Longitudinal TCR repertoire profiling predicts refractory cytomegalovirus reactivation after haploidentical...

Cytomegalovirus (CMV) reactivation is a major cause of morbidity and mortality after haploidentical hematopoietic stem cell transplantation (Haplo-HSCT). While T cells are crucial for controlling CMV, the dynamic characteristics of the T-cell receptor (TCR) repertoire and their predictive value for refractory CMV reactivation remain poorly characterized. This study aimed to longitudinally monitor the TCR repertoire in Haplo-HSCT patients to identify features associated with CMV reactivation and to explore predictive biomarkers for refractory and recurrent CMV disease.

We enrolled 164 Haplo-HSCT patients, of which a subset of 28 was prospectively monitored for TCR repertoire dynamics via multiplex PCR and high-throughput sequencing. Patients were stratified into control (no reactivation, n = 10), acute phase (first reactivation, n = 18), and resolution phase (after clearance, n = 18) groups. CMV-specific TCRs were identified by database matching.

The incidence of CMV reactivation was 48.17% (79/164). CMV reactivation induced significant perturbations in TCR repertoire architecture, characterized by increased CDR3 clonality, reduced clonotype distribution evenness, and a decreased number of unique Clonotypes (all P < 0.05) compared to controls, indicating a state of oligoclonal expansion. This skewed architecture persisted even after viral clearance, suggesting delayed CMV-specific immune reconstitution. Consistent with this, patients with CMV reactivation had significantly lower CD4⁺ T cell counts compared to those without reactivation (P = 0.003). Longitudinal analysis revealed that impaired CD4⁺ T cell recovery preceded CMV reactivation. Patients with secondary reactivation (6/18) exhibited reduced diversity of V-J gene pairings (lower Shannon index, P < 0.05) and significant downregulation of specific TRBV28/TRBJ1-1 and TRBV6-1/TRBJ2-7 combinations. Crucially, lower abundance and total frequency of CMV-specific TCRs at initial reactivation predicted secondary reactivation (P < 0.05) and were correlated with a longer disease course. Four patients who failed letermovir prophylaxis lacked CMV-specific TCRs in their top clonotypes.

The abundance of CMV-specific TCRs is a key predictor of refractory CMV reactivation post-Haplo-HSCT. Longitudinal TCR profiling offers a promising strategy for risk stratification, potentially guiding more personalized preemptive therapies.

Not applicable.