THE FORMATION AND PERSISTENCE OF CARBOPLATIN-DNA ADDUCTS IN RATS

The formation and persistence of platinum-DNA adducts were studied wit h immuno(cyto)chemical methods in male and female Sprague-Dawley rats treated with a single i.p. dose of carboplatin. Linear dose-effect cur ves were observed for kidney and liver with an immunocytochemical assa y using NKI-A59 antiserum that recognizes intrastrand cross-links. Wit h this method, no staining of the nuclei due to platinum-DNA damage co uld be observed in the spleen, testis, uterus, or ovary after administ ration of up to 80 mg/kg carboplatin. A homogeneous staining of the nu clei in the liver was observed. The nuclear staining in the kidney was somewhat more intense but less homogeneous, with small groups of inte nsely stained nuclei occasionally being seen in the outer cortex. An a pproximately 15 to 20-times lower dose of cisplatin than of carboplati n was needed to reach equal staining levels in the liver and kidney. P lateau staining levels in both tissues were reached at between approxi mately 8 and 48 h after administration of the carboplatin. This was fo llowed by a significant reduction in the kidney samples, whereas the s taining levels in the liver section seemed to be more persistent. No m ajor difference was observed between male and female rats in the forma tion and removal of DNA damage in these tissues. The levels of the var ious DNA adducts were measured with a competitive ELISA in liver, kidn ey, spleen, testis, and combined ovary/uterus samples collected at 8 a nd 48 h after carboplatin administration. At both 8 and 48 h, the high est platination levels were observed in the kidney, followed-in decrea sing order-by the liver, combined uterus and ovary samples, spleen, an d testis. At 8 h after administration of carboplatin, the relative occ urrence of the bifunctional adducts Pt-GG (34%), Pt-AG (27%), and G-Pt -G (32%), was similar in all tissues. The same held for the monoadduct s that amounted to about 7% of the total DNA platination. These data i ndicate that in the first few hours after carboplatin treatment, no pr eference for the formation of Pt-CG adducts was observed, which confir ms our earlier observations obtained with cultured cells. When the tot al DNA-platination levels (calculated from the sum of the adducts) see n at 8 and 48 h after treatment were compared, a substantial decrease in DNA platination was observed in the kidney (37%), liver (30%) and o vary/uterus (39%), whereas the repair levels in the testis (9%) and, p robably, the spleen (18%) were substantially lower. In all tissues stu died, only the relative occurrence of the Pt-GG adducts increased betw een 8 and 48 h, and as a result, at 48 h, after carboplatin administra tion the Pt-GG adduct was the major adduct persisting in the DNA sampl es.