In Vivo Hematopoietic Stem Cell Editing via AAV9 Delivery

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Solution Overview

Problem

Current methods for genetically editing stem cells, particularly hematopoietic stem cells, are complex, lengthy, and require life-threatening conditioning regimens, limiting their application to hospitals with stem cell transplantation facilities.

Innovation Solution

An in vivo method using adeno-associated virus (AAV) vectors encapsulating guide RNAs and RNA-guided endonucleases, such as Cas9, to directly edit genes in hematopoietic stem cells, mobilized by agents like plerixafor and G-CSF, allowing for simple, rapid, and affordable gene editing without the need for hospital-based treatments.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Reliability

If ex vivo stem cell editing methods are used, then genetic modification can be achieved, but the process becomes complex, lengthy, and requires life-threatening conditioning regimens

Engineering Contradiction:

Improvegenetic modification effectivenessVSAvoidediting process complexity

Core Design Contradiction:

ReliabilityVSDevice complexity

Solution Approach 1:

The patent uses AAV9 viral vectors as intermediaries to deliver CRISPR-Cas9 gene editing components directly to hematopoietic stem cells in vivo. The viral vector serves as a mediator that transports the genetic material through the bloodstream to the target cells, eliminating the need for complex ex vivo manipulation and conditioning regimens while maintaining effective genetic modification

Inventive Principle:

Principle #24Intermediary (Mediator)

Solution Approach 2:

The patent enables the patient's own hematopoietic stem cells to perform the gene editing function autonomously within their body. By delivering the CRISPR components in vivo, the cells self-edit their own genome without requiring external laboratory processing, thereby simplifying the overall process and eliminating the need for specialized hospital facilities

Inventive Principle:

Principle #25Self-service

2Reliability

If ex vivo stem cell editing is performed, then genetic editing can be achieved, but it can only be performed in hospitals equipped with stem cell transplantation facilities

Engineering Contradiction:

Improvegenetic editing capabilityVSAvoidtreatment accessibility

Core Design Contradiction:

ReliabilityVSEase of operation

Solution Approach 1:

The AAV9 viral vector acts as a mobile intermediary that delivers gene editing components through the circulatory system to target cells throughout the body. This intermediary approach transforms a facility-dependent procedure into a system that can be administered via simple intravenous infusion, making genetic editing accessible in ordinary outpatient clinics

Inventive Principle:

Principle #24Intermediary (Mediator)

Solution Approach 2:

Instead of bringing the patient to a specialized facility for complex cell manipulation, the patent inverts the approach by bringing the editing tools directly to the patient's cells in their own body. This inversion of the traditional ex vivo approach enables treatment in any setting where intravenous infusion can be administered

Inventive Principle:

Principle #13The other way round (Inversion)

3Reliability

If traditional gene therapy methods are used, then genetic modification is achieved, but the process is lengthy and requires multiple steps

Engineering Contradiction:

Improvegenetic modification outcomeVSAvoidtreatment duration

Core Design Contradiction:

ReliabilityVSLoss of time

Solution Approach 1:

The patent combines multiple gene editing components (guide RNA and Cas9 enzyme) into a single AAV9 viral vector delivery system. This merging of components into one administrable unit eliminates the need for multiple separate steps including cell collection, laboratory processing, and reinfusion, reducing the treatment to a single intravenous infusion that achieves the same genetic modification outcome

Inventive Principle:

Principle #5Merging (Combining)

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 efficiently edits genes in hematopoietic stem cells, reducing expression by at least 50%, minimizing off-target effects, and can be performed in outpatient settings, reducing costs and risks associated with ex vivo treatments.

Implementation Method 1

administering to the subject a plurality of adeno-associated virus 9 (AAV9) vectors encapsulating (a) at least one guide RNA (gRNA) that targets a genomic region of interest or a nucleic acid encoding the at least one gRNA, and (b) a nucleic acid encoding a RNA-guided endonuclease

Methodology Applied

Scientific EffectViral transduction:

Implementation Method 2

Type II CRISPR-Cas-based systems have been used for genome editing, and require a Cas polypeptide or variant thereof guided by a customizable guide RNA (gRNA) for programmable DNA targeting

Methodology Applied

Scientific EffectCRISPR-Cas9 gene editing: