Humanized IL-15 Animals with Endogenous Locus Regulation
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Solution Overview
Problem
Existing transgenic mice that express human IL-15 from randomly integrated transgenes exhibit abnormal immune cell function and a range of pathologies due to dysregulation, necessitating the development of non-human animals with human IL-15 sequences at an endogenous locus under regulatory control of endogenous elements to achieve physiological relevance and normal lymphocyte populations.
Innovation Solution
Genetically modified non-human animals are created with a humanized IL-15 locus at an endogenous site, regulated by non-human IL-15 regulatory elements, allowing for the expression of human IL-15 protein without abnormal lymphocyte populations or pathologies, and capable of passing this modification through the germline.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If transgenic mice are created with randomly inserted human IL-15 transgenes, then human IL-15 expression is achieved, but abnormal immune cell function and pathologies occur due to dysregulation
Solution Approach 1:
The patent uses endogenous regulatory elements as intermediaries to mediate between the human IL-15 coding sequence and the host genome. These regulatory elements (promoters, enhancers, silencers) act as natural control mechanisms that ensure the transgene is expressed in the correct tissues, at appropriate times, and at physiological levels, thereby preventing the dysregulation and pathologies seen in random integration approaches.
Solution Approach 2:
The patent merges the human IL-15 coding sequence with endogenous regulatory elements into a single functional unit. This combination ensures that the transgene is not only expressed but is regulated in a physiologically relevant manner, mimicking the natural control of the endogenous IL-15 gene while maintaining human sequence identity for proper protein function.
2Ease of manufacture
If human IL-15 sequences are integrated at random locations, then transgene expression is achieved, but dysregulation of immune cell function occurs
Solution Approach 1:
The patent changes the parameter of integration site specificity from random to targeted. By using homologous recombination or CRISPR/Cas9-mediated targeting to specific endogenous loci, the integration precision is dramatically improved. This targeted approach ensures that the human IL-15 transgene is placed in a controlled genomic environment with appropriate regulatory context, preventing the dysregulation associated with random integration while maintaining manufacturing feasibility.
3Quantity of substance
If human IL-15 is expressed without endogenous regulatory control, then expression levels are achieved, but pathologies develop due to lack of physiological regulation
Solution Approach 1:
The patent incorporates endogenous regulatory elements that provide feedback control over human IL-15 expression. These elements include negative feedback mechanisms such as repressor binding sites and microRNA target sites that naturally regulate IL-15 levels. When IL-15 expression reaches physiological thresholds, these feedback mechanisms automatically downregulate transcription or promote mRNA degradation, preventing overexpression-induced pathologies while maintaining adequate expression levels for therapeutic effect.
Data Source
AI summary
Genetically modified non-human animals comprising a humanized interleukin-15 (IL-15) gene. Cells, embryos, and non-human animals comprising a human IL-15 gene. Rodents that express humanized or human IL-15 protein.