Immune Reconstitution after Haploidentical Hematopoietic Cell Transplantation: Impact of Reduced Intensity Conditioning and CD3/CD19-Depleted Grafts

Haploidentical hematopoietic cell transplantation (HHCT) using CD3/CD19 depleted grafts may lead to faster engraftment and immune reconstitution since grafts contain also graft-facilitating-cells, CD34- progenitors, NK cells, and dendritic cells. Reduced intensity conditioning may also have a positive impact on immune reconstitution following HHCT.
26 adults received CD3/CD19 depleted HHCT after RIC (150-200 mg/m² fludarabine, 10mg/kg thiothepa, 120 mg/m² melphalan and 5mg/day OKT-3 (day -5 to +14)) at our institution between 2005-2008. We prospectively evaluated engraftment and immune reconstitution. B-, NK-, T- and T-cell subsets (CD3/8, CD4/8, CD4/45RA/RO), TCR-Vβ repertoire and NK-cell receptors (NKP30, NKP44, NKP46, NKG2D, CD158a/b/e, CD85j, NKG2A, CD161) were analyzed by FACS.
Grafts contained 8.8x10⁶ CD34+ (range, 4.3-18.0 x10⁶), 2.9x10⁴ CD3+ (range, 1.2-9.2x10⁴) and 3.6x10⁷ CD56+ (range, 0.02-23.0 x10⁷) cells/kg. Engraftment was rapid with a median time to >500 granulocytes/µl of 11 days (range, 9-15) and a median time to >20 000 platelets/µl of 11 days (range, 8-23). Full chimerism was reached on day 14 (median; range, 6-26). NK-cell engraftment was rapid, reaching normal values on day 20 (median of 247 CD16+CD56+CD3- cells/µl (range, 1-886)) with NK cells comprising up to 70% of lymphocytes. B-cell reconstitution was delayed with 81 (range, 0-280) and 335 (range, 11-452) CD19+20+ cells/µl on days 150 and 400, respectively. T-cell reconstitution was impaired with 49 (range, 0-586) and 364 (range, 35-536) CD3+ cells/µl on day 60 and day 150, respectively. We observed an increase of CD3+CD8+ cells in contrast to CD3+CD4+ cells early after HHCT with a median of 24 (range, 0-399) vs 16 (range, 0-257) and 159 (range, 1-402) vs 96 (range, 18-289) cells/µl on day 50 and day 200, respectively. CD4+CD45RA+ T cells increased slowly while CD4+CD45RO+ T cells reconstituted faster with a median of 61 CD4+CD45RO+ cells/µl (range, 0-310) vs 24 CD4+CD45RA+ (range, 0 to 152) on day 100. Within the CD4+CD25+ regulatory T cells there was a slow regeneration with median of 14 CD4+CD25+ cells/µl (range, 0-96) on day 100 and 28 CD4+CD25+ cells/µl (range, 19-160) on day 200. CD14+CD45+ monocytes did not reach normal values within the time of observation with 7 CD14+CD45+ cells/µl (range, 0-21) on day 120 and 7 CD14+CD45+ cells (range, 2-381) on day 400. TCR-Vβ repertoire and NK-cell receptor reconstitution was analyzed so far in 7 and 8 patients, respectively. We found a skewed T-cell repertoire with oligoclonal T-cell expansions to day 100 and normalization after day 200. An increased natural cytotoxicity receptor (NKP30, NKP44, NKP46) and NKG2A, but decreased NKG2D and KIR-expression was observed on NK-cells until day 100.
In conclusion, T- and B-cell reconstitution is delayed after HHCT using CD3/CD19 depleted grafts and RIC. However, T-cell reconstitution is faster compared to data published with CD34 selected grafts and myeloablative conditioning. A fast NK-cell reconstitution early after HHCT was observed. Thus a combination of reduced intensity conditioning with CD3/CD19 depleted grafts appears to accelerate the immune recovery after haploidentical stem cell transplantation.