Most patients with acute myeloid leukemia (AML) display an initial response to high dose chemotherapy but a majority develops resistance and relapse within 1–2 years. The targeted drug gemtuzumab ozogamicin (GO) has shown promising clinical effects. Consisting of a monoclonal CD33 antibody coupled to a toxin, calicheamicin, GO enters the cell after binding to the CD33 antigen, preferentially expressed on myeloid cells. Calicheamicin is then released from the antibody and causes multiple signaling effects resulting in cell death. We have previously shown that GO-induced apoptotic signaling in AML cells involves activation of the Bcl-2 proteins Bax and/or Bak. In this study we aimed to clarify if altered DNA repair or DNA damage signaling are molecular determinants of GO-sensitivity in AML. Moreover, attempts were made to connect the upstream DNA double stranded break (dsb) signaling to GO-induced activation of Bax and Bak. Therefore, we analyzed GO-induced DNA damage signaling in GO responsive (HL60) and resistant (KG1a) AML cells. We can show that GO-resistant cells display a higher basal activity of γH2AX and DNA-PKcs and hence a higher DNA repair capacity. Moreover, by treating AML cells with clinically relevant doses of GO we have observed an increased level of both γH2AX and phosphorylated DNA-PKcs which indicates the prevalence of DNA dsbs. This activation was observed in HL60 as well as in KG1a cells, in vitro. Thus, our data suggest that resistance to GO is not due to lack of DNA dsbs induction, but is rather influenced by the DNA dsbs signaling to the downstream apoptotic components. Key activators downstream of DNA-PK but upstream of Bak/Bax, including c-Abl, caspase-2, Bid, Bad, Bim, Bcl-2 and Bcl-xL, are therefore currently being analyzed. In conclusion, we show that GO induced DNA dsbs in both GO sensitive and GO resistant AML cells in vitro, and that resistance to GO treatment is at least in part due to factors downstream of DNA-PK but upstream of Bak/Bax activation.

Disclosures: No relevant conflicts of interest to declare.

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