Role of the Spleen In CML: Lessons from an Inducible Transgenic Mouse Model.
Authors: Schemionek Mirle , Elling Christian , Kamp Linda , Spieker Tilmann , Berdel Wolfgang E. , Mueller-Tidow Carsten , Koschmieder Steffen 
Published: October 2019

Abstract
Abstract 3384
Spleen size is one of the most important independent pre-treatment risk factors (Sokal and Hasford Scores) in patients with chronic myeloid leukemia (CML). However, the role of the spleen in the pathogenesis of CML remains poorly defined. We used an inducible transgenic mouse model of CML (ScltTA/Bcr-Abl) in which we have previously shown that the disease phenotype is transplantable using unfractionated BM cells or Lin−c-kit+Sca-1+ (LSK) cells. Here, we compared the effects of Bcr-Abl induction on cells derived from the bone marrow (BM) versus spleen and assessed primary mice as well as transplanted and serially transplanted congenic recipients by flow cytometry, clonogenic assays, Bcr-Abl expression levels, and histology. In addition, we assessed the effect of imatinib treatment on leukemic spleen cells in comparison to BM cells in transplanted mice. The results showed that splenomegaly and splenic neutrophilia occurred as early as seven days after induction of Bcr-Abl by tetracycline withdrawal, and this was followed by an increase of clonogenic growth in the spleen but not BM as well as disruption of the splenic architecture on day 14. The percentage of LSK cells was increased 4.9-fold in the spleen but only 1.5-fold in the BM on day 28 after Bcr-Abl induction. To address functional differences including the self-renewal potential between BM- and spleen-derived CML-initiating cells, we conducted comparative transplantations of induced vs. un-induced BM and spleen cells. While Bcr-Abl positive BM-transplanted recipients developed CML-like disease including neutrophilia and splenomegaly, there was no significant difference between recipients transplanted with induced vs- un-induced BM donor cells. Conversely, mice transplanted with induced spleen cells showed significantly increased percentages of granulocytic Gr1+ donor cells in BM and spleen and increased splenomegaly compared to recipients transplanted with un-induced spleen cells. Moreover, upon serial transplantation of spleen cells from these primary transplant recipients, peripheral blood analysis revealed an increased percentage of donor Gr1+ granulocytic cells in mice receiving spleen cells that had initially been induced before the first transplantation as compared to un-induced cells. Interestingly, upon serial transplantation of BM-derived cells, recipient mice developed neither an increase of granulocytic Gr1+ cells in the BM nor splenomegaly. However, percentages of immature granulocytes (Gr1low/CD11b+) were significantly increased in the spleen of mice transplanted with induced and un-induced BM cells but not in BM. Imatinib treatment in BM-transplanted recipients turned out to be less effective in spleen cells compared to BM cells. While percentages of mature donor granulocytes (Gr1high) in BM and numbers of granulocytes in PB were decreased to control levels upon 4 weeks of imatinib treatment, donor granulocytes cells were still significantly increased in the spleen. In conclusion, our data show significant differences in the dynamics and function of Bcr-Abl positive BM vs. spleen hematopoiesis. While our previous data showed that Bcr-Abl enhances differentiation of LT-HSCs in BM, our current findings suggest that leukemic, self-renewing cells are increased in the spleen thus implying that the spleen provides a reservoir for proliferating CML-initiating cells.

Disclosures:
Koschmieder: Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS: Membership on an entity's Board of Directors or advisory committees.