Chronic kidney disease (CKD) affects an estimated 7% of the US population and results in scarring and loss of peritubular fibroblasts which produce erythropoietin (EPO). EPO-deficient anemia of CKD is currently treated with recombinant EPO analog injections that have recently been associated with undesired side effects such as increased risk of stroke, heart attacks, and deep vein thrombosis which may preclude further use of this therapy. Although the mechanisms of these side effects are unclear, it is clear that bolus dosing of EPO analogs either weekly or monthly does not recapitulate the physiologic regulation of this important hormone and bolus dosing may alter EPO signaling pathways. Thus, there is a critical need to develop alternative therapies for anemia of CKD. Herein we describe an innovative experimental design using non-viral transposon-mediated gene transfer to develop a new strategy for therapy of anemia of CKD. Genetically modified T lymphocytes whose specificity is directed to persistent (latent) viruses such as Epstein-Barr virus (EBV) survive long-term (>8 years) in stable numbers in vivo due to chronic viral antigen stimulation. Moreover, preclinical and recent clinical studies have shown T cells can be readily induced to apoptose by activation of a co-transferred suicide gene, providing an additional layer of safety and control. We therefore hypothesize that virus specific T cells genetically modified to inducibly express EPO and a separately inducible suicide gene represent an ideal candidate cell population for sustained and safe treatment of anemia of CKD.
In specific aim 1, we propose to modify virus specific murine T cells to inducibly express EPO and a suicide gene and we will infuse them into wild type and CKD mice to measure their effectiveness in regulating hematocrit levels in vivo.
Specific aim 2 focuses on extending these genetic modifications to human T cells and testing them in vitro for their ability to be propagated long-term via chronic viral antigen stimulation, as well as inducibly express EPO and undergo selectively induced cell ablation if needed.
In specific aim 3, we will evaluate the functionality of genetically modified human T cells from patients with CKD and determine the frequency of EBV-specific T cells and their response to EBV antigen in the presence and absence of transgenically expressed EPO ex vivo.

Public Health Relevance

This project is focused on developing an efficient, safe, and novel cell therapy for anemia of chronic kidney disease. The proposed strategy could also be used for therapy for a variety of other human diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK093660-05
Application #
8880194
Study Section
Pathobiology of Kidney Disease Study Section (PBKD)
Program Officer
Kusek, John W
Project Start
2012-07-01
Project End
2017-06-30
Budget Start
2015-07-01
Budget End
2016-06-30
Support Year
5
Fiscal Year
2015
Total Cost
$341,475
Indirect Cost
$123,975
Name
Vanderbilt University Medical Center
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
004413456
City
Nashville
State
TN
Country
United States
Zip Code
37212
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Hamada, Motoharu; Nishio, Nobuhiro; Okuno, Yusuke et al. (2018) Integration Mapping of piggyBac-Mediated CD19 Chimeric Antigen Receptor T Cells Analyzed by Novel Tagmentation-Assisted PCR. EBioMedicine 34:18-26
Veach, Ruth Ann; Wilson, Matthew H (2018) CRISPR/Cas9 engineering of a KIM-1 reporter human proximal tubule cell line. PLoS One 13:e0204487
O'Neil, Richard T; Saha, Sunandan; Veach, Ruth Ann et al. (2018) Transposon-modified antigen-specific T lymphocytes for sustained therapeutic protein delivery in vivo. Nat Commun 9:1325
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Woodard, Lauren E; Downes, Laura M; Lee, Yi-Chien et al. (2017) Temporal self-regulation of transposition through host-independent transposase rodlet formation. Nucleic Acids Res 45:353-366
Nakazawa, Yozo; Matsuda, Kazuyuki; Kurata, Takashi et al. (2016) Anti-proliferative effects of T cells expressing a ligand-based chimeric antigen receptor against CD116 on CD34(+) cells of juvenile myelomonocytic leukemia. J Hematol Oncol 9:27
Galvan, Daniel L; O'Neil, Richard T; Foster, Aaron E et al. (2015) Anti-Tumor Effects after Adoptive Transfer of IL-12 Transposon-Modified Murine Splenocytes in the OT-I-Melanoma Mouse Model. PLoS One 10:e0140744

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