Anti-FLT3 monoclonal antibody, clone FQS3422(3)

Introduction: Hematopoietic stem cell transplantation (HSCT) is a proper treatment for many cancers, including malignant and non-malignant hematologic and immune disorders. However, due to the small quantity of HSCTs, their number should be increased ex vivo. Cell proliferation increase by animal serum can transmit certain diseases and immune problems to the recipient; therefore, replacement of culture media with media of human origin seems to be crucial. Acellular Wharton's jelly (AWJ) can be considered as an alternative medium in this regard. This study aimed to investigate the effect of AWJ substitution for fetal bovine serum (FBS) on the expression of myeloid and lymphoid specific markers and CD34(+) cells proliferation pattern following co-culture with mesenchymal stem cells. Materials and methods: In this study, CD34(+) umbilical cord blood HSCs were co-cultured with MSCs in the presence of 10% FBS or AWJ, stem cell factor (SCF), thrombopoietin (TPO), and FLT3-L and the proliferation pattern and expression of myeloid as well as lymphoid lineage markers were then evaluated. Results: The expression of myeloid and lymphoid specific markers was increased and decreased in AWJ and FBS containing culture medium, respectively. Conclusion: This study provided evidence for the differentiation of UCB-derived CD34(+) cells toward myeloid lineage and lack of their differentiation toward lymphoid lineage in AWJ containing culture medium.

Class III receptor tyrosine kinases control the development of hematopoietic stem cells. Constitutive activation of FLT3 by internal tandem duplications (ITD) in the juxtamembrane domain has been causally linked to acute myeloid leukaemia. Oncogenic FLT3 ITD is partially retained in compartments of the biosynthetic route and aberrantly activates STAT5, thereby promoting cellular transformation. The pool of FLT3 ITD molecules in the plasma membrane efficiently activates RAS and AKT, which is likewise essential for cell transformation. Little is known about features and mechanisms of FLT3 ligand (FL)-dependent internalization of surface-bound FLT3 or FLT3 ITD. We have addressed this issue by internalization experiments using human RS4-11 and MV4-11 cells with endogenous wild-type FLT3 or FLT3 ITD expression, respectively, and surface biotinylation. Further, FLT3 wild-type, or FLT3 ITD-GFP hybrid proteins were stably expressed and characterized in 32D cells, and internalization and stability were assessed by flow cytometry, imaging flow cytometry, and immunoblotting. FL-stimulated surface-exposed FLT3 WT or FLT3 ITD protein showed similar endocytosis and degradation characteristics. Kinase inactivation by mutation or FLT3 inhibitor treatment strongly promoted FLT3 ITD surface localization, and attenuated but did not abrogate FL-induced internalization. Experiments with the dynamin inhibitor dynasore suggest that active FLT3 as well as FLT3 ITD is largely endocytosed via clathrin-dependent endocytosis. Internalization of kinase-inactivated molecules occurred through a different yet unidentified mechanism. Our data demonstrate that FLT3 WT and constitutively active FLT3 ITD receptor follow, despite very different biogenesis kinetics, similar internalization and degradation routes.