4EGI-1 induces a high level of apoptosis in AML blast cells but not in normal CD34⁺ hematopoietic progenitors. Caspase-3 cleavage: AML blast cells from 3 different AML patients were cultured 24 hours in 10% FCS MEM without or with 50 or 100 μM 4EGI-1. The same experimental procedure was applied in 3 CD34⁺ samples, and 25 μM LY294002 was used as a control for apoptosis induction (1 representative sample is provided). Protein extracts from 10⁶ cells were resolved by SDS-PAGE electrophoresis, transferred to nitrocellulose, and probed with anti–caspase-3 and antiactin antibodies. Annexin V binding: 2 × 10⁵ cells were cultured without or with increasing concentrations of 4EGI-1 (from 25 to 100μM) during 24 or 48 hours, and then labeled with annexin V–PE for flow cytometry analysis. Results are expressed as a mean of experiments done on 5 primary AML samples and on 5 different CD34⁺ samples. Vertical bars represent SD. ***P < .001.