Universal Donor Cell Engineering for MHC Evasion and NK Survival
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Solution Overview
Problem
Existing methods for generating universal donor cells face challenges in effectively evading immune rejection and ensuring cell survival post-engraftment, with potential issues such as residual MHC-I expression and susceptibility to natural killer cells, and there is a need for improved strategies to generate cells that can survive and evade immune responses.
Innovation Solution
A method involving site-directed nucleases, such as CRISPR systems, are used to target genes encoding survival and tolerogenic factors like TXNIP, HLA-E, and PD-L1, along with MHC-I and MHC-II antigens, to generate universal donor cells with enhanced immune evasion and survival capabilities.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Object-affected harmful factors
If MHC-I and MHC-II genes are disrupted to reduce immunogenicity, then immune rejection is reduced, but cell survival becomes compromised due to increased susceptibility to natural killer cells
Solution Approach 1:
The patent segments the immune evasion strategy into multiple independent genetic modifications: disrupting MHC-I genes (HLA-A, HLA-B, HLA-C) and MHC-II genes separately, and independently introducing compensatory survival factors. This modular approach allows each modification to be optimized and controlled separately, resolving the contradiction between immune evasion and survival.
Solution Approach 2:
The patent combines multiple genetic modifications into a single integrated solution: simultaneous disruption of MHC-I and MHC-II genes together with introduction of survival factors (CXCL12, BCL2, MYC) and tolerogenic factors (HLA-E, PD-L1). This merging creates a synergistic effect where immune evasion and survival enhancement work together rather than oppose each other.
Solution Approach 3:
The patent introduces intermediary molecules that mediate between the disrupted MHC system and the immune system: HLA-E as an intermediary MHC class I molecule that provides NK cell resistance without triggering strong T-cell responses, and PD-L1 as an intermediary that dampens T-cell activation. These intermediaries resolve the contradiction by providing alternative protective mechanisms.
2Object-affected harmful factors
If multiple genetic modifications are introduced to enhance immune evasion, then immune rejection is reduced, but the complexity of the genetic engineering process increases
Solution Approach 1:
The patent performs preliminary actions by pre-designing and pre-testing multiple genetic modification combinations in vitro before clinical application. The extensive characterization of cell lines with various MHC disruptions and survival factor introductions beforehand allows for selection of optimal combinations, reducing the complexity of clinical implementation.
Solution Approach 2:
The patent systematically varies genetic parameters (which MHC genes to disrupt, which survival factors to introduce) to identify optimal combinations. By changing these parameters in a controlled manner and characterizing the results, the patent reduces the effective complexity by finding simplified optimal solutions among many possibilities.
3Adaptability or versatility
If MHC-I genes are completely disrupted to achieve universal donor status, then compatibility with any recipient is improved, but residual MHC-I expression may still occur causing immune rejection
Solution Approach 1:
The patent applies partial action by disrupting but not necessarily completely eliminating all MHC-I gene function. The use of HLA-E as a compensatory molecule provides sufficient immune evasion without requiring absolute elimination of all MHC-I activity. This partial approach achieves universal donor status while avoiding the pitfalls of incomplete suppression.
Solution Approach 2:
The patent converts the potential harm of residual MHC-I expression into a benefit by introducing HLA-E, which can be expressed from the disrupted MHC-I loci or independently. HLA-E provides protective function against NK cells while having reduced immunogenicity to T cells, thus converting the problem of residual MHC-I expression into a solution.
Data Source
AI summary
Genetically modified cells that are compatible with multiple subjects, e.g., universal donor cells, and methods of generating said genetic modified cells are provided herein. The universal donor cells comprise at least one genetic modification within or near at least one gene that encodes a survival factor, wherein the genetic modification comprises an insertion of a polynucleotide encoding a tolerogenic factor. The universal donor cells may further comprise at least one genetic modification within or near a gene that encodes one or more MHC-I or MHC-II human leukocyte antigens or a component or a transcriptional regulator of a MHC-I or MHC-II complex, wherein said genetic modification comprises an insertion of a polynucleotide encoding a second tolerogenic factor.