Adoptive transfer of CMV-specific TCR-T cells for the treatment of CMV infection after haploidentical hematopoietic stem cell transplantation

Abstract

Background: Cytomegalovirus (CMV) reactivation after unmanipulated haploidentical stem cell transplantation (SCT) frequently occurs, causing life-threatening morbidities and transplantation failure. Pre-emptive therapy upon the detection of CMV viremia using antiviral agents is currently the standard of care but it was associated with significant toxicity. The CMV antigen-specific cytotoxic T lymphocyte therapy was limited by the time-consuming manufacture process and relatively low success rate. More effective and safer approaches for the treatment of CMV reactivation after haploidentical SCT are in urgent need.

Methods: A single-arm, open-label, phase I clinical trial evaluating the safety and efficacy of CMV-targeting T cell receptor-engineered T (CMV-TCR-T) cell therapy as the first-line pre-emptive therapy for patients with CMV reactivation after haploidentical peripheral blood SCT (PBSCT) was conducted in the Chinese PLA General Hospital. Six patients with CMV reactivation after haploidentical SCT were adoptively transferred by one to three doses of SCT donors-derived CMV-TCR-T cells. This trial was a dose-escalation study with doses ranging from 1×10³ CMV-TCR-T cells/kg body weight per dose to 5×10⁵ CMV-TCR-T cells/kg per dose.

Results: Except for the grade 1 cytokine release syndrome observed in one patient and mild fever in two patients, no other adverse events were observed. Four patients had response within a month after CMV-TCR-T cell infusion without the administration of any antiviral agents. The other two patients who initially did not respond to CMV-TCR-T cell therapy had salvage ganciclovir and foscarnet administration and then had rapid CMV clearance. The CMV-TCR-T cells displayed overall robust expansion and persistence in the peripheral blood after infusion. The CMV-TCR-T cells were first detected in the peripheral blood of these patients 3-7 days after the first dose of CMV-TCR-T infusion, rapidly expanded and persisted for at least 1-4 months, providing long-term protection against CMV reactivation. In one patient, the CMV-TCR-T cells started to expand even when the anti-graft-versus-host disease reagents were still being used, further indicating the proliferation potential of CMV-TCR-T cells.

Conclusions: Our study first showed CMV-TCR-T cell as a highly feasible, safe and effective first-line pre-emptive treatment for CMV reactivation after haploidentical PBSCT.

Trial registration number: ClinicalTrials.gov Registry (NCT05140187).

Keywords: CD8-Positive T-Lymphocytes; Cell Engineering; Clinical Trials as Topic; Immunotherapy, Adoptive.

Figure 1

Isolation of CMV-specific T cells. The peripheral blood mononuclear cells from healthy donors were stimulated by CMV antigen peptides including NLVPMVATV (HLA-A*02:01-restricted), ATVQGQNLK (HLA-A*11:01-restricted), and QYDPVAALF (HLA-A*24:02-restricted). After stimulation, cells were stained by specific peptide–MHC tetramers. Stimulated T cells were compared with their unstimulated controls to define the tetramer⁺ populations which were then sorted for single-cell TCR sequencing. CMV, cytomegalovirus; HLA, human leukocyte antigen; MHC, major histocompatibility complex; TCR, T cell receptor; SSC, side scatter.

Figure 2

In vitro efficacy of CMV-TCR-T cells. (A–C) Transduction rate of CMV-specific TCRs with different HLA restrictions was represented by the percentage of tetramer⁺ population. Non-transduced T cells corresponding to each group of TCR-T cells were used as negative controls (NC). (D–F) In vitro cytolytic activity against HLA-matched/unmatched target cells of CMV-TCR-T cells with different HLA restrictions, compared with corresponding NC. Asterisks indicated statistical significance determined by paired Student’s t-tests between groups (*p<0.05). CMV, cytomegalovirus; HLA, human leukocyte antigen; N.S., not significant; TCRs, T cell receptors; SSC, side scatter; SSC-H, side scatter height; PE-H, phycoerythrin height.

Figure 3

Clinical responses of patients. (A) Dynamics of viral load of each individual patient after SCT. The time of TCR-T cell infusion and salvage treatment was also indicated. Day 0 is the day of SCT. (B) Pathological presentations of CMV colitis in patients 02 (left) and 03 (right). The CMV-infected cells were emphasized by the rectangle. CMV, cytomegalovirus; HSCT, hematopoietic stem cell transplantation; SCT, stem cell transplantation; TCR-T cell, T cell receptor-engineered T cell.

Figure 4

TCR-T cell expansion and persistence. Expansion and persistence of CMV-TCR-T cells in the peripheral blood of each individual patient were measured by real-time qPCR and represented by the copy number of CMV-TCR transgenes. The time of TCR-T cell infusion was also indicated. CMV, cytomegalovirus; HSCT, hematopoietic stem cell transplantation; qPCR, quantitative PCR; TCR-T cell, T cell receptor-engineered T cell.