Targeting FLT3-ITD in AML using TKI against FLT3 cannot prevent relapse even in the presence of complete remission, suggesting the resistance and/or the persistence of leukemic-initiating cells in the hematopoietic niche. By mimicking the hematopoietic niche condition with cultures at low oxygen concentrations, we demonstrate in vitro that FLT3-ITD AML cells decrease their repopulating capacity when Vps34 is inhibited. Ex vivo, AML FLT3-ITD blasts treated with Vps34 inhibitors recovered proliferation more slowly due to an increase an apoptosis. In vivo, mice engrafted with FLT3-ITD AML MV4-11 cells have the invasion of the bone marrow and blood in 2 weeks. After 4 weeks of FLT3 TKI treatment with gilteritinib, the leukemic burden had strongly decreased and deep remission was observed. When treatment was discontinued, mice relapsed rapidly. In contrast, Vps34 inhibition strongly decreased the relapse rate, and even more so in association with mobilization by G-CSF and AMD3100. These results demonstrate that remission offers the therapeutic window for a regimen using Vps34 inhibition combined with mobilization to target persistent leukemic stem cells and thus decrease the relapse rate.

Chronic lymphocytic leukemia (CLL) is an incurable indolent non-Hodgkin lymphoma characterized by tumor B-cells that weakly express a B-cell receptor (BCR). The mutational status of the variable region (IGHV) within the immunoglobulin heavy-chain (IGH) locus is an important prognosis indicator and raises the question of the CLL cell of origin (COO). Mutated IGHV gene CLLs (mCLLs) are genetically imprinted by activation induced-cytidine deaminase (AID). AID is also required for IGH rearrangements: class switch recombination (CSR) and recombination between switch Mu (Sμ) and the 3’ regulatory region (3’RR) (Sμ-3’RRrec). The great majority of CLL B-cells being unswitched led us to examine IGH rearrangement blockade in CLL. Our results separated CLLs into two groups on the basis of Sμ-3’RRrec counts per sample: Sμ-3’RRrecHigh cases (mostly umCLLs) and Sμ-3’RRrecLow cases (mostly mCLLs), but not based on CSR junction counts. Sμ-3’RRrec appeared to be ongoing in Sμ-3’RRrecHigh CLL cells and comparison of Sμ-3’RRrec junction structural features pointed to different B-cell origins for both groups. In accordance with IGHV mutational status and PIM1 mutation rate, Sμ-3’RRrecHigh CLLs harbor a non-GC experienced B-cell imprint while Sμ- 3’RRrecLow CLLs are from AID-experienced B-cells from a secondary lymphoid organ. In addition to the proposals already made concerning the CLL cell of origin, our study highlights that analysis of IGH recombinatory activity can identify CLL cases from different origins. Finally, on-going Sμ-3’RRrec in Sμ-3’RRrecHigh cells appeared to presumably be the consequence of high c-MYC expression, as c-MYC overexpression potentiated IGH rearrangements and Sμ-3’RRrec, even in the absence of AID for the latter.