Safety and efficacy of plerixafor dose escalation for the mobilization of CD34+ hematopoietic progenitor cells in patients with sickle cell disease: interim results.

Gene therapy for sickle cell disease is limited by the yield of hematopoietic progenitor cells that can be harvested for transduction or gene editing. We therefore performed a phase I dose-escalation study of the hematopoietic progenitor cell mobilizing agent plerixafor to evaluate the efficacy and safety of standard dosing on peripheral blood CD34+ cell mobilization. Of 15 patients enrolled to date, only one was chronically transfused and ten were on hydroxyurea. Of eight patients who achieved a CD34+ cell concentration >30 cells/µL, six were on hydroxyurea. There was no clear dose response to increasing plerixafor dosage. There was a low rate of serious adverse events; two patients developed vaso-occlusive crises, at the doses of 80 µg/kg and 240 µg/kg. Hydroxyurea may have contributed to the limited CD34+ mobilization by affecting baseline peripheral blood CD34 counts, which correlated strongly with peak peripheral blood CD34 counts. Plerixafor administration did not induce significant increases in the fraction of activated neutrophils, monocytes, or platelets. However, increased neutrophils positive for activated ß2 integrin and Mac-1 were associated with serious adverse events. In summary, plerixafor was well tolerated but did not achieve consistent CD34+ cell mobilization in this cohort of patients, most of whom were being actively treated with hydroxyurea and only one was chronically transfused. The study will continue with escalation of the dose of plerixafor and modification of hydroxyurea administration. Clinicaltrials.gov identifier: NCT02193191.

Elucidating parasite and host-cell factors enabling Babesia infection in sickle red cells under hypoxic/hyperoxic conditions.

Sickle red blood cells (RBCs) represent a naturally existing host-cell resistance mechanism to hemoparasite infections. We investigate the basis of this resistance using Babesia divergens grown in sickle (SS) and sickle trait (AS) cells. We found that oxygenation and its corresponding effect on RBC sickling, frequency of fetal hemoglobin positive (HbF+) cells, cellular redox environment, and parasite proliferation dynamics, all played a role in supporting or inhibiting Babesia proliferation. To identify cellular determinants that supported infection, an image flow cytometric tool was developed that could identify sickled cells and constituent Hb. We showed that hypoxic conditions impaired parasite growth in both SS and AS cells. Furthermore, cell sickling was alleviated by oxygenation (hyperoxic conditions), which decreased inhibition of parasite proliferation in SS cells. Interestingly, our tool identified HbF+-SS as host-cells of choice under both hypoxic and hyperoxic conditions, which was confirmed using cord RBCs containing high amounts of HbF+ cells. Uninfected SS cells showed a higher reactive oxygen species-containing environment, than AA or AS cells, which was further perturbed on infection. In hostile SS cells we found that Babesia alters its subpopulation structure, with 1N dominance under hypoxic conditions yielding to equivalent ratios of all parasite forms at hyperoxic conditions, favorable for growth. Multiple factors, including oxygenation and its impact on cell shape, HbF positivity, redox status, and parasite pleiotropy allow Babesia propagation in sickle RBCs. Our studies provide a cellular and molecular basis of natural resistance to Babesia, which will aid in defining novel therapies against human babesiosis.