Early Engraftment Kinetics of Leukocyte Subsets by Means of HLA-Specific Mabs After Double Umbilical Cord Blood Transplantation Show a Very Rapid Induction of Single Complete Donor Chimerism.
Authors: Somers Judith AE , van Hensbergen Yvette , Brand Anneke , Meijer Ellen , Braakman Eric , Oudshoorn Machteld , Sintnicolaas Kees , Mulder Arend , Cornelissen Jan J. 
Published: October 2019

Abstract
Abstract 3338
Poster Board III-226
Double umbilical cord blood transplantation (UCBT) results in higher engraftment rates as compared to single UCBT in adult patients. Sustained hematopoiesis is usually derived from a single cord blood unit (CBU). So far, the mechanism of predominance of a particular CBU is unresolved. Immunological rejection of one CBU has been proposed, as well as a selective growth advantage of one particular unit. Frequent serial chimerism studies in leukocyte subsets by use of HLA-specific monoclonal antibodies (mAbs) during the first month post transplant might contribute to unravel the mechanism of graft predominance.

Methods.
Seventeen consecutive patients (pts) with high risk hematological diseases received a double UCBT preceded by a non-myeloablative conditioning regimen (Cy 60 mg/kg/ Flu 160 mg/kg/ TBI 2×2 Gy). CBUs were selected by low resolution typing for HLA-A and –B loci and by allele typing for HLA-DRB1. The minimal required HLA-match grade was 4/6. If discriminating HLA mismatches between the 3 different parties were present, early analysis in leukocyte subpopulations was performed at day 11, 18, 25 and 32 post transplant by flowcytometry using lineage-specific (CD3, CD4, CD8, CD19, CD16/56, CD14, CD33) mAbs in combination with fluorochrome labeled HLA-antigen specific human mAbs. Results of day 32 were compared with routine chimerism analysis (VNTR) of peripheral blood TNC and T-cells.

Results.
Two pts were non-evaluable for engraftment due to early death and insufficient follow up, respectively. Donor engraftment in 15 pts occurred after a median of 30 days (range: 11-45). At 1 month post transplant, 9 pts showed complete single and 3 showed mixed chimerism by VNTR-analysis. In 3 patients, cells originating from both CBUs were present. Ultimately, complete single donor chimerism was established in 14 pts. Early chimerism studies with HLA-specific mAbs were performed in 8 pts. In all pts results at day 32 corresponded with chimerism analysis performed by VNTR. For 7 other pts no discriminating HLA-mAbs were available. Simultaneous 3-donor-origin detection of T-cells, monocytes and granulocytes based on HLA-disparities was possible in 4 pts. In 3 of them, all subsets showed a similar pattern: a clear predominance of a single CBU in all lineages as from day 11-18 onwards. Furthermore, the disappearing CBU was transiently detectable in these pts at day 11. In the fourth patient, T cells were of recipient origin at day 32 with predominance of a single CBU in all other subsets. The disappearing unit could be followed in another 4 pts, which revealed again a transient appearance of that unit in the various lineages at day 11. Although detectable in a total of 8 pts at day 11, all disappearing CBUs were completely lost within 18 days. The prefreeze TNC did not predict for the prevailing UCB, nor did the number of HLA-mismatches.

Conclusions.
These results show that double UCBT following a non-myeloablative regimen without ATG is associated with a rapid induction of complete single donor chimerism. The disappearing unit, although early detectable, was completely lost within 18 days in all patients, in whom it could be monitored by HLA-specific mAbs. We observed no disproportional increase of any leukocyte subset of the prevailing CBU nor of host leukocyte subset. This characteristic, uniform, early engraftment pattern in the presence of different HLA-disparities may rather argue for a difference in growth potential between both CBUs than an immunological rejection of the disappearing unit.

Disclosures.
No relevant conflicts of interest to declare.