Inducible AAV Rep Cap Expression Systems

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Solution Overview

Problem

There is a need for improved methods of recombinant AAV (rAAV) production that enhance AAV packaging efficiency and control protein expression to reduce metabolic burden and toxicity during cell growth.

Innovation Solution

The development of polynucleotides, vector systems, cells, and methods for amplifying and inducing the expression of AAV Rep and Cap proteins, which can be used in triple transfection of cells or for inducible expression of rAAV virions, thereby increasing total and packaged virions during rAAV production.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Productivity

If protein expression is increased during cell growth, then production of AAV Rep and Cap proteins is improved, but metabolic burden and toxicity to cell growth increase

Engineering Contradiction:

Improveproduction of AAV Rep and Cap proteinsVSAvoidmetabolic burden and toxicity to cell growth

Core Design Contradiction:

ProductivityVSObject-affected harmful factors

Solution Approach 1:

The patent employs inducible expression systems where AAV Rep and Cap proteins are expressed only after cell growth has been established, rather than continuously during cell growth. This periodic expression strategy allows cells to first establish robust growth, then produce the desired proteins when conditions are favorable, thereby reducing metabolic burden while maintaining high productivity

Inventive Principle:

Principle #19Periodic action

Solution Approach 2:

The patent implements a two-stage process where cell growth is established first as a preliminary step, followed by induction of protein expression. This preliminary action ensures that cells are healthy and metabolically capable before undertaking the burden of high-level protein production, thus resolving the contradiction between productivity and cellular health

Inventive Principle:

Principle #10Preliminary action

2Productivity

If AAV packaging efficiency is enhanced, then production of packaged virions is improved, but complexity of production methods increases

Engineering Contradiction:

Improveproduction of packaged virionsVSAvoidcomplexity of production methods

Core Design Contradiction:

ProductivityVSDevice complexity

Solution Approach 1:

The patent combines multiple functions into a single inducible expression system that simultaneously controls the expression of both AAV Rep and Cap proteins. This merging of control mechanisms simplifies the overall production method while maintaining enhanced packaging efficiency, as a single induction event coordinates the production of all necessary viral components

Inventive Principle:

Principle #5Merging (Combining)

Data Source

PatentUS20250129127A1Systems for Amplification of AAV Rep and Cap Proteins

Publication Date: 2025.04.24 SHAPE THERAPEUTICS INC

AI summary

The present disclosure provides polynucleotides, vector systems, cells, and methods for amplifying expression of proteins, such as AAV Rep and Cap proteins or therapeutic proteins (e.g., an antibody). Increased expression of AAV Rep and Cap proteins is useful in increasing production of recombinant AAV virions. Also provided herein are vector systems for inducing amplification of expression of AAV Rep and Cap proteins. Also provided herein are vector systems for inducing amplification of expression of therapeutic proteins. Inducible expression of therapeutic proteins is useful in increasing production of therapeutic proteins.