Influence Of Cd33 Expression Levels And Itim-Dependent Internalization On Gemtuzumab Ozogamicin-Induced Cytotoxicity.

Gemtuzumab ozogamicin (GO; Mylotarg), a novel immunoconjugate used for treat- ment of acute myeloid leukemia (AML), contains the humanized anti-CD33 anti- body (hP67.6) as a carrier to facilitate cellular uptake of the toxic calicheami- cin-1 derivative. By use of lentivirus- mediated gene transfer to manipulate CD33 expression in myeloid cell lines that normally lack CD33 (murine 32D cells) or have very low levels of CD33 (human OCI-AML3 and KG-1a cells), we here show a quantitative relationship between CD33 expression and GO-induced cytotoxicity. The CD33 cytoplasmic immunoreceptor tyrosine-based inhibitory motifs (ITIMs) control internalization of antibody bound to CD33. Disruption of the ITIMs by intro- duction of point mutations not only pre- vented effective internalization of anti- body-bound CD33 but also significantly reduced GO-induced cytotoxicity. To- gether, our data imply a pivotal role of both the number of CD33 molecules ex- pressed on the cell surface and the amount of internalization of CD33 follow- ing antibody binding for GO-induced cyto- toxicity and suggest novel therapeutic approaches for improvement of clinical outcome of patients treated with GO. (Blood. 2005;105:1295-1302)