To the Editor:
In a recent article by Guenechea et al,1 they describe the engraftment of NOD/SCID mice with ex vivo-expanded CD34+ cord blood (CB) cells. In this report, they demonstrate that no changes in the long-term repopulation of NOD/SCID are produced after the transplantation of ex vivo-expanded CB cells. However, the ex vivo-expanded cells result in a delay in the engraftment of the recipients. Although the NOD/SCID model has been developed to evaluate long-term repopulating cells and no published data has correlated the early time of engraftment in NOD/SCID mice to the time of engraftment of neutrophils and/or platelets in patients, the investigators propose that this result has direct implications in the design of clinical protocols.
Recent data presented at the ASH meeting in Miami, by our own group2 and Stiff et al,3 demonstrated that ex vivo-expanded CB cells may be useful in transplantation of CB in adults. Compared with previously reported data4 of the use of unexpanded CB in adults, the expanded CB resulted in faster neutrophil engraftment. The culture conditions used by Guenechea et al1 differ from those used in the clinical studies2 3 and could result in different short-term engraftment. However, it is important to consider the use of the NOD/SCID model in evaluation of short-term engraftment. Firstly, the model was developed as a surrogate assay of human long-term engrafting cells. In clinical settings, patients achieving rapid neutrophil and platelet engraftment can become neutropenic as a result of loss of graft that could be due to the lack of long-term engrafting cells. Also, basic stem cell biology teaches that early engraftment results from committed progenitor cells and the long-term engraftment results from stem cells. This suggests that a stem cell model would not be predictive for early engraftment.
In addition, primate studies by Andrews et al5 demonstrated that rapid neutrophil engraftment with ex vivo-expanded cells was dependent on treatment with recombinant human granulocyte colony-stimulating factor (rhG-CSF) posttransplant. Patients undergoing transplant receive rhG-CSF posttransplant; so, the use of the NOD/SCID model as described is suboptimal if animals are not treated with rhG-CSF.
In summary, the conclusions presented in this manuscript are based on a mouse model that has not been shown to predict early engraftment in patients and is therefore questionable. Investigators should be cautious in using such data in the design of clinical studies until the models are shown in a rigorous manner to be predictive of clinical outcome.
Response
We thank Dr McNiece for his comments on our report, which give us the opportunity of reinforcing some of the main findings of our study.
The main argument used by Dr McNiece to question our conclusions is that no published data have shown in a rigorous manner that the NOD/SCID model is predictive of early engraftment in patients. Other indirect arguments have also been proposed to question our study.
(1) It is important to remember that, although the NOD/SCID model is certainly one of the best procedures for evaluating human repopulating cells, no formal validation of this assay to predict long-term repopulation in patients has been reported to date. Therefore, we wonder why our long-term engraftment results have not been questioned at all by Dr McNiece, whereas the short-term results have been considered questionable. In our opinion, both the short- and the long-term data offer new evidences regarding the engrafting ability of ex vivo-expanded samples.
(2) The direct assumption of the NOD/SCID model as a stem cell assay looks a little risky for us. In his own letter, Dr McNiece mentions the relevance of doing analysis in the early and late posttransplantation for assessing the functionality of different repopulating cells. From the careful reading of our report, Dr McNiece will see this constitutes the basis of our study.
(3) We agree that, after a rapid engraftment, the graft can be lost due to the lack of long-term engrafting cells. In fact, we demonstrated this with ex vivo-expanded mouse bone marrow cells 1 year ago.1-1 Now, with human cells transplanted into NOD/SCID mice, the evidence is different, but according to our results, and to those obtained by other laboratories,1-2 it seems to be very reproducible.
(4) The question of whether or not the administration of HGFs to recipients is critical for the engraftment of ex vivo-expanded samples constitutes an interesting issue, although this was not addressed in our study. Of significance, however, is the following statement from the baboon study mentioned by Dr McNiece1-3: “Transplantation of expanded cells without post-transplant growth factors resulted in prolonged neutrophil and platelet recovery. . . . .” It is worth mentioning in this respect another study in baboons in which a delayed engraftment was associated with the ex vivo expansion process,1-4 an observation that is in complete agreement with our study.
(5) The statement “Recent data demonstrated that expanded CB cells may be useful in transplantation of CB in adults” is unclear for us. Although we agree with Dr McNiece that ex vivo expansion could be very useful in hematopoietic transplantation, in our opinion the two mentioned studies do not formally demonstrate the clinical advantage of this approach. Also, the argument related to the use of different growth factors in our and their studies is inconsistent, given that we never claimed all culture conditions should reproduce our findings. Rather, we indicated that our novel conclusions should be taken into account in the field of ex vivo expansion.
Apart from these observations, we are sorry that Dr McNiece has not considered one of the main suggestions made in our study, namely that “other biological properties apart from the content in hematopoietic progenitors are modulating the grafting ability of the sample.” In a recent report published in Science,1-5 it has now been shown that the modulated expression of CXCR4 is critical for the engraftment of human hematopoietic precursors. Significantly, these investigators also used short-term engraftments in NOD/SCIDs as one of the main biological end points.
In summary, the clinical relevance of our observations will only be known in the future. In the meantime, we would support the concept that investigators may like to consider all experimental studies made with rigor, including those generating apparently unexpected data, in their protocol designs. We believe this will not only limit the risks potentially associated with ex vivo expansion, but will also facilitate the rapid optimization of these protocols to improve the outcome of transplanted patients.
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