Marked inactivation of O6-alkylguanine-DNA alkyltransferase activity with protracted temozolomide schedules

Abstract

Temozolomide, an oral DNA methylator that inactivates the DNA repair enzyme O(6)-alkylguanine-DNA alkyltransferase (AGAT), has demonstrated anticancer activity on protracted schedules. Protracted schedules may lead to an 'autoenhancement' of temozolomide's inherent cytotoxic potential by cumulative reduction of the cell's capacity for AGAT-mediated DNA repair and resistance. This study was undertaken to characterise AGAT inactivation and regeneration in the peripheral blood mononuclear cells (PBMCs) of patients treated on two protracted temozolomide schedules. O(6)-alkyl guanine-DNA alkyltransferase activity was measured in the PBMCs of patients treated on two phase I protracted temozolomide studies. Patients were treated daily for either 7 days every 2 weeks (Schedule A) or 21 days every 4 weeks (Schedule B). The effects of various temozolomide doses (75-175 mg m(-2)), treatment duration (7-21 days), and temozolomide plasma levels on AGAT inactivation and regeneration, as well as the relation between AGAT inactivation and toxicity, were studied. O(6)-alkyl guanine-DNA alkyltransferase activity in PBMCs was measured serially in 52 patients. Marked inactivation of AGAT occurred following 7 days of temozolomide treatment, with mean AGAT activity decreasing by 72% (P<0.0001). Similarly, mean AGAT activity decreased by 63 and 73% after 14 and 21 days of treatment, respectively (P<0.001 for both comparisons). O(6)-alkyl guanine-DNA alkyltransferase inactivation was greater after 7 days of treatment with higher doses of temozolomide than lower doses and remained markedly reduced 7 days post-treatment. However, AGAT inactivation following temozolomide treatment for 14 and 21 days was similar at all doses. On the continuous 21-day schedule, AGAT inactivation was significantly greater in patients who experienced severe thrombocytopenia than those who did not (90.3+/-5.5 vs 72.5+/-16.1%, P<0.045). In conclusion, protracted administration of temozolomide, even at relatively low daily doses, leads to significant and prolonged depletion of AGAT activity, which may enhance the antitumour activity of the agent.

Figure 1

Scatterplots of AGAT activity in PBMCs sampled (A), pretreatment and after 7 days of daily temozolomide treatment administered (at doses of 75–175 mg m⁻² day⁻¹) on Schedule A; and (B) after 14 and 21 days of temozolomide treatment (at doses of 85–125 mg m⁻² day⁻¹) on Schedule B.

Figure 2

Scatterplots of the percentage values of PBMC AGAT inactivation for Schedules A (day 8 values) and B (day 15 and 22 values) at each temozolomide dose: (A) 75 mg m⁻² day⁻¹ (day 8 only) and 85 mg m⁻² day⁻¹ (days 15 and 22); (B) 100 mg m⁻² day⁻¹; (C) 125 mg m⁻² day⁻¹; (D) 150 mg m⁻² day⁻¹; and (E) 175 mg m⁻² day⁻¹. Dose-limiting toxicity precluded daily treatment with temozolomide for longer than 7 days at the 150 and 175 mg m⁻² day⁻¹ dose levels.

Figure 3

(A) Scatterplot of the percentage values of PBMC AGAT inactivation as a function of the total cumulative temozolomide dose administered up to the time of PBMC sampling (R²=0.102). (B) Scatterplot of temozolomide exposure as the product of AUC_0-t multiplied by total days temozolomide administered prior to PBMC sampling (R²=0.188).

Figure 4

Mean (s.d.) AGAT activity at baseline and day 8, and recovery at day 15 following administration of temozolomide on days 1–8.