Effective Ex Vivo-Generation and Quantification of Leukemia-Derived DC Has To Precede a Specific T-Cell Stimulation for Adoptive Immunotherapy in AML and MDS.

The presentation of leukemic antigens can be improved in AML and MDS by in vitro conversion of leukemic cells in leukemia-derived DC (DC_leu), thereby forming a platform for the generation of leukemia-specific cytotoxic lymphocytes (CTL). In preliminary analyses we could already define optimal serum-free culture conditions, generate and characterize DC from 55 MDS- and 99 AML-cases and characterize their T-cell activating function (Kufner S. 2005 I-III). DC/DC_leu were quantified according to their surface DC /blast -marker profiles. However in 6/30% of cases with AML/MDS less than 10% of DC could be generated and only 50–60% of leukemic cells were convertible to DC_leu. Therefore we tested 5 alternative methods to improve the harvest of DC/DC_leu in 99 AML and 55 MDS-patients (1) ‘MCM-mimic’ (Lee, 2003), 2) ‘Ca-Ionophore’ (Houtenbos, 2003), 3) ‘Picibanil’ (Sato, 2003), 4) ‘Cytokines’ (Westers, 2003) and 5) ‘Poly I:C’ (Rouas, 2004)). 1. Although the percentual harvest of DC/DCleu was comparable in all of the methods (28–40% DC, with 45–65% of those DC being DC_leu and about 47% of blasts being convertible to DC_leu) not every method was successful in individual patients. 2. However we could demonstrate that DC can be generated from every sample with at least one of the following methods: MCM-mimic, Ca-Ionophore and Picibanil. 3. Highest DC/DC_leu yield was seen in monocytoid differentiated FAB-types and was independent of the cytogenetic risk. 4. Many ‘DC-surface markers’ can be aready expressed on naive blasts in AML/MDS-patients before there conversion to DC_leu. Moreover the ‘DC-marker expression’ is variable in individual patients and culture methods.

In summary our data show 1. that the generation of DC/DC_leu is possible independent from the karyotype in every patient under serum-free conditions with at least one of the 3 methods MCM-mimic, Ca-Ionophore and Picibanil. 2. The overall percentual harvest of DC_leu from MNC-fractions after culture in the 5 methods compared is low. Possibly a combination of the optimal method with optimal ‘DC- and blast markers’ in every single case could improve the detectability and quantification of DC/DC_leu and residual blasts. This contributes to improve the generability and quality of DC_leu to stimulate and expand anti-leukemia-directed T-cells for the immunotherapy of AML and MDS.