ALDH^br Hematopoietic Progenitors Promote Short-Term Engraftment in Experimental Models for Cord Blood Transplantation.

Abstract 2433

Poster Board II-410

We previously developed robust methods to purify human hematopoietic progenitors based on their high expression of aldehyde dehydrogenase (ALDH^br cells). These cells are enriched with both short- and long-term NOD/SCID repopulating cells. One early clinical trial suggested that augmenting a conventional cord blood transplant (CBT) with ALDH^br cells accelerated both neutrophil and platelet engraftment. We now describe experimental models for these clinical transplants, performed in two immunologically disparate strains of NOD/SCID mice. In the clinical study, pediatric patients first received 80% of an unmanipulated cord blood (CB) graft. 4 hours later they received the ALDH^br cells purified from the remaining 20% of the graft. In the experimental model, the mice were divided among three cohorts. Some mice were transplanted with 4,000 purified ALDH^br cells, alone. Other mice were transplanted with total CB using a cell dose that contained 4,000 ALDH^br cells. In the final group, mice first received the same dose of unmanipulated CB and, after 4 hours, 4,000 purified ALDH^br cells were also administered. After 4 weeks, the mice were sacrificed to determine their levels of human hematopoietic chimerism. When these transplants were performed using NOD/SCID-IL2Rγ^null (NSγ) mice, the ALDH^br cells demonstrated strong short-term engraftment to the bone marrow (12 ± 4.9%; n = 5) that was characterized by human CD19⁺ B cells and CD33⁺ myeloid cells. The latter included CD15⁺ cells that indicate neutrophil engraftment. In addition, the peripheral blood of these mice contained low levels of human CD41⁺ CD61⁺ platelets. Unmanipulated CB also engrafted the bone marrow of NSγ mice (7.4 ± 4.7%; n = 8); however, >95% of the human cells appeared to be mature CD3⁺ T cells. Engraftment by either B cells or myeloid cells was consistently low to undetectable. Similarly, human platelets were not detected in the peripheral blood. When NSγ mice were transplanted first with total CB and subsequently with purified ALDH^br cells, the level of engraftment to the bone marrow increased >2-fold over what had been observed in mice transplanted with CB alone (18.9 ± 9.3%; n = 10; P = 0.006). However, nearly all of the human cells present within the bone marrow were T cells, as had been observed in animals that received only unmanipulated CB. In addition, human platelets were not observed in the peripheral blood of NSγ mice that had received both CB and ALDH^br cells. When similar transplants were performed using NOD/SCID-β2-microglobulin^null (NSβ) mice, the ALDH^br cells again demonstrated strong short-term engraftment to the bone marrow (5.6 ± 4.7%; n = 5) that was characterized by human CD19⁺ B cells and CD33⁺ myeloid cells. The peripheral blood of these mice also contained low levels of human platelets. In contrast, total CB achieved only very low engraftment to the bone marrow of NSβ mice (0.32 ± 0.19%; n = 4) and human platelets were not detected in the peripheral blood. However, when NSβ mice first received total CB which was augmented 4 hours later with purified ALDH^br progenitors, the level of human hematopoietic chimerism in the bone marrow increased >10-fold (4.3 ± 1.9%; n = 5), with engraftment of human B cells and myeloid cells. The co-transplanted NSβ mice also demonstrated low-level engraftment of human platelets in the peripheral blood. Continuing studies will resolve the relative contributions of the ALDH^br cells and the unmanipulated CB by using two HLA-matched (6/6), but sex-mismatched, CBs. In total, these studies confirmed that ALDH^br progenitors by themselves provided efficient short-term myeloid engraftment in both NOD/SCID strains. This was in contrast to what was observed after transplantation of bulk CB, which by itself did not provide efficient myeloid engraftment in either strain. Finally, in both mouse strains, ALDH^br cells altered the outcome of CB transplants. In NSγ mice, ALDH^br progenitors appeared to facilitate either the engraftment or proliferation of mature CB T cells. Most importantly, the studies in NSβ mice strongly suggested that ALDH^br progenitors directly augment short-term myeloid and platelet engraftment by total CB. The latter studies in particular mirror the experience of early clinical CBT studies that use the same strategy.