Convergent clonal selection of donor- and recipient-derived CMV-specific T cells in hematopoietic stem cell transplant patients.

Competition between antigen-specific T cells for peptide:MHC complexes shapes the ensuing T cell response. Mouse model studies provided compelling evidence that competition is a highly effective mechanism controlling the activation of naïve T cells. However, assessing the effect of T cell competition in the context of a human infection requires defined pathogen kinetics and trackable naïve and memory T cell populations of defined specificity. A unique cohort of nonmyeloablative hematopoietic stem cell transplant patients allowed us to assess T cell competition in response to cytomegalovirus (CMV) reactivation, which was documented with detailed virology data. In our cohort, hematopoietic stem cell transplant donors and recipients were CMV seronegative and positive, respectively, thus providing genetically distinct memory and naïve T cell populations. We used single-cell transcriptomics to track donor versus recipient-derived T cell clones over the course of 90 d. We found that donor-derived T cell clones proliferated and expanded substantially following CMV reactivation. However, for immunodominant CMV epitopes, recipient-derived memory T cells remained the overall dominant population. This dominance was maintained despite more robust clonal expansion of donor-derived T cells in response to CMV reactivation. Interestingly, the donor-derived T cells that were recruited into these immunodominant memory populations shared strikingly similar TCR properties with the recipient-derived memory T cells. This selective recruitment of identical and nearly identical clones from the naïve into the immunodominant memory T cell pool suggests that competition is in place but does not interfere with rejuvenating a memory T cell population. Instead, it results in selection of convergent clones to the memory T cell pool.

Cytomegalovirus-specific T-cell reconstitution following letermovir prophylaxis after hematopoietic cell transplantation.

Decreased cytomegalovirus (CMV)-specific immunity after hematopoietic cell transplantation (HCT) is associated with late CMV reactivation and increased mortality. Whether letermovir prophylaxis-associated reduction in viral exposure influences CMV-specific immune reconstitution is unknown. In a prospective cohort of allogeneic HCT recipients who received letermovir, we compared polyfunctional CMV-specific T-cell responses to those of controls who received PCR-guided preemptive therapy before the introduction of letermovir. Thirteen-color flow cytometry was used to assess T-cell responses at 3 months after HCT following stimulation with CMV immediate early-1 (IE-1) antigen and phosphoprotein 65 (pp65) antigens. Polyfunctionality was characterized by combinatorial polyfunctionality analysis of antigen-specific T-cell subsets. Use of letermovir and reduction of viral exposure were assessed for their association with CMV-specific T-cell immunity. Polyfunctional T-cell responses to IE-1 and pp65 were decreased in letermovir recipients and remained diminished after adjustment for donor CMV serostatus, absolute lymphocyte count, and steroid use. Among letermovir recipients, greater peak CMV DNAemia and increased viral shedding were associated with stronger CD8+ responses to pp65, whereas the CMV shedding rate was associated with greater CD4+ responses to IE-1. In summary, our study provided initial evidence that letermovir may delay CMV-specific cellular reconstitution, possibly related to decreased CMV antigen exposure. Evaluating T-cell polyfunctionality may identify patients at risk for late CMV infection after HCT.

Cytomegalovirus Humoral Response Against Epithelial Cell Entry-Mediated Infection in the Primary Infection Setting After Hematopoietic Cell Transplantation.

BACKGROUND: The influence of humoral immunity on the prevention of primary cytomegalovirus (CMV) infection after hematopoietic cell transplantation (HCT) is poorly understood. METHODS: To determine whether neutralizing antibodies (nAbs) against CMV pentameric complex (PC)-mediated epithelial cell entry decrease CMV infection after HCT, samples were analyzed from a randomized controlled trial of CMV intravenous immunoglobulin (IVIG) prophylaxis. Weekly serum from 61 CMV donor-positive/recipient-negative (D+/R-) HCT patients (33 control, 28 CMV IVIG) was tested using a PC-entry nAb assay and quantitative CMV polymerase chain reaction (PCR). RESULTS: There was a trend toward higher weekly PC-entry nAb titers (P = .07) and decreased CMV infection by PCR at viral load cutoffs of ≥1000 and ≥10 000 IU/mL in the CMV IVIG arm. High nAb titers were not significantly protective against CMV infection later after HCT in both study arms. Among CMV-infected patients, each log2 increase in nAb titer was associated with an average 0.2 log10 decrease in concurrent CMV viral load after infection (P = .001; adjusted for study arm). CONCLUSIONS: This study provides initial support that CMV IVIG prophylaxis moderately enhances PC-entry nAB activity in D+/R- HCT recipients.

Effect of Preemptive Therapy vs Antiviral Prophylaxis on Cytomegalovirus Disease in Seronegative Liver Transplant Recipients With Seropositive Donors: A Randomized Clinical Trial.

Importance: Despite the use of a cytomegalovirus (CMV) prevention strategy of antiviral prophylaxis for high-risk CMV-seronegative liver transplant recipients with seropositive donors, high rates of delayed-onset postprophylaxis CMV disease occur. An alternate approach, preemptive therapy (initiation of antiviral therapy for early asymptomatic CMV viremia detected by surveillance testing), has not previously been directly compared with antiviral prophylaxis in these patients. Objective: To compare preemptive therapy with antiviral prophylaxis in CMV-seronegative liver transplant recipients with seropositive donors for the prevention of CMV disease. Design, Setting, and Participants: Randomized clinical trial of preemptive therapy vs antiviral prophylaxis in 205 CMV-seronegative liver transplant recipients with seropositive donors aged older than 18 years. The trial was conducted at 6 academic transplant centers in the United States between October 2012 and June 2017, with last follow-up in June 2018. Interventions: Patients were randomized 1:1 to receive either preemptive therapy (valganciclovir, 900 mg, twice daily until 2 consecutive negative tests a week apart) for viremia detected by weekly plasma CMV polymerase chain reaction for 100 days (n = 100) or valganciclovir, 900 mg, daily for 100 days as antiviral prophylaxis (n = 105). Main Outcomes and Measures: The primary outcome was incidence of CMV disease by 12 months, defined as CMV syndrome (CMV viremia and clinical or laboratory findings) or end-organ disease. Secondary outcomes included acute allograft rejection, opportunistic infections, graft and patient survival, and neutropenia. Results: Among 205 patients who were randomized (mean age, 55 years; 62 women [30%]), all 205 (100%) completed the trial. The incidence of CMV disease was significantly lower with preemptive therapy than antiviral prophylaxis (9% [9/100] vs 19% [20/105]; difference, 10% [95% CI, 0.5% to 19.6%]; P = .04]). The incidence of allograft rejection (28% vs 25%; difference, 3% [95% CI, -9% to 15%]), opportunistic infections (25% vs 27%; difference, 2% [95% CI, -14% to 10%]), graft loss (2% vs 2%; difference, <1% [95% CI, -4% to 4%]), and neutropenia (13% vs 10%; difference, 3% [95% CI, -5% to 12%]) did not differ significantly for the preemptive therapy vs antiviral prophylaxis group, respectively. All-cause mortality at last follow-up was 15% in the preemptive therapy vs 19% in the antiviral prophylaxis group (difference, 4% [95% CI, -14% to 6%]; P = .46). Conclusions and Relevance: Among CMV-seronegative liver transplant recipients with seropositive donors, the use of preemptive therapy, compared with antiviral prophylaxis, resulted in a lower incidence of CMV disease over 12 months. Further research is needed to replicate these findings and assess long-term outcomes. Trial Registration: ClinicalTrials.gov Identifier: NCT01552369.

Prospective Assessment of Cytomegalovirus Immunity in High-Risk Donor-Seropositive/Recipient-Seronegative Liver Transplant Recipients Receiving Either Preemptive Therapy or Antiviral Prophylaxis.

The differential impact of preemptive therapy (PET) and antiviral prophylaxis (AP) on development of cytomegalovirus (CMV)-specific neutralizing antibody (nAb) and T-cell responses have not previously been directly compared in high-risk donor-seropositive/recipient-seronegative (D+R-) organ transplant recipients. We prospectively assessed T-cell and nAb responses 3 months after transplantation in cohorts of high-risk D+R- liver transplant recipients who received either PET (n = 15) or AP (n = 25) and a control group of CMV-seropositive transplant recipients (R+) (AP; n = 24). CMV phosphoprotein 65 (pp65)- and immediate early protein 1-specific multifunctional T-cell responses were determined by means of intracellular cytokine staining and nAbs against BADrUL131-Y4 CMV in adult retinal pigment epithelial cell line-19 human epithelial cells; nAbs were detected in 8 of 12 (67%) in the PET group, none of 17 in the AP group, and 20 of 22 (91%) in the R+ group. Multifunctional CD8 and CD4 T-cell responses to pp65 were generally similar between PET and R+ groups, and lower for the AP group; multifunctional CD4 responses were similar across all groups. Among D+R- liver transplant recipients, PET was associated with the development of greater nAb and multifunctional CD8 T-cell responses compared with AP, providing a potential mechanism to explain the relative protection against late-onset disease with PET. Future studies are needed to define specific immune parameters predictive of late-onset CMV disease with AP.

Cytomegalovirus immunity in high-risk liver transplant recipients following preemptive antiviral therapy vs. prophylaxis.

Cytomegalovirus (CMV)-specific T-cells, NK cells, and neutralizing antibodies (nAb) were assessed in a randomized trial of CMV prevention with preemptive antiviral therapy (PET) vs. prophylactic antiviral therapy (PRO) in donor seropositive/recipient seronegative (D+R-) liver transplant recipients (LTxR), at 100 days (end of intervention), and at 6 and 12 months post-transplant. The PET group had significantly increased numbers of circulating polyfunctional T-cells, NK cells, and nAb compared to the PRO group at day 100 and several CMV immune parameters remained significantly higher by 12 months post-transplant. Among PET recipients, preceding CMV viremia (vs. no preceding viremia) was associated with significantly higher levels of most CMV immune parameters at day 100. Higher numbers of CMV-specific polyfunctional T-cells and NKG2C+ NK cells at day 100 were associated with a decreased incidence of CMV disease in multivariable Cox regression. The strongest associations with protection against CMV disease were with increased numbers of CMV-specific polyfunctional CD4 T-cells, CD3negCD56dimCD57negNKG2Cpos, and CD3negCD56dimCD57posNKG2Cpos NK cells. PET is superior to PRO for CMV disease prevention by allowing low-level CMV replication and associated antigen exposure that is promptly controlled by antiviral therapy and facilitates enhanced CMV protective immunity in D+R- LTxR.