Diazoketone and chloromethylketone analogs of methotrexate as potential antitumor agents

The synthesis of 4‐amino‐4‐deoxy‐N¹⁰‐methylpteroyl‐(6‐diazo‐5‐oxo)‐L‐norleucine and 4‐amino‐4‐deoxy‐N¹⁰‐methylpteroyl‐(6‐chloro‐5‐oxo)‐L‐norleucine, analogs of methotrexate in which the γ‐carboxyl group is replaced by a diazoketone and a chloromethylketone, respectively, was carried out. The analogs inhibited the growth of leukemia L‐1210 cells in culture by 50% at 4 × 10⁻⁷ M and 2 × 10⁻⁷ M, respectively, and were effective inhibitors of the synthesis of thymidylate in L‐1210 cells in vitro (I₅₀ = 3 × 10⁻⁶ M), exhibiting significant antifolate activity. The results demonstrated the feasibility of introducing chemically reactive groups at the γ‐position of pteroyl glutamates with retention of biological activity. However, in the systems investigated thus far, there was no evidence of covalent bond formation due to these reactive groups at the active sites of the enzymes.