Clonal Dynamics and Cellular Responses to Stress-Induced Toxicity in Autologous Stem Cell Transplantation

Autologous stem cell transplantation (ASCT) is a standard therapy for hematologic malignancies that involves harvesting hematopoietic stem and progenitor cells (HSPCs) prior to chemotherapy and repopulating them in the bone marrow, creating a genetic bottleneck shaping blood reconstitution dynamics. We integrated bulk error-corrected targeted sequencing of clonal hematopoiesis (CH) associated genes and mitochondrial single-cell assay for transposase-accessible chromatin with sequencing (mtscATAC-seq) to resolve mutational dynamics of commonly altered hematological genes and trace clonal evolution via somatic mitochondrial DNA (mtDNA) mutations, providing insights into cellular heterogeneity and clonal behavior during hematopoietic recovery. In a cohort of 60 patients, CH-associated mutations were detected in 59% at baseline, with DNMT3A and TET2 most frequently affected. Longitudinal analysis presented a transient spike in mutation count and gene diversity post-transplant, normalizing by three months, indicating temporary genetic instability. Tandem ASCT and extended follow-up revealed conserved mutational dynamics, with progressive clonal complexity and sustained clonal expansion. Mitochondrial single-cell profiling showed patient-specific immune reconstitution trajectories and clonal dynamics, with overall balanced multilineage output from graft HSPCs with a T-cell-biased expansion post-ASCT. Our findings provide a firsthand comprehensive view of ASCT-induced clonal dynamics and immune reconstitution, paving the way for more targeted and effective post-transplant therapies.