Targeting the pentose phosphate pathway mitigates graft-versus-host disease by rewiring alloreactive T cell metabolism
Saeed Daneshmandi, Eun Ko, Qi Yan, Jee Eun Choi, Prashant K. Singh, Richard M. Higashi, Andrew N. Lane, Teresa W.M. Fan, Jingxin Qiu, Sophia Hani, Keli L. Hippen, Jianmin Wang, Philip L. McCarthy, Bruce R. Blazar, Hemn Mohammadpour
Saeed Daneshmandi, Eun Ko, Qi Yan, Jee Eun Choi, Prashant K. Singh, Richard M. Higashi, Andrew N. Lane, Teresa W.M. Fan, Jingxin Qiu, Sophia Hani, Keli L. Hippen, Jianmin Wang, Philip L. McCarthy, Bruce R. Blazar, Hemn Mohammadpour
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Research Article Immunology Oncology

Targeting the pentose phosphate pathway mitigates graft-versus-host disease by rewiring alloreactive T cell metabolism

Abstract

Glycolysis fuels cytotoxic allogeneic T cells in acute graft-versus-host disease (aGvHD), but the downstream role of glucose metabolism in modulating aGvHD remains unclear. Targeting glycolysis or glucose receptors is toxic. Therefore, we explored alternative glucose-dependent pathways, focusing on the pentose phosphate pathway (PPP). Single-cell RNA sequencing revealed PPP upregulation in allogeneic T cells during allogeneic hematopoietic cell transplantation (allo-HCT). We showed that donor T cell deficiency in 6-phosphogluconate dehydrogenase (6PGD), the second rate-limiting enzyme in the PPP, significantly reduced aGvHD severity and mortality in murine models. Functional assays demonstrated that PPP blockade led to proliferation arrest without inducing apoptosis. PPP blockade shifted T cell metabolism away from T cell dependency on glycolysis for rapid T cell proliferation. Pharmacological inhibition of the PPP through 6PGD blockade with 6-aminonicotinamide (6AN) effectively reduced aGvHD severity, like donor 6PGD-deficient T cells in an allogeneic aGvHD model. Similarly, 6AN reduced xenogeneic GvHD lethality. 6PGD inhibition preserved the graft-versus-tumor (GvT) effect, with the generation of a small subset of granzyme Bhi effector T cells with potent antitumor activity. These findings highlight the PPP as a key regulator of allogeneic T cell proliferation and differentiation and identify 6PGD as a promising therapeutic target to mitigate aGvHD severity while preserving beneficial GvT effects.

Authors

Saeed Daneshmandi, Eun Ko, Qi Yan, Jee Eun Choi, Prashant K. Singh, Richard M. Higashi, Andrew N. Lane, Teresa W.M. Fan, Jingxin Qiu, Sophia Hani, Keli L. Hippen, Jianmin Wang, Philip L. McCarthy, Bruce R. Blazar, Hemn Mohammadpour

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Figure 1

Blockade of oxidative PPP at the 6PGD metabolic checkpoint ameliorates the severity of and the mortality due to aGvHD.

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Blockade of oxidative PPP at the 6PGD metabolic checkpoint ameliorates t...
(A and B) BALB/c (H-2d) mice were lethally irradiated (8.5 Gy) on day –1 and transplanted with 3.5 × 106 T cell–depleted bone marrow (TCD-BM) cells and 0.7 × 106 purified splenic WT T cells from C57BL/6 (H-2b) mice on day 0. Splenic donor T cells (CD45+H2Kb+H2Kd–TCRβ+) were sorted and examined by scRNA-seq (A). The expression of oxidative PPP gene signatures was detected within the T cell clusters (B). T cells were isolated and pooled for analysis from 5 spleens. (C and D) Allo-HCT model was generated as in A and B using 0.2 × 106 purified splenic WT naive T cells. The expression levels of oxidative PPP pathway genes (C) and the Pgd gene (6PGD) (D) were examined in sorted alloreactive T cells (Allo: CD45+H2Kb+H2Kd–TCRβ+) and from BALB/c mice transplanted with syngeneic mice T cells (Syn: CD45+H2Kb–H2Kd+TCRβ+) on day +7. T cells were pooled from 5 spleens (1-way ANOVA). (E–G) BALB/c (H-2d) mice were lethally irradiated (8.5 Gy) on day –1 and transplanted with 3.5 × 106 TCD-BM cells with or without 0.2 × 106 splenic naive T cells from WT (Pgdfl/fl) or 6PGD-deficient (Pgdfl/flCd4Cre) C57BL/6 (H-2b) mice on day 0. Body weight loss (E) and clinical GvHD scores (F) are presented as mean ± SEM. n = 5 mice per group (2-way ANOVA). Survival data (G) are presented as percentage survival (log-rank Mantel-Cox test). (H and I) Allo-HCT model was generated as in E–G. GvHD progression was examined in the GI tract on days +7, +14, and +21. Representative images of H&E staining demonstrate infiltration of intraepithelial lymphocytes (blue arrows) and apoptosis (red arrows) (H). Original magnification, ×200. Semiquantitative GvHD scoring of small (top) and large (bottom) intestines are shown. n = 5 mice per group (Student’s t test). (J–L) C3H/SW (H-2b) mice were lethally irradiated (11.5 Gy) on day –1 and transplanted with 3.5 × 106 TCD-BM cells with or without 0.5 × 106 splenic naive T cells from Pgdfl/fl or Pgdfl/flCd4Cre C57BL/6 (H-2b) mice on day 0 as minor-mismatched model. Body weight loss (J) and clinical GvHD score (K) data are presented as mean ± SEM. Data are representative of 2 independent experiments (2-way ANOVA). Survival data are presented as percentage survival (log-rank Mantel-Cox test) (L). Data are shown as mean ± SEM. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001.