Standard Guide for Performance of Chinese Hamster Ovary Cell/Hypoxanthine Guanine Phosphoribosyl Transferase Gene Mutation Assay

Abstract

2.1 The CHO/HGPRT assay detects forward mutations of the X-linked hypoxanthine-guanine phosphoribosyl transferase (hgprt) locus (coding for the enzyme, HGPRT) in Chinese hamster ovary (CHO) cells. Cells originally derived from Chinese hamster ovary tissue are exposed to a test article and, following an appropriate cell culture regimen, descendants of the original treated population are monitored for the loss of functional HGPRT, presumably due to mutations. Resistance to a purine analogue, 6-thioguanine (6TG) (or less desirably, 8-azaguanine (8AG)), is employed as the genetic marker. HGPRT catalyzes the conversion of the nontoxic 6TG to its toxic ribophosphorylated derivative. Loss of the enzyme or its activity therefore leads to cells resistant to 6TG. 2.2 Because HGPRT is an enzyme of the purine nucleotide salvage pathway, loss of the enzyme is not a lethal event. Different types of mutational events (base substitutions, frame shifts, deletions, and so forth) can be detectable at the hgprt locus. The assay does not appear to detect large-scale chromosomal damage. The CHO/HGPRT assay has been used to study a wide range of mutagens, including radiations (2-4), and a wide variety of chemicals (1) and complex chemical mixtures (5).