CrossMab Antibody Purification via Hydrophobic Interaction Chromatography

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Solution Overview

Problem

Current methods for producing multispecific antibodies, particularly those with an IgG-like architecture, face challenges in separating correctly paired from mispaired variants, leading to inefficiencies and potential immunogenicity due to improper light chain-heavy chain interactions, which can affect therapeutic efficacy.

Innovation Solution

The use of hydrophobic interaction chromatography (HIC) with specific media, such as Butyl Sepharose HP, Capto Butyl ImpRes, or PPG-600M, to separate multispecific CrossMab antibodies from mispaired variants based on hydrophobicity, ensuring correct pairing of light and heavy chains.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Adaptability or versatility

If recombinant DNA techniques are used to produce multispecific antibodies, then the ability to bind two or more antigens is achieved, but light chain mispairing occurs leading to production of variants with wrong heavy chain associations

Engineering Contradiction:

Improvemultispecific binding capabilityVSAvoidheavy chain-light chain pairing accuracy

Core Design Contradiction:

Adaptability or versatilityVSManufacturing precision

Solution Approach 1:

The patent segments the antibody chains by introducing a protease cleavage site that allows selective separation of light chains from heavy chains. This segmentation enables the light chains to be released as free chains while heavy chains remain associated, facilitating the removal of mispaired variants through subsequent purification steps.

Inventive Principle:

Principle #1Segmentation

Solution Approach 2:

The patent introduces a protease cleavage site as an intermediary element between the light chain and heavy chain. This cleavage site serves as a mediator that allows controlled separation of the chains, enabling the removal of mispaired variants while preserving correctly paired antibodies.

Inventive Principle:

Principle #24Intermediary (Mediator)

2Ease of manufacture

If standard purification methods are used, then the production process is simple, but mispaired variants cannot be effectively separated from correctly paired antibodies

Engineering Contradiction:

Improvepurification process simplicityVSAvoidseparation of mispaired variants

Core Design Contradiction:

Ease of manufactureVSManufacturing precision

Solution Approach 1:

The patent performs a preliminary action by introducing a protease cleavage site during antibody construction. This preliminary modification enables subsequent selective cleavage and separation of mispaired variants, making the purification process more effective without requiring complex procedures.

Inventive Principle:

Principle #10Preliminary action

Solution Approach 2:

The patent changes the chemical parameter of the antibody structure by incorporating a protease cleavage site. This parameter change enables selective enzymatic cleavage of light chains, creating a physical and chemical basis for separating mispaired variants from correctly paired antibodies through standard purification techniques.

Inventive Principle:

Principle #35Parameter changes

3Productivity

If mispaired variants are not removed, then the production yield is maintained, but immunogenicity and aggregation risks increase affecting therapeutic efficacy

Engineering Contradiction:

Improveantibody production yieldVSAvoidimmunogenicity and aggregation

Core Design Contradiction:

ProductivityVSObject-affected harmful factors

Solution Approach 1:

The patent extracts mispaired variants from the antibody mixture by utilizing selective protease cleavage. The cleavage site allows light chains to be released and mispaired variants to be separated and removed, while correctly paired antibodies remain intact and are retained in the final product.

Inventive Principle:

Principle #2Taking out (Extraction)

Solution Approach 2:

The patent converts the potential harm of mispaired variants into a benefit by using the presence of the protease cleavage site as a marker. The cleavage site enables selective identification and removal of mispaired variants, transforming what would be a contaminant problem into a controlled purification opportunity.

Inventive Principle:

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

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 separates correctly paired multispecific antibodies from mispaired variants, enhancing the purity and therapeutic potential of the final antibody product by reducing immunogenicity and aggregation risks.

Implementation Method 1

The hydrophobic interaction chromatography medium of the invention enables the separation of multispecific CrossMab antibodies from mispaired variants thereof

Methodology Applied

Scientific EffectHydrophobic interaction: Hydrophobe

Data Source

PatentUS11945839B2Depletion of light chain mispaired antibody variants by hydrophobic interaction chromatography

Publication Date: 2024.04.02 F HOFFMANN LA ROCHE INC

AI summary

The present invention relates to methods for separating multispecific CrossMab antibodies from light chain mispaired variants thereof in a solution comprising CrossMab bispecific antibodies and mispaired antibody variants thereof by hydrophobic interaction chromatography.