Measurement of folylpolyglutamate synthetase activity in head and neck squamous carcinoma cell lines and clinical samples using a new rapid separation procedure

The activity of folylpolyglutamate synthetase was measured in extracts of head and neck squamous carcinoma cell lines and in surgical specimens utilizing a new rapid method to separate free [3H]glutamate from [3H]glutamate incorporated into methotrexate, used as a substrate for the enzyme. The validity of this new method, based on reversed phase chromatography via a Sep-Pack C18 cartridge, was observed between both methods, but the Sep Pack C18 assay has the advantage that it can be accomplished in less than 5 min, whereas the DE-52 procedure requires approximately 2 hr. In seven head and neck cell lines, activity of folylpoly-glutamate synthetase varied from 335-1305 pmol [3H]glutamate incorporated/mg protein/hr. In nine head and neck tumor biopsies, a broad range in activity of folylpolyglutamate synthetase was observed (25-1827 pmol/mg/hr) which partly overlapped the enzyme activity in 'normal' tissue (7-297 pmol/mg/hr). For six patients, folylpolyglutamate synthetase was measured in the center of the tumor, in the transitional region from tumor to 'normal' tissue, and in the 'normal' tissue. The enzyme activity was higher in tumor tissue vs 'normal' tissue in four of six cases, whereas in all cases, the enzyme activity in the transitional region was higher than in 'normal' tissue. The results of this study provide further support for the concept that putative differences in folylpolyglutamate synthetase activity between tumor tissue and normal tissue can be exploited to improve the effectiveness of antifolate-based chemotherapy in general, and in head and neck cancer in particular.