FIGURE 2 from Single-Cell CD4 and CD8 T-Cell Secretome Profiling Reveals Temporal and Niche Differences in Acute Myeloid Leukemia Following Immune Checkpoint Blockade Therapy

Baseline T-cell functional activity in PB and BM. A, Heat map of T-cell cytokine expression values in PB (n = 20) and BM (n = 16) samples at baseline split by cell subset and scaled by row with clinical characteristic and functional group annotations. ComplexHeatmap was used for data analysis and visualization. B, ComplexHeatmap comparing cytokine expression based on functional groups of CD4 and CD8 cells between TMEs at baseline in the PB and BM. C, Bar plot representation of the significant functional groups from B. Pairwise analysis using ggpaired shows differences based on functional group expression compared between TMEs (P < 0.05).</p

Follicular lymphoma patients with KIR2DL2 and KIR3DL1 and their ligands (HLA-C1 and HLA-Bw4) show improved outcome when receiving rituximab

Abstract Background The ECOG-ACRIN Cancer Research Group evaluated rituximab treatment schedules for patients with newly-diagnosed low-tumor-burden follicular-lymphoma (FL). All patients received 4-weekly rituximab treatments as induction therapy. Clinically-responding patients were randomized to receive rituximab every 13 weeks (“maintenance”) vs. no additional rituximab until progression (“non-maintenance”). Based on “time-to-rituximab-failure (TTRF)”, the study-committee reported there was no overall-benefit for maintenance rituximab in this setting. Tumor-reactive mAbs, like rituximab, trigger natural killer (NK) cells. NK-cell responses are regulated, in part, by interactions between killer immunoglobulin-like receptors (KIRs) on NK cells and their interactions with KIR-ligands. In a separate study of children with neuroblastoma treated with a different mAb, we found certain KIR/KIR-ligand genotypes associated with improved outcome. Here, we assessed whether a subset of FL patients show improved outcome from the maintenance rituximab based on these same KIR/KIR-ligand genotypes. Methods Genotypes for KIR/KIR-ligand were determined and assessed for associations with outcome [duration of response, TTRF and % tumor shrinkage] as a post-hoc analysis of this phase III trial. Our primary objective was to assess specific KIR/KIR-ligand genotype associations, followed by separate prespecified KIR/KIR-ligand genotype associations in follow-up analyses. Statistical analyses for association of genotype with clinical outcome included: Log-rank tests and Cox proportional hazards regression models to assess duration of response and TTRF; analysis of variance (ANOVA) was used for assessment of % tumor shrinkage. Results We found that patients inheriting KIR2DL2 and its ligand (HLA-C1) along with KIR3DL1 and its ligand (HLA-Bw4) had improved outcome over patients without this genotype. In addition, patients with KIR2DL2 and HLA-C1 along with KIR3DL1 and HLA-Bw4 also showed improved duration of response and tumor shrinkage if they received maintenance, while patients without this genotype showed no such improvement when receiving maintenance. Conclusions The data presented here indicate that a subset of FL patients, identified by certain KIRs/KIR-ligands, have improved outcome and may benefit from additional rituximab treatment. Taken together, this suggests that the efficacy of tumor-reactive mAb treatment for some patients is influenced by KIRs on NK cells. However, prior to considering these genotypes in a clinically-actionable manner, these findings need independent validation in other studies

Follicular lymphoma patients with KIR2DL2 and KIR3DL1 and their ligands (HLA-C1 and HLA-Bw4) show improved outcome when receiving rituximab

BACKGROUND: The ECOG-ACRIN Cancer Research Group evaluated rituximab treatment schedules for patients with newly-diagnosed low-tumor-burden follicular-lymphoma (FL). All patients received 4-weekly rituximab treatments as induction therapy. Clinically-responding patients were randomized to receive rituximab every 13 weeks ( maintenance ) vs. no additional rituximab until progression ( non-maintenance ). Based on time-to-rituximab-failure (TTRF) , the study-committee reported there was no overall-benefit for maintenance rituximab in this setting. Tumor-reactive mAbs, like rituximab, trigger natural killer (NK) cells. NK-cell responses are regulated, in part, by interactions between killer immunoglobulin-like receptors (KIRs) on NK cells and their interactions with KIR-ligands. In a separate study of children with neuroblastoma treated with a different mAb, we found certain KIR/KIR-ligand genotypes associated with improved outcome. Here, we assessed whether a subset of FL patients show improved outcome from the maintenance rituximab based on these same KIR/KIR-ligand genotypes. METHODS: Genotypes for KIR/KIR-ligand were determined and assessed for associations with outcome [duration of response, TTRF and % tumor shrinkage] as a post-hoc analysis of this phase III trial. Our primary objective was to assess specific KIR/KIR-ligand genotype associations, followed by separate prespecified KIR/KIR-ligand genotype associations in follow-up analyses. Statistical analyses for association of genotype with clinical outcome included: Log-rank tests and Cox proportional hazards regression models to assess duration of response and TTRF; analysis of variance (ANOVA) was used for assessment of % tumor shrinkage. RESULTS: We found that patients inheriting KIR2DL2 and its ligand (HLA-C1) along with KIR3DL1 and its ligand (HLA-Bw4) had improved outcome over patients without this genotype. In addition, patients with KIR2DL2 and HLA-C1 along with KIR3DL1 and HLA-Bw4 also showed improved duration of response and tumor shrinkage if they received maintenance, while patients without this genotype showed no such improvement when receiving maintenance. CONCLUSIONS: The data presented here indicate that a subset of FL patients, identified by certain KIRs/KIR-ligands, have improved outcome and may benefit from additional rituximab treatment. Taken together, this suggests that the efficacy of tumor-reactive mAb treatment for some patients is influenced by KIRs on NK cells. However, prior to considering these genotypes in a clinically-actionable manner, these findings need independent validation in other studies

FIGURE 4 from Single-Cell CD4 and CD8 T-Cell Secretome Profiling Reveals Temporal and Niche Differences in Acute Myeloid Leukemia Following Immune Checkpoint Blockade Therapy

Single-cell analysis of IsoPlexis data for CD4 cells (left) and CD8 cells (right). A, UMAP visualization colored by sample location (orange for PB, purple for BM). B, UMAP colored by time point (green for baseline, pink for post-therapy). C, Neighborhood graph depicting differential abundance testing results obtained from miloR. Colors represent the log fold-change between baseline (red) and post-IO (blue) cells. White neighborhoods are nondifferential (FDR 10%). The edges depict links between cells shared by neighborhoods. D, Beeswarm plot of the distribution of neighborhoods by timepoint. E, Beeswarm plot of the distribution of neighborhoods by UMAP-based cluster.</p

Supplementary Figure 3 from Single-Cell CD4 and CD8 T-Cell Secretome Profiling Reveals Temporal and Niche Differences in Acute Myeloid Leukemia Following Immune Checkpoint Blockade Therapy

Baseline T cell individual cytokine activity in PB and BM.</p

Supplementary Figure 1 from Single-Cell CD4 and CD8 T-Cell Secretome Profiling Reveals Temporal and Niche Differences in Acute Myeloid Leukemia Following Immune Checkpoint Blockade Therapy

Exploratory analysis of baseline T cell cytokine dynamics in PB and BM.</p

FIGURE 5 from Single-Cell CD4 and CD8 T-Cell Secretome Profiling Reveals Temporal and Niche Differences in Acute Myeloid Leukemia Following Immune Checkpoint Blockade Therapy

Single-cell cytokine and polyfunctional group analysis at baseline. Heat map of cytokine expression by cluster for A. CD4 T cells. Annotation bars show cluster breakdown by sample location and timepoint. Hierarchical clustering was used to create new polyfunctional groups (CD4-G1-5), which are highlighted by black boxes. B, Boxplots for CD4 T cells show the comparison of complete responders (CR, brown) and nonresponders (NR, blue) at baseline by sample location in these newly defined polyfunctional groups for T cells. C, Heat map of cytokine expression by cluster for CD8 T cells with hierarchical clustering creating new polyfunctional groups (CD8-G1-6). D, Boxplots for CD8 T cells show comparison of CR (brown) and NR (blue) at baseline.</p

Supplementary Figure 2 from Single-Cell CD4 and CD8 T-Cell Secretome Profiling Reveals Temporal and Niche Differences in Acute Myeloid Leukemia Following Immune Checkpoint Blockade Therapy

Baseline correlation analysis of age with individual cytokines and functional groups.</p

FIGURE 3 from Single-Cell CD4 and CD8 T-Cell Secretome Profiling Reveals Temporal and Niche Differences in Acute Myeloid Leukemia Following Immune Checkpoint Blockade Therapy

Exploratory analysis of CR and NR patients at baseline. CRs have significantly higher CD4 effector cytokine expression than NRs in the BM. No significant difference in functional group expression from CD8 cells in either PB and BM between CR and NR.</p