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Synthetic closed-loop gene circuit for phenylalanine regulation.

Silvia Galvan1, Yu-Qing Xie1, Ana P Teixeira1

  • 1Department of Biosystems Science and Engineering, ETH Zurich, Klingelbergstrasse 48, Basel CH-4056, Switzerland.

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Engineered cells (PRO) can sense and degrade excess phenylalanine, a key issue in phenylketonuria (PKU). This closed-loop system shows promise for treating metabolic diseases by restoring normal amino acid levels.

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Area of Science:

  • Synthetic biology
  • Genetic engineering
  • Cell-based therapies

Background:

  • Cell-based therapies offer potential for chronic disease treatment.
  • Closed-loop systems that self-regulate in response to biomarkers are highly desirable.
  • Phenylketonuria (PKU) is a metabolic disorder characterized by elevated phenylalanine levels.

Purpose of the Study:

  • To engineer a cell-autonomous genetic system (PRO) for sensing and degrading phenylalanine.
  • To develop a self-regulating therapeutic system for PKU treatment.
  • To demonstrate the efficacy of PRO cells in reducing phenylalanine levels in vitro and in vivo.

Main Methods:

  • Engineered a transcriptional switch using human phenylalanine hydroxylase regulatory domain.
  • Optimized the sensing module via random mutagenesis for phenylalanine responsiveness.
  • Expressed phenylalanine-degrading enzymes in engineered human cells (PRO cells).
  • Tested PRO cell efficacy in human whole blood and in a PKU mouse model.

Main Results:

  • PRO cells effectively sensed and degraded excess phenylalanine in a dose-dependent manner.
  • Microencapsulated PRO cells normalized phenylalanine levels in human whole blood.
  • Alginate-encapsulated PRO cells significantly reduced blood phenylalanine in PKU mice.

Conclusions:

  • Synthetic self-regulating systems are promising for treating metabolic diseases.
  • PRO cells demonstrate potential as a novel therapeutic for phenylketonuria.
  • Genetic programming of cells offers a viable strategy for next-generation therapies.