CD4-Positive T Cell Production from Pluripotent Stem Cells

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Solution Overview

Problem

Existing methods for inducing CD4-positive T cells from pluripotent stem cells are hindered by the lack of CD4 molecule expression, preventing sufficient helper function and activation of dendritic cells.

Innovation Solution

Introducing a CD4 gene or its product into T cells induced from pluripotent stem cells using a retrovirus, followed by specific culturing and co-culturing steps with FLT3L, IL-7, IL-15, and mitogens, to achieve CD4-positive T cell production and activation of dendritic cells.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Reliability

If T cells are induced from pluripotent stem cells using conventional methods, then T cells can be produced, but CD4 molecules are not expressed and helper function is insufficient

Engineering Contradiction:

ImproveCD4 molecule expressionVSAvoiddifficulty in preparing cells expressing CD4 molecules

Core Design Contradiction:

ReliabilityVSEase of manufacture

Solution Approach 1:

The patent introduces the CD4 gene into pluripotent stem cells before differentiating them into T cells. This preliminary genetic modification ensures that when the T cells differentiate, they will express CD4 molecules, thereby resolving the contradiction between producing T cells and ensuring CD4 expression.

Inventive Principle:

Principle #10Preliminary action

Solution Approach 2:

The patent changes the genetic parameters of the pluripotent stem cells by introducing the CD4 gene, which fundamentally alters the expression profile of the resulting T cells. This parameter change enables CD4 expression in T cells that would otherwise lack this molecule, solving the reliability issue while maintaining the ease of manufacturing through established differentiation protocols.

Inventive Principle:

Principle #35Parameter changes

2Reliability

If CD4-positive T cells are produced from pluripotent stem cells, then dendritic cell activation and immune function are enhanced, but the production process becomes more complex

Engineering Contradiction:

Improvedendritic cell activation capabilityVSAvoidproduction process complexity

Core Design Contradiction:

ReliabilityVSDevice complexity

Solution Approach 1:

By introducing the CD4 gene into pluripotent stem cells at the beginning of the differentiation process, the patent ensures that CD4 expression is established before T cell maturation. This preliminary action simplifies the overall process compared to attempting to add CD4 later, as it integrates the modification into the natural differentiation timeline.

Inventive Principle:

Principle #10Preliminary action

Solution Approach 2:

The patent uses a retroviral vector as an intermediary to deliver the CD4 gene into the pluripotent stem cells. This intermediary mechanism efficiently transfers the genetic material without requiring complex direct gene editing methods, thereby enhancing dendritic cell activation capability while keeping the production process relatively straightforward.

Inventive Principle:

Principle #24Intermediary (Mediator)

3Reliability

If conventional T cell induction methods are used, then T cells can be produced, but sufficient helper function cannot be achieved due to lack of CD4 molecules

Engineering Contradiction:

Improvehelper functionVSAvoiddifficulty in preparing CD4-expressing cells

Core Design Contradiction:

ReliabilityVSEase of manufacture

Solution Approach 1:

The patent fundamentally changes the expression parameters of the T cells by introducing the CD4 gene into the source pluripotent stem cells. This ensures that the resulting T cells inherently express CD4 and possess full helper function, eliminating the need for complex post-differentiation modifications and making the manufacturing process as straightforward as conventional methods.

Inventive Principle:

Principle #35Parameter changes

Solution Approach 2:

The CD4 gene introduction is performed as a preliminary step before T cell differentiation, ensuring that the helper function is built into the cell lineage from the start. This approach achieves sufficient helper function without adding manufacturing complexity, as the gene modification occurs during the pluripotent state when cells are most receptive to genetic manipulation.

Inventive Principle:

Principle #10Preliminary action

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

Functional CD4-positive T cells are produced, capable of activating dendritic cells, thereby enhancing immune function and providing a therapeutic agent for cancer treatment.

Implementation Method 1

introducing a CD4 gene or a gene product thereof into T cells induced from pluripotent stem cells

Methodology Applied

Scientific EffectGene transfer:

Implementation Method 2

culturing the CD34-positive hematopoietic progenitor cells obtained in the Step (1), in the presence of FLT3L (Flt3 Ligand) and IL (Interleukin)-7

Methodology Applied

Scientific EffectCytokine signaling:

Implementation Method 3

co-culturing the cells obtained in the Step (2), with peripheral blood mononuclear cells (PBMCs), in the presence of IL-7 and IL-15

Methodology Applied

Scientific EffectCytokine signaling:

Implementation Method 4

bringing the cells obtained in the Step (2) into contact with mitogen

Methodology Applied

Scientific EffectMitogenic stimulation:

Data Source

PatentUS11987811B2Method for producing CD4-positive T cells from pluripotent stem cells

Publication Date: 2024.05.21 KYOTO UNIV

AI summary

An object of the present invention is induction of CD4-positive T cells from pluripotent stem cells. This object is achieved by production of CD4-positive T cells by introducing a CD4 gene or a gene product thereof into T cells induced from pluripotent stem cells.