Type of T Cells Key in cGVHD Bronchiolitis Obliterans Syndrome After Allo-HSCT

Granzyme K (GZMK)-expressing CD8⁺ T cells were identified as an important component of the pathogenesis of bronchiolitis obliterans syndrome (BOS) that can occur following allogeneic hematopoietic stem cell transplantation (allo-HSCT) as a late-stage complication of chronic graft-versus-host disease (cGVHD), according to study results presented at ASH Annual Meeting 2023.

In patients who undergo allo-HSCT, cGVHD is the main cause of nonrelapse mortality and persists as a complicating factor contributing to worsening patient-reported quality of life and a higher incidence of secondary malignancies. In particular, BOS is a late-stage complication of cGVHD with a 5-year survival rate of 40% to 50%. Therapeutic regimens for BOS are severely lacking, due in part to poorly elucidated pathogenesis.

This research, presented by by Haowen Xiao, MD, PhD, a visiting investigator at Memorial Sloan Kettering Cancer Center in New York, first generated a comprehensive immune map of patients with cGVHD. In total, 22 patients with cGVHD and 11 patients without cGVHD after allo-HSCT have been enrolled, with collections of peripheral blood mononuclear cell samples. Of 22 patients with cGVHD, 10 had cGVHD with only skin involvement, and 12 had lung involvement with diagnosed BOS.

Via mass cytometry, bulk RNA-sequencing, and cytokine array, researchers showed circulating GZMK⁺CD8⁺ T cells accumulated in patients with cGVHD BOS. In vitro analysis of human embryonic lung fibroblast HFL-1 cells showed increased expression of genes related to fibrosis when human GZMK was added, suggesting a role for GZMK⁺CD8⁺ T cells in the development of lung fibrosis in cGVHD-BOS. Additional molecular assessments with RT-qPCR and Western blotting confirmed increased expression of fibrosis-related genes in the presence of GZMK.

To understand the role of GZMK⁺CD8⁺ T cells in cGVHD-BOS, researchers used a post-allo-HSCT mouse model, single-cell RNA sequencing, and paired single-cell TCR sequencing. This study analyzed engrafted immune cells from lung tissues of 3 cGVHD-BOS mice and 3 control mice, specifically isolating living donor mouse immune cells (FVS510- H2b+ H2k- CD45+).

This analysis identified 3 distinct T cell clusters expressing high levels of GZMK and CCR5, categorized as CD8 EX GZMK, CD8 EM CCL4, and CD8 EFF GZMA. Notably, the total presence of GZMK⁺CD8⁺ T cells significantly increased in cGVHD-BOS mouse lungs compared with controls (P <.01), particularly featuring dominant populations of CD8 EX GZMK and CD8 EM CCL4 cells.

Additionally, researchers assessed potential novel drugs to address cGVHD-BOS by targeting the expression of GZMK in T cells. In a screening process involving 40 small molecular drugs selected from a pool of 932 compounds targeting epigenetic regulators, bosutinib, a second-generation BCR-ABL1-targeting tyrosine kinase inhibitor (TKI), emerged as a candidate capable of inhibiting GZMK expression in CD8⁺ T cells in vitro.

Bosutinib showed promising outcomes in cGVHD-BOS mice, demonstrating reduced lung stiffness and restored lung compliance, indicators of diminished pulmonary fibrosis and improved lung function. Further validation through multiple immunofluorescence staining assays confirmed bosutinib’s ability to decrease the infiltration of GZMK⁺CD8⁺T cells and collagen deposition in the lungs of cGVHD-BOS mice. This research illuminates a potential new therapeutic avenue for managing cGVHD-BOS, offering hope for future treatment strategies.

Reference

1. Xiao H, Gao Y, Liu R, et al. Clonal GZMK⁺CD8⁺ T cells are identified as a hallmark of pathogenesis of Cgvhd-induced bronchiolitis obliterans syndrome after allogeneic hematopoietic stem cell transplantation. Presented at: 2023 ASH Annual Meeting. San Diego, CA. Abstract 476.