GCN2iB

The treatment of acute lymphoblastic leukemia (ALL) cells with GCN2iB renders ALL cells sensitive to ASNase by preventing the induction of ASNS, resulting in reduced levels of de novo protein synthesis. Combined treatment with ASNase and GCN2iB induces the stress-activated MAPK pathway, thereby triggering apoptosis. By using cell-panel analyses, we also shows that acute myelogenous leukemia and pancreatic cancer cells are highly sensitive to the combined treatment. ^[1]

The combination of ASNase treatment with GCN2iB synergistically blocks in vivo tumor growth in ALL, AML, and pancreatic xenograft models. ^[1]

• GCN2 kinase assay

  Recombinant GCN2 (1 nmol/L) protein is pre-incubated with GCN2 inhibitors for 60 min and then incubated with ATP (K_M value of GCN2 = 190 μmol/L) and the green fluorescent protein-eIF2α substrate (130 nmol/L) at 25°C. The amount of phosphorylated substrate is determined using the LanthaScreen Tb-anti-p-eIF2α (pSer52) antibody kit. The IC50 value of eIF2α kinase is measured using the XLfit software.

• Cell lines

  Pancreatic cancer cells, CCRF-CEM cells, MEF cells (GCN2-WT or -KO), CCRF-CEM, MV-4-11, SU.86.86 cells

• Concentrations

  37 nM, 110 nM, 330 nM, 1.0 μM

• Incubation Time

  4 h, 72 h

• Method

  CCRF-CEM cells are treated with GCN2iB as indicated for 4 h. Cell lysates are analyzed by western blot; MEF cells (GCN2-WT or -KO) are treated with GCN2 inhibitors as indicated for 72 h. Cell viability is measured; CCRF-CEM, MV-4-11, and SU.86.86 cells are treated with 1 μmol/L GCN2iB as indicated for 72 h. Cell viability is measured at day 0 and day 3; CCRF-CEM and SU.86.86 cells are treated with GCN2 inhibitors as indicated for 72 h. Cell viability is measured.

• Animal Models

  6-week-old female SCID mice

• Dosages

  1 mg/kg, 3 mg/kg, 10 mg/kg

• Administration

  Oral gavage, IP