Controlling transgene expression in subcutaneous implants using a skin lotion containing the apple metabolite phloretin

Proc Natl Acad Sci U S A. 2009 Jun 30;106(26):10638-43. doi: 10.1073/pnas.0901501106. Epub 2009 Jun 22.

Abstract

Adjustable control of therapeutic transgenes in engineered cell implants after transdermal and topical delivery of nontoxic trigger molecules would increase convenience, patient compliance, and elimination of hepatic first-pass effect in future therapies. Pseudomonas putida DOT-T1E has evolved the flavonoid-triggered TtgR operon, which controls expression of a multisubstrate-specific efflux pump (TtgABC) to resist plant-derived defense metabolites in its rhizosphere habitat. Taking advantage of the TtgR operon, we have engineered a hybrid P. putida-mammalian genetic unit responsive to phloretin. This flavonoid is contained in apples, and, as such, or as dietary supplement, regularly consumed by humans. The engineered mammalian phloretin-adjustable control element (PEACE) enabled adjustable and reversible transgene expression in different mammalian cell lines and primary cells. Due to the short half-life of phloretin in culture, PEACE could also be used to program expression of difficult-to-produce protein therapeutics during standard bioreactor operation. When formulated in skin lotions and applied to the skin of mice harboring transgenic cell implants, phloretin was able to fine-tune target genes and adjust heterologous protein levels in the bloodstream of treated mice. PEACE-controlled target gene expression could foster advances in biopharmaceutical manufacturing as well as gene- and cell-based therapies.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alkaline Phosphatase / genetics
  • Alkaline Phosphatase / metabolism
  • Animals
  • CHO Cells
  • COS Cells
  • Cell Line
  • Cell Line, Tumor
  • Cell Transplantation / methods*
  • Chlorocebus aethiops
  • Cricetinae
  • Cricetulus
  • Dermatologic Agents / pharmacology*
  • Dose-Response Relationship, Drug
  • Flavonoids / pharmacology
  • Gene Expression Regulation / drug effects*
  • Genetic Vectors / genetics
  • Humans
  • Malus / metabolism
  • Mice
  • NIH 3T3 Cells
  • Operon / genetics
  • Phloretin / pharmacology*
  • Pseudomonas putida / genetics
  • Response Elements / genetics
  • Transfection
  • Transgenes / genetics*

Substances

  • Dermatologic Agents
  • Flavonoids
  • Alkaline Phosphatase
  • Phloretin