title: Differential response of human acute myeloid leukemia cells to gemtuzumab ozogamicin in vitro: role of Chk1 and Chk2 phosphorylation and caspase 3.
abstract: Gemtuzumab ozogamicin (GO) is a humanized anti-CD33 antibody conjugated to the anticancer agent calicheamicin, approved for the treatment of CD33+-relapsed acute myeloid leukemia. We have investigated the effects of GO on 4 human myeloid leukemia lines of different French-American-British (FAB) types (KG-1, THP-1, HL-60, and NB-4), observing 3 different types of response. Exposure to GO (10-1000 ng/mL) induced G2 arrest (up to 80% of the cells) followed by apoptosis (45% of the cells) in HL-60 and NB-4 cells. By contrast, in THP-1 cells we observed a strong G2 arrest (up to 75% of the cells) with little apoptosis. Finally, the KG-1 line was completely resistant to the same concentrations of GO. These different responses did not correlate with the levels of expression of either CD33 or multiple-drug resistance proteins, although the higher cyclosporin A (CsA)-inhibitable efflux activity of KG-1 cells may play a role in the resistance of this line to the drug. We could show that Chk1 and Chk2 phosphorylation, but not p53 or p21 expression, correlated with G2 arrest, implicating the ataxia-telangiectasia mutated/ataxia-telangiectasia related (ATM/ATR)-Chk1/Chk2 pathway in the cell cycle response to GO. However, apoptosis was associated with caspase 3 activation. Freshly isolated acute myeloid leukemia (AML) cells showed patterns of response to GO in vitro similar to those observed with the cell lines, including phosphorylation of Chk2 and caspase 3 activation. Our results suggest that the different molecular pathways induced by the drug in vitro may reflect, at least in part, the variable response to GO obtained in vivo.
authors.affiliation: Laboratory of Molecular Immunohaematology, Istituto Ricerche Farmacologiche Mario Negri, via Eritrea 62, 20157 Milan, Italy. golay@marionegri.it
authors.forename: Donatella
authors.initials: D
authors.lastname: Amico
authors.forename: Anna Maria
authors.initials: AM
authors.lastname: Barbui
authors.forename: Eugenio
authors.initials: E
authors.lastname: Erba
authors.forename: Alessandro
authors.initials: A
authors.lastname: Rambaldi
authors.forename: Martino
authors.initials: M
authors.lastname: Introna
authors.forename: Josee
authors.initials: J
authors.lastname: Golay
mesh_terms: D000208:Acute Disease; D000617:Aminoglycosides; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D000911:Antibodies, Monoclonal / Q000494:pharmacology*; D061067:Antibodies, Monoclonal, Humanized; D017209:Apoptosis / Q000187:drug effects; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism*; D000071877:Checkpoint Kinase 1; D064447:Checkpoint Kinase 2; D019008:Drug Resistance, Neoplasm; D016195:G2 Phase / Q000187:drug effects; D000079982:Gemtuzumab; D006801:Humans; D007399:Interphase / Q000187:drug effects; D007951:Leukemia, Myeloid / Q000188:drug therapy / Q000473:pathology*; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D014407:Tumor Cells, Cultured
chemical_list: D000617:Aminoglycosides; D000900:Anti-Bacterial Agents; D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; D000079982:Gemtuzumab; D011494:Protein Kinases; D064447:Checkpoint Kinase 2; C000605867:CHEK1 protein, human; C578634:CHEK2 protein, human; D000071877:Checkpoint Kinase 1; D017346:Protein Serine-Threonine Kinases; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases