Abstract 217: BCR-ABL tyrosine kinase inhibition combined with a novel pan BCL2 family inhibitor abrogates leukemia stem cell survival in a niche-dependent manner.
Authors: Ma Wenxue , Low-Marchelli Janine , Leu Heather S. , Court Recart Angela , Wei Jun , Huang Xianshu , Krajewska Maryla , Pellecchia Maurizio , Reed John C. , Jamieson Catriona H.M. 
Published: August 2014

Leukemia stem cells (LSC) play a crucial role in initiation, maintenance, and progression of chronic myeloid leukemia (CML). Targeted therapy with a BCR-ABL targeted tyrosine kinase inhibitor (e.g. dasatinib) has improved survival of patients with chronic phase CML. However, like blast crisis (BC) CML, survival remains only 30% over 5 years, and treatment frequently fails to eliminate dormant LSC that are hypothesized to drive CML relapse. Overexpression of BCL2 family genes has been observed in human BC CML and may fuel LSC survival. Our previous data demonstrated that pro-survival long isoforms of BCL2 and MCL1 were preferentially expressed in BC CML LSC compared to normal stem cells. These data showed that the bone marrow (BM) is enriched for BCL2 and MCL1-expressing LSC and serves as a reservoir for therapeutic resistance, thereby underscoring the importance of pan-BCL2 inhibition in the BM. A potent pan-BCL2 inhibitor, sabutoclax, has been identified that inhibits the binding of BH3 peptides to Bcl-XL, Bcl-2, Mcl-1 and Bfl1-1 at nanomolar concentrations.

To demonstrate whether sabutoclax or the combination of sabutoclax and dasatinib could synergize to eradicate LSC in the BM, humanized BC CML LSC mouse models were established by using 50,000 CD34+ cells selected from primary BC CML patient samples and transplanted into neonatal RAG2-/-γc-/- mice. Mice were screened for peripheral blood (PB) engraftment of human cells at 6 weeks, randomized, and dosed with (1) vehicle, (2) sabutoclax (10mg/kg, i.v., twice a week for 2 weeks), (3) dasatinib (50mg/kg, p.o., daily for 2 weeks), or (4) combination. Mice were sacrificed one day after last dose; PB, spleen (SP) and BM were collected for FACS analysis to measure engraftment and cell cycle status of LSC in different hematopoietic niches. BCL2 and MCL1 expression was measured in BM and SP by isoform specific qRT-PCR and immunohistochemistry. Treatment of CML LSC-transplanted mice with sabutoclax alone led to a significant reduction in LSC burden in the SP, but only slightly in the BM. Meanwhile, Single agent sabutoclax reduced the engraftment of quiescent LSC. The effect of sabutoclax synergized with dasatinib to further reduce LSC burden in the BM to eradicate LSC that drive therapeutic resistance. Importantly, these treatments spare normal human stem cells in both BM and SP thereby providing in vivo evidence of a reasonable therapeutic index. Sabutoclax not only reduced the expression of BCL2 and MCL1 mRNA long isoform in the BM, but also significantly reduced the number of human CD34+, BCL2+ and MCL1+ cells in both BM and SP. These results demonstrate that BM and SP niche BC CML LSC survival is driven by overexpression of multiple pro-survival BCL2 family isoforms rendering them susceptible to a novel pan-BCL2 inhibitor, sabutoclax, at doses that spare normal hematopoietic progenitors.

Citation Format: Wenxue Ma, Janine Low-Marchelli, Heather S. Leu, Angela Court Recart, Jun Wei, Xianshu Huang, Maryla Krajewska, Maurizio Pellecchia, John C. Reed, Catriona H.M. Jamieson. BCR-ABL tyrosine kinase inhibition combined with a novel pan BCL2 family inhibitor abrogates leukemia stem cell survival in a niche-dependent manner. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 217. doi:10.1158/1538-7445.AM2013-217