Rapid memory T-cell reconstitution recapitulating CD45RA-depleted haploidentical transplant graft content in patients with hematologic malignancies

Abstract

T-cell depletion of an HLA-haploidentical graft is often used to prevent GVHD, but the procedure may lead to increased graft failure, relapse and infections due to delayed immune recovery. We hypothesized that selective depletion of the CD45RA+ subset can effectively reduce GVHD through removal of naive T cells, while providing improved donor immune reconstitution through adoptive transfer of CD45RA- memory T cells. Herein, we present results from the first 17 patients with poor-prognosis hematologic malignancy, who received haploidentical donor transplantation with CD45RA-depleted progenitor cell grafts following a novel reduced intensity conditioning regimen without TBI or serotherapy. Extensive depletion of CD45RA+ T cells and B cells, with preservation of abundant memory T cells, was consistently achieved in all 17 products. Neutrophil engraftment (median day +10) and full donor chimerism (median day +11) was rapidly achieved post transplantation. Early T-cell reconstitution directly correlated with the CD45RA-depleted graft content. T-cell function recovered rapidly with broad TCR Vβ spectra. There was no infection-related mortality in this heavily pretreated population, and no patient developed acute GVHD despite infusion of a median of >100 million per kilogram haploidentical T cells.

Figure 1

Flow cytometry evaluation of CD45RA-depleted product and lymphocyte reconstitution on Day 30 and Day 60 Representative evaluation of samples from patient 6 (the patient with median tempo of immune cell reconstitution). Gates were set for CD14–, CD4+, or CD8+ cells as indicated at the top of the panels. See supplemental figure 1 for post-transplant flow cytometry gating strategies from a representative umbilical cord blood recipient.

Figure 2

Engraftment, immune recovery, and outcomes (A)Time to neutrophil engraftment (ANC >500/μL), platelet engraftment (>20 × 10⁹/L), and onset of acute GvHD. (B)Time to full donor chimerism, normalization of NK cell or CD8+ T cell counts. (C) Time to relapse or transplant-related mortality (TRM).

Figure 3

Quantitative subset analysis Values are from all patients who were alive without relapse at each time point. Data from donors are prior to G-CSF mobilization. (A) Absolute number of T, B, and NK-lymphocytes, red line denotes median value. (B) Absolute number of CD8+CD45RA– and CD4+CD45RA– memory T cells, red line denotes median value. (C) Median percentage of CD25^brightCD127⁻ regulatory T cells in the CD3+CD4+ fraction. (D-F) Average proportion of the following subsets: effector memory T cells (CD45RA⁻CD27⁻, CD45RA⁻ CCR7⁻, or CD45RA⁻CD62L⁻); central memory T cells (CD45RA⁻CD27⁺, CD45RA⁻CCR7⁺, or CD45RA⁻CD62L⁺); terminal effector memory RA+ cells (CD45RA⁺CD27⁻, CD45RA⁺CCR7⁻, or CD45RA⁺CD62L⁻); and naïve T cells (CD45RA⁺CD27⁺, CD45RA⁺CCR7⁺, or CD45RA⁺CD62L⁺)

Figure 3

Quantitative subset analysis Values are from all patients who were alive without relapse at each time point. Data from donors are prior to G-CSF mobilization. (A) Absolute number of T, B, and NK-lymphocytes, red line denotes median value. (B) Absolute number of CD8+CD45RA– and CD4+CD45RA– memory T cells, red line denotes median value. (C) Median percentage of CD25^brightCD127⁻ regulatory T cells in the CD3+CD4+ fraction. (D-F) Average proportion of the following subsets: effector memory T cells (CD45RA⁻CD27⁻, CD45RA⁻ CCR7⁻, or CD45RA⁻CD62L⁻); central memory T cells (CD45RA⁻CD27⁺, CD45RA⁻CCR7⁺, or CD45RA⁻CD62L⁺); terminal effector memory RA+ cells (CD45RA⁺CD27⁻, CD45RA⁺CCR7⁻, or CD45RA⁺CD62L⁻); and naïve T cells (CD45RA⁺CD27⁺, CD45RA⁺CCR7⁺, or CD45RA⁺CD62L⁺)

Figure 4

Functional and TCR studies (A)Median lymphocyte proliferation as measured by Brdu incorporation at day +30 and +60 post-transplant. Patients with CMV seropositive and seronegative donors are presented separately. (B)TCR V_β spectratyping score and TREC per mL of blood (log-scale) from all donors and patients who were alive without relapse at each time point, red line denotes median value. (C) Representative V_β spectratyping from the patient with a median V_β spectratyping score