Exposure-response analysis of rilotumumab in gastric cancer: the role of tumour MET expression.

BACKGROUND: Rilotumumab, an investigational, monoclonal antibody, inhibits MET-mediated signalling. In a randomized phase 2 trial of rilotumumab±epirubicin/cisplatin/capecitabine in gastric or oesophagogastric junction cancer, patients receiving rilotumumab showed a trend towards improved survival, especially in MET-positive patients, but no clear dose-response relationship was observed. Exposure-response and biomarker analyses were used for dose selection and to differentiate patient subpopulations that may benefit most from treatment. Here, we analyse rilotumumab exposure-survival and exposure-safety and the impact of MET expression on these relationships. METHODS: Individual rilotumumab exposure parameters were generated using population pharmacokinetic modelling. Relationships among rilotumumab dose (7.5 and 15 mg kg(-1)), exposure, and clinical outcomes (progression-free survival (PFS) and overall survival (OS)) were evaluated with Cox regression models and Kaplan-Meier plots. MET status and other baseline covariates were evaluated in subgroup and multivariate analyses. Treatment-emergent adverse events were summarised by exposure. RESULTS: Among MET-positive patients, higher rilotumumab exposure, vs placebo and low exposure, was associated with improved median PFS (80% CI: 7.0 (5.7-9.7) vs 4.4 (2.9-4.9) and 5.5 (4.2-6.8) months) and OS (13.4 (10.6-18.6) vs 5.7 (4.7-10.2) and 8.1 (6.9-11.1) months) without increased toxicity. No rilotumumab benefit was seen among MET-negative patients. CONCLUSIONS: Rilotumumab had an exposure-dependent treatment effect in patients with MET-positive gastric or oesophagogastric junction cancer.

A phase I trial of vertical inhibition of IGF signalling using cixutumumab, an anti-IGF-1R antibody, and selumetinib, an MEK 1/2 inhibitor, in advanced solid tumours.

BACKGROUND: We completed a phase I clinical trial to test the safety and toxicity of combined treatment with cixutumumab (anti-IGF-1R antibody) and selumetinib (MEK 1/2 inhibitor). METHODS: Patients with advanced solid tumours, refractory to standard therapy received selumetinib hydrogen sulphate capsules orally twice daily, and cixutumumab intravenously on days 1 and 15 of each 28-day cycle. The study used a 3+3 design, with a dose-finding cohort followed by an expansion cohort at the maximally tolerated dose that included pharmacokinetic and pharmacodynamic correlative studies. RESULTS: Thirty patients were enrolled, with 16 in the dose-finding cohort and 14 in the expansion cohort. Grade 3 or greater toxicities included nausea and vomiting, anaemia, CVA, hypertension, hyperglycaemia, and ophthalmic symptoms. The maximally tolerated combination dose was 50 mg twice daily of selumetinib and 12 mg kg(-1) every 2 weeks of cixutumumab. Two patients achieved a partial response (one unconfirmed), including a patient with BRAF wild-type thyroid carcinoma, and a patient with squamous cell carcinoma of the tongue, and six patients achieved time to progression of >6 months, including patients with thyroid carcinoma, colorectal carcinoma, and basal cell carcinoma. Comparison of pre- and on-treatment biopsies showed significant suppression of pERK and pS6 activity with treatment. CONCLUSIONS: Our study of anti-IGF-1R antibody cixutumumab and MEK 1/2 inhibitor selumetinib showed that the combination is safe and well-tolerated at these doses, with preliminary evidence of clinical benefit and pharmacodynamic evidence of target inhibition.

Phase I study of troxacitabine administered by continuous infusion in subjects with advanced solid malignancies.

BACKGROUND: Troxacitabine is a novel L-nucleoside analogue. Preclinical studies showed improved activity with infusions of at least 3 days compared with bolus regimens, especially at concentrations >20 ng/ml. This phase I study tested the feasibility of achieving a troxacitabine steady-state concentration of 20 ng/ml for at least 72 h in patients with solid tumors. PATIENTS AND METHODS: Patients with solid tumors received troxacitabine as a progressively longer infusion on days 1-4 of a 28-day cycle. The initial length of infusion and infusion rate were 48 h and 3 mg/m(2)/day. RESULTS: Twenty-one patients were treated at infusion lengths that increased from 48 to 72 h and then 96 h. The infusion rate was decreased from 3 to 1.88 mg/m(2)/day due to toxicity. Dose-limiting toxicities consisted of grade 4 neutropenia (three) and grade 3 constipation (one). The maximum tolerated dose of continuous infusion troxacitabine in patients with solid tumors is 7.5 mg/m(2) administered over 96 h. This dose level resulted in steady-state drug concentration of at least 20 ng/ml for 72 h. CONCLUSIONS: Administration of troxacitabine by continuous infusion achieved the prospectively defined target plasma concentration. Pharmacokinetics (PK) modeling coupled with real-time PK assessment was an efficient approach to conduct hypothesis-driven phase I trials.

Taxol: a novel investigational antimicrotubule agent.

Microtubules are among the most strategic subcellular targets of anticancer chemotherapeutics. Despite this fact, new antimicrotubule agents that possess unique mechanisms of cytotoxic action and have broader antineoplastic spectra than the vinca alkaloids have not been introduced over the last several decades--until the recent development of taxol. Unlike classical antimicrotubule agents like colchicine and the vinca alkaloids, which induce depolymerization of microtubules, taxol induces tubulin polymerization and forms extremely stable and nonfunctional microtubules. Taxol has demonstrated broad activity in preclinical screening studies, and antineoplastic activity has been observed in several classically refractory tumors. These tumors include cisplatin-resistant ovarian carcinoma in phase II trials and malignant melanoma and non-small cell lung carcinoma in phase I studies. Taxol's structural complexity has hampered the development of feasible processes for synthesis, and its extreme scarcity has limited the use of a conventional, broad-scale screening approach for evaluation of clinical antitumor activity. However, taxol's unique mechanism of action, its spectrum of preclinical antitumor activity, and tumor responses in early clinical trials have generated renewed interest in pursuing its development.

The current status of camptothecin analogues as antitumor agents.

The nuclear enzyme topoisomerase I (topo I) has been recently recognized as the target for the anticancer drug camptothecin (CPT) and its derivatives. Two of the agents that target this enzyme--topotecan (TPT) and CPT-11--appear to be active against a broad range of human tumors. In the following presentation, we review 1) the role of topo I in normal cells, 2) the chemistry and proposed mechanism of action of CPT and its analogues, 3) the results of preclinical and clinical testing of TPT and CPT-11, and 4) mechanisms of resistance to these agents. In normal cells, topo I is thought to be involved in gene transcription and DNA replication. During the course of its normal catalytic cycle, topo I transiently forms a covalent bond with DNA. CPT and its derivatives slow the religation step of the enzyme and stabilize the covalent adduct between topo I and DNA. In S-phase cells, advancing replication forks convert these topo I-DNA adducts into double-strand breaks that appear to be responsible for the cytotoxicity of these agents. Preclinical studies demonstrate antineoplastic activity for TPT and CPT-11 in a variety of tumor models. Phase I studies have identified neutropenia as the dose-limiting toxicity for both drugs. Gastrointestinal effects might also be dose-limiting for CPT-11 administered on some schedules. CPT-11 has shown antitumor activity in phase II trials for patients with carcinomas of lung, cervix, ovary, colon, and rectum and for patients with non-Hodgkin's lymphoma. Phase II studies of TPT are in progress. Resistance to the cytotoxic effects of these agents might result from decreased production of topo I or from production of a mutated form of topo I. In addition, decreased metabolic activation of CPT-11 (which is a pro-drug) and active efflux of TPT by P-glycoprotein-mediated transport might contribute to resistance. As agents with a novel mechanism of action, tolerable toxicity, and encouraging antitumor activity in early clinical trials, TPT and CPT-11 are undergoing further clinical development. If these agents can be successfully combined with other active chemotherapy agents, the topo I-directed agents offer the potential for significant advances in the treatment of patients with a variety of malignancies.

Phase I-phase II trial of N-phosphonacetyl-L-aspartic acid given by intravenous infusion and 5-fluorouracil given by bolus injection.

A phase I clinical trial of N-phosphonacetyl-L-aspartic acid (PALA) and 5-fluorouracil (FUra) was performed on 30 patients. PALA was given as a 15-minute iv infusion once daily for 5 days, and FUra was given as a bolus injection on days 2, 3, 4, and 5. Cycles of treatment were repeated every 3 weeks. Dose-limiting toxicity was manifested by stomatitis and diarrhea. Skin rash was observed also but was not dose limiting. No consistent hematopoietic or renal toxicity was observed. Seventeen patients with disseminated metastatic melanoma and measurable disease were evaluated for response. One partial response was seen; however, the response was associated with significant toxicity, and the treatment could not be repeated. Stable disease was observed in 3 patients with melanoma, 1 patient with colon carcinoma, and 1 patient with ovarian carcinoma. Our findings suggest that the clinical activity of PALA and FUra given according to the above schedule for melanoma is less than 25% (P less than 0.05). Pharmacokinetic studies of FUra revealed no consistent effect of PALA pretreatment on FUra disappearance in plasma. The mean FUra elimination half-line in plasma was 7.11 +/- 0.84 minutes (SEM), which is no different from that reported for FUra alone. The recommended doses on this schedule for phase II studies are 1,000 mg PALA/m2/day iv daily for 5 days and 200 mg FUra/m2/day iv on days 2, 3, 4, and 5.

Cytotoxic effects of topotecan combined with various anticancer agents in human cancer cell lines.

BACKGROUND: Topotecan (TPT) is a topoisomerase I poison that exhibits antineoplastic activity. Analysis of the cytotoxic effects of combinations of TPT and other anticancer agents has been limited. PURPOSE: We assessed the cytotoxic effects produced by combinations of TPT and other antineoplastic agents in experiments involving multiple human cancer cell lines of diverse histologic origins. METHODS: The cytotoxic effects of various antimetabolites (fluorouracil, methotrexate, or cytarabine), antimicrotubule agents (vincristine or paclitaxel [Taxol]), DNA alkylating agents (melphalan, bis[chloroethyl]nitrosourea [BCNU], or 4-hydroperoxycyclophosphamide [4HC]), and a DNA-platinating agent (cisplatin), alone and in combination with TPT, were measured in clonogenic (i.e., colony-forming) assays. HCT8 ileocecal adenocarcinoma, A549 non-small-cell lung carcinoma, NCI-H82ras(H) lung cancer, T98G glioblastoma, and MCF-7 breast cancer cell lines were used in these assays. The data were analyzed by the median effect method, primarily under the assumption that drug mechanisms of action were mutually nonexclusive (i.e., completely independent of one another). For each level of cytotoxicity (ranging from 5% to 95%), a drug combination index (CI) was calculated. A CI less than 1 indicated synergy (i.e., the effect of the combination was greater than that expected from the additive effects of the component agents), a CI equal to 1 indicated additivity, and a CI greater than 1 indicated antagonism (the effect of the combination was less than that expected from the additive effects of the component agents). RESULTS: When the mechanisms of drug action were assumed to be mutually nonexclusive, virtually all CIs for combinations of TPT and either antimetabolites or antimicrotubule agents revealed cytotoxic effects that were less than additive. The CIs calculated at low-to-intermediate levels of cytotoxicity for combinations of TPT and the DNA alkylating agents melphalan, BCNU, and 4HC also showed drug effects that were less than additive; in most cases, however, nearly additive or even synergistic effects were observed with these same drug combinations at high levels of cytotoxicity (i.e., at > or = 90% inhibition of colony formation). Results obtained with combinations of TPT and cisplatin varied according to the cell line examined. With A549 cells, less than additive effects were seen at low-to-intermediate levels of cytotoxicity, and more than additive effects were seen at high levels of cytotoxicity. With NCI-H82ras(H) cells, synergy was observed over most of the cytotoxicity range. CONCLUSIONS AND IMPLICATIONS: TPT cytotoxicity appears to be enhanced more by combination with certain DNA-damaging agents than by combination with antimetabolites or antimicrotubule agents. Interactions between TPT and other drugs can vary depending on the cell type examined. Further investigation is required to determine the basis of the observed effects and to determine whether these in vitro findings are predictive of results obtained in vivo.

Phase I trial of trimetrexate glucuronate on a five-day bolus schedule: clinical pharmacology and pharmacodynamics.

Trimetrexate glucuronate (TMTX), a nonclassical folate antagonist, has been evaluated clinically on several schedules. We have studied TMTX administered as an iv bolus for 5 consecutive days every 3 weeks in 35 patients with advanced solid tumors. Drug was given at doses ranging from 7.6 to 18.8 mg/m2. The maximal tolerated dose was 13.1 mg/m2 per day x 5 for patients without prior myelotoxic treatment and 7.6 mg/m2 per day x 5 for previously treated patients. Because of wide individual differences in drug tolerance, dose escalation in 25% increments is recommended for patients not experiencing toxic effects. The dose-limiting toxicity was neutropenia. Rash and mucositis were also significant. TMTX concentrations were measured 1 and 24 hours after each dose, and the data were fit by use of a one-compartment pharmacokinetic model. With this simplified sampling and modeling scheme, the mean total body clearance (+/- SD) of trimetrexate was 31 +/- 20 mL/min per m2 and the volume of distribution was 13 +/- 7 L/m2. Mean plasma concentrations 1 hour after a dose were 1.12, 2.43, 3.33, and 3.25 mumol/L at 7.6, 9.1, 10.9, and 13.1 mg/m2, respectively. The mean TMTX concentration (+/- SD) 24 hours after a dose was 114 +/- 87 nmol/L. The mean area under the concentration-versus-time curve at 13.1 mg/m2 was 2,266 mumol.min/L. The drug concentration 1 hour after the first dose and the area under the concentration-versus-time curve were highly correlated with leukopenia and thrombocytopenia (r = .6 and .65 and P = .0007 and .0001, respectively). The maximal tolerated dose on the daily x 5 schedule was 30% of the dose tolerated on an iv bolus schedule. The choice of drug schedule for clinical trials when murine and human pharmacokinetics differ is discussed. Phase II trials are under way with both the iv bolus and the daily x 5 schedules.

Clinical and pharmacologic reappraisal of dichloromethotrexate.

Dichloromethotrexate (DCMTX) has been the subject of sporadic clinical development for the last 30 years. Although DCMTX was developed in hopes of discovering a more potent antifolate, the potential pharmacologic and toxicologic advantages of the analog have become of greater interest. This phase I and pharmacokinetic trial of DCMTX given on days 1, 8, and 15 every 28 days was undertaken to test these potential advantages. The maximally tolerated dose on this schedule was 980 mg/m2. Hepatic toxicity was dose limiting. Malaise, myelosuppression, and mucositis were also major toxic effects. The recommended dose for subsequent phase II studies of DCMTX administered on this schedule is 785 mg/m2 with a reduction to 625 mg/m2 for patients with a poor performance status or extensive prior therapy. Plasma disappearance curves for most patients were biphasic or triphasic, although several demonstrated more complex kinetic patterns that suggested significant enterohepatic circulation. The magnitude of the area under the plasma disappearance curve was related to the severity of DCMTX-induced hepatotoxicity. The elimination kinetics were linear, with a mean plasma clearance of 294 mL/min (range, 128-715). The pharmacokinetic behavior of DCMTX does not support its use over methotrexate in regional perfusion. DCMTX's primarily nonrenal elimination suggests that it may have an advantage over methotrexate when combined with nephrotoxic drugs such as cisplatin. However, there is little reason to commit major resources to further evaluation of DCMTX unless significant advantages in antineoplastic activity are identified.

Sixteen-week dose-intense chemotherapy in the adjuvant treatment of breast cancer.

Fifty-three women with breast cancer were treated with a new 16-week dose-intense, chemotherapy regimen. Patients with operable breast cancer with 10 or more histologically positive axillary nodes were treated with this five-drug regimen that incorporated the concepts of weekly chemotherapy, sequential administration of antimetabolites, and continuous infusion of fluorouracil (5-FU). The chemotherapy regimen consisted of eight cycles (each of 2 wk duration) of 100 mg of cyclophosphamide/m2 orally on days 1-7, 40 mg of doxorubicin/m2 intravenous (IV) on day 1, 100 mg of methotrexate/m2 IV on day 1 with 10 mg of leucovorin rescue/m2 every 6 hours for six oral doses on day 2, 1 mg of vincristine IV on day 1, and 600 mg of 5-FU/m2 IV at hour 20 over 2 hours. A continuous infusion of 300 mg of 5-FU/m2 per day was given IV on days 8-9 of each 2-week cycle. The doses and schedule of drug administration were designed to minimize dosage reduction and treatment delay. At a median follow-up of 17 months, there have been eight relapses in the 53 patients. The actuarial 3-year disease-free survival is 80% (95% confidence interval, 62% to 90%). The major side effects were attributable to myelosuppression. Absolute neutrophil counts less than 250/microL were noted in 12 (23%) patients; seven patients (13%) required hospitalization for management of neutropenic fever. No treatment-related deaths occurred. Ninety-four percent of the planned doses were administered, and only 5% of the courses were delayed because of toxic reactions. The encouraging therapeutic data, manageable side effects, and our ability to deliver over 90% of the planned doses provide the rationale for a phase III comparison of this new dose-intense regimen and standard chemotherapy in patients with operable disease and positive axillary nodes.

Effects of the differentiating agent hexamethylene bisacetamide on normal and myelodysplastic hematopoietic progenitors.

Hexamethylene bisacetamide (HMBA; NSC 95580) is a potent polar-planar differentiating agent of leukemia and solid tumor cell lines in vitro at clinically achievable concentrations. HMBA is currently being studied in patients with myelodysplastic syndrome. Previous phase I trials have demonstrated that HMBA produces hematologic toxicity in morphologically normal bone marrows of patients with solid tumors. Because of concern that HMBA may produce more severe myelotoxicity in patients with myelodysplastic syndrome since these patients have limited hematopoietic reserves, we studied the effects of HMBA on myelodysplastic and normal hematopoietic progenitors in vitro. HMBA concentrations that are optimal for differentiation in vitro (2 to 5 mmol/L) and HMBA concentrations that are being achieved in clinical trials (1 to 2 mmol/L) inhibited the growth of granulocyte-macrophage colony-forming units and erythroid burst-forming units from all 15 patients with myelodysplastic syndrome and all 4 normal subjects, HMBA did not induce proliferation of myelodysplastic or normal progenitors at any concentration; rather, it produced nearly identical inhibition of normal and myelodysplastic hematopoietic progenitors. HMBA also produced quantitatively similar inhibition of clonogenic leukemic growth of two myeloid leukemia cell lines. For a differentiating agent to be effective, it will likely have to either produce both differentiation and proliferation of abnormal hematopoietic progenitors or show selective inhibitory effects on abnormal as compared with normal progenitors. Although the mechanisms responsible for the antiproliferative effects of HMBA cannot be determined from this study, similar inhibitory effects of HMBA on normal and abnormal hematopoietic progenitors suggest that HMBA may be of limited utility in producing and sustaining elevations of peripheral blood cell counts in patients with myelodysplastic syndrome.

Clinical pharmacology of oral and i.v. N-methylformamide: a pharmacologic basis for lack of clinical antineoplastic activity.

N-Methylformamide (NMF) has been an agent of considerable interest to oncologists because of its broad spectrum of preclinical antitumor activity, tumor-differentiating abilities, and radiosensitizing and chemosensitizing properties. In this report, the pharmacokinetics of NMF are described, based on data from two phase I studies exploring both iv and oral routes of administration. Mean peak NMF plasma concentrations at recommended phase II doses were 0.46 mmol/L for NMF administered orally, 600 mg/m2 three times/week X 4 weeks every 6 weeks, and 2.78 mmol/L for NMF administered as a weekly iv bolus at 2,000 mg/m2 X 3 weeks every 4 weeks. These NMF concentrations were significantly lower than the concentrations that have been demonstrated to induce antineoplastic and relevant biologic effects in preclinical studies. Plasma disappearance curves were biphasic in the majority of patients; however, 25% of the curves were best fit by a monoexponential kinetic model. Mean alpha half-life and beta half-life values (+/- SE) were 10 +/- 2 and 732 +/- 93 min, respectively. Volumes of distribution for the theoretical central compartment (Vc) and at steady-state (Vss) were 13.8 +/- 1.1 L/m2 and 18.7 +/- 1.1 L/m2, respectively. The mean plasma clearance of NMF was 19.1 +/- 2.1 mL/min per square meter, and the relative contributions to parent compound disposition by respiratory and renal routes were insignificant. No metabolites were identified. Gastrointestinal absorption of oral NMF was rapid and nearly complete; oral bioavailability was calculated to be 0.87. Pharmacodynamic associations were observed between the magnitude of the area under the plasma disappearance curves and hepatotoxicity, the dose-limiting toxic effect of iv NMF, and the symptom complex of nausea, vomiting, and malaise, which precluded dose escalation of oral NMF.

Phase I and pharmacologic studies of topotecan in patients with impaired hepatic function.

BACKGROUND: Topotecan, a topoisomerase I inhibitor that has demonstrated anticancer activity toward leukemias and solid tumors in clinical trials, is eliminated via hepatic and renal routes. However, dosing guidelines for the administration of topotecan to patients with impaired hepatic function have not yet been established. PURPOSE: We compared the maximum tolerated doses (MTDs), the toxic effects, and the pharmacokinetics and pharmacodynamics of topotecan in patients who had refractory, malignant, solid tumors and who either had or lacked hepatic injury. The potential role of three substrate markers of liver function (indocyanin green [ICG]-- a marker of hepatic blood flow; lorazepam--a substrate marker of hepatic glucuronidation; and antipyrine--a substrate marker for cytochrome P450 activity) in optimizing topotecan doses for patients with liver injury was also evaluated. METHODS: Twenty-one cancer patients, 14 of whom had hepatic injury due to metastatic disease, biliary obstruction, or cirrhosis, were treated with intravenously delivered courses of topotecan consisting of 0.5, 1.0, or 1.5 mg/m2 of drug per day for 5 days. Most patients received more than one course of treatment, with new courses initiated at 3-week intervals. Patient responses (evaluated by tumor measurements) and treatment-induced toxic effects were assessed. Prior to the initiation of topotecan treatment, patients were given intravenous injections of ICG, lorazepam, and antipyrine to determine the plasma pharmacokinetics of these compounds. The pharmacokinetics of topotecan (both the lactone and the carboxylate forms) were determined by analysis of plasma and urine samples collected on the first day of the first course of drug treatment. Scatter plots of area under the plasma concentration versus time curves in relation to percent decreases in either absolute neutrophil count or platelet count were used to explore the pharmacodynamics of topotecan. The Student's t test and the Mann-Whitney U test were used to compare pharmacokinetic parameters between patients with and without abnormal hepatic function. Correlations were assessed using the Spearman's rank correlation coefficient (rs). Reported P values are based on two-tailed tests of significance. RESULTS: Patients with hepatic injury tolerated topotecan doses up to 1.5 mg/m2, i.e., the MTD of this drug established in previous studies. The nature and severity of treatment-induced toxic effects and the pharmacokinetics of topotecan were similar in patients with and without liver injury. No differences were observed in the urinary excretion of topotecan between the two patient groups. Clearances of total topotecan and its lactone species correlated only with clearance of ICG (rs = .64, P = .004; and rs = .68, P = .0017, respectively). The pharmacodynamic effects of topotecan were not altered by liver dysfunction. CONCLUSIONS AND IMPLICATIONS: Cancer patients with hepatic injury can be treated with topotecan at a starting dose of 1.5 mg/m2, given daily for 5 days and administered every 3 weeks. Topotecan dose modifications do not appear to be required for patients with hepatic dysfunction and normal renal function.

Microtubule changes and cytotoxicity in leukemic cell lines treated with taxol.

Taxol, a diterpenoid plant product that enhances the polymerization of tubulin, is currently entering clinical trials in the treatment of human leukemia. In order to develop an in vitro assay to predict tumor sensitivity to taxol, human leukemic cell lines were exposed to clinically achievable concentrations of taxol for relevant exposure periods. Changes in microtubules visualized by indirect immunofluorescence were compared to drug sensitivity measured by a clonogenic assay. Taxol produced either multiple mitotic asters in G2/M or microtubule bundling throughout the cell cycle. In cells that were relatively resistant to taxol, microtubule bundling was reversible while microtubule bundling in relatively sensitive cells persisted in the presence or absence of taxol. In contrast, aster formation was unrelated to cytotoxicity in any cell line. In the future, these microtubule effects may be useful in predicting the chemotherapeutic efficacy of taxol.

Development of polyploidization in taxol-resistant human leukemia cells in vitro.

The effects of taxol on antitubulin immunofluorescent staining patterns, cellular DNA content, and labeling with [3H]thymidine were measured for the taxol-sensitive HL60 and taxol-resistant K562 cell lines after exposures for 0, 4, 12, and 24 h. Taxol caused a relative increase in the fraction of 4C interphase and metaphase cells in both lines although the 4C interphase accumulation was greater for the resistant K562 line. Of the cells with S-phase DNA content, taxol-treated HL60 cells were less likely to incorporate [3H]thymidine than taxol-treated K562 cells. However, a decrease in percentage of S-phase labeling for both lines relative to control cells was seen. Finally, taxol induced the development of polyploid cells (cells with DNA contents greater than that of the 4C G2-M peak) in the relatively taxol-resistant K562 cells, an effect not seen in the relatively taxol-sensitive HL60 line. After 24 h of taxol exposure 70% of all K562 cells were polyploid while only 8% of the HL60 cells were polyploid. The capacity of K562 cells to generate polyploidy in response to taxol correlated with taxol resistance by previous assay and may be a useful indicator of drug resistance.

Direct relationship of marrow cell growth and 1-beta-D-arabinofuranosylcytosine metabolism.

To define the relationship between perturbed cell growth and intracellular metabolism of 1-beta-D-arabinofuranosylcytosine (ara-C) in sensitive human cells, growth kinetic, and biochemical pharmacological determinants were examined in normal human bone marrow populations in vitro in normal serum and in the presence of drug-induced humoral stimulatory activity (HSA). Cells cultured in HSA demonstrated both increased proliferation and greater ara-C-related inhibition of DNA synthesis than did cells maintained in normal serum, as measured by [3H]thymidine incorporation into DNA and [3H]thymidine granulocyte precursor labeling index. Parallel measurements of [3H]ara-C incorporation into DNA demonstrated similar behavior in HSA-perturbed cells. When these cultured cells were exposed to 1 and 10 microM ara-C, intracellular formation of 1-beta-D-arabinofuranosylcytosine 5'-triphosphate over 3 hr and retention of this active form during 1 subsequent hr in drug-free medium were both increased in HSA-stimulated cells relative to cells cultured in normal serum. These studies demonstrate coupling of induced cell growth kinetics with enhanced intracellular metabolism of the S-phase-specific antimetabolite ara-C in normal human marrow cells. The close direct relationship between growth kinetic perturbation and augmentation of intracellular ara-C activation in this normal hematopoietic model provides a basis for comparison with leukemic cell populations, in which uncoupling of growth kinetics and pharmacokinetics may signify divergence from normal drug-sensitive cell behavior and, thus, resistance to ara-C cytotoxicity.

Effect of P-glycoprotein expression on the accumulation and cytotoxicity of topotecan (SK&F 104864), a new camptothecin analogue.

Topotecan (TPT, 9-dimethylaminomethyl-10-hydroxycamptothecin) is the first topoisomerase I-directed cytotoxic agent to enter clinical trials in the United States in two decades. The effect of P-glycoprotein (Pgp) overexpression on TPT cytotoxicity was examined in CHRC5 (colchicine-resistant) and AuxB1 (parental) Chinese hamster ovary cells. Examination of the IC50 values observed in colony-forming assays revealed that the CHRC5 cells were 15-fold (SD, +/- 3; n = 3) resistant to TPT after a 1-h exposure and 3.2-fold (SD, +/- 1.4; n = 4) resistant in continuous exposure experiments. Band depletion immunoblotting revealed that 4-fold higher concentrations of extracellular TPT were required to induce the formation of topo I-DNA complexes in CHRC5 cells as compared to AuxB1 cells. To assess the role of Pgp in this resistance, drug accumulation and cytotoxicity assays were repeated in the absence and presence of quinidine. Addition of quinidine enhanced TPT accumulation (measured by high-performance liquid chromatography) and diminished the IC50 for TPT to a greater extent in CHRC5 cells than in AuxB1 cells. To examine whether similar effects could be detected in Pgp-expressing human cells, MCF-7/Adriar breast cancer cells and KG1a human acute myelogenous leukemia cells were examined. Quinidine or verapamil enhanced TPT accumulation in both of these cell lines but had no effect in parental MCF-7 cells or a variety of human leukemia cell lines that do not overexpress Pgp. Cytotoxicity measurements performed by counting the number of surviving cells (MCF-7/Adriar) or employing a modified, highly stable tetrazolium dye reduction assay (leukemia cell lines) revealed that quinidine diminished the IC50 for TPT in the Pgp-overexpressing cell lines but not in the control lines. These results suggest that Pgp overexpression diminishes TPT accumulation and TPT cytotoxicity in hamster and human cells. It should be stressed, however, that these effects were substantially smaller than the effects of Pgp overexpression on the accumulation and cytotoxicity of the anthracycline daunorubicin and the epipodophyllotoxin etoposide in the same cell lines.

Phase I and pharmacokinetic study of hepsulfam (NSC 329680).

Hepsulfam (NSC 329680), a bifunctional alkylating agent structurally related to busulfan, has entered clinical trial based on its broader preclinical antitumor activity compared with that of busulfan and its i.v. formulation which may circumvent the many problems arising from the p.o. administration of busulfan, such as significant individual differences in bioavailability. In this Phase I study, 53 patients received 95 courses of hepsulfam at doses ranging from 30 to 480 mg/m2 administered i.v. over 30 min every 28 days. Hematological toxicity was dose limiting. Leukopenia and thrombocytopenia were dose related, delayed in onset, and sustained for long durations. Toxicity was cumulative in most patients receiving more than one course. This pattern of myelosuppression suggests that hepsulfam is cytotoxic to hematopoietic stem cells. Although hematological toxicity was not particularly severe during most courses, its lengthly duration precluded the prompt administration of subsequent courses. Minimal nonhematological effects were observed. Pharmacokinetic studies revealed that the clearance rate of hepsulfam is linear over the dose range studied and that its plasma disposition is biphasic with mean alpha and beta half-lives of 19 +/- 18 (SE) min and 337 +/- 248 (SE) min, respectively. The area under the plasma clearance curve correlated with the percentage of change in WBC using a sigmoidal Emax model and with the duration of thrombocytopenia in patients with hematological toxicity. Based on the protracted duration of the toxicity of multiple doses that were greater than 210 mg/m2, the recommended starting dose for Phase II trials is 210 mg/m2. However, these trials should be pursued with caution because of the protracted nature of hepsulfam's myelosuppression. Because hepsulfam produced minimal nonhematological toxicity, substantial dose escalation above 480 mg/m2 may be possible with hematopoietic stem cell support.

Phase I and pharmacological study of the novel topoisomerase I inhibitor 7-ethyl-10-[4-(1-piperidino)-1-piperidino]carbonyloxycamptothecin (CPT-11) administered as a ninety-minute infusion every 3 weeks.

7-Ethyl-10-[4-(1-piperidino)-1-piperidino]carbonyloxycamptothecin (CPT-11; Irinotecan), a semisynthetic analogue of camptothecin (CPT) with broad preclinical antitumor activity, has demonstrated impressive activity in phase II trials in Japan in advanced small and non-small cell lung, colorectal, cervical, and ovarian carcinomas, as well as in refractory lymphomas and leukemias. In this phase I and pharmacological study, 90-min infusions of CPT-11 were administered every 3 weeks at doses ranging from 100 to 345 mg/m2 to patients with solid malignancies. Acute, severe, and refractory vomiting, diarrhea, and/or abdominal cramps associated with flushing, warmth, and diaphoresis occurred in the immediate posttreatment period at the 240-mg/m2 dose level in several patients who were not treated with premedications. The characteristics and temporal nature of these toxicities, the prompt resolution of symptoms following treatment with diphenhydramine, and the successful use of a premedication regimen consisting of ondansetron and diphenhydramine in preventing these acute effects suggest that vasoactive substances are involved in the mediation of these acute toxicities. With the routine use of these premedications, there was no single toxicity type that limited the escalation of CPT-11 doses. Instead, a constellation of severe hematological and gastrointestinal effects precluded the repetitive administration of CPT-11 at doses above 240 mg/m2, the maximum tolerated dose and recommended phase II dose on this schedule. Major responses were observed in patients with advanced colorectal, cervical, and renal cancers. The disposition of total CPT-11 in plasma was fit by a biexponential kinetic model with renal elimination accounting for 37 +/- 4% (SE) of total drug disposition. The Cmax for the active metabolite of CPT-11, 7-ethyl-10-hydroxycamptothecin (SN-38), was achieved at 2.2 +/- 0.1 h after treatment, and mean residence times for both CPT-11 and SN-38 were long, 9.1 and 10.0 h, respectively. Compared with topotecan, another CPT analogue under development, a larger proportion of total drug exposure was accounted for by the active lactone (closed-ring) forms of CPT-11 and SN-38; areas under the time-versus concentration curve for their respective lactone were 44 and 50% of areas under the time-versus-concentration curve for total CPT-11 and SN-38. Although intermittent dosing schedules appear to be superior to single dosing schedules for CPT and some CPT analogues in preclinical tumor models, the maintenance of biologically relevant concentrations of SN-38 for relatively long durations may negate the potential pharmacological benefits of intermittent and continuous administration schedules for CPT-11.(ABSTRACT TRUNCATED AT 400 WORDS)

Phase I and pharmacological study of the pulmonary cytotoxin 4-ipomeanol on a single dose schedule in lung cancer patients: hepatotoxicity is dose limiting in humans.

4-Ipomeanol (IPO), a naturally occurring pulmonary toxin, is the first cytotoxic agent to undergo clinical development based on a biochemical-biological rationale as an antineoplastic agent targeted specifically against lung cancer. This rationale is based on preclinical observations that metabolic activation and intracellular binding of IPO, as well as cytotoxicity, occurred selectively in tissues and cancers derived from tissues that are rich in specific P450 mixed function oxidase enzymes. Although tissues capable of activating IPO to cytotoxic intermediates in vitro include liver, lung, and kidney, IPO has been demonstrated in rodents and dogs to undergo in situ activation, bind covalently, and induce cytotoxicity preferentially in lung tissue at doses not similarly affecting liver or kidneys. Although the drug was devoid of antitumor activity in the conventional murine preclinical screening models, cytotoxic activity was observed in human lung cancers in vitro and in human lung cancer xenografts in vivo, adding to the rationale for clinical development. Somewhat unexpectantly, hepatocellular toxicity was the dose-limiting principal toxicity of IPO administered as a 30-min infusion every 3 weeks to patients with lung cancer. In this study, 55 patients received 254 courses at doses almost spanning 3 orders of magnitude, 6.5 to 1612 mg/m2. Transient and isolated elevations in hepatocellular enzymes, predominantly alanine aminotransferase, occurred in the majority of courses of IPO at 1032 mg/m2, which is the recommended IPO dose for subsequent phase II trials. At higher doses, hepatocellular toxicity was more severe and was often associated with right upper quadrant pain and severe malaise. Toxic effects were also noted in other tissues capable of activating IPO, including possible nephrotoxicity in a patient treated with one course of IPO at 154 mg/m2 and severe, reversible pulmonary toxicity in another patient who received nine courses of IPO at doses ranging from 202 to 826 mg/m2. Although individual plasma drug disposition curves were well described by a two-compartment first order elimination model, The relationship between IPO dose and area under the disposition curve was curvilinear, suggesting saturable elimination kinetics. At the maximum tolerated dose, the mean half-lives (lambda 1 and lambda 2) were 6.7 and 114.5 min, respectively. Renal excretion of parent compound accounted for less than 2% of the administered dose of IPO. An unidentified metabolite was detected in the plasma of patients treated at higher doses. No objective antitumor responses were observed; however, stable disease persisted for at least eight courses in 27% of patients.(ABSTRACT TRUNCATED AT 400 WORDS)