Ansamitocin p-3 (Maytansinol isobutyrate, NSC292222)

Ansamitocin p-3 at 5 μM completely inhibits the polymerization of tubulin isolated from bovine brains, but in contrast to VCR, Ansamitocin p-3 at a high concentration of 80 μM does not leads to the aggregation of tubulin. Ansamitocin p-3 at 16 μM also potently depolymerizes the polymerized tubulin (IC50 = 3.8 μM). The addition of Ansamitocin p-3 to culture cells blocks the morphological alteration of AC cells from fibroepithelioid to a glial cell type caused by the exposure to a certain concentration of dibutyryl cyclic adenosine 3':5'-monophosphate. In addition, Ansamitocin p-3 treatment at 16 nM causes the well-defined network of cytoplasmic microtubules of A31 cells rapidly dispersed. Short-term Ansamitocin p-3 treatment also inhibits the synthesis of DNA in A31 cells or KB cells. These results confirm that Ansamitocin p-3 acts by interfering with the microtubule assembly system, thus resulting in an inhibition of mitotic spindle fiber formation and, ultimately, cytokilling. ^[1] Ansamitocin p-3 displays potent cytotoxicity against A-549, HT-29, and MCF-7 cells in a dose-dependent manner with ED50 of 4 ×10^-7, 4 × 10^-7, and 2 × 10^-6 μg/mL, respectively. ^[2] Ansamitocin p-3 also exhibits cytotoxicity against HCT-116 cells with a much low EC50 of 0.081 nM. ^[3] Ansamitocin p-3 enhances the effect of radiation both in Drosophila cells and human cancer cells in a p53 dependent manner. ^[4]

Ansamitocin p-3 treatment (>1 μg) significantly suppresses the growth of leukemia SN36, and induces an increased arrest in metaphase of P388 leukemia cells. Ansamitocin p-3 treatment at 25 μg/kg/day significantly prolongs the survival time of mice bearing i.p. B16 melanoma by 130%. Ansamitocin p-3 treatment also significantly prolongs the survival time of mice bearing Ehrlich ascites carcinoma, Sarcoma 180, and P815 mastocytoma, while slightly prolongs the survival time of mice bearing ascites MOPC-104E myeloma,leukemia L1210, and leukemia C1498. ^[1]

• Polymerization inhibition assay

  After the addition of 100 μL of various concentrations of Ansamitocin p-3 solution (GTP minus MES buffer) or 1 M Tris buffer, pH 8.4 (for blank), to 400 μL of bovine tubulin solution (1 mg/mL in cold MES buffer), maintained at 0 °C for 10 to 15 minutes, the mixture is warmed in a water bath at 37 °C for 30 to 60 minutes. The polymerization of tubulin is followed by an increase in turbidity of the mixture during warming. The turbidity measurement is performed at 460 nm with a Hitachi type 101 spectrophotometer.

• Cell lines

  A31 and KB

• Concentrations

  Dissolved in DMSO, final concentrations ~10 μM

• Incubation Time

  ~24 hours

• Method

  Cells are synchronized, and then exposed to various concentrations of Ansamitocin p-3 for ~24 hours. Cells are labeled with [³H]thymidine (5 Ci/mM, 1 μCi/mL) in 1 mL of the medium. After pulse-labeling at 37 °C for 1 hour, the cells on coverslips are fixed with methanol:acetic acid (3:1). The acidsoluble fraction is washed out from the cells, and the radio activity of each coverslip is determined by a liquid scintillation counter.

• Animal Models

  Female DBA/2 mice bearing P388, L1210, or P815 cells, C57BL/6 mice bearing B16, or C1498 cells, ICR mice bearing sarcoma 180 and EAC, and BALB/c mice bearing MOPC-104E cells

• Dosages

  ~200 μg/kg

• Administration

  Administered i.p. or i.v. daily