This Article Abstract Full Text (PDF) Purchase Article View Shopping Cart Alert me when this article is cited Alert me if a correction is posted Services Email this article to a colleague Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Save to my personal folders Download to citation manager Rights & Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Tolcher, A. W. Articles by Rowinsky, E. K. Search for Related Content PubMed PubMed Citation Articles by Tolcher, A. W. Articles by Rowinsky, E. K. Social Bookmarking                            What's this?

Phase I and Pharmacokinetic Study of NSC 655649, a Rebeccamycin Analog With Topoisomerase Inhibitory Properties

From the Institute for Drug Development, Cancer Therapy and Research Center; Department of Pharmacology and Department of Medicine, Division of Medical Oncology, University of Texas Health Science Center at San Antonio; Brooke Army Medical Center; and Audie Murphy Veterans Administration Hospital, San Antonio, TX.

Address reprint requests to Anthony W. Tolcher, MD, FRCPC, Institute for Drug Development, Cancer Therapy and Research Center, 8122 Datapoint Dr, Ste 250, San Antonio, TX 78229; email: atolcher{at}saci.org

	ABSTRACT

TOP ABSTRACT INTRODUCTION PATIENTS AND METHODS RESULTS DISCUSSION REFERENCES

 
PURPOSE: To assess the feasibility of administering NSC 655649, a water-soluble, rebeccamycin analog with topoisomerase inhibitory properties, as a brief intravenous (IV) infusion once every 3 weeks and to determine the maximum-tolerated dose (MTD) of NSC 655649, characterize its pharmacokinetic behavior, and seek preliminary evidence of antitumor activity.

PATIENTS AND METHODS: Patients with advanced solid malignancies were treated with escalating doses of NSC 655649 administered over 30 to 60 minutes IV once every 3 weeks. An accelerated dose-escalation method was used to guide dose escalation. After three patients were treated at the first dose level, doses were escalated in increments that ranged up to 150% using single patient cohorts until moderate toxicity was observed, when a more conservative dose-escalation scheme was invoked. MTD was defined as the highest dose level at which the incidence of dose-limiting toxicity did not exceed 20%. MTD was determined for both minimally pretreated (MP) and heavily pretreated (HP) patients. Plasma and urine were sampled to characterize the pharmacokinetic and excretory behavior of NSC 655649.

RESULTS: Forty-five patients were treated with 130 courses of NSC 655649 at doses ranging from 20 mg/m² to 744 mg/m². Myelosuppression was the principal toxicity. Severe neutropenia, which was often associated with thrombocytopenia, was unacceptably high in HP and MP patients treated at 572 mg/m² and 744 mg/m², respectively. Nausea, vomiting, and diarrhea were common but rarely severe. The pharmacokinetics of NSC 655649 were dose dependent and fit a three-compartment model. The clearance and terminal elimination half-lives for NSC 655649 averaged 7.57 (SD = 4.2) L/h/m² and 48.85 (SD = 23.65) hours, respectively. Despite a heterogeneous population of MP and HP patients, the magnitude of drug exposure correlated well with the severity of myelosuppression. Antitumor activity was observed in two HP ovarian cancer patients and one patient with a soft tissue sarcoma refractory to etoposide and doxorubicin.

CONCLUSION: Recommended phase II doses are 500 mg/m² and 572 mg/m² IV once every 3 weeks for HP and MP patients, respectively. The absence of severe nonhematologic toxicities, the encouraging antitumor activity in HP patients, and the unique mechanism of antineoplastic activity of NSC 655649 warrant further clinical development.

	INTRODUCTION

TOP ABSTRACT INTRODUCTION PATIENTS AND METHODS RESULTS DISCUSSION REFERENCES

 
REBECCAMYCIN, AN N-glycoside fermentation product originally isolated from the actinomycete strain Saccharothrix aerocoligenes, demonstrated impressive cytotoxicity against a broad range of human cancers in vitro, including A549 lung, HCT-116 colon, and KB nasopharyngeal carcinomas.^1-3 However, the poor water solubility and pharmaceutical characteristics of rebeccamycin precluded its candidacy for clinical development. Water-soluble rebeccamycin analogs were synthesized and evaluated as potential lead compounds.^2,4 The glycosyl-dichloro-indolocarbazole rebeccamycin analog NSC 655649 (Fig 1) was selected for further clinical development because of its favorable pharmaceutical characteristics, water solubility, and striking antitumor activity against a broad range of experimental cancers.^5,6

View larger version (16K): [in this window] [in a new window]   Fig 1. Structures of (A) NSC 655649 and (B) rebeccamycin.

The toxicologic and pharmacologic profiles of NSC 655649 have been evaluated in mice, rats, and dogs.⁵ Rapidly proliferating hematopoietic, gastrointestinal, lymphoid, and reproductive organs were the most sensitive tissues to the toxic effects of NSC 655649. In both rodents and dogs, myelosuppression was the principal dose-limiting toxicity (DLT). NSC 655649 and a major metabolite were both detected in the plasma and urine.⁵ In mice treated with a single intravenous (IV) dose of NSC 655649, plasma clearance (Cl) was biphasic, with terminal half-life (t_1/2) values ranging from 128 to 409 minutes. In dogs treated with a single IV dose, a rapid distribution phase preceded a much slower elimination phase, with t_1/2 values ranging from 4.6 to 6.4 hours.⁵ Furthermore, drug exposure increased disproportionately as the dose of NSC 655649 was increased, suggesting nonlinear pharmacokinetics. In mice, plasma protein binding was negligible, but the volume of distribution was large. The urinary excretion of NSC 655649 and a major unidentified metabolite together accounted for less than 5% of the administered dose.⁵

The impetus for pursuing the clinical development of NSC 655649 included its structural uniqueness and novel mechanism of action and its broad spectrum of antitumor activity, particularly against multidrug-resistant tumors. The principal objectives of this phase I and pharmacokinetic study were to (1) determine the maximum-tolerated dose (MTD) of NSC 655649 administered as a brief IV infusion every 3 weeks, (2) determine the toxicities of NSC 655649 on this schedule, (3) characterize the pharmacokinetic behavior of NSC 655649, and (4) seek preliminarily evidence of antitumor activity in patients with advanced solid malignancies.

	PATIENTS AND METHODS

TOP ABSTRACT INTRODUCTION PATIENTS AND METHODS RESULTS DISCUSSION REFERENCES

 
Patient Selection
Patients with histologically or cytologically confirmed solid malignancies refractory to standard therapy or for whom no standard therapy existed were eligible. Patient entry criteria also included age 18 years old; life-expectancy of at least 12 weeks; a Southwest Oncology Group performance status of 0 to 2; no prior chemotherapy within 4 weeks (6 weeks for prior mitomycin C or a nitrosourea); adequate hematopoietic (hemoglobin 9 g/dL, absolute neutrophil count [ANC] 1,500/µL, platelet count 100,000/µL), hepatic (bilirubin < 1.5 mg/dL, AST, ALT, and alkaline phosphatase two times the upper limit of normal, or < five times the institutional upper limit of normal if the elevation was caused by hepatic metastases), and renal (serum creatinine 1.5 mg/dL) functions; measurable or assessable disease; no evidence of brain metastases; and no coexisting medical problem of sufficient severity to limit compliance with the study.

Drug Administration
The starting dose of NSC 655649, 20 mg/m² administered as a 30-minute IV infusion every 3 weeks represented one third of the toxic-dose-low in dogs. All toxicities were graded according to the National Cancer Institute’s common toxicity criteria, version 1. The MTD was defined as the highest dose at which fewer than two of six new patients experienced treatment-related DLT, defined as any grade 3 or higher nonhematologic toxicity (including grade 3 nausea or vomiting despite optimal antiemetics), grade 4 thrombocytopenia (< 25,000/µL), and grade 4 neutropenia (ANC < 500/µL) lasting for at least 4 days or accompanied by fever. An accelerated dose-escalation method was used to guide dose escalation in cohorts of new patients. This was, in part, guided by safety data obtained from another phase I study of NSC 655649 staggered ahead of this trial performed at the University of Wisconsin.¹⁰ Three patients were treated at the starting dose level. The dose of NSC 655649 was increased using single patient cohorts until moderate toxicity was observed. Additional patients were then treated, and a more conservative modified Fibonacci dose-escalation scheme was invoked. If one of three patients experienced DLT, the cohort was expanded to six patients. Because it was projected that the extent of prior therapy was likely to be a determinant of the principal hematological DLT of NSC 655649, patients were classified as either heavily pretreated (HP) or minimally pretreated (MP), and the MTD was defined separately for each group. HP patients were defined as those patients who had previously been treated with more than six courses of an alkylating agent–containing chemotherapy regimen, more than two courses of carboplatin or mitomycin C, any prior nitrosourea-containing regimen, irradiation to 25% of the bone marrow–containing areas, high-dose chemotherapy requiring hematopoietic stem-cell reinfusion, or widespread metastases to bone.

NSC 655649 was supplied by the National Cancer Institute in 20-mL vials containing 200 mg of NSC 655649, 22.4 mg/mL of tartaric acid, and sterile water (US Pharmacopoeia), for injection. The appropriate dose of the drug was then further diluted in 50 to 100 mL 0.9% saline solution, USP, for total doses < 400 mg, or 250 to 500 mL 0.9% saline solution, USP, for doses to 400 mg. At dose levels 247 mg/m², NSC 655649 was infused IV over 30 minutes, whereas at dose levels of 400 mg/m², NSC 655649 was infused over 60 minutes.

Pretreatment and Follow-Up Studies
Complete medical histories, physical examinations, concurrent medication profiles, assessments of performance status, and routine laboratory studies were performed pretreatment and weekly. Routine laboratory studies included a complete blood count, a differential WBC count, prothrombin and partial thromboplastin times, electrolytes, blood urea nitrogen, serum creatinine, uric acid, glucose, alkaline phosphatase, lactate dehydrogenase, ALT, AST, total bilirubin, calcium, total protein, albumin, cholesterol, and triglycerides. A urinalysis and a serum pregnancy test (as appropriate) were performed before treatment. Pretreatment studies also included an ECG, relevant radiologic studies to evaluate all measurable and assessable sites of disease, and an assessment of relevant tumor markers. Radiologic evaluations for disease status assessment were repeated after every other course. Patients were able to continue treatment if they did not develop progressive disease or experience intolerable toxicity. A complete response was scored if there was disappearance of all measurable and assessable disease for at least two measurements performed at least 4 weeks apart without worsening of disease-related symptomatology or declining performance status. A partial response required at least a 50% reduction in the sum of the product of the bidimensional measurements of all lesions documented by at least two measurements separated by at least 4 weeks. Any concurrent increase in the size of any lesion by 25% or the appearance of any new lesion was considered disease progression.

Plasma and Urine Pharmacokinetic Sampling and Assay
Blood samples were collected into heparinized tubes via an indwelling venous catheter placed in the contralateral arm before the infusion and immediately before the end of infusion. Samples were also collected at 15 and 30 minutes and 1, 1.5, 2, 3, 4, 6, 8, 12, 24, 48, 72, and 168 hours after the end of the infusion. The blood samples were centrifuged at 3,000 rpm for 15 minutes immediately after collection. Next the plasma was transferred to separate tubes and frozen to -20°C until assayed. Urine was collected continuously from 0 to 12 hours, and from 12 to 24 hours. The urine collections were mixed, and 2-mL aliquots were frozen at -20°C in labeled sample tubes.

The analytic standards for NSC 655649 and rebeccamycin (NSC 359079) were obtained from the Pharmaceutical Management Branch, National Cancer Institute (Bethesda, MD). All glassware used in the extraction and high-performance liquid chromatography (HPLC) analysis was siliconized. To each 200 µL of plasma, 20 µL (100 ng) of internal standard was added, followed by 40 µL of 1N NaOH. After extraction with methyl t-butyl ether, the solvent was removed from the combined organic layers, and the residue was resuspended in 200 µL of 40% acetonitrite containing 0.1M sodium acetate pH 4.0 before injection onto the HPLC.

The HPLC system included a Waters 590 isocratic solvent pump, a Waters 712 autosampler, and a Waters model 486 ultraviolet light detector (Waters Corp, Milford, MA) set at 318 nm. Chromatographic data were collected and stored using Waters Maxima chromatography data collection software (Waters Corp, Milford, MA). The mobile phase consisted of 35% acetonitrile containing 5 mmol/L ammonium phosphate (pH 4.0) and 1% triethylamine. For the plasma extracts, 75 µL were injected onto a 3.9 x 150 mm C₁₈ Novapak column (equipped with a C₁₈ precolumn) maintained at 35°C with a Waters column temperature control system (Waters Corp). With a mobile phase flow rate of 1.2 mL/min, the NSC 655649 eluted at 4 minutes and the internal standard at 10 minutes. The lower limit of quantification for NSC 655649 freebase in plasma was 0.016 µg/mL. Standard curves were constructed by plotting the ratio of NSC 655649 peak areas to those of the internal standard versus known plasma concentrations. Plasma concentrations of NSC 655649 (freebase) were determined from linear least-squares regression equations derived from calibrations curves prepared from known standard samples. The HPLC assay was validated at NSC 655649 concentration between 0.016 to 24.5 µg/mL for plasma and 0.64 to 81.6 µg/mL for urine. Calibration curves for both plasma and urine NSC 655649 were linear (R² = 0.99) over the respective concentration ranges. The interassay coefficient of variation for the standard curves was 8.6%.

Pharmacokinetic and Pharmacodynamic Analyses
Individual NSC 655649 plasma concentration data sets from days 1 to 7 were analyzed by both noncompartmental methods and compartmental methods using Win Nonlin, version 1.5, (Scientific Consulting, Inc, Mountain View, CA). For the noncompartmental parameters, peak concentrations were determined by inspection of each patient’s plasma concentration-time curve. Elimination rate constants were estimated by linear regression of the last three data points on the terminal log-linear portion of the concentration-time curves. t_1/2 were calculated by dividing 0.693 by the elimination rate constants. The area under curve (AUC) was calculated using the linear trapezoidal rule up to the last measurable data point (for AUC_0-t), then extrapolated to infinity (AUC_0-). The systemic Cl was determined by dividing the dose (in mg freebase NSC 655649 per m²) by the AUC. The apparent volume of distribution at steady-state (Vd_ss) was determined by the following relationships: Vd_ss = (dose x AUMC/AUC²) - (dose x duration of infusion)/(2 x AUC), where AUMC is the area under the moment curve extrapolated to infinity. The mean residence time was calculated by dividing Vd_ss by the systemic Cl. The fraction of NSC 655649 excreted in the urine over 24 hours was calculated by dividing the total amount of NSC 655649 excreted in the urine over a 24-hour period by the total dose administered.

NSC 655649 plasma concentration data at the 500 and 572 mg/m² dose levels were also analyzed using model-dependent methods. After visual inspection of plasma concentration-time curves, individual data sets were fit with either two- or three-compartment models using nonlinear least-squares regression. The pharmacokinetic analysis was performed using a weighting factor of 1/yr.² The goodness of fit of each model was assessed by inspecting the weighted sum of squares, dispersion of the residuals, standard errors of the fitted pharmacokinetic parameters, and the Akaike information criterion.¹¹

The relationships between NSC 655649 pharmacokinetic parameters reflecting drug exposure (eg, dose, AUC, and maximum concentration [C_max]) and indices reflecting myelosuppression in the first course were explored. Relevant parameters of myelosuppression that were evaluated included ANC nadir values and the percentage decrements in the ANC and platelet counts, which were calculated as follows: 100% x ([pretreatment counts - nadir counts]/pretreatment counts). The relationships between NSC 655649 dose, C_max, and AUC, and the hematologic toxicity were assessed using the sigmoidal maximum effect (E_max) model of drug action (ie, percentage change in hematologic parameter = E_max x AUC₀/AUC₅₀ + AUC), where the maximal effect (E_max) was fixed at 100% and the AUC₅₀ is the AUC at which the effect is 50% of the maximal effect. The exponent is a constant that describes the sigmoidicity of the curve. The sigmoidal E_max model was fit to the data by nonlinear least-squares regression. The coefficient of determination (R²) and the standard errors for the estimated parameters were used as measures of goodness of fit for the pharmacodynamics model. Parameter values were expressed as means and SD values. Mean AUC_0- values of patients who did and did not experience severe hematologic toxicity were compared using the Student’s t test (two-sided).

	RESULTS

TOP ABSTRACT INTRODUCTION PATIENTS AND METHODS RESULTS DISCUSSION REFERENCES

 
General
Forty-five patients were treated with 130 total courses of NCS 655649 at doses ranging from 20 mg/m² to 744 mg/m². Table 1 lists the pertinent demographic characteristics of the study patients. The total numbers of new patients treated and courses at each dose level, as well the overall dose-escalation scheme, are listed in Table 2. The median number of courses administered per patient was two (range, one to 10 courses). Three patients required dose reduction for severe myelosuppression, and the dose was inadvertently reduced in one patient.

Toxicity
Hematologic toxicity. Myelosuppression, particularly neutropenia, was the principal DLT of NSC 655649. The median time to the nadir ANC was 14 days (range, 6 to 20 days). Treatment delay was required because of unresolved neutropenia in five (11%) of 45 patients during course 1, but cumulative myelosuppression was not apparent.

The distributions of the relevant grades of neutropenia and thrombocytopenia as functions of both the dose and extent of prior therapy are listed in Table 3, and scatterplots of the percentage decrements in ANC and platelet counts as a function of the NSC 655649 dose are shown in Fig 2A and 2B. The relationship between the dose of NSC 655649 and the decrement in the ANC was steep over the entire dose range, but it was particularly steep at doses 440 mg/m², as demonstrated in Fig 2A. At 440 mg/m², grade 3 or 4 neutropenia was experienced by only one (10%) of 10 HP patients in one (6%) of 18 courses, and DLT did not occur. However, all three HP patients developed grade 4 neutropenia in three of six courses at the next higher dose level (572 mg/m²). One of these individuals experienced several DLT, including grade 4 neutropenia lasting longer than 4 days, febrile neutropenia, and grade 4 thrombocytopenia. Another subject required a 7-day treatment delay because of unresolved neutropenia. Therefore, five additional HP patients were treated at 440 mg/m², which was well tolerated, and 14 HP patients were treated at an intermediate dose level of 500 mg/m². At 500 mg/m², grade 3 or 4 neutropenia occurred in eight (57%) of 14 HP patients in 22 (51%) of 43 courses; however, only one DLT, which consisted of protracted (> 4 days) grade 4 neutropenia during course 1, was noted. With respect to MP patients treated at 572 mg/m², grade 3 or 4 neutropenia occurred in four (67%) of six patients and seven (39%) of 18 courses, but no MP patient experienced DLT during course 1. At the 744 mg/m² dose level, grade 3 or 4 neutropenia occurred in three (75%) of four patients and five (33%) of 15 courses. One of four MP patients experienced hematologic DLT, consisting of both protracted (> 4 days) grade 4 neutropenia and grade 4 thrombocytopenia. This patient experienced no further dose-limiting events during treatment with two subsequent courses of NSC 655649 at the 572-mg/m² dose level.

View larger version (15K): [in this window] [in a new window]   Fig 2. Scatterplots depicting the effects of the first course of NSC 655649 on (A) percentage change in ANC and (B) percentage change in platelet counts. The extent of prior treatment, as defined in Patients and Methods, is also indicated, (•) minimally pretreated and () heavily pretreated.

Nonhematologic toxicity. The most common nonhematologic effects were vomiting, diarrhea, and stomatitis, and the distributions of these toxicities as a function of dose level are depicted in Table 4. Twenty-nine (64%) patients experienced nausea and vomiting at some time during treatment. Nausea and vomiting were generally mild or moderate (grade 1 or 2); however, four patients experienced grade 3 vomiting at the 440 mg/m² (one patient in course 1), 500 mg/m² (one patient in course 2), and 572 mg/m² (two patients in courses 1 and 2) dose levels. These events occurred within 24 hours of treatment and appeared to be dose related. Nausea and vomiting were also prevented or managed successfully with prochlorperazine or serotonin 5HT₃ receptor antagonists, but routine premedication was not necessary because most events were nausea alone, mild in severity and sporadic. Fifteen patients (33%) experienced diarrhea at some time during treatment. The diarrhea was generally mild to moderate (grade 1 or 2) in severity. However, three patients experienced severe (grade 3) diarrhea, including one individual who developed severe toxicity during course 1 at the 500-mg/m² dose level. A single subject experienced grade 3 diarrhea during a second course of NSC 655649 at the 20-mg/m² dose level, but the event was thought to be caused by the patient’s underlying malignancy. Six patients also complained of mild or moderate stomatitis (grade 1 or 2) after treatment with NSC 655649 at doses 440 mg/m². Alopecia occurred in four (9%) of 45 individuals, but the true incidence of alopecia may be higher because assessment was confounded by the contribution of residual alopecia from prior chemotherapy in several patients. Four patients (140 mg/m² [one patient], 440 mg/m² [one patient], and 572 mg/m² [two patients]) experienced moderate discomfort in the vein (grade 2 phlebitis) when infused with NSC 655649 diluted in either 50 or 100 mL of 0.9% saline. NSC 655649 was subsequently diluted in 250 to 500 mL of 0.9% saline solution for patients treated at dose levels 400 mg/m², and central venous access devices were used, which markedly decreased the frequency and severity of phlebitis.

Antitumor Activity
No major objective antitumor responses were observed. However, objective evidence of antitumor activity was noted in two patients. A 74-year-old female with advanced ovarian cancer, who had developed progressive disease after treatment with three prior chemotherapy regimens (carboplatin/cyclophosphamide/altretamine, cisplatin/paclitaxel, and topotecan), experienced a marked and rapid resolution of her ascites, a concomitant decrease in CA-125 from 2,020 to 9 U/mL, and improvement in assessable disease involving the omentum and peritoneum on computerized tomographic scanning. The patient received nine courses of NSC 655649 at the 500-mg/m² dose level after which disease progression was evident. A 44-year-old female with ovarian cancer refractory to four prior chemotherapy regimens (cisplatin/paclitaxel, carboplatin/cyclophosphamide, thiotepa/cisplatin/cyclophosphamide, and topotecan) experienced an 83% reduction in CA-125 (361 to 62 U/mL) accompanied by stable measurable disease on computerized tomographic scan. This patient also received nine courses of NSC 655649 at 500 mg/m² before progressive disease was evident. In addition, a 60-year-old female with a retroperitoneal leiomyosarcoma that had progressed through two prior combination chemotherapy regimens (doxorubicin/ifosfamide/dacarbazine and cyclophosphamide/etoposide) experienced a 23% reduction in measurable disease. The patient was treated with one course of NSC 655649 at the 500-mg/m² dose level and seven courses at the 375-mg/m² does level. One patient each with metastatic renal cell carcinoma and hepatocellular carcinoma had stable disease for 5 and 7 months, respectively.

Pharmacokinetics and Pharmacodynamics
Forty-three of the 45 patients had plasma sampling performed for pharmacokinetic studies, and 23 subjects had complete plasma sampling until day 7 according to an amended, more protracted plasma sampling scheme. Most plasma concentration sets were characterized by the presence of unexpected NSC 655649 peaks from 1.5 to 8 hours after infusion, possibly caused by enterohepatic recirculation of drug. All data sets were analyzed by both noncompartmental and compartmental methods. There was significant intersubject variability in NSC 655659 pharmacokinetics, which is shown in the scatterplots of individual C_max and AUC_0- values as a function of dose (Fig 3). Although mean AUC_0- values were proportional to the dose of NSC 655659 (Pearson’s correlation coefficient r = 0.6231, P < .0001), there was substantial overlap in individual AUC values between all dose levels. There was no correlation with NSC 655649 clearance and interindividual body-surface area (BSA) (data not shown, R² = 0.002). The mean noncompartmental pharmacokinetic parameter estimates at each dose level, as well as overall mean values, are listed in Table 5. The Vd_ss of NSC 655659 was large (averaging 360.24 ± 211 L/m²), the mean plasma Cl was 7.57 ± 4.2 L/h/m², and the elimination t_1/2 averaged 48.85 ± 24 hours. The calculated mean elimination t_1/2 was somewhat longer at 61.25 ± 20.62 hours, and the mean AUC_0- just exceeded the conventional acceptable range (10% to 20%) in the subset of 23 patients in whom prolonged plasma sampling to 7 days posttreatment was performed. However, the calculated mean Cl and Vd_ss for the subset of 23 patients was not significantly different from the 20 patients with shorter sampling times (P = .595 and P = .729, respectively).

View larger version (14K): [in this window] [in a new window]   Fig 3. Scatterplots showing the distributions of the following noncompartmental pharmacokinetic parameter values reflecting NSC 655649 exposure: (A) Cmax and (B) AUC0- as a function of NSC 655649 dose. Horizontal lines represent mean values.

1/2

View larger version (8K): [in this window] [in a new window]   Fig 4. Representative patient’s plasma concentration data at the 572 mg/m2 dose level compared with the predicted model from the compartmental pharmacokinetic model.

Scatterplots of the percent decrements in ANCs and platelets as functions of AUC_0- and C_max are depicted in Fig 5A-5D. Sigmoidal E_max models were fit to both ANC and platelet data sets, as shown in Figs 5A and 5C. Using these models, AUC_0- values of 37.41 and 86.88 µg/mL·hr predict for 50% decrements in ANCs and platelets, respectively. Similar sigmoidal E_max models were derived for dose versus decrements in ANC and platelets, but the fit for these models was inferior to those derived for AUC (R² = 0.3596 and 0.2747 for ANC and platelets, respectively). In contrast, the relationship between C_max and decrements in neutrophil and platelet counts could be adequately described by neither linear nor nonlinear models, as shown in Fig 5B and 5D. Pooling toxicity and pharmacokinetic data from all 10 dose levels, NSC 655649 exposure (AUC_0-) was significantly greater in those patients who experienced severe (grade 4) neutropenia during their first course than those who did not experience severe neutropenia (AUC_0- 80.84 ± 29.66 µg/mL·hr v 49.69 ± 33.30 µg/mL·hr, P = .0195). Similarly, drug exposure was greater for patients who experienced severe neutropenia at the 500 mg/m² and 572 mg/m² dose levels compared with those without toxicity; however, this difference was not statistically significant, possibly because of the small numbers of patients in the analysis (AUC_0- 75.74 ± 31.28 µg/mL·hr v 55.80 ± 23.52 µg/mL·hr units, P = .12)

View larger version (23K): [in this window] [in a new window]   Fig 5. Scatterplots depicting the relationships between percentage decrease in ANC during the first course of NSC 655649 and (A) AUC and (B) Cmax and the percentage decrements in platelets during the first course and (C) AUC and (D) Cmax. The solid lines represent fits of sigmoidal Emax models to the data when appropriate.

	DISCUSSION

TOP ABSTRACT INTRODUCTION PATIENTS AND METHODS RESULTS DISCUSSION REFERENCES

As predicted based on preclinical studies in rodents and dogs, neutropenia was the principal DLT of NSC 655649. Although grades 3 and 4 neutropenia were common at the 500-mg/m² and 572-mg/m² dose levels, the incidence of dose-limiting neutropenic events was low. Overall, the rates of DLT experienced by MP and HP patients treated with 500 mg/m² and 572 mg/m² of NSC 655649, respectively, were acceptable. Dose-limiting hematologic events occurred in one of 18 courses of NSC 655649 administered to MP patients at the 572-mg/m² dose level and in one of 43 courses administered to HP patients at the 500-mg/m² dose level. Attempts to further escalate the doses of NSC 655649 in HP and MP patients above 500 and 572 mg/m², respectively, resulted in unacceptably high rates of DLT, particularly severe neutropenia and thrombocytopenia. Based on these results, NSC 655649 doses of 500 mg/m² and 572 mg/m² are recommended for MP and HP patients, respectively, participating in subsequent disease-directed phase II and III evaluations.

The starting dose for this phase I study, 20 mg/m², was equivalent to one third of the toxic-dose-low in dogs, which is a conventional algorithm for dose selection of cytotoxics in phase I studies. In retrospect, the extrapolation of toxicology data from dogs to humans greatly underestimated the level at which clinically relevant toxicity was observed in this study. In fact, NSC 655649 doses were escalated 25-fold before significant clinical toxicity was noted. This discrepancy was striking and illustrates the unpredictability and limitation of species-to-species dose extrapolation for selecting starting doses for phase I trials based on animal toxicology.¹⁵ Nevertheless, the use of an accelerated dose-escalation scheme and single-patient cohorts when either no or negligible toxicity is observed permitted a relatively rapid escalation of NSC 655649 doses and decreased the number of patients treated at potentially subtherapeutic doses. Thirteen patients were treated at potentially subtherapeutic doses in this study; it would have been necessary to treat 27 patients at potentially subtherapeutic doses if a conventional modified Fibonacci dose-escalation scheme had been used.

The effects of NSC 655649 on the neutrophils and platelets were directly related to drug exposure, with sigmoidal E_max models adequately describing the relationships between dose and AUC and the percent decrements in neutrophil and platelet counts. However, the relationship between AUC and decrements in neutrophils and platelets was not ideal for all patient values encountered. Possible explanations for the significant degree of deviation from the modeled relationship include the heterogeneity of the patient population with respect to the risk of developing neutropenia (HP v MP) and also to the wide spectrum of hepatic function that might affect NSC 655649 clearance among patients treated.

Although nausea, vomiting, and diarrhea occurred frequently, severe nonhematologic toxicity was uncommon. Nausea and vomiting were observed in the most patients treated with higher doses of NSC 655649, but treatment with prochlorperazine and/or serotonin 5HT₃ receptor antagonists generally resulted in successful management or prevention of these toxicities. Mild or moderate (grade 1 or 2) diarrhea that was thought to be drug related occurred in 12 patients (27%) at some time during treatment; however, only two patients (4%) experienced severe diarrhea. Gastrointestinal toxicity, particularly diarrhea, may be caused by a direct toxic effect of NSC 655649, active metabolites on the gastrointestinal mucosa, or both because biliary excretion, enterohepatic recirculation, and fecal elimination seem to be the principal mechanisms of drug disposition.

The principal mechanisms of NSC 655649 elimination seem to be hepatic metabolism and biliary excretion. Renal clearance of the parent compound accounted for approximately 2% of overall drug disposition, and a N-de-ethylated dichloro metabolite has been identified in the urine and in negligible amounts in the plasma using mass spectrometry.¹⁶ Furthermore, the chemical structures of the minor metabolites that have also been identified indicate that NSC 655649 and the N-de-ethylated dichloro metabolite undergo loss of the N-glycan ring, loss of the terminal substituted amino moiety, and cleavage of the succinamide ring.¹⁶ This pattern of fragmentation is indicative of cytochrome P-450 microsomal metabolism.¹⁶ Decreased NSC 655649 clearance in pediatric patients receiving either cyclosporine or clarithromycin, which are known to inhibit cytochrome CYP 3A (cyclosporine and clarithromycin) and CYP 2C8 (cyclosporine), has also been reported.¹⁷ Preliminary preclinical investigations of the mechanism(s) of NSC 655649 metabolism and excretion performed in multidrug-resistant 1a/1b (-/-) knockout mice indicate that NSC 655649 is not a substrate for either Pgp or the specific cytochrome P-450 isoenzyme, CYP3A4 (J. Kuhn, unpublished data).

Because of the structural similarities of NSC 655649 and rebeccamycin, a potential confounding factor in the pharmacokinetic analysis of NSC 655649 is the generation of a metabolite structurally identical to the internal standard used in the current study, rebeccamycin, which would co-elute in the HPLC assay. However, this event would not be expected to alter the estimate of the terminal half-life estimate of NSC 655649 and would be expected to alter clearance, C_max, and AUC estimates by less than 15%. Studies to identify the specific CYP isoenzymes responsible for NSC 655649 metabolism and elucidate the precise metabolic scheme are currently being performed.

Although dosing for this phase I study was based on BSA, there was no correlation between BSA and AUC. The absence of a relationship between AUC and BSA has also been observed with other chemotherapeutic agents.^18,19 This finding suggests that NSC 655649 dosing could be more appropriately based on the determinants of NSC 655649 clearance such as measures of hepatic function. Larger studies of NSC 6556439 should be employed to accurately identify the predictive factors of NSC 655649 clearance and to determine rationale dosing schemes for adult phase II and phase II testing.

The observation of antitumor activity with NSC 655649 in several HP patients with clear taxane-resistant and platinum-resistant ovarian cancer patients is encouraging and provides a rationale for further evaluations of the agent in patients with ovarian cancer. Furthermore, the relatively slow clearance of NSC 655649 (mean t_1/2, 49 hours), the ability to attain plasma concentrations clinically that greatly exceed the IC₅₀ in preclinical models, and the preliminary evidence of antitumor in the current study support the development of NSC 655649 on a single-dosing, tri-weekly schedule.⁵ Based on these observations in advanced ovarian carcinoma, a tumor in which cisplatin and paclitaxel are among the most active agents, phase I studies of NSC 655649 in combination with cisplatin or paclitaxel represent the next rational step in the development of this rebeccamycin derivative. To accomplish this development strategy, a trial is currently under way to evaluate the feasibility of administering NSC 655649 in combination with cisplatin and to explore the effects produced by alternate sequences of these two drugs in patients with advanced malignancies.

	ACKNOWLEDGMENTS

 
Supported in part by National Institutes of Health grant nos. UO1 CA69853, 5P30 CA54174, and MOI RR01346-19.

	REFERENCES

TOP ABSTRACT INTRODUCTION PATIENTS AND METHODS RESULTS DISCUSSION REFERENCES

 
1. Bush JA, Long BH, Catino JJ, et al: Production and biological activity of rebeccamycin: A novel antitumor agent. J Antibiot (Tokyo) 40: 668-678, 1987[Medline]

2. Anizon F, Belin L, Moreau P, et al: Syntheses and biological activities (topoisomerase inhibition and antitumor and antimicrobial properties) of rebeccamycin analogues bearing modified sugar moieties and substituted on the imide nitrogen with a methyl group. J Med Chem 40: 3456-3465, 1997[Medline]

3. Pereira ER, Belin L, Sancelme M, et al: Structure-activity relationships in a series of substituted indolocarbazoles: Topoisomerase I and protein kinase C inhibition and antitumoral and antimicrobial properties. J Med Chem 39: 4471-4477, 1996[Medline]

4. Moreau P, Anizon F, Sancelme M, et al: Syntheses and biological evaluation of indolocarbazoles, analogues of rebeccamycin, modified at the imide heterocycle. J Med Chem 41: 1631-1640, 1998[Medline]

5. Division of Cancer Treatment NCI: Rebeccamycin Analogue BMY-27557-14 Clinical Brochure. Bethesda MD: National Cancer Institute, May 1995

6. Krishnan BS, Moore ME, Lavoie CP, et al: Fluorescence polarization studies of the binding of BMS 181176 to DNA. J Biomol Struct Dyn 12: 625-636, 1994[Medline]

7. Bailly C, Riou JF, Colson P, et al: DNA cleavage by topoisomerase I in the presence of indolocarbazole derivatives of rebeccamycin. Biochemistry 36: 3917-3929, 1997[Medline]

8. Bailly C, Colson P, Houssier C, et al: Recognition of specific sequences in DNA by a topoisomerase I inhibitor derived from the antitumor drug rebeccamycin. Mol Pharmacol 53: 77-87, 1998[Abstract/Free Full Text]

9. Weitman S, Moore R, Barrera H, et al: In vitro antitumor activity of rebeccamycin analog (NSC: 655649) against pediatric solid tumors. J Pediatr Hematol Oncol 20: 136-139, 1998[Medline]

10. Cleary JF, Berlin JD, Tutsch KD, et al: Phase I clinical and pharmacologic study of a rebeccamycin analog (NSC 655649). Proc Am Soc Clin Oncol 16: 217a, 1997 (abstr 760)

11. Yamaoka K, Nakagawa T, Uno T: Application of Akaike’s information criterion (AIC) in the evaluation of linear pharmacokinetic equations. J Pharmacokinet Biopharm 6: 165-175, 1978[Medline]

12. Tricoli JV, Sahai BM, McCormick PJ, et al: DNA topoisomerase I and II activities during cell proliferation and the cell cycle in cultured mouse embryo fibroblast (C3H 10T1/2) cells. Exp Cell Res 158: 1-14, 1985[Medline]

13. Tewey KM, Chen GL, Nelson EM, et al: Intercalative antitumor drugs interfere with the breakage-reunion reaction of mammalian DNA topoisomerase II. J Biol Chem 259: 9182-9187, 1984[Abstract/Free Full Text]

14. Liu LF, Rowe TC, Yang L, et al: Cleavage of DNA by mammalian DNA topoisomerase II. J Biol Chem 258: 15365-15370, 1983[Abstract/Free Full Text]

15. Grieshaber CK, Marsoni S: Relation of preclinical toxicology to findings in early clinical trials. Cancer Treat Rep 70: 65-72, 1986[Medline]

16. Weintraub ST, Krywicki RH, Renouf JP, et al: LC/MS identification of a rebeccamycin analog metabolite. Proc Am Soc Mass Spect 303, 1998 (abstr)

17. Langevin A, Weitman S, Kuhn J, et al: A trial of rebeccamycin analogue (NSC 655649) in children with solid tumors: A pediatric oncology group phase I cooperative agreement study. Proc Am Soc Clin Oncol 18: 198a, 1999 (abstr 764)

18. Gurney HP, Ackland S, Gebski V, et al: Factors affecting epirubicin pharmacokinetics and toxicity: Evidence against using body-surface area for dose calculation. J Clin Oncol 16: 2299-2304, 1998[Abstract]

19. Ratain MJ: Body-surface area as a basis for dosing of anticancer agents: Science, myth, or habit. J Clin Oncol 16: 2297-2298, 1998[Medline]

Submitted October 1, 2000; accepted February 28, 2001.

CiteULike    Complore    Connotea    Del.icio.us    Digg    Facebook    Reddit    Technorati    Twitter    What's this?

This article has been cited by other articles:

				A. D. Ricart, L. A. Hammond, J. G. Kuhn, C. H. Takimoto, A. Goetz, B. Forouzesh, L. Forero, J. L. Ochoa-Bayona, K. Berg, A. W. Tolcher, et al. Phase I and Pharmacokinetic Study of Sequences of the Rebeccamycin Analogue NSC 655649 and Cisplatin in Patients with Advanced Solid Tumors Clin. Cancer Res., December 15, 2005; 11(24): 8728 - 8736. [Abstract] [Full Text] [PDF]

				D. P. Lee, J. M. Skolnik, and P. C. Adamson Pediatric Phase I Trials in Oncology: An Analysis of Study Conduct Efficiency J. Clin. Oncol., November 20, 2005; 23(33): 8431 - 8441. [Abstract] [Full Text] [PDF]

				D. W. Nyman, K. J. Campbell, E. Hersh, K. Long, K. Richardson, V. Trieu, N. Desai, M. J. Hawkins, and D. D. Von Hoff Phase I and Pharmacokinetics Trial of ABI-007, a Novel Nanoparticle Formulation of Paclitaxel in Patients With Advanced Nonhematologic Malignancies J. Clin. Oncol., November 1, 2005; 23(31): 7785 - 7793. [Abstract] [Full Text] [PDF]

				J. Merchant, K. Tutsch, A. Dresen, R. Arzoomanian, D. Alberti, C. Feierabend, K. Binger, R. Marnoccha, J. Thomas, J. Cleary, et al. Phase I Clinical and Pharmacokinetic Study of NSC 655649, a Rebeccamycin Analogue, Given in Both Single-Dose and Multiple-Dose Formats Clin. Cancer Res., July 1, 2002; 8(7): 2193 - 2201. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) Purchase Article View Shopping Cart Alert me when this article is cited Alert me if a correction is posted Services Email this article to a colleague Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Save to my personal folders Download to citation manager Rights & Permissions Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Tolcher, A. W. Articles by Rowinsky, E. K. Search for Related Content PubMed PubMed Citation Articles by Tolcher, A. W. Articles by Rowinsky, E. K. Social Bookmarking                            What's this?