Page 222 - Haematologica Vol. 110 - January 2025

LETTER TO THE EDITOR
any P2rx7 protein expression on TIB-49 cells in vitro, while after transplantation these cells were found to express higher levels of P2rx7 ex vivo. TIB-49 cells overexpressing P2rx7 with TdTomato as an indicator (TIB-P2rx7high) were generated. WT mice were inoculated with TIB-49, TIB- Cd39high and TIB-P2rx7high cells. Curiously, the enhanced pathogenicity of TIB-49 cells linked to Cd39 overexpression was replicated by transgenic overexpression of P2rx7 on
A
TIB-49 cells (Figure 2B).
P2rx7 in TIB-49 cells was then deleted by CRISPR (TIB- P2rx7-/- cells) (Online Supplementary Figure S2B), and inoc- ulated into WT mice. The engraftment of TIB-P2rx7-/- cells was significantly delayed compared with that of TIB-49 cells (Figure 2C), indicating a role of P2rx7 in AML pathogenicity. We tested the efficacy of a pharmacological P2rx7 inhibitor, A740003. Here too, the survival of standard TIB-49-bearing
BCD
EF
Figure 2. Modulation of P2rx7 signaling impacts Cd39-associated pro-leukemogenic effects in vivo. TIB-Cd39high and TIB-P2rx7high cells showed enhanced pathogenicity in wild-type mice, while P2rx7 deletion in TIB-49 and TIB-Cd39high cells delayed engraft- ment of leukemic cells. A pharmacological P2rx7 inhibitor prolonged the survival of TIB-49-bearing mice with acute myeloid leukemia, but not of TIB-Cd39high leukemic mice. (A) Quantification of selected P2 receptor expression in 151 human acute myeloid leukemia samples in The Cancer Genome Atlas database with high or low expression of CD39 relative to the median expression level. The Mann-Whitney test was used for the statistical analysis. *P<0.05, ****P<0.0001. (B) C57BL/6 wild-type (WT) mice were inoculated with TIB-49, TIB-Cd39high or TIB-P2rx7high cells through tail vein injection. The graph shows the time to euthanasia of mice inoculated with TIB-49high (N=11), TIB-Cd39high (N=12) or TIB-P2rx7high (N=6) cells. The log-rank test was used for statistical analyses. TIB-Cd39high and TIB-P2rx7high cells showed enhanced pathogenicity in WT mice. (C) C57BL/6 WT mice were inoculated with TIB-49 cells or TIB-P2rx7-/- cells through tail vein injection. Times to euthanasia of mice inoculated with TIB-49 (N=15) or TIB-P2rx7-/- (N=15) cells were then compared. The log-rank test was used for statistical analyses. P2rx7 deletion in TIB-49 cells delayed engraftment in WT mice. (D, E) C57BL/6 WT mice were inoculated with 1×106 TIB-49 cells (D) or TIB-Cd39high cells (E) through tail vein injection. Mice received 50 mg/kg P2rx7 antagonist A740003 or dimethylsulfoxide (DMSO) vehicle treatment from day 15 every other day for 14 days. Times to euthanasia of mice bearing TIB-49 cells (N=15 in each group) or TIB-Cd39high cells (N=10 in each group) were compared. The log-rank test was used for statistical analyses. Benefits potentially seen with A740003 alone for TIB-49 cells, were not noted in the TIB-Cd39high group. (F) C57BL/6 WT mice were inoculated with TIB-Cd39high cells or TIB-Cd39high-P2rx7-/- cells through tail vein injection. Times to euthanasia of mice inoculated with TIB-Cd39high cells (N=10) or TIB- Cd39high-P2rx7-/- cells (N=10) were compared. The log-rank test was used for statistical analyses and indicated a benefit from genetic deletion of P2rx7 in these cells.
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