Page 60 - Haematologica August 2018

R. Mani et al.
may be due to high potency of diphtheria toxin, as few molecules are sufficient for cytotoxicity and/or independ- ent binding sites of IL-3 and SL-401. However, the results we saw with decreased myeloid colony formation ability of cord blood derived CD34+ cells and normal bone mar- row derived CD34+CD38– cells with SL-401 suggests that SL-401 can impact normal stem cells and myeloid progen- itors. These findings suggest that SL-401 could be used to target leukemic progenitors and decrease leukemic burden with potential use as a “bridge-to-transplant” before allo- geneic hematopoietic cell transplantation.39 These results are consistent with previous studies showing reduction in normal hematopoiesis when CD123-targeted CAR-T cells were used in mouse models.16
Bone marrow MSC play a critical role in leukemic cell survival and drug resistance. Recently, bone marrow- derived MSC from primary AML samples were used to establish faithful PDX models by coating ossicle/bioscaf- fold with MSC before inoculation into the mice by inves- tigators.40-42 We therefore evaluated SL-401 in AML co-cul- tures using HS5 stromal cell lines and autologous MSC derived from bone marrow aspirates. Despite variabilities and inconsistencies in culturing patient marrow-derived MSCs, we were able to successfully propagate six AML MSC and co-cultured AML blasts on these monolayers. This method accounts for the unique MSC cytogenetic/mutational aberration-dependent survival of AML blasts. Despite this MSC protection, SL-401 retained its activity, supporting its therapeutic potential in patients.
MDS are a group of myeloid disorders characterized by dysplastic changes in the bone marrow and peripheral cytopenias, which are manifestations of ineffective hematopoiesis.6 The MDS-RAEB / MDS-EB groups repre- sent high risk MDS and often show high levels of CD34+ cells. Thus, we interrogated the expression of CD123 in these groups. Almost all MDS samples tested (6/7) were positive for CD123 expression. Due to limited number of cells, we did not perform CD123 MESF analysis on MDS samples. However, MDS blasts responded to SL-401 as shown by decreases in both the blast percentage and the absolute count. Moreover, we showed that SL-401 reduced the numbers of CD123+ myeloid cells but not CD123– lymphoid cells in long-term cultures, supporting the selectivity of SL-401.
Our evaluation of SL-401 in mice engrafted with pri- mary AML cells is particularly relevant to the therapeutic potential of this agent in AML. Although difficult to estab- lish, AML PDX models capture the patient heterogeneity and genomic landscape of patients and form an important tool for evaluation of therapies.43 Consistent with previ- ously described variability in engraftment capabilities of
primary AML samples, only 3 of 7 AML samples engrafted successfully in NRGS mice, although the inclusion of busulfan as conditioning regimen allowed accelerated leukemic engraftment. Recently it was shown that inclu- sion of OKT3 either in in vitro cultures or in mice prevent- ed T-cell mediated GvHD and improved human hematopoietic cell engraftment. Thus, for our in vivo stud- ies, we cultured AML with growth factors and OKT3 prior to engraftment to eliminate CD3+ cells. SL-401 treated mice had significantly longer mean survival compared to the vehicle-treated controls. However, there was no differ- ence in the proportion of human leukemic cells in the spleen, bone marrow or peripheral blood of mice treated with vehicle and SL-401 at end points in the survival study, presumably due to discontinuation of treatment in SL-401 group and repopulation of leukemic cells in subse- quent accruing days and late time points the SL-401 group mice died. To assess the effect of SL-401 on tumor burden, we designed another study using same AML donors, but sacrificed both the groups on week 7 where we found that SL-401 treated mice had reduced human AML cells in marrow.
A first-in-human clinical trial with SL-401 in patients with blastic plasmacytoid dendritic cell neoplasm (BPDCN) noted that patients’ humoral immune responses against SL-401 limited the treatment cycles.20 However, in myeloid neoplasms where memory immune responses are poor, this is less likely to be a limitation. Furthermore, SL-401 is not a substrate of any known drug efflux pumps, making resistance due to this mechanism unlikely.44 Capillary leak syndrome is a known, severe side effect of SL-401 and needs to be managed for successful utilization of this agent in MDS and AML.
Together, these data indicate that SL-401 is active against CD123+ AML/MDS and normal hematopoietic progenitors. These findings have translational relevance in regards to management of potential side effects such as marrow aplasia. This agent is currently being evaluated in a Phase 2 trial of AML patients in remission with minimal residual disease, a setting associated with the persistence of CD123+ LSCs (clinicaltrials.gov identifier 02270463) and a phase I trial of azacitidine and SL-401 in MDS (clinicaltri- als.gov identifier 03113643).
Acknwoledgments
The authors are grateful to the AML and MDS patients who contributed to these studies, the OSU Comprehensive Cancer Center Leukemia Tissue Bank Shared Resource (P30 CA016058), the Clinical Flow Cytometry facility of the OSU Wexner Medical Center, R01 CA197844, R35 CA197734-01, D Warren Brown Foundation and the Lauber AML fund.
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