Investigating Antigen Recognition Patterns and T Cell Dynamics in Neoantigen Vaccine-Treated Chronic Lymphocytic Leukemia

Abstract

Cancer vaccination in chronic lymphocytic leukemia (CLL) remains challenging because CLL has an ultra-low tumor mutational burden and a profoundly immunosuppressive microenvironment, limiting both targetable neoantigens and anti-tumor T cell function. To investigate how vaccination shapes tumor evolution and immune remodeling in this setting, we performed longitudinal genomic and single-cell immune profiling across two complementary CLL vaccine cohorts: NeoVax, a personalized neoantigen vaccine trial in treatment-naïve high-risk CLL, and GVAX, an autologous whole-tumor cell vaccine administered after allogeneic hematopoietic stem cell transplantation. Using whole-exome sequencing with PhylogicNDT-based clonal reconstruction, we tracked tumor clonal dynamics over time. In the NeoVax cohort, vaccine-targeted mutations spanned both clonal and subclonal tumor populations, meaning immune pressure against these targets would be expected to act broadly rather than selectively. Across most patients, clonal architecture remained stable during and after vaccination, and immunogenic vaccine-targeted mutations did not show consistent selective contraction. However, one progressing patient showed expansion of a non-immunogenic driver-harboring lineage, suggesting that intrinsic driver-mediated fitness can overcome immune pressure against immunogenic neoantigens. In the GVAX cohort, clonal analysis identified relapse-associated expansion of a TP53-mutant subclone with accompanying allele-specific copy-number changes, consistent with a strong fitness advantage at relapse. Single-cell RNA sequencing of longitudinal bone marrow samples from six GVAX-treated patients showed that long-term remission was associated with sustained expansion of cytotoxic GZMB+ CD8+ T cell states after vaccination, whereas relapse patients lacked consistent effector enrichment. Results show that clonal dynamics alone may not fully explain vaccine response in CLL and support integrating tumor evolution with longitudinal immune-state changes to identify mechanisms of response and resistance in low-mutational-burden malignancies.