improved, but safety and effectiveness for treating ALAS1-related diseases remains insufficient

Engineering Contradiction:

Improvecutting efficiencyVSAvoidsafety and effectiveness

Core Design Contradiction:

ProductivityVSReliability

Solution Approach 1:

The patent optimizes multiple parameters of the gRNA including spacer sequence length (20 nucleotides), chemical modifications (2′-O-methyl and phosphorothioate), and structural features to achieve both high cutting efficiency and improved safety profile for therapeutic applications

Inventive Principle:

Principle #35Parameter changes

Solution Approach 2:

The gRNA is designed as a composite structure combining spacer sequence, scaffold sequence, and multiple chemical modifications (2′-O-methyl nucleotides and phosphorothioate linkages) to simultaneously achieve high efficiency and safety

Inventive Principle:

Principle #40Composite materials

2Productivity

If gRNA with high cutting efficiency is designed, then ALAS1 expression reduction is improved, but off-target effects and immune response may increase

Engineering Contradiction:

ImproveALAS1 expression reductionVSAvoidoff-target effects and immune response

Core Design Contradiction:

ProductivityVSObject-affected harmful factors

Solution Approach 1:

The patent converts potential harmful off-target binding into beneficial on-target specificity by designing the spacer sequence to match the unique ALAS1 genomic locus and using chemical modifications to enhance discrimination against off-target sites

Inventive Principle:

Principle #22Blessing in disguise (Convert harm into benefit)

Solution Approach 2:

Chemical modifications including 2′-O-methyl and phosphorothioate linkages act as intermediaries to reduce immune system recognition of the gRNA while maintaining its targeting and cutting functionality

Inventive Principle:

Principle #24Intermediary (Mediator)

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 gRNAs achieve a cutting efficiency of at least 40% to 95% for targeting the ALAS1 genomic locus, effectively reducing ALAS1 expression and providing a therapeutic approach for ALAS1-related diseases and disorders, such as Porphyria.

Implementation Method 1

the gRNA comprises a spacer sequence comprising any one of the sequences of SEQ ID NOs: 25-48 and 83-112

Methodology Applied

Scientific EffectComplementary base pairing:

Implementation Method 2

a Cas9 endonuclease, to induce cutting efficiency and reduce ALAS1 expression

Methodology Applied

Scientific EffectDNA cleavage:

Data Source

PatentUS20250027087A1Modulating expression of alas1 (5'-aminolevulinate synthase 1) gene

Publication Date: 2025.01.23 CRISPR THERAPEUTICS AG

AI summary

The present disclosure relates to methods, compositions and kits for treating conditions that are related to the modulation of expression of 5′-Aminolevulinate Synthase 1 (ALAS1) by gene editing.