Phase I and Pharmacologic Study of Topotecan: A Novel Topoisomerase I Inhibitor By Eric K. Rowinsky, Louise B. Grochow, Carolyn B. Hendricks, David S. Ettinger, Arlene A. Forastiere, Lisa A. Hurowitz, William P. McGuire, Susan E. Sartorius, Barbara G. Lubejko, Scott H. Kaufmann, and Ross C. Donehower Purpose: A phase I and pharmacologic study was undertaken to determine the maximum-tolerated dose (MTD), describe the principal toxicities, and characterize the pharmacologic behavior of topotecan, which is a semisynthetic analog of camptothecin with broad preclinical antitumor activity and the first topoisomerase I-targeting agent to enter clinical development in the United States since studies of sodium camptothecin over 2 decades ago. Patientsand Methods: Thirty-minute infusions of topotecan were administered daily for 5 consecutive days every 3 weeks to patients with advanced solid malignancies at doses ranging from 0.5 to 2.5 mg/m 2 /d. Results: At doses of 1.5 and 2.0 mg/m', grade 3 and 4 neutropenia occurred in most courses; however, neutropenia was brief and rarely associated with fevers or treatment delays. Neutropenia was more severe in patients with extensive prior treatment than in minimally pretreated patients, but these differences were not substantial. At 2.5 mg/m2, topotecan induced profound and prolonged neutropenia that was frequently associated with fever and treatment delays in minimally pretreated patients. Topotecan also induced mild depressions in the
hematocrit level in the majority of courses; however, precipitous drops requiring transfusional therapy occurred in 14% of courses and suggested a drug-induced hemolytic effect. Unlike sodium camptothecin, hemorrhagic cystitis was not observed. Thrombocytopenia, skin rash, diarrhea, and vomiting occurred infrequently and were modest in severity. Responses were observed in non-small-cell lung carcinoma and platinum-refractory ovarian carcinoma. Drug disposition in plasma was described by a biexponential model, with renal elimination accounting for 38.7% of drug disposition. Topotecan was rapidly hydrolyzed in vivo to a less active, open-ring form. Conclusions: Neutropenia is the dose-limiting toxicity, and 1.5 mg/m' is the recommended starting dose of topotecan for both minimally and heavily pretreated patients in future phase II trials, with escalation to 2.0 mg/m 2 if treatment is well tolerated. Non-small-cell lung and platinum-refractory ovarian carcinomas should be among those evaluated in phase II trials of topotecan. J Clin Oncol 10:647-656. © 1992 by American Society of Clinical Oncology.
rTOPOTECAN (SKF 104864), a semisynthetic water-
nmol/L), it was superior to camptothecin against mice bearing L1210 leukemia at the maximum-tolerated dose (MTD), with increased life-spans (ILS) of 173 - 16% versus 118 + 6%, respectively. 12 Topotecan was also superior to camptothecin in several tumor models, including Lewis lung carcinoma and B16 melanoma, and equivalent to camptothecin in the HT 29 human colon tumor xenograft and in the murine colon adenocarcino-
soluble analog of camptothecin, is the first topoisomerase I-targeting agent to enter clinical cancer trials since the limited clinical development of sodium camptothecin in the early 1970s. Topoisomerase I-targeting drugs stabilize a covalent DNA-topoisomerase I complex to yield enzyme-linked DNA single-strand breaks."6 Topoisomerase I possesses several characteristics that
may be relevant to its potential use as a strategic target in cancer therapy. First, its presence in relatively high levels in both proliferating and quiescent cells suggests that its function may be independent of cellular growth rate and that topoisomerase I-targeting drugs may be active in slow-growing as well as rapidly proliferating tumors.7'" There is also evidence indicating that the regulation of topoisomerase I is altered in neoplastic cells, which suggests a selective cytotoxic advantage for these agents in malignant tissues compared with normal tissues. For example, cells from colon tumors have been shown to contain higher intracellular levels of topoisomerase I than normal mucosal cells."1
Topotecan demonstrated broad antitumor activity in preclinical studies. Although topotecan was twofold less
active than camptothecin against L1210 cells in vitro (concentration inhibiting 50% of growth [ICs0s], 23 v 56
mas 38 and 51.12,13 However, an optimal administration schedule could not be defined and varied according to
the specific tumor model used.
From The Johns Hopkins Oncology Center, The Johns Hopkins MedicalInstitutions, Baltimore,MD. SubmittedAugust 8, 1991; acceptedNovember 5, 1991. Supported by National Institutes of Health contractno. NO1-CM57738. S.H.K. is the recipient of an American Cancer Society Career DevelopmentAward. Presentedin part at the Annual Meeting of the American Society of Clinical Oncology, Houston, TX, May 1991. Address reprint requests to Eric K Rowinsky, MD, The Johns Hopkins Oncology Center, Division of Pharmacologyand Experimental Therapeutics, 1-121, 600 N Wolfe St, Baltimore, MD 21205. © 1992 by American Society of ClinicalOncology. 0732-183X/92/1004-0023$3.00/0
Journal of Clinical Oncology, Vol 10, No 4 (April), 1992: pp 647-656
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The decision to develop topotecan was based on a combination of favorable characteristics including the following: (1) enhanced water solubility; (2) intrinsic activity as a parent compound rather than as a watersoluble prodrug of camptothecin; (3) superior or equivalent antitumor activity compared with camptothecin against a broad spectrum of tumors; (4) accessibility via feasible semisynthetic processes from camptothecin, avoiding complex and inefficient total synthetic processes; and (5) increased potency in vivo at the MTD, minimizing the requirement for camptothecin-a rela13 tively scarce natural product-as a starting material. ,14 In addition, topotecan had a different spectrum of toxicity than sodium camptothecin. Whereas, sodium camptothecin induced profound nonhematologic effects in preclinical and early clinical trials including hemorrhagic enterocolitis and cystitis, which precluded the agent's further clinical development, the predominant and dose-limiting toxicity of topotecan in animal studies was myelosuppression, principally neutropenia.12' 15-19 Two species of topotecan, an active closed-ring lactone and a less active, open-ring form generated by pH-dependent hydrolysis exist in equilibrium in aqueous solutions (Fig 1).14,20,21 While acidic conditions favor the formation of the active lactone, more basic pHs, including physiologic pHs, favor the formation of the less
active, open-ring species. The purposes of this study were to (1) determine the MTD of topotecan given by a brief, daily intravenous (IV) infusion for 5 days, repeated every 3 weeks; (2) recommend a dose for phase II trials; (3) characterize the toxicities associated with this schedule of administration; (4) seek preliminary evidence for antitumor activity of topotecan; and (5) describe the pharmacology of topotecan administered on this schedule. PATIENTS AND METHODS Eligibility Only patients with histologically documented solid tumors refractory to conventional therapy or for which no effective therapy existed were candidates for this study. Eligibility criteria included
(1) age > 18 years; (2) an Eastern Cooperative Oncology Group (ECOG) performance status < 2 (ambulatory and capable of self-care); (3) a life expectancy of at least 6 weeks (enabling the completion of at least two courses of therapy); (4) no major surgery within 14 days or wide-field radiotherapy and/or chemotherapy within 28 days of entering onto protocol (or 6 weeks in those treated with a nitrosourea or mitomycin); (5) adequate hematopoietic (WBC count > 4,000/LL and platelet count > 100,000/pL), hepatic (total bilirubin level < 1.5 mg/dL), and renal (creatinine level < 1.5 mg/dL) functions; (6) no history of hemorrhagic cystitis and fewer than five RBCs per high power field in the urine; and (7) no other coexisting medical problems of sufficient severity to prevent full compliance with the study. All patients gave informed written consent according to federal and institutional guidelines before treatment. Dosage and DrugAdministration Due to significant interspecies differences in the dose that is lethal in 10% of animals (LDJ0) in preclinical toxicologic studies, the starting dose of topotecan was /3oof the murine LD, 0 or 0.5 mg/m2 given as a 30-minute infusion daily for 5 consecutive days. Initial dose escalations were made using a modified Fibonacci search method to 1.0 and 1.5 mg/m2; however, a more conservative scheme with subsequent dose escalations to 2.0 and 2.5 mg/m' was used after hematologic toxicity was observed at the lower doses. At least three topotecan-naive patients were entered at each escalated dose level. As the MTD was approached and potential doselimiting toxicity was observed, at least six topotecan-naive patients were entered. The MTD was defined as one dose level below the dose that induced dose-limiting toxicities, which were defined as at least one of the following: (1) absolute neutrophil count [ANC] less than 500/LL or platelet count less than 50,000/pL for more than 5 days; (2) ANC less than 500/p.L with fever requiring parenteral antibiotics, and/or nonhematologic toxicity > National Cancer Institute (NCI) grade 3 in greater than one third of topotecannaive patients (at least two of a maximum of six total patients). Dose escalations were permitted in the same patient if the patient received at least two courses at the lower dose level without doselimiting toxicity and if two new patients had already been treated at the next highest dose. Dose reductions by one dose level were permitted for dose-limiting toxicity. Topotecan was supplied by the Division of Cancer Treatment, NCI (Bethesda, MD) in vials containing a lyophilized mixture of 5 mg of topotecan and 100 mg mannitol. The pH was previously adjusted to 3 to 4 with hydrochloric acid and sodium hydroxide. Topotecan was reconstituted with 2 mL sterile water, diluted in 100 mL of 5% dextrose solution, and then administered over 30 minutes. The first two doses of the initial course of topotecan were administered on the inpatient units of The Johns Hopkins Oncology Center, and all other treatment was given in the outpatient clinic. Pretreatmentand Follow-Up Studies
Fig 1. Structures of the active topotecan lactone (left) undergoing reversible pH-dependent hydrolysis to its inactive open-ring form.
Histories, physical examinations, and routine laboratory studies were performed pretreatment and weekly. Pretreatment and routine laboratory studies included complete blood cell differential WBC counts; electrolyte, BUN, creatinine, glucose, total protein, albumin, calcium, phosphate, uric acid, alkaline phosphatase, total and direct bilirubin, SGOT, SGPT, amylase levels; prothrombin
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649
PHASE I AND PHARMACOLOGIC STUDY OF TOPOTECAN time; urinalysis; and a fecal hemocult test for occult blood. Complete blood cell counts were also obtained on day 10 and every 2 days if ANCs decreased to < 1,000/p L. ECGs were obtained pretreatment and before each course. Toxicities were evaluated according to the NCI Common Toxicity Criteria."2 Formal tumor measurements were performed after every two courses, and patients were able to continue treatment if they did not develop progressive disease. A complete response (CR) was scored if there was disappearance of all active disease on two measurements separated by a minimum period of 3 weeks. A partial response (PR) required a 50% or greater reduction in the sum of the product of the bidimensional measurements of all measurable lesions documented by two measurements separated by at least 3 weeks; and a minor response (MR) required a decrease in the sum of the product of bidimensional tumor measurements of less than 50%. For the analysis of hematologic toxicity, patients were considered heavily pretreated with therapies that may affect long-term hematopoietic function if they had received large-field radiation to bone marrow-containing areas such as pelvis or spine or a six courses of chemotherapy containing an alkylating agent (excluding multiagent regimens consisting of low doses of cisplatin) or Ž two courses of a nitrosourea or mitomycin.
PharmacologicAnalyses Blood samples in heparinized tubes were collected before the infusion; at 5, 10, and 15 minutes during the infusion; and at the end of infusion. Samples were also collected at 5, 15, and 30 minutes and 1, 2, 3, 4, 8, and 24 hours after the end of infusion. Blood was sampled for pharmacokinetic studies on days 1 and 5. Urine was collected continuously for 24 hours in three 8-hour aliquots after drug administration on day 1. Specimen processing, extraction, and chromatographic quantitation of both the lactone and open-ring species of topotecan were performed using a modified method that was originally developed by Beijnen et al.2 Briefly, samples were centrifuged at 600g for 8 minutes immediately after sample collection, and 0.6 mL of plasma or urine was then added to 3.6 mL of cold methanol. The mixture was next vortexed for 10 seconds, centrifuged at 2,500 rpm for 10 minutes, and either immediately analyzed or stored at -70 0 C for up to 1 week. Parallel stability studies have demonstrated that both the lactone and hydroxy acid are stable for at least 1 week when extracted and stored in this manner. For chromatographic analysis, 500 p.L of the methanol extract was added to each of two paired autovials. To quantitate total drug, 20 pL of a 20% phosphoric acid solution was added to one of the paired samples to measure all drug, and the unacidified sample was used to quantitate the lactone. Separation of the sample was accomplished by reverse-phase high-performance liquid chromatography (HPLC). Using an ISS100 autosampler (Perkin Elmer, Rockville, MD), 50-p.L samples were injected onto a Waters Model M6000 HPLC system (Waters, Milford, MA) equipped with a 150 mm x 3.9 mm (internal diameter) x 5 l.m C18 Resolve column (Waters) and a Guard-Pak precolumn, dry-packed with Bodapak Corasil C18 (Waters). The mobile phase consisted of 80 mL 0.250 mol/L sodium dioctylsulfosuccinate, 11.2 mL triethylamine, 624 mL water, 1,240 mL methanol, and 46 mL 1.0 mol/L phosphate buffer, pH 6.0. The pH of the mixture was adjusted to 6.0 with phosphoric acid. The flow rate was maintained at 1.3 mL/min, and the column effluent was monitored fluorimetrically using an LS40 Perkin Elmer fluorescence detector
with excitation and emission wavelengths set at 382 nm and 523 nm, respectively. Peak areas were quantitated using a Nelson 3000 integration system (Perkin Elmer Nelson System, Cupertino, CA), and drug concentrations were determined from linear regression equations derived from calibration curves prepared with samples between 2 and 400 nmol/L. Under these conditions, the retention time for the lactone was 6 minutes, whereas the open-ring form was eluted rapidly in the solvent front, with a retention time of approximately 30 seconds. The open-ring form was converted back to the lactone with acidification. To quantitate the concentration of the open-ring form, the concentration of the lactone in the unacidified sample was subtracted from the total drug concentration measured in the acidified sample. Individual plasma clearance curves were fit using a biexponential, linear, mamillary model of drug distribution and administration after visual inspection of the raw data and curves. Concentration data were weighted by 1/concentration. The values of the following kinetic parameters were estimated for each clearance curve: alpha and beta rate constants and associated half-lives, volumes of distribution for the central compartment (Va) and steady-state (VJ), clearance rate, and area under the curve (AUC). Pharmacokinetic modeling and parameter estimation was performed by the nonlinear regression program PC NONLIN (Statistical Consultants, Lexington, KY). The mean kinetic parameter values that were derived from analysis of the day-1 and -5 clearance curves were compared using Student's t test for correlated data pairs. The pharmacokinetics and pharmacodynamics of topotecan were explored using scatterplots of dose or AUC versus the percentage decrease in ANC, defined as 100 x (Pretreatment ANC - Nadir ANC) Pretreatment ANC These relationships were also modeled to a sigmoidal maximum effect (Em,) model": % change in ANC =
Maximum Effect (dose)k (Doses) + Dosek
Nonlinear least square regression using PC NONLIN was used to estimate k, which describes the shape of the curve, and doses0 , the dose that results in 50% of the maximal effect (eg, 100% reduction in ANC). RESULTS
Twenty-nine patients received 161 total courses of topotecan through five dose levels (Table 1). One course involving a 44-year-old female with non-small-cell lung carcinoma who had progressive dyspnea and died sudTable 1. Topotecan Dose Escalations Dose (mg/m'
New
Total
Assessable
daily x 5 days)
Patients
New Patients Subsequently Escalated
Reduced
Patients
Courses
0.5 1.0 1.5 2.0 2.5 Total
3 6 10 6 4 29
3 3 0 0 0
0 0 2 1 3
3 11 15 11 4
6 32 72 46 4 160
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ROWINSKY ET AL
denly 24 hours after receiving topotecan during her third course. She was not fully assessable. Postmortem examination showed extensive intrathoracic tumor and no evidence of drug toxicity. The median number of courses administered per patient was three and ranged from one to 21 as of November 1, 1991 (one course, one patient; two, 11 patients; three, six patients; four, two patients; five, one patient; six, one patient; seven, one patient; eight, one patient; 10, one patient; 16+, one patient; 19, one patient; 20+, one patient; 21+, one patient). Nine patients were treated at two doses; four patients had dose escalations due to negligible toxicity; and five patients had dose reductions due to profound neutropenia and fever. Two patients were treated at three doses due to neglible adverse effects at their two lower doses. Patient characteristics are listed in Table 2. Of the 29 patients, 28 had an ECOG performance status of either 0 or 1. All patients received cytotoxic therapies before entry onto study. Nineteen and 10 patients were considered minimally and heavily pretreated, respectively. Hematologic Toxicity Neutropenia was the principal dose-limiting toxicity of topotecan on this schedule. Table 3 lists the median and range of nadir ANCs, the grades of neutropenia, and the number of episodes of neutropenia and fever requiring hospitalization for parenteral antibiotics at each dose level. Infections were not documented in any of the five episodes of neutropenia and fever in this study. The onset of neutropenia typically occurred between days 8 and 10, and nadir ANCs were noted between days 8 and 12. More importantly, neutropenia Table 2. Patient Characteristics No. of patients Sex (male:female) Median age, years (range) ECOG performance status 0 1 2 Previous treatment Chemotherapy Radiotherapy Chemotherapy + radiotherapy Disease site Colorectal NSC lung Ovarian Unknown primary (adeno) Breast Melanoma Head and neck
29 13:16 51 (35-76) 13 15 1 20 2 7 14 7 3 2 1 1 1
Abbreviations: NSC, non-small-cell; adeno, adenocarcinoma.
resolved by day 22 in 91% of courses. Treatment delays of 1 week were required in 13 courses, and a delay of 2 weeks was required in one course due to unresolved neutropenia. Three episodes of brief grade 3 neutropenia were experienced by two heavily pretreated patients who received a total of 32 courses at the 1.0-mg/m 2 dose level; however, significant and consistent myelosuppression did not occur until the 1.5-mg/m 2 dose level. At 1.5 mg/m 2, grade 3 and 4 neutropenia occurred in 58% and 17% of courses, respectively. For minimally pretreated patients, grade 3 neutropenia was noted by 89% of patients in 67% of courses and grade 4 neutropenia was noted in 11% of patients in 9% of courses. For heavily pretreated patients, grade 3 neutropenia was observed in 33% of patients and 51% of courses; grade 4 neutropenia occurred in 67% of patients during 23% of courses. Although the median ANC nadir was lower for heavily pretreated than minimally pretreated patients receiving 1.5 mg/m2, the difference was not substantial (693 v 792 cells per microliter), and only one heavily pretreated patient developed neutropenia and fever. At the 2.0-mg/m2 dose level, a higher proportion of courses were associated with grade 4 neutropenia. Grade 3 neutropenia was experienced by 54% of minimally pretreated patients in 35% of courses, and grade 4 neutropenia was noted in 36% of patients during 35% of courses. One heavily pretreated patient experienced grade 3 and 4 neutropenia during 50% and 50% of courses, respectively. At 2.0 mg/m2, median nadir ANCs were 695 and 499 cells per microliter for minimally and heavily pretreated patients, respectively, and neutropenia and fever requiring parenteral antibiotics occurred in only one course in each of the minimally and heavily pretreated cohorts. Neutropenia was severe in three of the four patients who received topotecan at the 2.5mg/m 2 dose level, with ANC nadirs of 128, 161, 224, and 2,380/ tL. At 2.5 mg/m 2, treatment delays due to unresolved neutropenia were required in two of the four patients, and fever and neutropenia requiring antibiotics occurred in two of four courses. Although the percentages of total courses associated with grade 3 and 4 neutropenia at 1.5 mg/m 2 (75%) and 2.0 mg/m 2 (74%) were relatively high, both dose levels were well tolerated by the majority of patients due to the brevity of the severe neutropenia, the low percentage of courses requiring subsequent treatment delays (9%), the low incidence of febrile episodes (3%), and the low frequency of serious nonhematologic effects. There was no evidence of a cumulative effect of topotecan on the nadir ANCs in both minimally and heavily pretreated
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651
PHASE I AND PHARMACOLOGIC STUDY OF TOPOTECAN Table 3. Topotecan: Neutropenia
0.5
Grade
Median Nadir ANC (range)
No. of Courses
Dose (mg/m2)
0
1
2
3
4
6
4,550 (2,850-6,396)
6
1.0 Minimally pretreated Heavily pretreated
32 20 12
1,759 (612-6,622) 1,782 (1,127-6,622) 1,275 (612-2,088)
11 9 2
8 7 1
10 4 6
3
1.5 Minimally pretreated Heavily pretreated
72 33 39
740 (141-2,415) 792 (304-1,809) 693 (141-2,415)
1
3
1
3
14 8 6
42 22 20
12* 3 9
2.0 Minimally pretreated Heavily pretreated
46 40 6
681 (73-1,690) 695 (73-1,690) 499 (300-528)
1 1
11 11
17 14 3
17* 14 3
2.5 Minimally pretreated
4
276 (128-2,380)
3
1
3*
*Five courses at 1.5 mg/m2 (one), 2.0 mg/m' (two), and 2.5 mg/m2 (two) associated with fever and neutropenia requiring hospitalization and antibiotics.
patients. To illustrate the lack of cumulative neutropenia from topotecan, the mean percent decreases in ANCs in patients receiving more than one course of topotecan at the same dose are depicted as a function of the cumulative number of courses in Fig 2. Topotecan induced negligible effects on platelets. Nadir platelet counts were generally observed on day 15; however, one, five, and four courses at 1.0, 1.5, and 2.0 mg/m 2, respectively, were associated with grade 2 thrombocytopenia, and only one course in the study (2.5 mg/m 2) was associated with grade 3 thrombocytopenia (nadir platelet count, 45,000/liL). In contrast, topotecan's effects on RBCs were observed frequently. A 1007
900 z
80-
hematocrit level in the majority of courses; however, precipitous drops requiring transfusional therapy occurred in 14% of courses and suggested a drug-induced hemolytic effect. Unlike sodium camptothecin, hemorrhagic cystitis was not observed. Thrombocytopenia, skin rash, diarrhea, and vomiting occurred infrequently and were modest in severity. Responses were observed in non-small-cell lung carcinoma and platinum-refractory ovarian carcinoma. Drug disposition in plasma was described by a biexponential model, with renal elimination accounting for 38.7% of drug disposition. Topotecan was rapidly hydrolyzed in vivo to a less active, open-ring form. Conclusions: Neutropenia is the dose-limiting toxicity, and 1.5 mg/m' is the recommended starting dose of topotecan for both minimally and heavily pretreated patients in future phase II trials, with escalation to 2.0 mg/m 2 if treatment is well tolerated. Non-small-cell lung and platinum-refractory ovarian carcinomas should be among those evaluated in phase II trials of topotecan. J Clin Oncol 10:647-656. © 1992 by American Society of Clinical Oncology.
rTOPOTECAN (SKF 104864), a semisynthetic water-
nmol/L), it was superior to camptothecin against mice bearing L1210 leukemia at the maximum-tolerated dose (MTD), with increased life-spans (ILS) of 173 - 16% versus 118 + 6%, respectively. 12 Topotecan was also superior to camptothecin in several tumor models, including Lewis lung carcinoma and B16 melanoma, and equivalent to camptothecin in the HT 29 human colon tumor xenograft and in the murine colon adenocarcino-
soluble analog of camptothecin, is the first topoisomerase I-targeting agent to enter clinical cancer trials since the limited clinical development of sodium camptothecin in the early 1970s. Topoisomerase I-targeting drugs stabilize a covalent DNA-topoisomerase I complex to yield enzyme-linked DNA single-strand breaks."6 Topoisomerase I possesses several characteristics that
may be relevant to its potential use as a strategic target in cancer therapy. First, its presence in relatively high levels in both proliferating and quiescent cells suggests that its function may be independent of cellular growth rate and that topoisomerase I-targeting drugs may be active in slow-growing as well as rapidly proliferating tumors.7'" There is also evidence indicating that the regulation of topoisomerase I is altered in neoplastic cells, which suggests a selective cytotoxic advantage for these agents in malignant tissues compared with normal tissues. For example, cells from colon tumors have been shown to contain higher intracellular levels of topoisomerase I than normal mucosal cells."1
Topotecan demonstrated broad antitumor activity in preclinical studies. Although topotecan was twofold less
active than camptothecin against L1210 cells in vitro (concentration inhibiting 50% of growth [ICs0s], 23 v 56
mas 38 and 51.12,13 However, an optimal administration schedule could not be defined and varied according to
the specific tumor model used.
From The Johns Hopkins Oncology Center, The Johns Hopkins MedicalInstitutions, Baltimore,MD. SubmittedAugust 8, 1991; acceptedNovember 5, 1991. Supported by National Institutes of Health contractno. NO1-CM57738. S.H.K. is the recipient of an American Cancer Society Career DevelopmentAward. Presentedin part at the Annual Meeting of the American Society of Clinical Oncology, Houston, TX, May 1991. Address reprint requests to Eric K Rowinsky, MD, The Johns Hopkins Oncology Center, Division of Pharmacologyand Experimental Therapeutics, 1-121, 600 N Wolfe St, Baltimore, MD 21205. © 1992 by American Society of ClinicalOncology. 0732-183X/92/1004-0023$3.00/0
Journal of Clinical Oncology, Vol 10, No 4 (April), 1992: pp 647-656
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ROWINSKY ET AL
The decision to develop topotecan was based on a combination of favorable characteristics including the following: (1) enhanced water solubility; (2) intrinsic activity as a parent compound rather than as a watersoluble prodrug of camptothecin; (3) superior or equivalent antitumor activity compared with camptothecin against a broad spectrum of tumors; (4) accessibility via feasible semisynthetic processes from camptothecin, avoiding complex and inefficient total synthetic processes; and (5) increased potency in vivo at the MTD, minimizing the requirement for camptothecin-a rela13 tively scarce natural product-as a starting material. ,14 In addition, topotecan had a different spectrum of toxicity than sodium camptothecin. Whereas, sodium camptothecin induced profound nonhematologic effects in preclinical and early clinical trials including hemorrhagic enterocolitis and cystitis, which precluded the agent's further clinical development, the predominant and dose-limiting toxicity of topotecan in animal studies was myelosuppression, principally neutropenia.12' 15-19 Two species of topotecan, an active closed-ring lactone and a less active, open-ring form generated by pH-dependent hydrolysis exist in equilibrium in aqueous solutions (Fig 1).14,20,21 While acidic conditions favor the formation of the active lactone, more basic pHs, including physiologic pHs, favor the formation of the less
active, open-ring species. The purposes of this study were to (1) determine the MTD of topotecan given by a brief, daily intravenous (IV) infusion for 5 days, repeated every 3 weeks; (2) recommend a dose for phase II trials; (3) characterize the toxicities associated with this schedule of administration; (4) seek preliminary evidence for antitumor activity of topotecan; and (5) describe the pharmacology of topotecan administered on this schedule. PATIENTS AND METHODS Eligibility Only patients with histologically documented solid tumors refractory to conventional therapy or for which no effective therapy existed were candidates for this study. Eligibility criteria included
(1) age > 18 years; (2) an Eastern Cooperative Oncology Group (ECOG) performance status < 2 (ambulatory and capable of self-care); (3) a life expectancy of at least 6 weeks (enabling the completion of at least two courses of therapy); (4) no major surgery within 14 days or wide-field radiotherapy and/or chemotherapy within 28 days of entering onto protocol (or 6 weeks in those treated with a nitrosourea or mitomycin); (5) adequate hematopoietic (WBC count > 4,000/LL and platelet count > 100,000/pL), hepatic (total bilirubin level < 1.5 mg/dL), and renal (creatinine level < 1.5 mg/dL) functions; (6) no history of hemorrhagic cystitis and fewer than five RBCs per high power field in the urine; and (7) no other coexisting medical problems of sufficient severity to prevent full compliance with the study. All patients gave informed written consent according to federal and institutional guidelines before treatment. Dosage and DrugAdministration Due to significant interspecies differences in the dose that is lethal in 10% of animals (LDJ0) in preclinical toxicologic studies, the starting dose of topotecan was /3oof the murine LD, 0 or 0.5 mg/m2 given as a 30-minute infusion daily for 5 consecutive days. Initial dose escalations were made using a modified Fibonacci search method to 1.0 and 1.5 mg/m2; however, a more conservative scheme with subsequent dose escalations to 2.0 and 2.5 mg/m' was used after hematologic toxicity was observed at the lower doses. At least three topotecan-naive patients were entered at each escalated dose level. As the MTD was approached and potential doselimiting toxicity was observed, at least six topotecan-naive patients were entered. The MTD was defined as one dose level below the dose that induced dose-limiting toxicities, which were defined as at least one of the following: (1) absolute neutrophil count [ANC] less than 500/LL or platelet count less than 50,000/pL for more than 5 days; (2) ANC less than 500/p.L with fever requiring parenteral antibiotics, and/or nonhematologic toxicity > National Cancer Institute (NCI) grade 3 in greater than one third of topotecannaive patients (at least two of a maximum of six total patients). Dose escalations were permitted in the same patient if the patient received at least two courses at the lower dose level without doselimiting toxicity and if two new patients had already been treated at the next highest dose. Dose reductions by one dose level were permitted for dose-limiting toxicity. Topotecan was supplied by the Division of Cancer Treatment, NCI (Bethesda, MD) in vials containing a lyophilized mixture of 5 mg of topotecan and 100 mg mannitol. The pH was previously adjusted to 3 to 4 with hydrochloric acid and sodium hydroxide. Topotecan was reconstituted with 2 mL sterile water, diluted in 100 mL of 5% dextrose solution, and then administered over 30 minutes. The first two doses of the initial course of topotecan were administered on the inpatient units of The Johns Hopkins Oncology Center, and all other treatment was given in the outpatient clinic. Pretreatmentand Follow-Up Studies
Fig 1. Structures of the active topotecan lactone (left) undergoing reversible pH-dependent hydrolysis to its inactive open-ring form.
Histories, physical examinations, and routine laboratory studies were performed pretreatment and weekly. Pretreatment and routine laboratory studies included complete blood cell differential WBC counts; electrolyte, BUN, creatinine, glucose, total protein, albumin, calcium, phosphate, uric acid, alkaline phosphatase, total and direct bilirubin, SGOT, SGPT, amylase levels; prothrombin
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649
PHASE I AND PHARMACOLOGIC STUDY OF TOPOTECAN time; urinalysis; and a fecal hemocult test for occult blood. Complete blood cell counts were also obtained on day 10 and every 2 days if ANCs decreased to < 1,000/p L. ECGs were obtained pretreatment and before each course. Toxicities were evaluated according to the NCI Common Toxicity Criteria."2 Formal tumor measurements were performed after every two courses, and patients were able to continue treatment if they did not develop progressive disease. A complete response (CR) was scored if there was disappearance of all active disease on two measurements separated by a minimum period of 3 weeks. A partial response (PR) required a 50% or greater reduction in the sum of the product of the bidimensional measurements of all measurable lesions documented by two measurements separated by at least 3 weeks; and a minor response (MR) required a decrease in the sum of the product of bidimensional tumor measurements of less than 50%. For the analysis of hematologic toxicity, patients were considered heavily pretreated with therapies that may affect long-term hematopoietic function if they had received large-field radiation to bone marrow-containing areas such as pelvis or spine or a six courses of chemotherapy containing an alkylating agent (excluding multiagent regimens consisting of low doses of cisplatin) or Ž two courses of a nitrosourea or mitomycin.
PharmacologicAnalyses Blood samples in heparinized tubes were collected before the infusion; at 5, 10, and 15 minutes during the infusion; and at the end of infusion. Samples were also collected at 5, 15, and 30 minutes and 1, 2, 3, 4, 8, and 24 hours after the end of infusion. Blood was sampled for pharmacokinetic studies on days 1 and 5. Urine was collected continuously for 24 hours in three 8-hour aliquots after drug administration on day 1. Specimen processing, extraction, and chromatographic quantitation of both the lactone and open-ring species of topotecan were performed using a modified method that was originally developed by Beijnen et al.2 Briefly, samples were centrifuged at 600g for 8 minutes immediately after sample collection, and 0.6 mL of plasma or urine was then added to 3.6 mL of cold methanol. The mixture was next vortexed for 10 seconds, centrifuged at 2,500 rpm for 10 minutes, and either immediately analyzed or stored at -70 0 C for up to 1 week. Parallel stability studies have demonstrated that both the lactone and hydroxy acid are stable for at least 1 week when extracted and stored in this manner. For chromatographic analysis, 500 p.L of the methanol extract was added to each of two paired autovials. To quantitate total drug, 20 pL of a 20% phosphoric acid solution was added to one of the paired samples to measure all drug, and the unacidified sample was used to quantitate the lactone. Separation of the sample was accomplished by reverse-phase high-performance liquid chromatography (HPLC). Using an ISS100 autosampler (Perkin Elmer, Rockville, MD), 50-p.L samples were injected onto a Waters Model M6000 HPLC system (Waters, Milford, MA) equipped with a 150 mm x 3.9 mm (internal diameter) x 5 l.m C18 Resolve column (Waters) and a Guard-Pak precolumn, dry-packed with Bodapak Corasil C18 (Waters). The mobile phase consisted of 80 mL 0.250 mol/L sodium dioctylsulfosuccinate, 11.2 mL triethylamine, 624 mL water, 1,240 mL methanol, and 46 mL 1.0 mol/L phosphate buffer, pH 6.0. The pH of the mixture was adjusted to 6.0 with phosphoric acid. The flow rate was maintained at 1.3 mL/min, and the column effluent was monitored fluorimetrically using an LS40 Perkin Elmer fluorescence detector
with excitation and emission wavelengths set at 382 nm and 523 nm, respectively. Peak areas were quantitated using a Nelson 3000 integration system (Perkin Elmer Nelson System, Cupertino, CA), and drug concentrations were determined from linear regression equations derived from calibration curves prepared with samples between 2 and 400 nmol/L. Under these conditions, the retention time for the lactone was 6 minutes, whereas the open-ring form was eluted rapidly in the solvent front, with a retention time of approximately 30 seconds. The open-ring form was converted back to the lactone with acidification. To quantitate the concentration of the open-ring form, the concentration of the lactone in the unacidified sample was subtracted from the total drug concentration measured in the acidified sample. Individual plasma clearance curves were fit using a biexponential, linear, mamillary model of drug distribution and administration after visual inspection of the raw data and curves. Concentration data were weighted by 1/concentration. The values of the following kinetic parameters were estimated for each clearance curve: alpha and beta rate constants and associated half-lives, volumes of distribution for the central compartment (Va) and steady-state (VJ), clearance rate, and area under the curve (AUC). Pharmacokinetic modeling and parameter estimation was performed by the nonlinear regression program PC NONLIN (Statistical Consultants, Lexington, KY). The mean kinetic parameter values that were derived from analysis of the day-1 and -5 clearance curves were compared using Student's t test for correlated data pairs. The pharmacokinetics and pharmacodynamics of topotecan were explored using scatterplots of dose or AUC versus the percentage decrease in ANC, defined as 100 x (Pretreatment ANC - Nadir ANC) Pretreatment ANC These relationships were also modeled to a sigmoidal maximum effect (Em,) model": % change in ANC =
Maximum Effect (dose)k (Doses) + Dosek
Nonlinear least square regression using PC NONLIN was used to estimate k, which describes the shape of the curve, and doses0 , the dose that results in 50% of the maximal effect (eg, 100% reduction in ANC). RESULTS
Twenty-nine patients received 161 total courses of topotecan through five dose levels (Table 1). One course involving a 44-year-old female with non-small-cell lung carcinoma who had progressive dyspnea and died sudTable 1. Topotecan Dose Escalations Dose (mg/m'
New
Total
Assessable
daily x 5 days)
Patients
New Patients Subsequently Escalated
Reduced
Patients
Courses
0.5 1.0 1.5 2.0 2.5 Total
3 6 10 6 4 29
3 3 0 0 0
0 0 2 1 3
3 11 15 11 4
6 32 72 46 4 160
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ROWINSKY ET AL
denly 24 hours after receiving topotecan during her third course. She was not fully assessable. Postmortem examination showed extensive intrathoracic tumor and no evidence of drug toxicity. The median number of courses administered per patient was three and ranged from one to 21 as of November 1, 1991 (one course, one patient; two, 11 patients; three, six patients; four, two patients; five, one patient; six, one patient; seven, one patient; eight, one patient; 10, one patient; 16+, one patient; 19, one patient; 20+, one patient; 21+, one patient). Nine patients were treated at two doses; four patients had dose escalations due to negligible toxicity; and five patients had dose reductions due to profound neutropenia and fever. Two patients were treated at three doses due to neglible adverse effects at their two lower doses. Patient characteristics are listed in Table 2. Of the 29 patients, 28 had an ECOG performance status of either 0 or 1. All patients received cytotoxic therapies before entry onto study. Nineteen and 10 patients were considered minimally and heavily pretreated, respectively. Hematologic Toxicity Neutropenia was the principal dose-limiting toxicity of topotecan on this schedule. Table 3 lists the median and range of nadir ANCs, the grades of neutropenia, and the number of episodes of neutropenia and fever requiring hospitalization for parenteral antibiotics at each dose level. Infections were not documented in any of the five episodes of neutropenia and fever in this study. The onset of neutropenia typically occurred between days 8 and 10, and nadir ANCs were noted between days 8 and 12. More importantly, neutropenia Table 2. Patient Characteristics No. of patients Sex (male:female) Median age, years (range) ECOG performance status 0 1 2 Previous treatment Chemotherapy Radiotherapy Chemotherapy + radiotherapy Disease site Colorectal NSC lung Ovarian Unknown primary (adeno) Breast Melanoma Head and neck
29 13:16 51 (35-76) 13 15 1 20 2 7 14 7 3 2 1 1 1
Abbreviations: NSC, non-small-cell; adeno, adenocarcinoma.
resolved by day 22 in 91% of courses. Treatment delays of 1 week were required in 13 courses, and a delay of 2 weeks was required in one course due to unresolved neutropenia. Three episodes of brief grade 3 neutropenia were experienced by two heavily pretreated patients who received a total of 32 courses at the 1.0-mg/m 2 dose level; however, significant and consistent myelosuppression did not occur until the 1.5-mg/m 2 dose level. At 1.5 mg/m 2, grade 3 and 4 neutropenia occurred in 58% and 17% of courses, respectively. For minimally pretreated patients, grade 3 neutropenia was noted by 89% of patients in 67% of courses and grade 4 neutropenia was noted in 11% of patients in 9% of courses. For heavily pretreated patients, grade 3 neutropenia was observed in 33% of patients and 51% of courses; grade 4 neutropenia occurred in 67% of patients during 23% of courses. Although the median ANC nadir was lower for heavily pretreated than minimally pretreated patients receiving 1.5 mg/m2, the difference was not substantial (693 v 792 cells per microliter), and only one heavily pretreated patient developed neutropenia and fever. At the 2.0-mg/m2 dose level, a higher proportion of courses were associated with grade 4 neutropenia. Grade 3 neutropenia was experienced by 54% of minimally pretreated patients in 35% of courses, and grade 4 neutropenia was noted in 36% of patients during 35% of courses. One heavily pretreated patient experienced grade 3 and 4 neutropenia during 50% and 50% of courses, respectively. At 2.0 mg/m2, median nadir ANCs were 695 and 499 cells per microliter for minimally and heavily pretreated patients, respectively, and neutropenia and fever requiring parenteral antibiotics occurred in only one course in each of the minimally and heavily pretreated cohorts. Neutropenia was severe in three of the four patients who received topotecan at the 2.5mg/m 2 dose level, with ANC nadirs of 128, 161, 224, and 2,380/ tL. At 2.5 mg/m 2, treatment delays due to unresolved neutropenia were required in two of the four patients, and fever and neutropenia requiring antibiotics occurred in two of four courses. Although the percentages of total courses associated with grade 3 and 4 neutropenia at 1.5 mg/m 2 (75%) and 2.0 mg/m 2 (74%) were relatively high, both dose levels were well tolerated by the majority of patients due to the brevity of the severe neutropenia, the low percentage of courses requiring subsequent treatment delays (9%), the low incidence of febrile episodes (3%), and the low frequency of serious nonhematologic effects. There was no evidence of a cumulative effect of topotecan on the nadir ANCs in both minimally and heavily pretreated
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651
PHASE I AND PHARMACOLOGIC STUDY OF TOPOTECAN Table 3. Topotecan: Neutropenia
0.5
Grade
Median Nadir ANC (range)
No. of Courses
Dose (mg/m2)
0
1
2
3
4
6
4,550 (2,850-6,396)
6
1.0 Minimally pretreated Heavily pretreated
32 20 12
1,759 (612-6,622) 1,782 (1,127-6,622) 1,275 (612-2,088)
11 9 2
8 7 1
10 4 6
3
1.5 Minimally pretreated Heavily pretreated
72 33 39
740 (141-2,415) 792 (304-1,809) 693 (141-2,415)
1
3
1
3
14 8 6
42 22 20
12* 3 9
2.0 Minimally pretreated Heavily pretreated
46 40 6
681 (73-1,690) 695 (73-1,690) 499 (300-528)
1 1
11 11
17 14 3
17* 14 3
2.5 Minimally pretreated
4
276 (128-2,380)
3
1
3*
*Five courses at 1.5 mg/m2 (one), 2.0 mg/m' (two), and 2.5 mg/m2 (two) associated with fever and neutropenia requiring hospitalization and antibiotics.
patients. To illustrate the lack of cumulative neutropenia from topotecan, the mean percent decreases in ANCs in patients receiving more than one course of topotecan at the same dose are depicted as a function of the cumulative number of courses in Fig 2. Topotecan induced negligible effects on platelets. Nadir platelet counts were generally observed on day 15; however, one, five, and four courses at 1.0, 1.5, and 2.0 mg/m 2, respectively, were associated with grade 2 thrombocytopenia, and only one course in the study (2.5 mg/m 2) was associated with grade 3 thrombocytopenia (nadir platelet count, 45,000/liL). In contrast, topotecan's effects on RBCs were observed frequently. A 1007
900 z
80-
