Clinical Validation of a Novel Elispot-Based Diagnostic Assay: Monitoring Cytomegalovirus-Specific Cell-Mediated Immunity and Risk Stratification in Hematopoietic Stem Cell Transplant Recipients

Increasing evidence suggests that impaired cytomegalovirus (CMV)-specific cell-mediated immunity (CMV-CMI) is a major cause of uncontrolled CMV reactivations and associated complications in hematopoietic stem cell transplantation (HSCT). No reliable test exists to predict patients at risk of primary and/or recurrent CMV reactivations following HSCT. Accurately assessing CMV-CMI might therefore improve the risk stratification of patients and allow optimizing and individualizing patient care. This study aimed to evaluate the suitability of a novel IFN-γ ELISpot assay (T-Track ® CMV), based on the stimulation of peripheral blood mononuclear cells with T-activated® pp65 and IE-1 CMV proteins, to predict protection from recurrent CMV reactivation following the resolution of a treatment-requiring primary CMV reactivation. A prospective, longitudinal, observational, multicenter study was conducted in 175 intermediate- and high-risk (Donor (D)+/Recipient (R)+, D+/R-, D-/R+) HSCT recipients ( ClinicalTrials.gov ID: NCT02156479). Patients underwent preemptive antiviral therapy per institutional guidelines. CMV DNAemia was analyzed by quantitative PCR. CMV-CMI was measured at day 45, 60, 80, 100 and 120 post-transplantation, as well as at onset and following the end of preemptive treatment. Occurrence of recurrent CMV reactivation was monitored up to 7.5 months post-transplantation. 154/175 patients fulfilling the inclusion/exclusion criteria and having at least one valid T-Track ® CMV test result were included in the final analysis. 101/154 (66%) patients experienced at least one (treatment-requiring) CMV reactivation during the observational period. Out of 74 patients (24 D+/R+, 3 D+/R-, 47 D-/R+) who experienced a first CMV reactivation and had a valid ELISpot test result at the end of this primary reactivation, 30 (41%) faced a recurrent CMV reactivation during the observational period. Interestingly, 41/44 patients free of recurrent reactivation had a positive test result (i.e. positive for at least one of pp65- and/or IE-1-specific result) after resolution of the primary CMV reactivation, resulting in a 93% specificity in diagnostic accuracy. Accordingly, a time-to-event analysis indicated a significantly lower incidence of recurrent CMV reactivation in patients with a positive test result (Figure 1; Hazard ratio=5.68; Log-Rank Test, p Altogether, this novel standardized IFN-γ ELISpot assay allows an improved risk stratification of CMV-related clinical complications, and can support clinicians in the identification and management of patients with increased risk of recurrent CMV reactivation following HSCT.