Microbiota dictate T cell clonal selection to augment graft-versus-host disease after stem cell transplantation.

Allogeneic T cell expansion is the primary determinant of graft-versus-host disease (GVHD), and current dogma dictates that this is driven by histocompatibility antigen disparities between donor and recipient. This paradigm represents a closed genetic system within which donor T cells interact with peptide-major histocompatibility complexes (MHCs), though clonal interrogation remains challenging due to the sparseness of the T cell repertoire. We developed a Bayesian model using donor and recipient T cell receptor (TCR) frequencies in murine stem cell transplant systems to define limited common expansion of T cell clones across genetically identical donor-recipient pairs. A subset of donor CD4+ T cell clonotypes differentially expanded in identical recipients and were microbiota dependent. Microbiota-specific T cells augmented GVHD lethality and could target microbial antigens presented by gastrointestinal epithelium during an alloreactive response. The microbiota serves as a source of cognate antigens that contribute to clonotypic T cell expansion and the induction of GVHD independent of donor-recipient genetics.

HA-1-targeted T cell receptor (TCR) T cell therapy for recurrent leukemia after hematopoietic stem cell transplantation.

Relapse is the leading cause of death after allogeneic hematopoietic stem cell transplantation (HCT) for leukemia. T cells engineered by gene transfer to express T cell receptors (TCR; TCR-T) specific for hematopoietic-restricted minor histocompatibility (H) antigens may provide a potent selective anti-leukemic effect post-HCT. We conducted a phase I clinical trial employing a novel TCR-T product targeting the minor H antigen HA-1 to treat or consolidate treatment of persistent or recurrent leukemia and myeloid neoplasms. The primary objective was to evaluate the feasibility and safety of administration of HA-1 TCR-T post-HCT. CD8+ and CD4+ T cells expressing the HA-1 TCR and a CD8-co-receptor were successfully manufactured from HA-1 disparate HCT donors. One or more infusions of HA-1 TCR-T following lymphodepleting chemotherapy were administered to nine HCT recipients who had developed disease recurrence post-HCT. TCR-T cells expanded and persisted in vivo after adoptive transfer. No dose-limiting toxicities occurred. Although the study was not designed to assess efficacy, four patients achieved or maintained complete remissions following lymphodepletion and HA-1 TCR-T, with one ongoing at >2 years. Single-cell RNA sequencing of relapsing/progressive leukemia after TCR-T therapy identified upregulated molecules associated with T cell dysfunction or cancer cell survival. HA-1 TCR-T therapy appears feasible and safe and shows preliminary signals of efficacy. This clinical trial is registered at clinicaltrials.gov as NCT03326921.

Optimizing Autologous Stem Cell Transplantation in Multiple Myeloma: The Impact of Intensive Chemomobilization.

Most transplant-eligible multiple myeloma (MM) patients undergo autologous peripheral blood stem cell collection (PBSC) using G-CSF with on-demand plerixafor (G ± P). Chemomobilization (CM) can be used as a salvage regimen after G ± P failure or for debulking residual tumor burden ahead of autologous peripheral blood stem cell transplantation (ASCT). Prior studies utilizing cyclophosphamide-based CM have not shown long-term benefits. At our center, intensive CM (ICM) using a PACE- or HyperCVAD-based regimen has been used to mitigate "excessive" residual disease based on plasma cell (PC) burden or MM-related biomarkers. Given the lack of efficacy of non-ICM, we sought to determine the impact of ICM on event-free survival (EFS), defined as death, progressive disease, or unplanned treatment escalation. We performed a retrospective study of newly diagnosed MM patients who collected autologous PBSCs with the intent to proceed immediately to ASCT at our center between 7/2020 and 2/2023. Patients were excluded if they underwent a tandem autologous or sequential autologous-allogeneic transplant, had primary PC leukemia, received non-ICM treatment (i.e., cyclophosphamide and/or etoposide), or had previously failed G ± P mobilization. To appropriately evaluate the impact of ICM among those who potentially could have received it, we utilized a propensity score matching (PSM) approach whereby ICM patients were compared to a cohort of non-CM patients matched on pre-ASCT factors most strongly associated with the receipt of ICM. Of 451 patients identified, 61 (13.5%) received ICM (PACE-based, n = 45; hyper-CVAD-based, n = 16). Post-ICM/pre-ASCT, 11 patients (18%) required admission for neutropenic fever and/or infection. Among 51 evaluable patients, the overall response rate was 31%; however, 46 of 55 evaluable patients (84%) saw a reduction in M-spike and/or involved free light chains. Among those evaluated with longitudinal peripheral blood flow cytometry (n = 8), 5 patients (63%) cleared circulating blood PCs post-ICM. Compared to patients mobilized with non-CM, ICM patients collected a slightly greater median number of CD34+ cells (10.8 versus 10.2 × 106/kg, P = .018). The median follow-up was 30.6 months post-ASCT. In a PSM multivariable analysis, ICM was associated with significantly improved EFS (hazard ratio [HR] 0.30, 95% CI 0.14 to 0.67, P = .003), but not improved OS (HR 0.38, 95% CI 0.10 to 1.44, P = .2). ICM was associated with longer post-ASCT inpatient duration (+4.1 days, 95% CI, 2.4 to 5.8, P < .001), more febrile days (+0.96 days, 95% CI 0.50 to 1.4, P < .001), impaired platelet engraftment (HR 0.23, 95% CI 0.06 to 0.87, P = .031), more bacteremia (OR 3.41, 95% CI 1.20 to 9.31, P = .018), and increased antibiotic usage (cefepime: +2.3 doses, 95% CI 0.39 to 4.1, P = .018; vancomycin: +1.0 doses, 95% CI 0.23 to 1.8, P = .012). ICM was independently associated with improved EFS in a matched analysis involving MM patients with excessive disease burden at pre-ASCT workup. This benefit came at the cost of longer inpatient duration, more febrile days, greater incidence of bacteremia, and increased antibiotic usage in the immediate post-ASCT setting. Our findings suggest that ICM could be considered for a subset of MM patients, but its use must be weighed carefully against additional toxicity.

Calcineurin inhibition rescues alloantigen-specific central memory T cell subsets that promote chronic GVHD.

Calcineurin inhibitors (CNIs) constitute the backbone of modern acute graft-versus-host disease (aGVHD) prophylaxis regimens but have limited efficacy in the prevention and treatment of chronic GVHD (cGVHD). We investigated the effect of CNIs on immune tolerance after stem cell transplantation with discovery-based single-cell gene expression and T cell receptor (TCR) assays of clonal immunity in tandem with traditional protein-based approaches and preclinical modeling. While cyclosporin and tacrolimus suppressed the clonal expansion of CD8+ T cells during GVHD, alloreactive CD4+ T cell clusters were preferentially expanded. Moreover, CNIs mediated reversible dose-dependent suppression of T cell activation and all stages of donor T cell exhaustion. Critically, CNIs promoted the expansion of both polyclonal and TCR-specific alloreactive central memory CD4+ T cells (TCM) with high self-renewal capacity that mediated cGVHD following drug withdrawal. In contrast to posttransplant cyclophosphamide (PT-Cy), CSA was ineffective in eliminating IL-17A-secreting alloreactive T cell clones that play an important role in the pathogenesis of cGVHD. Collectively, we have shown that, although CNIs attenuate aGVHD, they paradoxically rescue alloantigen-specific TCM, especially within the CD4+ compartment in lymphoid and GVHD target tissues, thus predisposing patients to cGVHD. These data provide further evidence to caution against CNI-based immune suppression without concurrent approaches that eliminate alloreactive T cell clones.