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Adoptive T-cell therapy improves treatment of canine non-Hodgkin lymphoma post chemotherapy.

O'Connor CM, Sheppard S, Hartline CA, Huls H, Johnson M, Palla SL, Maiti S, Ma W, Davis RE, Craig S, Lee DA, Champlin R, Wilson H, Cooper LJ - Sci Rep (2012)

Bottom Line: Graded doses of autologous T cells were infused after CHOP chemotherapy and persisted for 49 days, homed to tumor, and significantly improved survival.Serum thymidine kinase changes predicted T-cell engraftment, while anti-tumor effects correlated with neutrophil-to-lymphocyte ratios and granzyme B expression in manufactured T cells.The companion canine model has translational implications for human immunotherapy which can be readily exploited since clinical-grade canine and human T cells are propagated using identical approaches.

View Article: PubMed Central - PubMed

Affiliation: Division of Pediatrics, The University of Texas MD Anderson Cancer Center, Houston, Texas, USA.

ABSTRACT
Clinical observations reveal that an augmented pace of T-cell recovery after chemotherapy correlates with improved tumor-free survival, suggesting the add-back of T cells after chemotherapy may improve outcomes. To evaluate adoptive immunotherapy treatment for B-lineage non-Hodgkin lymphoma (NHL), we expanded T cells from client-owned canines diagnosed with NHL on artificial antigen presenting cells (aAPC) in the presence of human interleukin (IL)-2 and IL-21. Graded doses of autologous T cells were infused after CHOP chemotherapy and persisted for 49 days, homed to tumor, and significantly improved survival. Serum thymidine kinase changes predicted T-cell engraftment, while anti-tumor effects correlated with neutrophil-to-lymphocyte ratios and granzyme B expression in manufactured T cells. Therefore, chemotherapy can be used to modulate infused T-cell responses to enhance anti-tumor effects. The companion canine model has translational implications for human immunotherapy which can be readily exploited since clinical-grade canine and human T cells are propagated using identical approaches.

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Tracking infused T cells.(a) Ratio of CD4:CD8 T cells in PB after CHOP, but before adoptive transfer of T cells and compared with measurements taken three hours after each T-cell infusion. The grey shaded area represents the mean CD4:CD8 ratio (1.6:1) in healthy subjects (n = 4). (b) Mean T-cell counts in PB from 6 canines before and after adoptive transfer of T cells. The grey shaded area represents the range for CD8+ T cells in healthy subjects (n = 4, 581 to 958 cells/µL). Arrows represent days T cells were infused. (c) Mean expression of CD3+ T cells pre-stained with red fluorescent dye, PKH-26, in the PB of 6 canines. Arrows represent days T cells were infused. (d) Evaluation of fluorescence and staining of CD3 from a LN biopsy. Frozen tissues were viewed with fluorescent microscopy to detect (top) PKH-26+ (red) T cells and (bottom) co-stained with anti-CD3 (black) to validate T-cell presence.
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f4: Tracking infused T cells.(a) Ratio of CD4:CD8 T cells in PB after CHOP, but before adoptive transfer of T cells and compared with measurements taken three hours after each T-cell infusion. The grey shaded area represents the mean CD4:CD8 ratio (1.6:1) in healthy subjects (n = 4). (b) Mean T-cell counts in PB from 6 canines before and after adoptive transfer of T cells. The grey shaded area represents the range for CD8+ T cells in healthy subjects (n = 4, 581 to 958 cells/µL). Arrows represent days T cells were infused. (c) Mean expression of CD3+ T cells pre-stained with red fluorescent dye, PKH-26, in the PB of 6 canines. Arrows represent days T cells were infused. (d) Evaluation of fluorescence and staining of CD3 from a LN biopsy. Frozen tissues were viewed with fluorescent microscopy to detect (top) PKH-26+ (red) T cells and (bottom) co-stained with anti-CD3 (black) to validate T-cell presence.

Mentions: The ratio of CD4:CD8 T cells in PB was serially assessed as a measure of persistence of infused predominately CD8+ T cells (Fig 4A). The CD4:CD8 ratio in healthy subjects was 1.6:1 ± 0.2 (n = 4) while the mean pre-infusion ratio in canines with NHL was 3:1. After each infusion (predominately containing CD8+ T cells), the average ratio decreased to 1:1.2 after the third dose (n = 6), which was significantly reduced compared to pre-infusion measurements (p = 0.05). We also observed that the percentage of CD3+CD8+ T cells in PB increased after each T-cell infusion. The mean number of CD3+CD4+ and CD3+CD8+ T cells pre-infusion was 585 ± 223 cells/µL and 188 ± 44 cells/µL, respectively. By study day 35, the mean CD3+CD4+ and CD3+CD8+ T-cell counts in PB were 305 ± 166 cells/µL and 469 ± 142 cells/µL, respectively. Similarly, the mean absolute lymphocyte count (ALC) increased after the T-cell infusions (Supplementary Table S3) which reflected an increase (from 64% to 79%) in CD3+ T cells above pre-infusion levels over the same 35 day time frame (Fig 4B). To directly examine the persistence of adoptively transferred T cells, the infusion products were pre-labeled with PKH-26, a red fluorescent dye. PKH-26+CD3+ T cells incrementally increased in the PB after each of the three graded T-cell infusions and were detectable for up to 49 days (Fig 4C). Toxicities attributed to T cells were confined within a 72-hour period after the second T-cell infusion with the majority of the seven recorded adverse events being Grade I or II (Table 2). One canine with a Grade III gastrointestinal adverse event required hospitalization for dehydration (24 hours). In aggregate, these data support that multiple doses of ex vivo propagated T cells can be safely administered to canines with NHL and persist after infusion.


Adoptive T-cell therapy improves treatment of canine non-Hodgkin lymphoma post chemotherapy.

O'Connor CM, Sheppard S, Hartline CA, Huls H, Johnson M, Palla SL, Maiti S, Ma W, Davis RE, Craig S, Lee DA, Champlin R, Wilson H, Cooper LJ - Sci Rep (2012)

Tracking infused T cells.(a) Ratio of CD4:CD8 T cells in PB after CHOP, but before adoptive transfer of T cells and compared with measurements taken three hours after each T-cell infusion. The grey shaded area represents the mean CD4:CD8 ratio (1.6:1) in healthy subjects (n = 4). (b) Mean T-cell counts in PB from 6 canines before and after adoptive transfer of T cells. The grey shaded area represents the range for CD8+ T cells in healthy subjects (n = 4, 581 to 958 cells/µL). Arrows represent days T cells were infused. (c) Mean expression of CD3+ T cells pre-stained with red fluorescent dye, PKH-26, in the PB of 6 canines. Arrows represent days T cells were infused. (d) Evaluation of fluorescence and staining of CD3 from a LN biopsy. Frozen tissues were viewed with fluorescent microscopy to detect (top) PKH-26+ (red) T cells and (bottom) co-stained with anti-CD3 (black) to validate T-cell presence.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3278154&req=5

f4: Tracking infused T cells.(a) Ratio of CD4:CD8 T cells in PB after CHOP, but before adoptive transfer of T cells and compared with measurements taken three hours after each T-cell infusion. The grey shaded area represents the mean CD4:CD8 ratio (1.6:1) in healthy subjects (n = 4). (b) Mean T-cell counts in PB from 6 canines before and after adoptive transfer of T cells. The grey shaded area represents the range for CD8+ T cells in healthy subjects (n = 4, 581 to 958 cells/µL). Arrows represent days T cells were infused. (c) Mean expression of CD3+ T cells pre-stained with red fluorescent dye, PKH-26, in the PB of 6 canines. Arrows represent days T cells were infused. (d) Evaluation of fluorescence and staining of CD3 from a LN biopsy. Frozen tissues were viewed with fluorescent microscopy to detect (top) PKH-26+ (red) T cells and (bottom) co-stained with anti-CD3 (black) to validate T-cell presence.
Mentions: The ratio of CD4:CD8 T cells in PB was serially assessed as a measure of persistence of infused predominately CD8+ T cells (Fig 4A). The CD4:CD8 ratio in healthy subjects was 1.6:1 ± 0.2 (n = 4) while the mean pre-infusion ratio in canines with NHL was 3:1. After each infusion (predominately containing CD8+ T cells), the average ratio decreased to 1:1.2 after the third dose (n = 6), which was significantly reduced compared to pre-infusion measurements (p = 0.05). We also observed that the percentage of CD3+CD8+ T cells in PB increased after each T-cell infusion. The mean number of CD3+CD4+ and CD3+CD8+ T cells pre-infusion was 585 ± 223 cells/µL and 188 ± 44 cells/µL, respectively. By study day 35, the mean CD3+CD4+ and CD3+CD8+ T-cell counts in PB were 305 ± 166 cells/µL and 469 ± 142 cells/µL, respectively. Similarly, the mean absolute lymphocyte count (ALC) increased after the T-cell infusions (Supplementary Table S3) which reflected an increase (from 64% to 79%) in CD3+ T cells above pre-infusion levels over the same 35 day time frame (Fig 4B). To directly examine the persistence of adoptively transferred T cells, the infusion products were pre-labeled with PKH-26, a red fluorescent dye. PKH-26+CD3+ T cells incrementally increased in the PB after each of the three graded T-cell infusions and were detectable for up to 49 days (Fig 4C). Toxicities attributed to T cells were confined within a 72-hour period after the second T-cell infusion with the majority of the seven recorded adverse events being Grade I or II (Table 2). One canine with a Grade III gastrointestinal adverse event required hospitalization for dehydration (24 hours). In aggregate, these data support that multiple doses of ex vivo propagated T cells can be safely administered to canines with NHL and persist after infusion.

Bottom Line: Graded doses of autologous T cells were infused after CHOP chemotherapy and persisted for 49 days, homed to tumor, and significantly improved survival.Serum thymidine kinase changes predicted T-cell engraftment, while anti-tumor effects correlated with neutrophil-to-lymphocyte ratios and granzyme B expression in manufactured T cells.The companion canine model has translational implications for human immunotherapy which can be readily exploited since clinical-grade canine and human T cells are propagated using identical approaches.

View Article: PubMed Central - PubMed

Affiliation: Division of Pediatrics, The University of Texas MD Anderson Cancer Center, Houston, Texas, USA.

ABSTRACT
Clinical observations reveal that an augmented pace of T-cell recovery after chemotherapy correlates with improved tumor-free survival, suggesting the add-back of T cells after chemotherapy may improve outcomes. To evaluate adoptive immunotherapy treatment for B-lineage non-Hodgkin lymphoma (NHL), we expanded T cells from client-owned canines diagnosed with NHL on artificial antigen presenting cells (aAPC) in the presence of human interleukin (IL)-2 and IL-21. Graded doses of autologous T cells were infused after CHOP chemotherapy and persisted for 49 days, homed to tumor, and significantly improved survival. Serum thymidine kinase changes predicted T-cell engraftment, while anti-tumor effects correlated with neutrophil-to-lymphocyte ratios and granzyme B expression in manufactured T cells. Therefore, chemotherapy can be used to modulate infused T-cell responses to enhance anti-tumor effects. The companion canine model has translational implications for human immunotherapy which can be readily exploited since clinical-grade canine and human T cells are propagated using identical approaches.

Show MeSH
Related in: MedlinePlus