Ames, Matthew M.

Purpose Polymorphisms in tamoxifen metabolizing genes affect the plasma concentration of tamoxifen metabolites, but their effect on clinical outcome is unknown. Methods We determined cytochrome P450 (CYP)2D6 (*4 and *6) and CYP3A5 (*3) genotype from paraffin-embedded tumor samples and buccal cells (living patients) in tamoxifen-treated women enrolled onto a North Central Cancer Treatment Group adjuvant breast cancer trial. The relationship between genotype and disease outcome was determined using the log-rank test and Cox proportional hazards modeling. Results Paraffin blocks were obtained from 223 of 256 eligible patients, and buccal cells were obtained from 17 living women. CYP2D6 (*4 and *6) and CYP3A5 (*3) genotypes were determined from 190, 194, and 205 patient samples and in 17 living women. The concordance rate between buccal and tumor genotype was 100%. Women with the CYP2D6 *4/*4 genotype had worse relapse-free time (RF-time; P = .023) and disease-free survival (DFS; P = .012), but not overall survival (P = .169) and did not experience moderate to severe hot flashes relative to women heterozygous or homozygous for the wild-type allele. In the multivariate analysis, women with the CYP2D6 *4/*4 genotype still tended to have worse RFS (hazard ratio [HR], 1.85; P = .176) and DFS (HR, 1.86; P = .089). The CYP3A5*3 variant was not associated with any of these clinical outcomes. Conclusion In tamoxifen-treated patients, women with the CYP2D6 *4/*4 genotype tend to have a higher risk of disease relapse and a lower incidence of hot flashes, which is consistent with our previous observation that CYP2D6 is responsible for the metabolic activation of tamoxifen to endoxifen.

Purpose To determine the dose-limiting toxicities, maximum-tolerated dose, and pharmacokinetics of the cyclin-dependent kinase inhibitor flavopiridol (NSC 649890) when administered as a 1-hour infusion over 3 consecutive days to children with recurrent or refractory solid tumors. Patients and Methods Flavopiridol was administered as a 1-hour intravenous infusion daily for 3 consecutive days every 21 days, or when hematologic toxicity or any grade 2 or greater nonhematologic toxicity resolved. The starting dose was 37.5 mg/m2/d. Dose escalation was in cohorts of three patients in a standard fashion until dose-limiting toxicity and the maximum-tolerated dose were determined. Flavopiridol levels were measured on days 1, 2, and 3. Results Twenty-five children received flavopiridol at doses of 37.5 to 80 mg/m2/day over 3 consecutive days. The maximum-tolerated dose was 62.5 mg/m2/d. The primary dose-limiting toxicities were neutropenia and diarrhea. No antitumor effect was observed in this population. Mean peak plasma concentrations of 3.71 and 9.11 μmol/L were achieved at the end of the 1-hour infusion, following dose escalation from 37.5 mg/m2 to 80 mg/m2, respectively. The median flavopiridol plasma clearance was 8.0 L/h/m2 (range, 2.6 to 17.1 L/h/m2). Conclusion The maximum-tolerated dose of flavopiridol in children, and the recommended phase II dose for pediatric studies, was 62.5 mg/m2/day when administered as a 1-hour infusion for 3 consecutive days. Dose-limiting toxicities of neutropenia and diarrhea were similar to those in adult studies.

Abstract Pyrazoloacridine (PZA) is an experimental antitumor agent presently under investigation for treatment of solid tumors on the basis of its unique mechanism of action and selectivity for human solid tumor xenograft in mice. Using capillary electrophoresis coupled with electrospray ionization mass spectrometry, we have identified three oxidative PZA metabolites, 9-desmethyl-PZA, N-demethyl-PZA, and PZA N-oxide. The cytochrome P450 (CYP) isoforms involved in PZA metabolism were characterized by studies with CYP chemical inhibitors, correlation of marker activities for selected CYPs with formation of the metabolites using a human liver panel, and PZA metabolism by cDNA-expressed CYPs. 9-Desmethyl-PZA formation was catalyzed by CYP1A2, whereas N-demethyl-PZA formation was catalyzed by CYP3A4. PZA N-oxide formation was catalyzed by flavin monooxygenase (FMO) rather than CYP, as determined by studies with chemical inhibitors of FMO and metabolism by cDNA-expressed human flavin monooxygenase. After administration of [10b-14C]PZA to mice, six urinary metabolites were detected by high-performance liquid chromatography UV and radiochromatograms including 9-desmethyl-PZA, N-demethyl-PZA, and PZA N-oxide. Trace concentrations of 9-desmethyl-PZA and PZA N-oxide were detected in mouse plasma. PZA N-oxide and N-demethyl-PZA were detected in urine from patients after PZA administration. PZA, 9-desmethyl-PZA, and PZA N-oxide inhibited growth of A375 human melanoma cells. IC50 values were 0.17, 0.11, and 7.0 μm, respectively, for the three molecules.

Abstract Purpose: To assess the maximum tolerated dose, toxicities, pharmacokinetics, and antileukemic activity of topotecan and carboplatin in adults with recurrent or refractory acute leukemias. Experimental Design: Patients received topotecan and carboplatin by 5-day continuous infusion at nine dose levels. Patients achieving a complete remission received up to two additional courses for consolidation. Plasma topotecan and ultrafilterable platinum were assayed on days 1 to 5. In addition, pretreatment levels of various polypeptides in leukemic cells were examined by immunoblotting to assess possible correlations with response. Results: Fifty-one patients received a total of 69 courses of therapy. Dose-limiting toxicity consisted of grade 4/5 typhlitis and grade 3/4 mucositis after one course of therapy or grade 4 neutropenia and thrombocytopenia lasting >50 days when a second course was administered on day 21. Among 45 evaluable patients, there were 7 complete remissions, 2 partial remissions, 1 incomplete complete remission, and 1 reversion to chronic-phase chronic myelogenous leukemia. Topotecan steady-state plasma concentrations increased with dose. No accumulation of topotecan or ultrafilterable platinum occurred between days 1 and 5 of therapy. Leukemic cell levels of topoisomerase I, checkpoint kinase 1, checkpoint kinase 2, and Mcl-1 correlated with proliferating cell nuclear antigen but not with response. In contrast, low Bcl-2 expression correlated with response (P = 0.014, Mann-Whitney U test). Conclusions: The maximum tolerated dose was 1.6 mg/m2/d topotecan plus 150 mg/m2/d carboplatin. The complete remission rate in a heavily pretreated population was 16% (33% at the highest three dose levels). Responses seem to correlate with low pretreatment blast cell Bcl-2 expression.

Abstract Purpose: Flavopiridol, a cyclin-dependent kinase inhibitor, transcription inhibitor, and DNA-interacting agent, was combined with cisplatin or carboplatin to establish toxicities, evaluate pharmacokinetics, and examine its effects on patient cancers and levels of selected polypeptides in patient peripheral blood mononuclear cells (PBMC). Experimental Design: Therapy was given every 3 weeks. Stage I: cisplatin was fixed at 30 mg/m2 with escalating flavopiridol. Stage II: flavopiridol was fixed at the stage I maximum tolerated dose (MTD) with escalation of cisplatin. Stage III: flavopiridol was fixed at the stage I MTD with escalation of carboplatin. Results: Thirty-nine patients were treated with 136 cycles of chemotherapy. Neutropenia was seen in only 11% of patients. Grade 3 flavopiridol/CDDP toxicities were nausea (30%), vomiting (19%), diarrhea (15%), dehydration (15%), and neutropenia (10%). Flavopiridol combined with carboplatin resulted in unexpectedly high toxicities and one treatment-related death. Stable disease (>3 months) was seen in 34% of treated patients, but there were no objective responses. The stage II MTD was 60 mg/m2 cisplatin and 100 mg/m2/24 hours flavopiridol. As given, CDDP did not alter flavopiridol pharmacokinetics. Flavopiridol induced increased p53 and pSTAT3 levels in patient PBMCs but had no effects on cyclin D1, phosphoRNA polymerase II, or Mcl-1. Conclusions: Flavopiridol and cisplatin can be safely combined in the treatment of cancer patients. Unexpected toxicity in flavopiridol/carboplatin-treated patients attenuates enthusiasm for this alternative combination. Analysis of polypeptide levels in patient PBMCs suggests that flavopiridol may be affecting some, but not all, of its known in vitro molecular targets in vivo.

Purpose We determined the maximum-tolerated dose (MTD) and the dose-limiting toxicities (DLT) of 17-allylamino-17-demethoxygeldanamycin (17-AAG) when infused on days 1, 8, and 15 of a 28-day cycle in advanced solid tumor patients. We also characterized the pharmacokinetics of 17-AAG, its effect on chaperone and client proteins, and whether cytochrome P450 (CYP) 3A5 and NAD(P)H:quinone oxidoreductase 1 (NQO1) polymorphisms affected 17-AAG disposition or toxicity. Patients and Methods An accelerated titration design was used. Biomarkers were measured in peripheral-blood mononuclear cells (PBMCs) at baseline and on days 1 and 15, and pharmacokinetic analysis was performed on day 1 of cycle 1. CYP3A5*3 and NQO1*2 genotypes were determined and correlated with pharmacokinetics and toxicity. Results Twenty-one patients received 52 courses at 11 dose levels. DLTs at 431 mg/m2 were grade 3 bilirubin (n = 1), AST (n = 1), anemia (n = 1), nausea (n = 1), vomiting (n = 1), and myalgias (n = 1). No tumor responses were seen. 17-AAG consistently increased heat shock protein (Hsp) 70 levels in PBMCs. At the MTD, the clearance and half-life (t1/2) of 17-AAG were 11.6 L/h/m2 and 4.15 hours, respectively; whereas the active metabolite 17-aminogeldanamycin had a t1/2 of 7.63 hours. The CYP3A5*3 and NQO1*2 polymorphisms were not associated with 17-AAG toxicity. The CYP3A5*3 polymorphism was associated with higher 17-AAG clearance. Conclusion The MTD of weekly 17-AAG is 308 mg/m2. 17-AAG induced Hsp70 in PBMCs, indicating that Hsp90 has been affected. Further evaluation of 17-AAG is ongoing using a twice-weekly regimen, and this schedule of 17-AAG is being tested in combination with chemotherapy.

Purpose To determine the maximum tolerated dose, toxicity, and pharmacokinetics of gemcitabine in children with refractory solid tumors. Patients and Methods Gemcitabine was given as a 30-minute infusion for 2 or 3 consecutive weeks every 4 weeks, to 42 patients aged 1 to 21 years. Doses of 1,000, 1,200 and 1,500 mg/m2 were administered for 3 weeks. Subsequently, gemcitabine was given for only 2 consecutive weeks at 1,500, 1,800, and 2,100 mg/m2. Plasma concentrations of gemcitabine and its metabolite, 2′2′-difluorodeoxyuridine, were measured in 28 patients. Results Forty patients who received 132 courses of gemcitabine were assessable for toxicity. The maximum tolerated dose of gemcitabine given weekly for 3 weeks was 1,200 mg/m2. Dose-limiting toxicity was not seen in one-third of children treated at any doses given for 2 weeks. The major toxicity was myelosuppression in three of five patients at 1,500 mg/m2 for 3 weeks, and one of seven patients at 1,800 mg/m2 for 2 weeks. Other serious adverse events were somnolence, fever and hypotension, and rash in three patients. Gemcitabine plasma concentration–time data were fit to a one- (n = 5) or two-compartment (n = 23) open model. Mean gemcitabine clearance and half-life values were 2,140 mL/min/m2 and 13.7 minutes, respectively. One patient with pancreatic cancer had a partial response. Seven patients had stable disease for 2 to 17 months. Conclusion Gemcitabine given by 30-minute infusion for 2 or 3 consecutive weeks every 4 weeks was tolerated well by children at doses of 2,100 mg/m2 and 1,200 mg/m2, respectively.

Purpose: We sought to determine the maximum-tolerated dose (MTD) and evaluate the toxicities and clinical activity of two irinotecan (CPT-11), fluorouracil (FU), leucovorin (LV), and oxaliplatin schedules in patients with advanced solid tumors. Additionally, we investigated the effect of CPT-11 on oxaliplatin pharmacokinetics. Patients and Methods: Thirteen patients (cohort 1) received intravenous CPT-11 (infusion) and FU/LV (bolus) on days 1, 8, 15, and 22 and oxaliplatin (infusion) on days 1 and 15 every 6 weeks for a total 37 courses (median, three courses) at three dose levels. Twenty-two cohort 2 patients received intravenous CPT-11/oxaliplatin (infusion, day 1) and FU/LV (90-minute bolus infusion, days 2 to 5) every 3 weeks for a total of 122 courses (median, four courses) at three dose levels. Pharmacokinetic and neurotoxicity assessments were performed at the cohort 2 MTD. Results: Dose-limiting toxicity (DLT) seen in both cohorts at the starting dose required dose de-escalation. Cohort 1 DLT included diarrhea and neutropenia. In cohort 2, diarrhea, vomiting, dehydration, neutropenia, febrile neutropenia, and paresthesias were DLTs. Antitumor activity was seen in both cohorts. In cohort 2, the total platinum area under the curve of patients increased 17% in cycle 2 (P = .048), but objective neurotoxicity was not seen. Conclusion: The toxicities resulting from the addition of oxaliplatin to CPT-11/FU/LV are significant but manageable. The MTDs for the weekly schedule are CPT-11 (75 mg/m2), oxaliplatin (50 mg/m2), FU (320 mg/m2), and LV (20 mg/m2); and, for the 3-weekly schedule, the MTDs are CPT-11 (175 mg/m2), oxaliplatin (85 mg/m2), FU (240 mg/m2), and LV (20 mg/m2). Second-cycle platinum accumulation raises the possibility for enhanced cumulative neurotoxicity with CPT-11/oxaliplatin combinations.