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Genetic control of T-cell proliferation with synthetic RNA regulatory systems

Citation

Chen, Yvonne Yu-Hsuan (2011) Genetic control of T-cell proliferation with synthetic RNA regulatory systems. Dissertation (Ph.D.), California Institute of Technology. http://resolver.caltech.edu/CaltechTHESIS:12172010-180148854

Abstract

Adoptive T-cell therapy, or the use of autologous T cells to seek and destroy diseased cells, is a promising treatment option for opportunistic diseases, virus-associated malignancies, and cancers. However, the safety and efficacy of adoptive T-cell therapy depend, in part, on the ability to sustain and tightly regulate the proliferation of transferred T cells in vivo. The emerging field of synthetic biology provides powerful conceptual and technological tools for the construction of regulatory systems that can interface with and reprogram complex biological processes such as cell growth. Here, we present the development of RNA-based regulatory systems that can control T-cell proliferation in a ligand-dependent manner, and examine the construction of integrated control systems capable of fine-tuned programming of cellular behavior. We systematically investigate the translation of ribozyme-based regulatory devices from yeast to mammalian cells and identify design parameters critical to the portability of regulatory devices across host organisms. We report the construction of ligand-responsive ribozyme switch systems capable of modulating the transgenic expression of growth-stimulatory cytokines in mammalian lymphocytes. We demonstrate the ability of ribozyme switch systems to regulate T-cell proliferation in primary human central memory T cells and in animal models. We further develop ligand-responsive, miRNA-based devices to regulate the endogenous expression of cytokine receptor chains and the functional output of cytokine signaling pathways, highlighting the ability to construct integrated T-cell proliferation control systems employing various regulatory mechanisms to modulate multiple components in relevant signaling pathways. Finally, we describe efforts in the generation of novel RNA aptamers to clinically suitable molecules, which can serve as the molecular inputs for ligand-responsive, RNA-based control systems in therapeutic applications. The regulatory systems developed in this work are designed to be modular and transportable across host organisms and application contexts, thus providing a template for future designs in RNA-based genetic regulation. This work demonstrates the capability of RNA-based regulatory systems to advance next-generation treatment options for critical diseases, and highlights the potential of synthetic biological systems to achieve novel and practical functions in diverse applications.

Item Type:Thesis (Dissertation (Ph.D.))
Subject Keywords:nucleic acid therapies, RNA controller, synthetic biology, synthetic riboswitch, immunotherapy
Degree Grantor:California Institute of Technology
Division:Chemistry and Chemical Engineering
Major Option:Chemical Engineering
Minor Option:Biology
Thesis Availability:Mixed availability, specified at file level
Research Advisor(s):
  • Smolke, Christina D.
Thesis Committee:
  • Jensen, Michael C.
  • Davis, Mark E.
  • Tirrell, David A.
  • Smolke, Christina D. (chair)
Defense Date:10 December 2010
Funders:
Funding AgencyGrant Number
National Science Foundation (NSF)Graduate Research Fellowship
City of Hope National Cancer InstituteCancer Center Support Grant
National Institutes of Health (NIH)RC1GM091298
Alfred P. Sloan FoundationFellowship
Record Number:CaltechTHESIS:12172010-180148854
Persistent URL:http://resolver.caltech.edu/CaltechTHESIS:12172010-180148854
Default Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:6215
Collection:CaltechTHESIS
Deposited By: Yvonne Chen
Deposited On:23 Dec 2010 19:28
Last Modified:26 Dec 2012 04:32

Thesis Files

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PDF (Cover, Acknowledgment, Abstract, Table of Contents) - Final Version
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PDF (Chapter 1--Introduction) - Final Version
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PDF (Chapter 2--Translation of Ribozyme-Based Gene Expression Control Devices from Yeast to Mammalian Cells) - Final Version
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PDF (Chapter 3--T-Cell Proliferation Control with Ribozyme-Based Regulatory Systems) - Final Version
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PDF (Chapter 6--In Vitro Selection of RNA Aptamers to Clinically Applicable Small-Molecule Ligands) - Final Version
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PDF (Chapter 4--Proliferation Control of Human Lymphocytes with Ribozyme-Based Regulatory Systems) - Final Version
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PDF (Appendix 1--Small-Molecule Toxicity Curves in CTLL-2 Cells) - Final Version
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PDF (Appendix 2--Small-Molecule Toxicity Curves in NK-92 Cells) - Final Version
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PDF (Chapter 7--Conclusions) - Final Version
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