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.
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.
|Luo, Wentian; Galvan, Daniel L; Woodard, Lauren E et al. (2017) Comparative analysis of chimeric ZFP-, TALE- and Cas9-piggyBac transposases for integration into a single locus in human cells. Nucleic Acids Res 45:8411-8422|
|Woodard, Lauren E; Cheng, Jizhong; Welch, Richard C et al. (2017) Kidney-specific transposon-mediated gene transfer in vivo. Sci Rep 7:44904|
|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|
|Saha, Sunandan; Woodard, Lauren E; Charron, Elizabeth M et al. (2015) Evaluating the potential for undesired genomic effects of the piggyBac transposon system in human cells. Nucleic Acids Res 43:1770-82|
|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|
|Woodard, Lauren E; Wilson, Matthew H (2015) piggyBac-ing models and new therapeutic strategies. Trends Biotechnol 33:525-33|
|Saito, Shoji; Nakazawa, Yozo; Sueki, Akane et al. (2014) Anti-leukemic potency of piggyBac-mediated CD19-specific T cells against refractory Philadelphia chromosome-positive acute lymphoblastic leukemia. Cytotherapy 16:1257-69|
|Galvan, Daniel L; Kettlun, Claudia S; Wilson, Matthew H (2014) Targeting piggyBac transposon integrations in the human genome. Methods Mol Biol 1114:143-61|
|Nakazawa, Yozo; Saha, Sunandan; Galvan, Daniel L et al. (2013) Evaluation of long-term transgene expression in piggyBac-modified human T lymphocytes. J Immunother 36:3-10|
Showing the most recent 10 out of 13 publications