7-Hydroxymethotrexate cytotoxicity and selectivity in a human Burkitt's lymphoma cell line versus human granulocytic progenitor cells: Rescue by folinic acid and nucleosides

Abstract

The cytotoxicity of 7-hydroxymethotrexate (7-OH-MTX), the primary plasma metabolite of methotrexate (MTX) in humans, was assessed by inhibition of colony formation in agar, using human bone marrow granulocyte-macrophage stem cells (CFU) from healthy volunteers and RAJI cells, a human Burkitt's lymphoma cell line. After a 2 hr exposure of cells to 7-OH-MTX, the concentrations necessary to produce a 50% inhibition of colony formation were 180 μ M and 10 μ M for bone marrow cells and for RAJI cells respectively. A continuous incubation with 20 μ M folinic acid (CF) protected the RAJI cells from 7-OH-MTX cytotoxicity at concentrations below 5 μ M but was not able to completely reverse 7-OH-MTX effects at higher doses. Continuous incubation of 7-OH-MTX-preloaded cells (2 hr, id 90) with the end products of folate-dependent reactions, adenosine (100 μ M) and thymidine (10 μ M), completely rescued RAJI cells from the 7-OH-MTX cytotoxic effects. Moreover, while thymidine alone had no effect on the 7-OH-MTX response curve, both adenosine alone or CF-adenosine combination produced 75% and 90% protection respectively. CF and adenosine concentrations necessary to achieve 90% protection were 20 and 100 μ M respectively. This study demonstrates that ( i) 7-OH-MTX can exhibit a cytotoxic selectivity for this human Burkitt's lymphoma cell line as compared to human bone marrow stem cells and ( ii) the cytotoxicity of 7-OH-MTX cannot be reversed by CF alone. These data suggest that 7-OH-MTX and/or its polyglutamylated derivatives may play an important role on different enzyme(s) involved in the interconversion of tetrahydrofolate cofactors necessary for the de novo purine biosynthesis.