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Anti-Tumor Effects after Adoptive Transfer of IL-12 Transposon-Modified Murine Splenocytes in the OT-I-Melanoma Mouse Model.

Galvan DL, O'Neil RT, Foster AE, Huye L, Bear A, Rooney CM, Wilson MH - PLoS ONE (2015)

Bottom Line: Adoptive transfer of gene modified T cells provides possible immunotherapy for patients with cancers refractory to other treatments.We have previously used the non-viral piggyBac transposon system to gene modify human T cells for potential immunotherapy.We next gene-modified OT-I cells to express mIL-12.

View Article: PubMed Central - PubMed

Affiliation: Center for Cell and Gene Therapy, Baylor College of Medicine, Houston, Texas, United States of America.

ABSTRACT
Adoptive transfer of gene modified T cells provides possible immunotherapy for patients with cancers refractory to other treatments. We have previously used the non-viral piggyBac transposon system to gene modify human T cells for potential immunotherapy. However, these previous studies utilized adoptive transfer of modified human T cells to target cancer xenografts in highly immunodeficient (NOD-SCID) mice that do not recapitulate an intact immune system. Currently, only viral vectors have shown efficacy in permanently gene-modifying mouse T cells for immunotherapy applications. Therefore, we sought to determine if piggyBac could effectively gene modify mouse T cells to target cancer cells in a mouse cancer model. We first demonstrated that we could gene modify cells to express murine interleukin-12 (p35/p40 mIL-12), a transgene with proven efficacy in melanoma immunotherapy. The OT-I melanoma mouse model provides a well-established T cell mediated immune response to ovalbumin (OVA) positive B16 melanoma cells. B16/OVA melanoma cells were implanted in wild type C57Bl6 mice. Mouse splenocytes were isolated from C57Bl6 OT-I mice and were gene modified using piggyBac to express luciferase. Adoptive transfer of luciferase-modified OT-I splenocytes demonstrated homing to B16/OVA melanoma tumors in vivo. We next gene-modified OT-I cells to express mIL-12. Adoptive transfer of mIL-12-modified mouse OT-I splenocytes delayed B16/OVA melanoma tumor growth in vivo compared to control OT-I splenocytes and improved mouse survival. Our results demonstrate that the piggyBac transposon system can be used to gene modify splenocytes and mouse T cells for evaluating adoptive immunotherapy strategies in immunocompetent mouse tumor models that may more directly mimic immunotherapy applications in humans.

No MeSH data available.


Related in: MedlinePlus

Homing of piggyBac-modified mouse splenocytes to tumor sites in vivo.OT-I mouse splenocytes were transfected with pCMV-m7pB and pT-effLuc-Thy1.1. 5 X 105 B16/OVA cells into the flank of C57Bl6 mice (day –8). piggyBac-modified splenocytes were adoptively transferred via tail vein injection on day 0 and day +8. Localization of infused splenocytes was visualized via in vivo imaging of luciferase expression on day +11. Show are 3 of 6 representative animals.
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pone.0140744.g004: Homing of piggyBac-modified mouse splenocytes to tumor sites in vivo.OT-I mouse splenocytes were transfected with pCMV-m7pB and pT-effLuc-Thy1.1. 5 X 105 B16/OVA cells into the flank of C57Bl6 mice (day –8). piggyBac-modified splenocytes were adoptively transferred via tail vein injection on day 0 and day +8. Localization of infused splenocytes was visualized via in vivo imaging of luciferase expression on day +11. Show are 3 of 6 representative animals.

Mentions: The Thy1.1 transgene permitted detection of transfection efficiency using flow cytometry, whereas the luciferase transgene (Fig 1) permitted in vivo imaging after adoptive transfer of transfected splenocytes. We implanted 5 X 105 B16/OVA cells into the flank of wild-type C57Bl6 mice (day –8). Splenocytes were isolated from OT-I C57Bl6 mice and transfected with pCMV-PB and pT-effLuc-Thy1.1. Twenty four hours after transfection, activated transgenic OT-I splenocytes were adoptively transferred via tail vein injection on day 0 and day +8. We performed in vivo imaging of luciferase expression to evaluate localization of stably transfected OT-I mouse T cells on day +11. We observed localization of transgenic OT-I splenocytes at sites of tumor eleven days post adoptive transfer (Fig 4). Transgene-modified cells exhibited concentrated localization at the melanoma tumor transplant site. These results confirmed that piggyBac-transgene modified OT-1 splenocytes could be adoptively transferred, imaged with in vivo imaging, and confirmed homing to tumor sites in vivo. However, cells exhibited limited persistence in vivo based on imaging at day 18 post transfer (Fig 4).


Anti-Tumor Effects after Adoptive Transfer of IL-12 Transposon-Modified Murine Splenocytes in the OT-I-Melanoma Mouse Model.

Galvan DL, O'Neil RT, Foster AE, Huye L, Bear A, Rooney CM, Wilson MH - PLoS ONE (2015)

Homing of piggyBac-modified mouse splenocytes to tumor sites in vivo.OT-I mouse splenocytes were transfected with pCMV-m7pB and pT-effLuc-Thy1.1. 5 X 105 B16/OVA cells into the flank of C57Bl6 mice (day –8). piggyBac-modified splenocytes were adoptively transferred via tail vein injection on day 0 and day +8. Localization of infused splenocytes was visualized via in vivo imaging of luciferase expression on day +11. Show are 3 of 6 representative animals.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4608718&req=5

pone.0140744.g004: Homing of piggyBac-modified mouse splenocytes to tumor sites in vivo.OT-I mouse splenocytes were transfected with pCMV-m7pB and pT-effLuc-Thy1.1. 5 X 105 B16/OVA cells into the flank of C57Bl6 mice (day –8). piggyBac-modified splenocytes were adoptively transferred via tail vein injection on day 0 and day +8. Localization of infused splenocytes was visualized via in vivo imaging of luciferase expression on day +11. Show are 3 of 6 representative animals.
Mentions: The Thy1.1 transgene permitted detection of transfection efficiency using flow cytometry, whereas the luciferase transgene (Fig 1) permitted in vivo imaging after adoptive transfer of transfected splenocytes. We implanted 5 X 105 B16/OVA cells into the flank of wild-type C57Bl6 mice (day –8). Splenocytes were isolated from OT-I C57Bl6 mice and transfected with pCMV-PB and pT-effLuc-Thy1.1. Twenty four hours after transfection, activated transgenic OT-I splenocytes were adoptively transferred via tail vein injection on day 0 and day +8. We performed in vivo imaging of luciferase expression to evaluate localization of stably transfected OT-I mouse T cells on day +11. We observed localization of transgenic OT-I splenocytes at sites of tumor eleven days post adoptive transfer (Fig 4). Transgene-modified cells exhibited concentrated localization at the melanoma tumor transplant site. These results confirmed that piggyBac-transgene modified OT-1 splenocytes could be adoptively transferred, imaged with in vivo imaging, and confirmed homing to tumor sites in vivo. However, cells exhibited limited persistence in vivo based on imaging at day 18 post transfer (Fig 4).

Bottom Line: Adoptive transfer of gene modified T cells provides possible immunotherapy for patients with cancers refractory to other treatments.We have previously used the non-viral piggyBac transposon system to gene modify human T cells for potential immunotherapy.We next gene-modified OT-I cells to express mIL-12.

View Article: PubMed Central - PubMed

Affiliation: Center for Cell and Gene Therapy, Baylor College of Medicine, Houston, Texas, United States of America.

ABSTRACT
Adoptive transfer of gene modified T cells provides possible immunotherapy for patients with cancers refractory to other treatments. We have previously used the non-viral piggyBac transposon system to gene modify human T cells for potential immunotherapy. However, these previous studies utilized adoptive transfer of modified human T cells to target cancer xenografts in highly immunodeficient (NOD-SCID) mice that do not recapitulate an intact immune system. Currently, only viral vectors have shown efficacy in permanently gene-modifying mouse T cells for immunotherapy applications. Therefore, we sought to determine if piggyBac could effectively gene modify mouse T cells to target cancer cells in a mouse cancer model. We first demonstrated that we could gene modify cells to express murine interleukin-12 (p35/p40 mIL-12), a transgene with proven efficacy in melanoma immunotherapy. The OT-I melanoma mouse model provides a well-established T cell mediated immune response to ovalbumin (OVA) positive B16 melanoma cells. B16/OVA melanoma cells were implanted in wild type C57Bl6 mice. Mouse splenocytes were isolated from C57Bl6 OT-I mice and were gene modified using piggyBac to express luciferase. Adoptive transfer of luciferase-modified OT-I splenocytes demonstrated homing to B16/OVA melanoma tumors in vivo. We next gene-modified OT-I cells to express mIL-12. Adoptive transfer of mIL-12-modified mouse OT-I splenocytes delayed B16/OVA melanoma tumor growth in vivo compared to control OT-I splenocytes and improved mouse survival. Our results demonstrate that the piggyBac transposon system can be used to gene modify splenocytes and mouse T cells for evaluating adoptive immunotherapy strategies in immunocompetent mouse tumor models that may more directly mimic immunotherapy applications in humans.

No MeSH data available.


Related in: MedlinePlus