NK Cell CD38 Knockout and CD16a Overexpression

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Solution Overview

Problem

NK cells overexpressing CD16a have a low survival rate and insufficient killing power against tumor cells, with additional challenges of mutual killing among NK cells.

Innovation Solution

NK cells are engineered by knocking out the CD38 gene and overexpressing CD16a using CRISPR-Cas editing and lentiviral transfection systems, with a specific protocol involving multiple cell culture media and transfection reagents to enhance survival and killing efficiency.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Strength

If CD16a is overexpressed on NK cells to enhance ADCC and tumour cell killing, then the killing power against tumour cells is improved, but the survival rate of NK cells decreases and mutual killing among NK cells increases

Engineering Contradiction:

Improvekilling power against tumour cellsVSAvoidsurvival rate of NK cells

Core Design Contradiction:

StrengthVSReliability

Solution Approach 1:

The patent removes the harmful factor (CD38) that causes mutual killing among NK cells through CRISPR-Cas9 gene knockout. By extracting this detrimental element, the system maintains enhanced killing power while improving cell survival rate, resolving the contradiction between strength and reliability.

Inventive Principle:

Principle #2Taking out (Extraction)

Solution Approach 2:

The patent changes the expression level parameter of CD16a to high overexpression through lentiviral transfection. This parameter change enhances ADCC activity and tumour cell killing while, in combination with CD38 knockout, improves survival rate, thereby resolving the contradiction between killing power and reliability.

Inventive Principle:

Principle #35Parameter changes

2Strength

If CD16a is overexpressed to enhance ADCC effects, then the killing function against tumour cells is improved, but intracellular depletion and mutual killing among NK cells occur

Engineering Contradiction:

ImproveADCC effectsVSAvoidmutual killing and intracellular depletion

Core Design Contradiction:

StrengthVSObject-generated harmful factors

Solution Approach 1:

The patent extracts and removes the harmful factor CD38 through gene knockout, which is responsible for mutual killing and intracellular depletion. This eliminates the harmful effects while preserving and enhancing the desired ADCC function through CD16a overexpression.

Inventive Principle:

Principle #2Taking out (Extraction)

Solution Approach 2:

The patent converts the harmful interaction between CD38 and CD16a into a beneficial outcome by removing CD38. This eliminates the mutual killing mechanism while allowing CD16a to function optimally for ADCC, transforming a harmful system into a beneficial one.

Inventive Principle:

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

3Strength

If gene transfer techniques are used to overexpress CD16a, then the cytotoxicity and killing effect are enhanced, but the survival rate remains insufficient

Engineering Contradiction:

ImprovecytotoxicityVSAvoidsurvival rate

Core Design Contradiction:

StrengthVSReliability

Solution Approach 1:

The patent removes the limiting factor (CD38) through CRISPR-Cas9 knockout, which was preventing high survival rates. By extracting this harmful element, the system achieves both enhanced cytotoxicity through CD16a overexpression and improved survival rate, resolving the contradiction between strength and reliability.

Inventive Principle:

Principle #2Taking out (Extraction)

Solution Approach 2:

The patent creates a composite genetic modification system combining two elements: CRISPR-Cas9 mediated CD38 knockout and lentiviral CD16a overexpression. This composite approach synergistically enhances cytotoxicity while improving survival rate, achieving both goals simultaneously.

Inventive Principle:

Principle #40Composite materials

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 modified NK cells exhibit enhanced survival rate, reduced self-killing, and improved killing performance against tumor cells, while maintaining stable overexpression of CD16a even after multiple subcultures.

Implementation Method 1

constructing a CRISPR-Cas editing system for knocking out the CD38 gene

Methodology Applied

Scientific EffectCRISPR-Cas gene editing:

Implementation Method 2

overexpressing CD16a using CRISPR-Cas editing and lentiviral transfection systems

Methodology Applied

Scientific EffectLentiviral transfection:

Data Source

PatentUS20250066731A1NK cell and preparation method therefor

Publication Date: 2025.02.27 GUANGDONG PROCAPZOOM BIOSCIENCES CO LTD

AI summary

Provided in the present disclosure is an NK cell, including a polynucleotide knocked out of CD38 and overexpressing CD16a. Compared to conventional NK cells, the NK cells of the present disclosure have been inserted with a CD16a overexpression gene and simultaneously knocked out with a CD38 gene, which improves the killing function against tumor cell lines, while simultaneously reducing the mutual killing of NK cells, improving cell survival rate and growth rate, and reducing intracellular consumption.