Page 86 - Haematologica May 2020

A. Magnani et al.
the recipient. In order to investigate this putative selective advantage over the course of erythroid development, we compared donor chimerism among compartments at dif- ferent stages of development. Our results show that not only the peripheral donor RBC have a selective advantage related to the shortened lifetime of SCD RBC but also the earlier erythroid progenitor/precursors. Despite the results of studies suggesting that the selective advantage is restricted to RBC in the periphery,23,28 our hypothesis has already been proposed in small series of patients.17,20 Indeed, the selective advantage of donor erythroid pro- genitor/precursors could be due to the ineffective erythro- poiesis in SCD, as shown by the loss of SCD erythroblasts reported in patients with MC.24 Interestingly, in a SCD patient with MC, the expression of the apoptotic regulator Fas was significantly higher in recipient than in donor ery- throblasts and RBC,29 suggesting that SCD “ineffective” erythroid cells undergo apoptosis, while donor cells have a survival advantage.
clinical remission in SCD, the present study provides important information for patients undergoing autolo- gous transplantation of genetically modified hematopoi- etic stem cells (HSC). It is important to note that today’s lentiviral-based gene addition strategies and genome-edit- ing approaches aimed at reactivating the expression of the anti-sickling fetal γ-globin at best generate a heterozy- gous phenotype in vitro; therapeutic hemoglobin accounts for at most 60% of the total Hb types, e.g. in RBC derived from HSC harboring multiple copies of the vector.30 Under these optimal conditions (i.e. generation of AS-like HSC), our analyses of SCD patients with MC suggested that an HSC genetic modification rate below 30% would not be sufficient to ameliorate the SCD clinical pheno- type, whereas a myeloid donor chimerism level ≥30% would probably lead to clinical improvements. Furthermore, our present results suggest that in order to fully control hemolysis, this threshold should be raised to >50%. Under suboptimal conditions (i.e. therapeutic Hb <60%), we predict that a donor chimerism level largely exceeding 30% would be required to correct the SCD clinical phenotype.
Consistently, the initial clinical data from gene therapy trials highlighted the absence of clinical benefit in patients with SCD harboring 10-30% of transduced HSC and low therapeutic Hb levels (i.e. 0.1-1.2 g/dL, which probably accounts for <10% of the total Hb types [31]. In contrast, clinical remission was observed in a SS patient with a mean vector copy number per cell of ~2, >60% of genetically modified HSC, and therapeutic Hb levels accounting for ~50% of the total Hb.14
In conclusion, these findings show that the degree of myeloid donor engraftment (rather than T cells) is a better predictor of the disease control in these patients - as shown by the inverse correlation between HbS levels and the chimerism in the myeloid lineage. According to these observations the whole blood chimerism could be appro- priate for evaluating engraftment when donor chimerism is higher than 50%, but it looks unsatisfactory for patients developing lower donor chimerism; in this case a lineage- specific chimerism analysis should be preferred.
Our results provide a new insight into the selective advantage of donor erythroid cells in SCD patients. In par- ticular, we show that not only donor peripheral RBC but also progenitors/precursors have a selective advantage over the recipient cells. Lastly, our study provides impor- tant clues for future gene therapy trials, and suggests that higher levels of gene correction will be needed to obtain full disease control.
Acknowledgments
The authors would like to thank patients and their families.
Funding
This work was funded by grants from “Cordons de Vie” char- ity and Agence Nationale de la Recherche (ANR-10-IAHU-01 “Investissements d’Avenir” program). This project has received funding from the European Union’s Horizon 2020 research and innovation program under grant agreement No 693762 - Gene For Cure.
We further focused on a group of patients with low lev- els of donor chimerism (<70% for total WBC), and in par- ticular on a subgroup of six patients who presented a donor chimerism level ≤50%. Indeed, as reported in the overall French cohort,22 donor chimerism values as low as 16%, 18% and 21% for total WBC were recorded; these patients were the only ones to display high (>45%) HbS levels, hemolytic anemia and (in one case) a hepatic VOC. All three patients had an AS donor. These patients also presented the lowest level of donor chimerism in CD15+ cells (5.4%, 10% and 16%), which was more closely cor- related than WBC chimerism with the HbS level.
In contrast, patients with total WBC chimerism ≤50% and AA donors had low HbS levels and no anemia, although high reticulocyte counts were recorded. In the study population as a whole, there were no significant differences between patients with an AA donor and those with an AS donor. However, when the level of donor chimerism was low, having an AA donor was an advan- tage.
Three patients with the lowest levels of donor myeloid chimerism received donor lymphocyte infusions during follow up in order to stabilize donor cells engraftment. Of note no modification in donor chimerism was observed after infusion. The use of donor lymphocyte infusion in this context is debated as the risks might exceed the ben- efits.
Although our results came from a small number of patients, they show that individuals with a myeloid donor chimerism level above 30% have stable, sufficient levels of total Hb and no longer display the clinical signs of SCD - regardless of the donor’s Hb genotype. In contrast, the full correction of hemolytic anemia requires higher myeloid donor chimerism levels (i.e. >50%) - particularly for patients with an AS donor. A myeloid donor chimerism level as low as 30% was sufficient to prevent VOC in patients with AA donors but not constantly in those with AS donors. For patients with very low degree of myeloid donor engraftment, an AS graft is associated with a higher risk of SCD symptoms.
By defining the level of correction needed to obtain
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