M S Rhee1, T J Ryan, J Galivan. 1. Wadsworth Center for Laboratories and Research, New York State Department of Health, Albany, New York 12201-0509, USA.
Abstract
PURPOSE: To examine the activity of glutamyl hydrolase (GH) on the poly-gamma-glutamates of multitargeted antifolate (MTA) (LY231514) and the effect of enhanced GH on the pharmacological activity of MTA. METHODS: Expressed and purified GH were used to study the enzymatic cleavage of MTA poly-gamma-glutamates and wild-type and GH-enhanced H35 hepatoma cell lines to evaluate growth inhibition. RESULTS: MTA tri- and penta-gamma-glutamates were good substrates for human GH, having higher rates than MTX tri- and penta-gamma-glutamates. Preferential hydrolysis with human enzyme occurred at the two gamma-glutamyl bonds at the carboxyl end of the molecule, whereas the rat enzyme preferred the innermost gamma-linkage. Incubation of rat H35 hepatoma cell lines with MTA resulted in the intracellular accumulation of primarily tetra-, penta-, and hexa-gamma-glutamate. The formation of these were markedly reduced in H35D cells, which is a line resistant to antifolates chiefly through enhanced cellular levels of GH activity. CONCLUSIONS: MTA poly-gamma-glutamates are effective substrates for GH and their pharmacological effectiveness bears an inverse relationship to cellular GH activity. This observation, along with enhanced resistance to MTA of thymidylate synthase-amplified cells, substantiates the importance of the poly-gamma-glutamates of MTA inhibiting TS as the primary target. Further evidence for the inverse relationship of GH to classical antifolate pharmacological activity is established.
PURPOSE: To examine the activity of glutamyl hydrolase (GH) on the poly-gamma-glutamates of multitargeted antifolate (MTA) (LY231514) and the effect of enhanced GH on the pharmacological activity of MTA. METHODS: Expressed and purified GH were used to study the enzymatic cleavage of MTA poly-gamma-glutamates and wild-type and GH-enhanced H35 hepatoma cell lines to evaluate growth inhibition. RESULTS:MTA tri- and penta-gamma-glutamates were good substrates for human GH, having higher rates than MTX tri- and penta-gamma-glutamates. Preferential hydrolysis with human enzyme occurred at the two gamma-glutamyl bonds at the carboxyl end of the molecule, whereas the rat enzyme preferred the innermost gamma-linkage. Incubation of rat H35 hepatoma cell lines with MTA resulted in the intracellular accumulation of primarily tetra-, penta-, and hexa-gamma-glutamate. The formation of these were markedly reduced in H35D cells, which is a line resistant to antifolates chiefly through enhanced cellular levels of GH activity. CONCLUSIONS: MTA poly-gamma-glutamates are effective substrates for GH and their pharmacological effectiveness bears an inverse relationship to cellular GH activity. This observation, along with enhanced resistance to MTA of thymidylate synthase-amplified cells, substantiates the importance of the poly-gamma-glutamates of MTA inhibiting TS as the primary target. Further evidence for the inverse relationship of GH to classical antifolate pharmacological activity is established.
Authors: Stephen J Bagley; Steven Vitale; Suhong Zhang; Charu Aggarwal; Tracey L Evans; Evan W Alley; Roger B Cohen; Corey J Langer; Ian A Blair; Anil Vachani; Alexander S Whitehead Journal: Clin Lung Cancer Date: 2016-10-26 Impact factor: 4.785
Authors: Sung-Eun Kim; Toshinori Hinoue; Michael S Kim; Kyoung-Jin Sohn; Robert C Cho; Peter D Cole; Daniel J Weisenberger; Peter W Laird; Young-In Kim Journal: Genes Nutr Date: 2014-12-13 Impact factor: 5.523
Authors: Allen L Cohn; J William Myers; Steven Mamus; Charles Deur; Steven Nicol; Karen Hood; Muhammad M Khan; Des Ilegbodu; Lina Asmar Journal: Invest New Drugs Date: 2008-02-28 Impact factor: 3.850
Authors: Yanyun Gao; Patrick Dorn; Shengchen Liu; Haibin Deng; Sean R R Hall; Ren-Wang Peng; Ralph A Schmid; Thomas M Marti Journal: Cancer Cell Int Date: 2019-11-29 Impact factor: 5.722