Work

Differential Gene Expression Analysis Indicates Elevated Presence of Terminal Effector T-Cells in CD22 CAR T-Cell Infusion Products from B-Cell Acute Lymphoblastic Leukemia (B-ALL) Patients with High Baseline Bone Marrow (BM) Disease Burden

Year: 2025

Type: article

Abstract: Introduction High BM disease burden in B-ALL pre-CAR T-cell treatment is associated with lower remission rates , more prolonged cytopenias , increased risk of severe cytokine release syndrome , and worse overall survival . Whether high disease burden also impacts T-cell fitness and CAR T-cell quality which may additionally impair outcomes is not known. We investigated transcriptional differences between pre-infusion CD22 CAR T-cell products from patients with relapsed/refractory B-ALL, stratifying by BM disease burden at time of apheresis to assess for differences in CAR T-cell products. Methods Utilizing Nanostring gene expression analysis performed on 43 frozen CAR T-cell infusion products ( Lichtenstein, Blood, 2021 ) from patients treated on a phase I clinical trial targeting CD22 (NCT02315612), we conducted a secondary analysis of the gene expression data by grouping the samples by baseline BM disease , based on percentage of blasts by morphology (low: < 25%; high: ≥ 25%). After normalization, principal component analysis (PCA) was performed to cluster samples by variance. We assessed for differentially expressed genes (DEGs) between low and high BM disease groups using DESeq2 (log2(fold change) > 0.5; p value < 0.01). Fisher's exact test was used to compare overall response rate (ORR). Wald test was used to compare DEGs and Benjamini-Hochberg was used to calculate p adjusted (padj). Results Across 43 CAR T-cell products, 13 (30%) were from patients with low BM disease burden and 30 (70%) samples were from patients with high disease burden. ORR was similar between the two groups (low BM ORR 85%; high BM ORR 86%; p >.99). A PCA showed no clear grouping by BM disease alone (Figure 1). We next identified 8 DEGs between CAR T-cells manufactured from patients with high versus low BM disease (Figure 2). Notably, CD45RA, a marker expressed both on naïve and terminally differentiated T-cells, had a significantly increased expression in the high BM disease CAR T-cell group (padj=0.02, Figure 2). Interestingly, CCR7, CD28, and CD27 were not differentially expressed, suggesting that the CD45RA increase represents terminally differentiated effector T-cells (Figure 3). Conclusion We investigated how baseline BM disease may impact the transcriptional profile of CAR T-cell products. Interestingly, there is an increase in CD45RA expression in products from patients with high BM disease, which may indicate more terminally differentiated cells in these products. Future analysis to build upon these findings should be conducted to explore T-cell phenotype differences as a function of disease burden as well as identify other patient specific factors that may impact CAR T-cell quality. Figure legends: 1) Principal component analysis by baseline BM disease. 2) Volcano plot of differential gene expression by baseline BM disease. 3) T-cell maturation marker normalized counts by baseline BM disease

Source: Transplantation and Cellular Therapy

DOI: 10.1016/j.jtct.2025.01.345