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. 2010 Dec;12(8):1056-62.
doi: 10.3109/14653249.2010.506506. Epub 2010 Aug 24.

Adoptive transfer of PR1 cytotoxic T lymphocytes associated with reduced leukemia burden in a mouse acute myeloid leukemia xenograft model

Qing Ma  1 , Changqing Wang, Dan Jones, Kathryn E Quintanilla, Dan Li, Yang Wang, Eric D Wieder, Karen Clise-Dwyer, Gheath Alatrash, You Mj, Mark F Munsell, Sijie Lu, Muzaffar H Qazilbash, Jeffrey J Molldrem
Affiliations

Adoptive transfer of PR1 cytotoxic T lymphocytes associated with reduced leukemia burden in a mouse acute myeloid leukemia xenograft model

Qing Ma et al. Cytotherapy. 2010 Dec.

Abstract

Background aims: Tumor antigen-specific cytotoxic T lymphocytes (CTL) have been used in the treatment of human cancer, including leukemia. Several studies have established PR1 peptide, an HLA-A2.1-restricted peptide derived from proteinase 3 (P3), as a human leukemia-associated antigen. PR1-specific CTL elicited in vitro from healthy donors have been shown to lyse P3-expressing AML cells from patients. We investigated whether PR1-CTL can be adoptively transferred into NOD/SCID mice to eliminate human leukemia cells.

Methods: PR1-CTL were generated in bulk culture from peripheral blood mononuclear cells (PBMC) stimulated with autologous dendritic cells. Human acute myeloid leukemia (AML) patient samples were injected and engrafted in murine bone marrow at 2 weeks post-transfer.

Results: Following adoptive transfer, bone marrow aspirate from mice that received AML alone had 72-88% blasts in a hypercellular marrow, whereas mice that received AML plus PR1-CTL co-infusion had normal hematopoietic elements and only 3-18% blasts in a hypocellular marrow. The PR1-CTL persisted in the bone marrow and liver and maintained a CD45RA⁻CD28+ effector phenotype.

Conclusions: We found that adoptive transfer of PR1-CTL generated in vitro is associated with reduced AML cells in NOD/SCID mice. PR1-CTL can migrate to the sites of disease and maintain their capacity to kill the AML cells. The surface phenotype of PR1-CTL was consistent with their trafficking pattern in both vascular and end-organ tissues.

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Conflict of interest statement

Declaration of interest: we have no conflict of interests.

Figures

Figure 1
Figure 1
Characterization of pre-transplant PR1-CTL. CTL elicited with PR1 from PBMC were characterized for cytotoxicity. (A) PR1-CTL were incubated for 4 h at 37°C with PR1 or pp65 peptide-pulsed T2 cells at the indicated peptide concentrations at an effector:target (E:T) ratio of 10:1, and the percentage specific lysis determined. Three replicate wells were used for each dilution of effector cells. (B) PR1-CTL and PR2-CTL were elicited simultaneously from the same donor in parallel. CTL were incubated for 4 h at 37°C with AML cells from UPN1 patient at an E:T ratio of 10:1 and percentage specific lysis determined. Results are the means and SD from three replicate wells.
Figure 2
Figure 2
Statistical analysis of AML blast counts in NOD/SCID bone marrow. Comparisons between PR1-CTL-treated and AML-alone samples were performed using a two-tailed t-test. Significant differences were noted in both UPN1 (P = 0.0002) and UPN2 (P < 0.0001) groups. An anova test showed significance (P < 0.0001) between PR1-CTL-treated, PR2- or pp65-CTL-treated and AML-alone groups, and the Tukey range test showed significance among all three groups (P < 0.05).
Figure 3
Figure 3
PR1-CTL reduce AML burden in NOD/SCID mice. (A) Giemsa stain of bone marrow (upper panel) and hematoxylin and eosin stain of liver (lower panel). The panels are representative slides from the mice 2 weeks post-transfer of PBS, AML alone, AML plus PR1-CTL, AML plus pp65-CTL or AML plus PR2-CTL. (B) FACS of representative bone marrow and blood from the mice 2 weeks post-transfer of AML plus PR1-CTL (upper panel) and AML alone (lower panel). Human CD45 Ab and mouse CD45.2 Ab were used to identify cells of human origin. The percentage of gated human cells (69.6%) in the bone marrow of AML alone is displayed.
Figure 4
Figure 4
Post-transplant CTL surface phenotype. (A) Cells from representative mice post-transplant with AML plus PR1-CTL were stained with Ab for the surface phenotype of CTL in blood, bone marrow, spleen and liver. The upper panel of each sample displays the percentage of cells positive for human CD8 and PR1 tetramer. The surface marker of these cells, including CD45RA and CD28, are displayed on the lower panel of each sample. (B) Cells from the bone marrow and spleen of mice post-transplant with AML plus pp65-CTL or PR2-CTL were stained with human CD8 and tetramer. Each sample displays the percentage of cells positive for CD8 and pp65 or PR2 tetramer.
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