Universal Donor Cells With MHC Editing and Tolerogenic Factors
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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 risks from residual MHC-I expression and susceptibility to natural killer cells.
Innovation Solution
Genetic modification of cells by introducing deletions and insertions in MHC-I and MHC-II genes, combined with the introduction of tolerogenic factors like PD-L1 and HLA-E, to reduce immunogenicity and enhance survival through the use of RNA-guided endonucleases such as CRISPR-Cas9.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If MHC-I and MHC-II genes are disrupted to reduce immunogenicity, then immune evasion is improved, but residual MHC-I expression may still cause immune rejection
Solution Approach 1:
The patent changes the molecular parameters of the cell by disrupting MHC-I and MHC-II genes through CRISPR-Cas9-mediated homology-directed repair, introducing specific mutations (deletions, insertions, or substitutions) that prevent proper MHC molecule expression while maintaining cell function
Solution Approach 2:
The patent extracts or removes the immunogenic components (MHC-I and MHC-II molecules) from the cell surface through genetic disruption, eliminating the target structures that trigger immune rejection while preserving the cell's therapeutic function
2Reliability
If MHC-I expression is minimized to evade CD8+ T cells, then cellular immune rejection is reduced, but susceptibility to natural killer cells increases
Solution Approach 1:
The patent changes the immunophenotypic parameters of the cells by disrupting MHC-I genes to reduce CD8+ T cell recognition, while simultaneously introducing tolerogenic factors (HLA-E, HLA-G, PD-L1) that modify NK cell interaction parameters to maintain resistance
Solution Approach 2:
The patent creates a composite immunophenotype by combining multiple genetic modifications (MHC disruption plus tolerogenic factor introduction) to achieve a cell surface profile that simultaneously evades both CD8+ T cells and NK cells
3Reliability
If multiple genetic modifications are introduced to enhance immune evasion, then allogeneic rejection is reduced, but cell complexity and potential off-target effects increase
Solution Approach 1:
The patent applies a universal CRISPR-Cas9 platform that can deliver multiple guide RNAs and donor templates simultaneously, enabling multiple genetic modifications through a single transfection event, thereby reducing procedural complexity while achieving comprehensive immune evasion
Solution Approach 2:
The patent uses CRISPR-Cas9 as an intermediary system that coordinates multiple genetic modifications through a unified mechanism, where the Cas9 enzyme guided by multiple gRNAs simultaneously targets different genes, simplifying the delivery and execution of complex genetic edits
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 one or more MHC-I or MHC-II human leukocyte antigens or component or transcriptional regulator of the MHC-I or MHC-II complex, at least one genetic modification that increases the expression of at least one polynucleotide that encodes a tolerogenic factor, and optionally at least one genetic modification that increases or decreases the expression of at least one gene that encodes a survival factor.