Nonviral integrating vectors can be used for expression of therapeutic genes. piggyBac (PB), a transposon/transposase system, has been used to efficiently generate induced pluripotent stems cells from somatic cells, without genetic alteration. In this paper, we apply PB transposition to express a chimeric antigen receptor (CAR) in primary human T cells. We demonstrate that T cells electroporated to introduce the PB transposon and transposase stably express CD19-specific CAR and when cultured on CD19+ artificial antigen-presenting cells, numerically expand in a CAR-dependent manner, display a phenotype associated with both memory and effector T cell populations, and exhibit CD19-dependent killing of tumor targets. Integration of the PB transposon expressing CAR was not associated with genotoxicity, based on chromosome analysis. PB transposition for generating human T cells with redirected specificity to a desired target such as CD19 is a new genetic approach with therapeutic implications.
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References
1.
BachmannM.F., WolintP., SchwarzK., JagerP., OxeniusA.2005. Functional properties and lineage relationship of CD8+ T cell subsets identified by expression of IL-7 receptor α and CD62L. J. Immunol., 175:4686–4696.
2.
BergerC., JensenM.C., LansdorpP.M., GoughM., ElliottC., RiddellS.R.2008. Adoptive transfer of effector CD8+ T cells derived from central memory cells establishes persistent T cell memory in primates. J. Clin. Invest., 118:294–305.
3.
BiagiE., MarinV., Giordano AttianeseG.M., DanderE., D'AmicoG., BiondiA.2007. Chimeric T-cell receptors: New challenges for targeted immunotherapy in hematologic malignancies. Haematologica, 92:381–388.
4.
CadinanosJ., BradleyA.2007. Generation of an inducible and optimized piggyBac transposon system. Nucleic Acids Res., 35:e87.
5.
CaryL.C., GoebelM., CorsaroB.G., WangH.G., RosenE., FraserM.J.1989. Transposon mutagenesis of baculoviruses: Analysis of Trichoplusia ni transposon IFP2 insertions within the FP-locus of nuclear polyhedrosis viruses. Virology, 172:156–169.
6.
CooperL.J., ToppM.S., SerranoL.M., GonzalezS., ChangW.C., NaranjoA., WrightC., PopplewellL., RaubitschekA., FormanS.J., JensenM.C.2003. T-cell clones can be rendered specific for CD19: Toward the selective augmentation of the graft-versus-B-lineage leukemia effect. Blood, 101:1637–1644.
7.
CorthayA., SkovsethD.K., LundinK.U., RosjoE., OmholtH., HofgaardP.O., HaraldsenG., BogenB.2005. Primary antitumor immune response mediated by CD4+ T cells. Immunity, 22:371–383.
8.
DingS., WuX., LiG., HanM., ZhuangY., XuT.2005. Efficient transposition of the piggyBac (PB) transposon in mammalian cells and mice. Cell, 122:473–483.
9.
ElickT.A., LoboN., FraserM.J.Jr. 1997. Analysis of the cis-acting DNA elements required for piggyBac transposable element excision. Mol. Gen. Genet., 255:605–610.
10.
FewellJ.G., MatarM., SlobodkinG., HanS.O., RiceJ., HovanesB., LewisD.H., AnwerK.2005. Synthesis and application of a non-viral gene delivery system for immunogene therapy of cancer. J. Control. Release, 109:288–298.
11.
FraserM.J., CaryL., BoonvisudhiK., WangH.G.1995. Assay for movement of lepidopteran transposon IFP2 in insect cells using a baculovirus genome as a target DNA. Virology, 211:397–407.
12.
FraserM.J., CiszczonT., ElickT., BauserC.1996. Precise excision of TTAA-specific lepidopteran transposons piggyBac (IFP2) and tagalong (TFP3) from the baculovirus genome in cell lines from two species of Lepidoptera. Insect Mol. Biol., 5:141–151.
13.
GalvanD.L., NakazawaY., KajaA., KettlunC., CooperL.J., RooneyC.M., WilsonM.H.2009. Genome-wide mapping of PiggyBac transposon integrations in primary human T cells. J. Immunother, 32:837–844.
14.
GonzalezS., NaranjoA., SerranoL.M., ChangW.C., WrightC.L., JensenM.C.2004. Genetic engineering of cytolytic T lymphocytes for adoptive T-cell therapy of neuroblastoma. J. Gene Med., 6:704–711.
15.
GuoG., YangJ., NicholsJ., HallJ.S., EyresI., MansfieldW., SmithA.2009. Klf4 reverts developmentally programmed restriction of ground state pluripotency. Development, 136:1063–1069.
16.
HuangX., GuoH., KangJ., ChoiS., ZhouT.C., TammanaS., LeesC.J., LiZ.Z., MiloneM., LevineB.L., TolarJ., JuneC.H., McIvorR.S., WagnerJ.E., BlazarB.R., ZhouX.2008. Sleeping Beauty transposon-mediated engineering of human primary T cells for therapy of CD19+ lymphoid malignancies. Mol. Ther., 16:580–589.
17.
JensenM.C., PopplewellL., DiGiustoD.L., KalosM., CooperL.J.N., RaubitschekA., FormanS.J.2007. A first-in-human clinical trial of adoptive therapy using CD19-specific chimeric antigen receptor re-directed T cells for recurrent/refractory follicular lymphoma. Mol. Ther., 15:S142.
18.
JonesS., PengP.D., YangS., HsuC., CohenC.J., ZhaoY., AbadJ., ZhengZ., RosenbergS.A., MorganR.A.2009. Lentiviral vector design for optimal T cell receptor gene expression in the transduction of peripheral blood lymphocytes and tumor-infiltrating lymphocytes. Hum. Gene Ther., 20:630–640.
19.
LiX., LoboN., BauserC.A., FraserM.J.Jr. 2001. The minimum internal and external sequence requirements for transposition of the eukaryotic transformation vector piggyBac. Mol. Genet. Genomics, 266:190–198.
20.
LiX., HarrellR.A., HandlerA.M., BeamT., HennessyK., FraserM.J.Jr. 2005. piggyBac internal sequences are necessary for efficient transformation of target genomes. Insect Mol. Biol., 14:17–30.
21.
LiuZ., NohH.S., ChenJ., KimJ.H., FaloL.D.Jr., YouZ.2008. Potent tumor-specific protection ignited by adoptively transferred CD4+ T cells. J. Immunol., 181:4363–4370.
22.
LuZ.Y., CondominesM., TarteK., NadalL., DelteilM.C., RossiJ.F., FerrandC., KleinB.2007. B7-1 and 4-1BB ligand expression on a myeloma cell line makes it possible to expand autologous tumor-specific cytotoxic T cells in vitro. Exp. Hematol., 35:443–453.
23.
LundinK.U., HofgaardP.O., OmholtH., MuntheL.A., CorthayA., BogenB.2003. Therapeutic effect of idiotype-specific CD4+ T cells against B-cell lymphoma in the absence of anti-idiotypic antibodies. Blood, 102:605–612.
24.
MaragathavallyK.J., KaminskiJ.M., CoatesC.J.2006. Chimeric Mos1 and piggyBac transposases result in site-directed integration. FASEB J., 20:1880–1882.
25.
MitraR., Fain-ThorntonJ., CraigN.L.2008. piggyBac can bypass DNA synthesis during cut and paste transposition. EMBO J., 27:1097–1109.
26.
MumbergD., MonachP.A., WanderlingS., PhilipM., ToledanoA.Y., SchreiberR.D., SchreiberH.1999. CD4+ T cells eliminate MHC class II-negative cancer cells in vivo by indirect effects of IFN-γProc. Natl. Acad. Sci. U.S.A., 96:8633–8638.
27.
NewmanJ.C., BaileyA.D., FanH.Y., PavelitzT., WeinerA.M.2008. An abundant evolutionarily conserved CSB–PiggyBac fusion protein expressed in Cockayne syndrome. PLoS Genet., 4:e1000031.
28.
OchsenbeinA.F., RiddellS.R., BrownM., CoreyL., BaerlocherG.M., LansdorpP.M., GreenbergP.D.2004. CD27 expression promotes long-term survival of functional effector-memory CD8+ cytotoxic T lymphocytes in HIV-infected patients. J. Exp. Med., 200:1407–1417.
29.
ParkJ.R., DiGiustoD.L., SlovakM., WrightC., NaranjoA., WagnerJ., MeechoovetH.B., BautistaC., ChangW.C., OstbergJ.R., JensenM.C.2007. Adoptive transfer of chimeric antigen receptor re-directed cytolytic T lymphocyte clones in patients with neuroblastoma. Mol. Ther., 15:825–833.
30.
RolleC.E., CarrioR., MalekT.R.2008. Modeling the CD8+ T effector to memory transition in adoptive T-cell antitumor immunotherapy. Cancer Res., 68:2984–2992.
31.
RossigC., BrennerM.K.2003. Chimeric T-cell receptors for the targeting of cancer cells. Acta Haematol., 110:154–159.
32.
SallustoF., LenigD., ForsterR., LippM., LanzavecchiaA.1999. Two subsets of memory T lymphocytes with distinct homing potentials and effector functions. Nature, 401:708–712.
33.
SarkarA., SimC., HongY.S., HoganJ.R., FraserM.J., RobertsonH.M., CollinsF.H.2003. Molecular evolutionary analysis of the widespread piggyBac transposon family and related “domesticated” sequences. Mol. Genet. Genomics, 270:173–180.
34.
SchertzerJ.D., LynchG.S.2008. Plasmid-based gene transfer in mouse skeletal muscle by electroporation. Methods Mol. Biol., 433:115–125.
35.
SchmiederA.H., GrabskiL.E., MooreN.M., DempseyL.A., Sakiyama-ElbertS.E.2007. Development of novel poly(ethylene glycol)-based vehicles for gene delivery. Biotechnol. Bioeng., 96:967–976.
36.
SerranoL.M., PfeifferT., OlivaresS., NumbenjaponT., BennittJ., KimD., SmithD., McNamaraG., Al-KadhimiZ., RosenthalJ., FormanS.J., JensenM.C., CooperL.J.2006. Differentiation of naive cord-blood T cells into CD19-specific cytolytic effectors for posttransplantation adoptive immunotherapy. Blood, 107:2643–2652.
37.
SinghH., ManuriP.R., OlivaresS., DaraN., DawsonM.J., HulsH., HackettP.B., KohnD.B., ShpallE.J., ChamplinR.E., CooperL.J.2008. Redirecting specificity of T-cell populations for CD19 using the Sleeping Beauty system. Cancer Res., 68:2961–2971.
38.
SmitsE., PonsaertsP., LenjouM., NijsG., Van BockstaeleD.R., BernemanZ.N., Van TendelooV.F.2004. RNA-based gene transfer for adult stem cells and T cells. Leukemia, 18:1898–1902.
39.
Van TendelooV.F., PonsaertsP., BernemanZ.N.2007. mRNA-based gene transfer as a tool for gene and cell therapy. Curr. Opin. Mol. Ther., 9:423–431.
40.
WieheJ.M., PonsaertsP., RojewskiM.T., HomannJ.M., GreinerJ., KronawitterD., SchrezenmeierH., HombachV., WiesnethM., ZimmermannO., TorzewskiJ.2007. mRNA-mediated gene delivery into human progenitor cells promotes highly efficient protein expression. J. Cell. Mol. Med., 11:521–530.
41.
WilliamsD.A.2008. Sleeping beauty vector system moves toward human trials in the United States. Mol. Ther., 16:1515–1516.
42.
WilsonM.H., CoatesC.J., GeorgeA.L.Jr. 2007. PiggyBac transposon-mediated gene transfer in human cells. Mol. Ther., 15:139–145.
WuS.C., MeirY.J., CoatesC.J., HandlerA.M., PelczarP., MoisyadiS., KaminskiJ.M.2006. piggyBac is a flexible and highly active transposon as compared to Sleeping Beauty, Tol2, and Mos1 in mammalian cells. Proc. Natl. Acad. Sci. U.S.A., 103:15008–15013.
45.
XueX., HuangX., NodlandS.E., MatesL., MaL., IzsvakZ., IvicsZ., LebienT.W., McIvorR.S., WagnerJ.E., ZhouX.2009. Stable gene transfer and expression in cord blood-derived CD34+ hematopoietic stem and progenitor cells by a hyperactive Sleeping Beauty transposon system. Blood, 114:1319–1330.
46.
ZanzonicoP., KoehneG., GallardoH.F., DoubrovinM., DoubrovinaE., FinnR., BlasbergR.G., RiviereI., O'ReillyR.J., SadelainM, LarsonS.M.2006. [131I]FIAU labeling of genetically transduced, tumor-reactive lymphocytes: Cell-level dosimetry and dose-dependent toxicity. Eur. J. Nucl. Med. Mol. Imaging, 33:988–997.
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