Phase I Safety, Pharmacokinetic, and Pharmacodynamic Study of the Thrombospondin-1–Mimetic Angiogenesis Inhibitor ABT-510 in Patients With A
http://www.100md.com
《临床肿瘤学》
the Department of Medical Oncology, Erasmus MC, University Medical Center, Rotterdam
Department of Medical Oncology and Pulmonology, University Hospital Groningen, Groningen, the Netherlands
Abbott Laboratories, Chicago, IL.
ABSTRACT
PURPOSE: ABT-510 is an angiogenesis inhibitor derived from thrombospondin-1, a naturally occurring inhibitor of angiogenesis. We investigated ABT-510, which was administered subcutaneously in patients with advanced solid malignancies, to assess safety, pharmacokinetics, and serum markers of angiogenesis.
PATIENTS AND METHODS: ABT-510 was administered subcutaneously as a continuous infusion (100 mg/24 h) and bolus injections (100, 200, and 260 mg once daily; 50 and 100 mg twice daily) in 28-day cycles.
RESULTS: Thirty-nine patients received a total of 144 treatment cycles. Administration by continuous infusion was hampered by the onset of painful skin infiltrates at the injection site. In the bolus injection regimens, the most common toxicities observed were mild injection-site reactions and fatigue. Maximum-tolerated dose was not defined, but 260 mg was defined as the maximum clinically practical dose. ABT-510 pharmacokinetics were linear across the dosage ranges tested, and the potential therapeutic threshold (plasma concentrations > 100 ng/mL > 3 h/d) was achieved with all dose regimens. Median serum basic fibroblast growth factor (bFGF) levels decreased from 14.1 pg/mL (range, 0.5 to 77.7 pg/mL) at baseline to 3.2 pg/mL (range, 0.2 to 29.4 pg/mL) after 56 days of treatment (P = .003). No correlations with time on study or ABT-510 dose or exposure were observed for individual changes in bFGF. Stable disease lasting for six cycles or more was seen in six patients.
CONCLUSION: ABT-510 demonstrated a favorable toxicity profile and linear and time-independent pharmacokinetics with biologically relevant plasma concentrations. The significant number of patients with prolonged stable disease and the convenient method of dosing merit further studies with this angiogenesis inhibitor.
INTRODUCTION
Cancer progression is characterized by cell growth, tissue invasion, and metastasis. Angiogenesis is essential for these processes. Thrombospondin-1 (TSP-1) is a large adhesive glycoprotein that is activated by the tumor suppressor gene p53 and has an inhibitory effect on angiogenesis.1,2 It inhibits the activity of multiple pro-angiogenic factors including vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), and interleukin-8 (IL-8).3 The antiangiogenic activity of TSP-1 depends on activation of p59fyn and p38MAPK through interaction with the CD36 receptor or related proteins. Endothelial cell apoptosis induced by TSP-1 is mediated by the Fas/Fas ligand interaction and is accompanied by activation of caspase-3 and FAK fragmentation.4-6
The angiogenesis-inhibiting activity of TSP-1 has been mapped to the 50,000-dalton N-terminal third of the molecule and, more specifically, to the properdin type-1 repeats in this region. A single D-amino acid replacement in one properdin-region heptapeptide leads to a 1,000-fold increase of angiogenesis-inhibitory activity in preclinical models.3 ABT-510 is a nonapeptide analog of this active heptapeptide that mimics the natural angiogenesis-inhibiting activity of TSP-1 (Fig 1). ABT-510 competes with TSP-1 for binding to the endothelial cells, induces Fas ligand expression in endothelial cells, and inhibits VEGF- and bFGF-stimulated migration of human microvascular endothelial cells.
In vivo, ABT-510 inhibits VEGF-induced corneal neovascularization in mice and inhibits tumor growth in several mouse xenograft models in a dose-dependent manner. ABT-510 also inhibits B16 F10 melanoma lung metastases formation.7 Studies in rats, with ABT-510 administered intravenously daily for 1 week, revealed renal dose-limiting toxicity (DLT) at a dose of 75 mg/kg/d and a dose with no adverse effects of 25 mg/kg/d. In monkeys, doses up to 75 mg/kg/d intravenously for 1 month were tolerated without DLTs.7 Preclinical studies indicate that cytochrome P-450 enzymes are not involved in metabolism of ABT-510 and that ABT-510 has no inhibitory effects on cytochrome P-450 enzymes. ABT-510 is metabolized primarily by cleavage of peptide bonds to form M-1, M-2, and M-3 peptides, which are mainly excreted in bile and urine (Fig 1). In vivo, M-1, a five–amino acid peptide formed by peptide bond hydrolysis between threonine-5 and norvaline-6, predominates. ABT-510 is not extensively bound to plasma proteins.
Modeling of pharmacokinetic and pharmacodynamic data from preclinical experiments in 11 different murine models was performed to identify a pharmacokinetic target for clinical studies. Efficacy measures varied by model and included tumor volume, number of metastases, and VEGF- or bFGF-stimulated new vessel density. As a result, the clinical pharmacokinetic target was aimed at reaching plasma concentrations greater than 100 ng/mL for at least 3 hours per day, which, on average, achieved 75% of maximum efficacy in these models.8
We performed a phase I safety, pharmacokinetic, and pharmacodynamic study with ABT-510 administered subcutaneously to patients with advanced solid malignancies. The principal objectives of this study were to determine single- and multiple-dose plasma pharmacokinetics, with special emphasis on correlation with preclinical pharmacologic data, and to establish the safety profile and determine the maximum-tolerated dose (MTD) of ABT-510 when administered by subcutaneous continuous infusion and by subcutaneous injection once daily or twice daily for 28 consecutive days.
PATIENTS AND METHODS
Eligibility Criteria
Patients with a histologically confirmed diagnosis of an advanced solid malignancy refractory for standard therapy were eligible. Additional eligibility criteria included the following: age 18 years; Eastern Cooperation Oncology Group performance status less than 3; an estimated life expectancy of 3 months; and no radiotherapy, chemotherapy, or hormonal therapy within 4 weeks before study start, with the exception of small-field radiation. Specific exclusion criteria included a known human immunodeficiency virus–positive status, a diagnosis of primary brain tumor or known CNS metastases, and evidence of uncontrolled clinically significant disease unrelated to the primary malignancy. The study was approved by the local ethics boards of the two participating centers, and all patients gave written informed consent.
Drug Administration
ABT-510 (Abbott Laboratories, Chicago, IL) was supplied in vials containing ABT-510 1.1 mL, with 100 mg/mL dissolved in dextrose 5%. The vials were stored at 2 to 8°C and brought to room temperature 1 hour before dosing. The following three methods of drug administration were studied: subcutaneous continuous infusion and once-daily and twice-daily subcutaneous bolus injection. For continuous infusion, a MiniMed 470C micro-infusion pump (MiniMed, Sylmar, CA) was used. This pump is specifically developed for the subcutaneous delivery of liquid medications. It uses syringes with a maximal capacity of 3 mL and can deliver volumes from 0 to 0.350 mL/h in 0.001-mL increments, with an accuracy of ± 2%. Patients used the abdominal wall as the injection site and changed the injection site every 3 days. The reservoir was changed once daily, preferably in the morning. Patients in the subcutaneous bolus regimen cohorts injected themselves in the abdominal wall. The maximum volume of one injection was set at 1.3 mL, with a maximum of two injections a day; this accounted for a maximum clinically practical daily dose of 260 mg. Patients in the group administering bolus injections once daily were instructed to inject themselves in the morning, preferably at the same time each day, and patients in the group administering bolus injections twice daily were instructed to inject themselves in the morning and evening, with an interval of 12 hours in between doses. Times of injection were recorded in a diary. The starting doses were 100 mg every 24 hours as a continuous infusion or bolus injections of 100 mg once daily or 50 mg twice daily, based on safety and pharmacokinetic data obtained in the phase I study of healthy volunteers.7 No adjustments for body-surface area or weight were made. Medication was administered daily without interruption; 28 days of treatment defined a treatment cycle.
Cohorts of three to six patients were studied. Dose-escalation decisions were made after review of observed toxicities and pharmacokinetic data and discussion between investigators and sponsor. Toxicity was assessed according to National Cancer Institute Common Toxicity Criteria version 2.0. Escalations were pursued until either the MTD was identified or the maximum clinically practical daily dose of 260 mg, as defined earlier, was reached. The MTD was defined as the highest dose of ABT-510 administered for at least one treatment cycle at which no more than one of six patients experienced DLT. A DLT was defined as a grade 3 or 4 adverse event (except inadequately treated nausea or vomiting) or any grade 2 adverse event requiring dose modification or treatment delay occurring in the first treatment cycle. Once the MTD was determined or the maximum clinically practical dose of 260 mg daily was reached for each schedule, the cohort size would be expanded up to a total of 10 patients.
Pretreatment and Follow-Up Studies
Before therapy, a complete medical history was taken, and a physical examination, ECG, and chest x-ray were performed. The following were performed at baseline and at each scheduled visit: CBC, including WBC differential; serum biochemistry, including sodium, potassium, chloride, magnesium, bicarbonate, creatinine, blood urea nitrogen, albumin, total protein, AST, ALT, lactate dehydrogenase, total bilirubin, calcium, phosphate, glucose, alkaline phosphatase, and amylase; prothrombin time; activated partial thromboplastin time (aPTT); and urinalysis. Plasminogen, fibrinogen, and factor VIII were collected at baseline and as clinically indicated thereafter. Urine was collected over 24 hours at baseline and on day 22 for albumin excretion. Weekly evaluations during the first treatment cycle included history, physical examination, toxicity assessment, CBC, prothrombin time, aPTT, and serum biochemistry. The same evaluations were also performed after 2 months and every 3 months thereafter. Additional visits were allowed at the discretion of the responsible physicians. Tumor measurements were performed after two cycles and then after every three cycles thereafter. Tumor response was assessed using WHO criteria.9 Patients were allowed to continue treatment in the absence of progressive disease or unacceptable toxicity.
Pharmacokinetic Sampling and Assays
Blood samples (5 mL) for pharmacokinetic analysis were collected from the continuous infusion patients using an indwelling intravenous canula before switching the reservoir and 0.5, 1, 2, 4, 6, 8, and 10 hours after the reservoir changes on days 1 and 22 of the first cycle. For the bolus injection patients, blood samples were collected before dosing and 5, 15, and 30 minutes and 1, 2, 4, 6, 8, and 10 hours after the morning dose on days 1 and 22 of the first cycle. Samples were collected in EDTA-containing tubes and then placed on ice and centrifuged at 2,000 x g for 10 minutes within 1 hour using a refrigerated (4°C) centrifuge. Plasma was stored in polypropylene tubes at –20°C until analysis. Plasma concentrations of ABT-510 and its major metabolite M-1 were determined using a validated liquid chromatography with mass spectrometry method. Sample preparation involved a plasma aliquot supplemented with internal standard (ABT-818) and solid-phase extraction with SPEC C18AR discs in a 96-well plate format (Varian, Lake Forest, CA). Extracts were injected via autosampler into a BHK ODS-W/S C18 5 μm 100 x 3.0 mm analytic column (BHK Laboratories Inc, Naperville, IL), with mobile phase of acetonitrile/0.1% formic acid in water (pH 4.0) at a flow rate of 0.3 mL/min. Analysis was performed on an API III+ mass spectrometer (Perkin Elmer Sciex, Concord, Ontario, Canada), with positive ionization using the Turbo IonSpray source. Multiple reaction monitoring with the following nominal transitions, in chromatographic elution order, was m/z (mass-charge ratio) 502 72 for ABT-388 (M-1 metabolite), m/z 1210 249 for internal standard, and m/z 995 270 for ABT-510. Peak areas were integrated using MacQuan v1.6 (Perkin Elmer Sciex). Watson v6.2.0.02 LIMS (PSS, Inc, Wayne, PA) was used for regression (weighted 1/x2) and quantitation. The lower limit of quantitation in plasma was approximately 0.5 ng/mL for ABT-510 and 3 ng/mL for the M-1 metabolite.
Urine was collected on days 1 and 22 of the first cycle in the bolus injection cohorts only. Two samples of at least 15 mL were collected immediately before dosing on day 1 for the baseline drug assay. Urine was collected over 24 hours after dosing on days 1 and 22 for the patients dosed once daily and from 0 to 12 and 12 to 24 hours after dosing on days 1 and 22 for the patients dosed twice daily. The urine samples were refrigerated during the collection period, and total volumes were measured. Two 15-mL samples from each sampling period were stored at –20°C until analysis. For measurement of urine concentrations of ABT-510 and its major metabolite M-1, 200 μL of urine was combined with internal standard and diluted. In addition, a trapping column (20 x 2.0 mm, 5 μm; ODS-W/S C18; BHK Laboratories) was used on line for cleanup of the diluted urine samples. The analytes were eluted from the trapping column and onto the analytic column for chromatographic separation and introduction to the source of the mass spectrometer. The mass spectrometer detection and quantification were the same as for the plasma assay. The lower limits of quantification were approximately 6 ng/mL for ABT-510 and 99 ng/mL for the M-1 metabolite.
Pharmacodynamic Sampling and Assays
Blood samples (5 mL) for determination of VEGF, bFGF, and IL-8 levels were drawn at baseline, on day 22, at the end of cycle 2, after every three cycles thereafter, and at the final study visit. Samples were preferably drawn in the morning before the injection of ABT-510 or change of the reservoir. Samples were collected in Becton Dickinson Vacutainer serum separation tubes (Becton Dickinson, Mountain View, CA). After collection, samples were allowed to clot and were centrifuged within 30 minutes at 800 x g for 15 minutes. Serum was stored in polypropylene tubes at –20°C until analysis. At the same collection times, urine samples of at least 10 mL were collected. After collection, urine was stored in the refrigerator and, within 4 hours, the following protease inhibitors were added: aminoethyl benzenesulfonic acid 80 μg (Pefabloc SC; Pentafarm, AG, Basel, Switzerland), EDTA-sodium 200 μg, leupeptin 0.2 μg (Roche Molecular Biochemicals, Basel, Switzerland), and pepstatin 0.2 μg (Roche). Thereafter, the urine specimen was centrifuged at 4°C at 3,000 x g for 10 minutes. The supernatant was stored in polypropylene tubes at –20°C until analysis. Serum and urine VEGF, bFGF, and IL-8 were measured using commercially available kits (Human VEGF Quantikine Immunoassay Kit, Human FGF Basic Quantikine Immunoassay Kit, and Human IL-8/CXLX8 Quantikine ELISA Kit; R&D Systems Inc, Minneapolis, MN). The lower limits of quantitation in serum were 0.9, 0.22, and 0.5 pg/mL for VEGF, bFGF, and IL-8, respectively. The lower limits of quantitation in urine were 1.0 and 0.1 ng/g creatinine for VEFG and bFGF, respectively.
Pharmacokinetic Analysis
Noncompartmental methods were used to determine values of pharmacokinetic variables of ABT-510 after administration by continuous infusion or subcutaneous bolus injection using WINNonlin (Scientific Consultant, Apex, NC) -Pro, version 4.1 (Pharsight Corporation, Cary, NC). Maximum measured concentration, minimum measured concentration, and time of maximum observed concentration were calculated from the concentration-time curves. Additional parameters estimated were terminal elimination rate constant (?), the corresponding half-life (t1/2), apparent clearance, and apparent volume of distribution. The trapezoidal rule was used to calculate area under the blood concentration–time curve (AUC) to infinity on day 1 and AUC for dosing interval for day 22. The percentage of dose recovered in urine as ABT-510 and M-1 was calculated as the amount recovered in urine divided by the dose and multiplied by 100. The amount of M-1 recovered in urine was converted to the equivalent ABT-510 amount by multiplying by the ratio of molecular weights (994:501).
Statistics
The sample size was based on clinical justification and patient numbers historically used for testing of new antineoplastic compounds. In models for the analysis of safety data, dose was treated as a factor with discrete levels or as a continuous variable. The nonparametric Friedman's test was performed to compare pharmacodynamic data on days 1, 22, and 56. Two-sided P < .05 was considered significant. For the pharmacokinetic analysis, descriptive statistics of parameters were determined with a breakdown by regimen and dose level on days 1 and 22.
RESULTS
A total of 39 patients were enrolled onto six dosing cohorts. The patient characteristics are listed in Table 1. Four patients received ABT-510 by continuous infusion at a dose of 100 mg/24 h. The remaining 35 patients received ABT-510 by bolus injection (50 and 100 mg twice daily and 100, 200, and 260 mg once daily). A total of 144 cycles of ABT-510 were administered, with a median of two cycles per patient (range, one to 19 cycles; Table 2).
Toxicity
The incidence of the observed side effects, possibly or probably related to ABT-510, as a function of the dose and schedule are listed in Table 3. All patients in the continuous infusion cohort developed grade 2 skin infiltrates at the site of the ABT-510 infusion after 48 hours. These infiltrates consisted of erythema and edema of the skin with a maximum diameter of 5 cm. The infiltrates were sometimes painful and persisted for 7 to 21 days after discontinuation of therapy. A skin biopsy of one of these infiltrates revealed an influx of neutrophils around the smooth vascular endothelium without any signs of vasculitis. These skin infiltrates did not reoccur when the infusion site was changed daily instead of once every 3 days. On the basis of the observed skin reactions and the inconvenience of daily changing of infusion site, dosing by continuous infusion was discontinued. In the bolus injection cohorts, only mild to moderate (grade 1 and 2) skin reactions were observed, consisting of redness, slight edema, and sporadic pain at the injection site. These symptoms were of short duration and disappeared within a few minutes to 1 hour after the injection. Mild to moderate skin reactions and fatigue were the most common side effects observed with ABT-510 administration. There was no correlation between side effects observed and ABT-510 dose. MTD could not be defined in the bolus injection schedules. A total of 17 severe adverse events were reported in 10 different patients. Only three severe adverse events were considered to be possibly related to ABT-510 (a fatal intracranial hemorrhage, a transient ischemic attack, and new-onset diabetes mellitus).
The intracranial hemorrhage occurred in a 57-year-old male patient with non–small-cell lung cancer. He was hospitalized on day 4 of cycle 2 of treatment with ABT-510 100 mg once daily because of progressive headache, accompanied by dizziness and nausea. Neurologic examination revealed cerebellar ataxia, and magnetic resonance imaging evaluation showed cerebellar bleeding in a previously undetected cerebellar metastasis. The patient's medical history was positive for hypertension but negative for cardiovascular events, diabetes mellitus, hyperlipidemia, hemorrhagic diatheses, and anticoagulant medication. Laboratory evaluation did not indicate a coagulopathy. This patient died the next day. Although a causal relationship with ABT-510 cannot be ruled out, hemorrhage in brain metastases is considered not uncommon in non–small-cell lung cancer.10,11
The transient ischemic attack occurred in a 53-year-old woman with an advanced leiomyosarcoma receiving ABT-510 260 mg once daily. The patient had received extensive prior therapy. She experienced motor aphasia and facial nerve palsy at the clinic before dosing for pharmacokinetic sampling on day 22 of cycle 1. She had no history of cardiovascular events, hypertension, diabetes mellitus, or hyperlipidemia, but she used oral contraceptives. The brain magnetic resonance imaging scan and ECG were normal. Additional laboratory investigations revealed a platelet count of 709 x 109/L, slightly elevated D-dimers, and normal antithrombin III and aPTT. The elevated platelet count was already present at baseline. The patient was treated with an oral platelet aggregation-inhibiting agent and subcutaneous low molecular weight heparin. Because a relationship with ABT-510 could not be ruled out, this drug was discontinued. In the following months, no new cardiovascular events were observed. Both platelets and D-dimers remained elevated after discontinuation of ABT-510 and were considered related to the advanced malignancy.
A 54-year-old male patient with an advanced liposarcoma receiving ABT-510 100 mg twice daily was diagnosed with diabetes mellitus and grade 4 hyperglycemia on day 7 of cycle 1. The patient was known to have lung and bone metastases and an infiltrating mass in the pancreatic region. No other risk factors for diabetes mellitus were present. The patient was treated with subcutaneous insulin twice daily and continued receiving ABT-510 injections for two cycles. The diabetes was most likely caused by an infiltrating mass in the pancreatic region, although a relationship with ABT-510 could not be ruled out.
Pharmacokinetics
Plasma sampling for pharmacokinetic studies was performed on day 1 of the first cycle in 38 patients and on day 22 in 36 patients. Because plasma concentration data were not available at all time points from some subjects, a complete pharmacokinetic analysis could be performed using only 28 subjects on day 1 and 27 subjects on day 22. The values for the pharmacokinetic parameters after dosing with continuous infusion or subcutaneous bolus injections are listed in Table 4. The ABT-510 plasma concentration versus time curves on days 1 and 22 for all dose levels studied are shown in Figures 2 and 3, respectively.
The continuous infusion cohort achieved a steady-state ABT-510 concentration of 242 ± 19 ng/mL on day 22 (n = 4). After subcutaneous bolus injection, ABT-510 was rapidly absorbed and eliminated with overall (n = 47) values of 0.7 ± 0.3 hour for time of maximum observed concentration, 22.5 ± 6.5 L/h for apparent clearance, 36.8 ± 10.8 L for apparent volume of distribution, and 1.1 ± 0.2 hour for t1/2; these values appeared to be similar across all the bolus injection regimens and across days 1 and 22. ABT-510 plasma concentrations did not accumulate with the once-daily or twice-daily dosing regimens.
Pharmacokinetic evaluation of the major metabolite M-1 showed that the continuous infusion regimen produced a steady-state concentration of 273 ± 66 ng/mL on day 22 (n = 4). After subcutaneous bolus injections, the overall mean t1/2 of M-1 was 2.9 hours; this was similar across all dosing groups for both days 1 and 22. On average, 58% ± 19% (n = 10) of the ABT-510 dose was recovered in the urine as M-1, and approximately 1% was recovered as unchanged drug. The percentage of dose excreted in the urine as M-1 was similar after the morning and evening doses in the groups dosed twice daily. There was no consistent trend across doses or days in the percentage of dose excreted as M-1.
Pharmacodynamics
Serum bFGF, VEGF, and IL-8 results are listed in Table 5 and presented in Figures 4, 5, and 6. Median serum bFGF concentrations decreased between days 1 and 56 (P = .003), whereas median serum VEGF and IL-8 concentrations did not change significantly. The duration of treatment was not correlated with the change in serum concentrations of bFGF. Changes in serum concentrations of bFGF did not correlate with changes in serum concentrations of VEGF and IL-8, and the changes in serum concentrations of bFGF, VEGF, or IL-8 over time were not correlated across individuals with ABT-510 dose, maximum measured concentration, or AUC values.
Urine concentrations of bFGF and VEGF did not change significantly (n = 23) and did not correlate with changes in serum concentrations of bFGF. Median urine concentrations of bFGF on days 1 and 56 were 0.1 ng/g creatinine (range, 0.1 to 0.7 ng/g creatinine) and 0.2 ng/g creatinine (range, 0.1 to 0.8 ng/g creatinine; P = .99), respectively; and urine concentrations of VEGF on days 1 and 56 were 41 ng/g creatinine (range, 4 to 412 ng/g creatinine) and 48 ng/g creatinine (range, 1 to 222 ng/g creatinine; P = .32), respectively.
Antitumor Activity
There were no partial or complete responses observed. Stable disease lasting more than two cycles was observed in 13 patients, with six of the patients experiencing stable disease lasting six cycles. One female patient with a recurrent angiosarcoma on the upper left leg, who developed new metastatic skin lesions every week before the start of ABT-510, had a period of stable disease without development of new lesions for 10 cycles. One male patient with renal cell cancer, who had bone metastases and three new primary tumors and/or metastases in the other kidney, experienced stable disease for 17 cycles. One male patient with a myxoid chondrosarcoma and multiple lymph node and pulmonary metastases has been treated for 19 cycles without clinical and radiologic signs of progression. There was no apparent relationship between the occurrence of prolonged stable disease and the ABT-510 dose or treatment schedule.
DISCUSSION
This phase I, two-center, open-label, multiple dose–escalation study of ABT-510 is the first clinical study in cancer patients with an agent that mimics the naturally occurring angiogenesis inhibitor TSP-1. This study demonstrates that ABT-510 has linear, time-independent pharmacokinetics and a favorable toxicity profile.
In preclinical mouse tumor models, ABT-510 inhibits tumor growth and metastasis formation but does not induce tumor regression. Efficacious doses range from 0.1 to 200 mg/kg/d, depending on tumor model and mode of administration (intraperitoneal, subcutaneous bolus, or continuous infusion). A limited number of tumor regressions, along with prolonged disease stabilization, were observed in a study involving tumor-bearing companion dogs evaluating ABT-510 at doses of 0.5 mg/kg/d twice daily. Regressions were observed in dogs with soft tissue sarcomas, epithelial tumors, and lymphomas.12 These composite preclinical data suggest that ABT-510 would primarily exhibit cytostatic activity with occasional tumor responses. In addition, ABT-510 exhibited a favorable safety profile in preclinical efficacy and toxicology studies, with renal DLT being observed at a dose of 75 mg/kg/d in rats. Given the low likelihood of observing tumor responses with ABT-510, particularly in a phase I study population, and the potential that an MTD would not be defined or that the efficacious dose would be significantly lower than the MTD, potential biomarkers for antitumor activity were being looked for to assist in the selection of an optimal biologically active phase II dose.
A potential pharmacokinetic end point was defined as a plasma concentration exceeding 100 ng/mL for at least 3 hours per day because preclinical models have shown that this time over threshold produced 75% of maximally observed efficacy in 11 different models.8 This finding is consistent with in vitro studies that demonstrated that an exposure of more than 20 nmol/L (approximately 20 ng/mL) for approximately 4 hours is required to induce Fas ligand expression in endothelial cells (unpublished data). Increases in time over threshold beyond 3 hours and further increases in overall exposure (AUC) correlated with increased efficacy in some models. With all doses of ABT-510 administered by bolus subcutaneous injection, the pharmacokinetic target of 100 ng/mL for at least 3 hours per day was achieved (Table 4). On the basis of the concept of time over threshold, it is obvious that increasing the frequency of injections from once daily to twice daily results in more prolonged time above threshold than doubling the dose. The 100-mg twice-daily dose regimen, for example, results in approximately 11 hours above the threshold of 100 ng/mL compared with approximately 7 hours for the 200-mg once-daily dose regimen. For this reason, twice-daily administration is being recommended for evaluation in phase II studies. However, it remains to be seen whether time above threshold will correlate with clinical outcome in humans and can be used as a surrogate end point.
Dose escalation was halted at 260 mg/d once daily because this involved two injections of 1.3 mL, which was the predefined maximum volume and number of injections per day. Also, the incremental exposure over the target threshold projected for increased doses was determined to be of modest relative value. This defined maximum dose is relative because, even at this volume of injection, the toxicities generally were mild, and the local skin reactions were acceptable. On the basis of this study, the recommended phase II dose of ABT-510 is 100 mg twice daily subcutaneously, although additional evaluation of the dose-response effect (dose ranging) with ABT-510 might be of value.
Dose escalation in the 24-hour continuous subcutaneous infusion arm was discontinued at 100 mg/d because of the skin toxicity observed at the injection site in the four subjects treated with this regimen. Although the skin infiltrates were not dose limiting according to the defined criteria, further dose escalation was not considered feasible. The pharmacokinetic data from this dosing cohort also predicted drug exposure less than 100 ng/mL with a 50% dose reduction; therefore, no further administration using this dosing method was pursued.
The evaluation of ABT-510 effects on serum markers of angiogenesis, such as VEGF, bFGF, and IL-8, was performed in this phase I study strictly as an exploratory exercise to investigate the value of the serum markers as potential biomarkers. The evaluation of serum angiogenesis markers identified a significant decrease in median serum bFGF levels on day 56 compared with day 1. VEGF and IL-8 levels did not show significant changes. The cause of the observed bFGF decrease remains speculative. It could be related to direct effects of ABT-510 and/or changes in tumor status. However, the clinical significance of circulating bFGF in relation to tumor status is still controversial.13 The fact that no changes in VEGF levels were observed during treatment with ABT-510 is consistent with preclinical studies showing that inhibition of carcinogenesis and angiogenesis by TSP-1 was not caused by changes in VEGF expression, receptor binding, or receptor activation.14 Associations between bFGF, VEGF, and IL-8 and the angiogenic state in cancer patients are complex because this state is controlled by the angiogenic switch in such a way that predominance of inducers results in angiogenesis and predominance of inhibitors results in vascular quiescence.15 It is still far from clear whether serial measurements of these pro-angiogenic factors during treatment with an angiogenesis inhibitor will be useful as a marker of their activity. In previous studies with other angiogenesis inhibitors, monitoring of urinary or plasma VEGF and bFGF provided no significant information.16-19
No tumor regressions were observed among the patients treated with ABT-510 in this study; this was not unexpected given the preclinical efficacy profile. However, in a number of patients, prolonged stable disease was observed, and of these patients, six patients had stable disease for more than six cycles; tumor types included sarcoma (n = 2), renal cell carcinoma, cervix carcinoma, colorectal carcinoma, and germ cell tumor. Although from this uncontrolled trial setting it cannot be concluded whether this is a drug effect or a result of indolent growth patterns, these data potentially add to the fact that relevant plasma concentrations were achieved.
Currently, ABT-510 is being tested in phase II studies, either as single agent or in combination with cytotoxic chemotherapy, in patients with soft tissue sarcoma, renal cell cancer, lymphoma, and non–small-cell lung cancer. Several of these trials are randomized, comparing different doses of ABT-510. The absence of cytochrome P-450 interactions and the favorable safety profile make ABT-510 well suited for combination therapy, both with chemotherapeutic agents and with other antiangiogenic agents. Several preclinical models have demonstrated that both elevated VEGF expression and downregulation of TSP-1 are necessary for tumor angiogenesis.20,21 Blockade of pro-angiogenic signaling accompanied by simultaneous augmentation of a suppressed inhibitory signal may be an attractive approach to inhibit tumor angiogenesis. Preclinical studies evaluating this approach have been initiated. Phase I studies combining ABT-510 with several standard chemotherapy regimens (fluorouracil/leucovorin and cisplatin/gemcitabine) have recently been completed. The combinations seemed feasible, without pharmacokinetic interactions and without additional toxicity.22
Authors' Disclosures of Potential Conflicts of Interest
Althoguh all authors have completed the disclosure declaration, the following authors or their immediate family members have indicated a financial interest. No conflict exists for drugs or devices used in a study if they are not being evaluated as part of the investigation. For a detailed description of the disclosure categories, or for more information about ASCO's conflict of interest policy, please refer to the Author Disclosure Declaration and the Disclosures of Potential Conflicts of Interest section in Information for Contributors.
Acknowledgment
The authors would like to thank the following individuals for their participation in this clinical trial: Dan Daszkowski, Chetan Karyekar, Susan Glad-Anderson, and Leny van Doorn.
NOTES
Supported by Abbott Laboratories, Chicago, IL.
Presented in part at the 38th Annual Meeting of the American Society of Clinical Oncology, Orlando, FL, May 18-21, 2002.
Authors' disclosures of potential conflicts of interest are found at the end of this article.
REFERENCES
Lawler J: The structural and functional properties of thrombospondin. Blood 67:1197-1209, 1986
Dameron KM, Volpert OV, Tainsky MA, et al: Control of angiogenesis of fibroblasts by p53 regulation of thrombospondin-1. Science 265:1582-1584, 1994
Volpert OV, Tolsma SS, Pellerin S, et al: Inhibition of angiogenesis by thrombospondin-2. Biochem Biophys Res Commun 217:326-332, 1995
Jimenez B, Volpert OV, Crawford SE, et al: Signals leading to apoptosis dependent of neovascularization by thrombospondin-1. Nat Med 6:41-48, 2000
Crawford SE, Stellmach V, Murphey-Ullrich JE, et al: Thrombospondin-1 is a major activator of TGF-B1 in vivo. Cell 93:1159-1170, 1998
Volpert OV, Zaichuk T, Zhou W, et al: Inducer-stimulated Fas targets activated endothelium for destruction by anti-angiogenic thrombospondin-1 and pigment epithelium-derived factor. Nat Med 8:349-357, 2002
Abbott Laboratories: Information for Clinical Investigators ABT-510. Global Pharmaceutical Research and Development of Abbott Laboratories, Abbott Park, Illinois 60064-3500, USA (ed 4). Abbott Park, IL, Abbott Laboratories, 2002
Carr R, Marsh K, Schneider A, et al: Pharmacokinetic/pharmacodynamic relationships for the angiogenesis inhibitor ABT-510 in preclinical efficacy models. Eur J Cancer 38:S79, 2002 (abstr 250)
World Health Organization. WHO Handbook for Reporting Results of Cancer Treatment. WHO Offset Publication No. 4, Geneva, Switzerland, World Health Organization, 1979
Bitoh S, Hasegawa H, Ohtsuki H, et al: Cerebral neoplasms initially presenting with massive intracerebral hemorrhage. Surg Neurol 22:57-62, 1984
Satoh H, Yamashita YT, Isjikawa H, et al: Intracranial hemorrhage due to cerebral metastases of lung-cancer: A case report. Oncol Rep 6:371-372, 1999
Khanna C, Rusk T, Haviv F et al: Antiangiogenic thrombospondin-1 peptides result in regression of naturally occurring cancers in pet dogs. Proc Am Soc Clin Oncol 21:22a, 2002 (abstr 85)
Poon RTT, Fan ST, Wong J, et al: Clinical implications of circulating angiogenic factors in cancer patients. J Clin Oncol 19:1207-1225, 2001
Hawighorst T, Oura H, Streit M, et al: Thrombospondin-1 selectively inhibits early-stage carcinogenesis and angiogenesis but not tumor lymphangiogenesis and lymphatic metastasis in transgenic mice. Oncogene 21:7945-7956, 2002
Volpert OV: Modulation of endothelial cell survival by an inhibitor of angiogenesis thrombospondin-1: A dynamic balance. Cancer Metastasis Rev 19:87-92, 2000
Voest EE, Beerepoot LV, Groenewegen G, et al: Phase I trial of recombinant human angiostatin by twice-daily subcutaneous injection in patients with advanced cancer. Proc Am Soc Clin Oncol 21:81a, 2002 (abstr 322)
Eder JP, Supko JG, Clark JW, et al: Phase I clinical trial of recombinant human endostatin administered as a short intravenous infusion repeated daily. J Clin Oncol 20:3772-3784, 2002
Herbst RS, Hess KR, Tran HT, et al: Phase I study of recombinant human endostatin in patients with advanced solid tumors. J Clin Oncol 20:3792-3803, 2002
Thomas JP, Arzoomanian RZ, Alberti D, et al: Phase I pharmacokinetic and pharmacodynamic study of recombinant human endostatin in patients with advanced solid tumors. J Clin Oncol 21:223-231, 2003
Watnick R, Cheng Y, Rangarajan A, et al: Ras modulates Myc activity to repress thrombospondin-1 expression and increase tumor angiogenesis. Cancer Cell 3:219-321, 2003
Filleur S, Courtin A, Ait-Si-Ali S, et al: SiRNA-mediated inhibition of vascular endothelial growth factor severely limits tumor resistance to antiangiogenic thrombospondin-1 and slows tumor vascularization and growth. Cancer Res 63:3919-3922, 2003
Vos de FY, Hoekstra R, Eskens FALM et al: Dose-finding and pharmacokinetic study of ABT-510 with gemcitabine and cisplatin in patients with advanced cancer. Proc Am Soc Clin Oncol 22:214s, 2004 (abstr 3077)(Ronald Hoekstra, Filip Y.)
Department of Medical Oncology and Pulmonology, University Hospital Groningen, Groningen, the Netherlands
Abbott Laboratories, Chicago, IL.
ABSTRACT
PURPOSE: ABT-510 is an angiogenesis inhibitor derived from thrombospondin-1, a naturally occurring inhibitor of angiogenesis. We investigated ABT-510, which was administered subcutaneously in patients with advanced solid malignancies, to assess safety, pharmacokinetics, and serum markers of angiogenesis.
PATIENTS AND METHODS: ABT-510 was administered subcutaneously as a continuous infusion (100 mg/24 h) and bolus injections (100, 200, and 260 mg once daily; 50 and 100 mg twice daily) in 28-day cycles.
RESULTS: Thirty-nine patients received a total of 144 treatment cycles. Administration by continuous infusion was hampered by the onset of painful skin infiltrates at the injection site. In the bolus injection regimens, the most common toxicities observed were mild injection-site reactions and fatigue. Maximum-tolerated dose was not defined, but 260 mg was defined as the maximum clinically practical dose. ABT-510 pharmacokinetics were linear across the dosage ranges tested, and the potential therapeutic threshold (plasma concentrations > 100 ng/mL > 3 h/d) was achieved with all dose regimens. Median serum basic fibroblast growth factor (bFGF) levels decreased from 14.1 pg/mL (range, 0.5 to 77.7 pg/mL) at baseline to 3.2 pg/mL (range, 0.2 to 29.4 pg/mL) after 56 days of treatment (P = .003). No correlations with time on study or ABT-510 dose or exposure were observed for individual changes in bFGF. Stable disease lasting for six cycles or more was seen in six patients.
CONCLUSION: ABT-510 demonstrated a favorable toxicity profile and linear and time-independent pharmacokinetics with biologically relevant plasma concentrations. The significant number of patients with prolonged stable disease and the convenient method of dosing merit further studies with this angiogenesis inhibitor.
INTRODUCTION
Cancer progression is characterized by cell growth, tissue invasion, and metastasis. Angiogenesis is essential for these processes. Thrombospondin-1 (TSP-1) is a large adhesive glycoprotein that is activated by the tumor suppressor gene p53 and has an inhibitory effect on angiogenesis.1,2 It inhibits the activity of multiple pro-angiogenic factors including vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), and interleukin-8 (IL-8).3 The antiangiogenic activity of TSP-1 depends on activation of p59fyn and p38MAPK through interaction with the CD36 receptor or related proteins. Endothelial cell apoptosis induced by TSP-1 is mediated by the Fas/Fas ligand interaction and is accompanied by activation of caspase-3 and FAK fragmentation.4-6
The angiogenesis-inhibiting activity of TSP-1 has been mapped to the 50,000-dalton N-terminal third of the molecule and, more specifically, to the properdin type-1 repeats in this region. A single D-amino acid replacement in one properdin-region heptapeptide leads to a 1,000-fold increase of angiogenesis-inhibitory activity in preclinical models.3 ABT-510 is a nonapeptide analog of this active heptapeptide that mimics the natural angiogenesis-inhibiting activity of TSP-1 (Fig 1). ABT-510 competes with TSP-1 for binding to the endothelial cells, induces Fas ligand expression in endothelial cells, and inhibits VEGF- and bFGF-stimulated migration of human microvascular endothelial cells.
In vivo, ABT-510 inhibits VEGF-induced corneal neovascularization in mice and inhibits tumor growth in several mouse xenograft models in a dose-dependent manner. ABT-510 also inhibits B16 F10 melanoma lung metastases formation.7 Studies in rats, with ABT-510 administered intravenously daily for 1 week, revealed renal dose-limiting toxicity (DLT) at a dose of 75 mg/kg/d and a dose with no adverse effects of 25 mg/kg/d. In monkeys, doses up to 75 mg/kg/d intravenously for 1 month were tolerated without DLTs.7 Preclinical studies indicate that cytochrome P-450 enzymes are not involved in metabolism of ABT-510 and that ABT-510 has no inhibitory effects on cytochrome P-450 enzymes. ABT-510 is metabolized primarily by cleavage of peptide bonds to form M-1, M-2, and M-3 peptides, which are mainly excreted in bile and urine (Fig 1). In vivo, M-1, a five–amino acid peptide formed by peptide bond hydrolysis between threonine-5 and norvaline-6, predominates. ABT-510 is not extensively bound to plasma proteins.
Modeling of pharmacokinetic and pharmacodynamic data from preclinical experiments in 11 different murine models was performed to identify a pharmacokinetic target for clinical studies. Efficacy measures varied by model and included tumor volume, number of metastases, and VEGF- or bFGF-stimulated new vessel density. As a result, the clinical pharmacokinetic target was aimed at reaching plasma concentrations greater than 100 ng/mL for at least 3 hours per day, which, on average, achieved 75% of maximum efficacy in these models.8
We performed a phase I safety, pharmacokinetic, and pharmacodynamic study with ABT-510 administered subcutaneously to patients with advanced solid malignancies. The principal objectives of this study were to determine single- and multiple-dose plasma pharmacokinetics, with special emphasis on correlation with preclinical pharmacologic data, and to establish the safety profile and determine the maximum-tolerated dose (MTD) of ABT-510 when administered by subcutaneous continuous infusion and by subcutaneous injection once daily or twice daily for 28 consecutive days.
PATIENTS AND METHODS
Eligibility Criteria
Patients with a histologically confirmed diagnosis of an advanced solid malignancy refractory for standard therapy were eligible. Additional eligibility criteria included the following: age 18 years; Eastern Cooperation Oncology Group performance status less than 3; an estimated life expectancy of 3 months; and no radiotherapy, chemotherapy, or hormonal therapy within 4 weeks before study start, with the exception of small-field radiation. Specific exclusion criteria included a known human immunodeficiency virus–positive status, a diagnosis of primary brain tumor or known CNS metastases, and evidence of uncontrolled clinically significant disease unrelated to the primary malignancy. The study was approved by the local ethics boards of the two participating centers, and all patients gave written informed consent.
Drug Administration
ABT-510 (Abbott Laboratories, Chicago, IL) was supplied in vials containing ABT-510 1.1 mL, with 100 mg/mL dissolved in dextrose 5%. The vials were stored at 2 to 8°C and brought to room temperature 1 hour before dosing. The following three methods of drug administration were studied: subcutaneous continuous infusion and once-daily and twice-daily subcutaneous bolus injection. For continuous infusion, a MiniMed 470C micro-infusion pump (MiniMed, Sylmar, CA) was used. This pump is specifically developed for the subcutaneous delivery of liquid medications. It uses syringes with a maximal capacity of 3 mL and can deliver volumes from 0 to 0.350 mL/h in 0.001-mL increments, with an accuracy of ± 2%. Patients used the abdominal wall as the injection site and changed the injection site every 3 days. The reservoir was changed once daily, preferably in the morning. Patients in the subcutaneous bolus regimen cohorts injected themselves in the abdominal wall. The maximum volume of one injection was set at 1.3 mL, with a maximum of two injections a day; this accounted for a maximum clinically practical daily dose of 260 mg. Patients in the group administering bolus injections once daily were instructed to inject themselves in the morning, preferably at the same time each day, and patients in the group administering bolus injections twice daily were instructed to inject themselves in the morning and evening, with an interval of 12 hours in between doses. Times of injection were recorded in a diary. The starting doses were 100 mg every 24 hours as a continuous infusion or bolus injections of 100 mg once daily or 50 mg twice daily, based on safety and pharmacokinetic data obtained in the phase I study of healthy volunteers.7 No adjustments for body-surface area or weight were made. Medication was administered daily without interruption; 28 days of treatment defined a treatment cycle.
Cohorts of three to six patients were studied. Dose-escalation decisions were made after review of observed toxicities and pharmacokinetic data and discussion between investigators and sponsor. Toxicity was assessed according to National Cancer Institute Common Toxicity Criteria version 2.0. Escalations were pursued until either the MTD was identified or the maximum clinically practical daily dose of 260 mg, as defined earlier, was reached. The MTD was defined as the highest dose of ABT-510 administered for at least one treatment cycle at which no more than one of six patients experienced DLT. A DLT was defined as a grade 3 or 4 adverse event (except inadequately treated nausea or vomiting) or any grade 2 adverse event requiring dose modification or treatment delay occurring in the first treatment cycle. Once the MTD was determined or the maximum clinically practical dose of 260 mg daily was reached for each schedule, the cohort size would be expanded up to a total of 10 patients.
Pretreatment and Follow-Up Studies
Before therapy, a complete medical history was taken, and a physical examination, ECG, and chest x-ray were performed. The following were performed at baseline and at each scheduled visit: CBC, including WBC differential; serum biochemistry, including sodium, potassium, chloride, magnesium, bicarbonate, creatinine, blood urea nitrogen, albumin, total protein, AST, ALT, lactate dehydrogenase, total bilirubin, calcium, phosphate, glucose, alkaline phosphatase, and amylase; prothrombin time; activated partial thromboplastin time (aPTT); and urinalysis. Plasminogen, fibrinogen, and factor VIII were collected at baseline and as clinically indicated thereafter. Urine was collected over 24 hours at baseline and on day 22 for albumin excretion. Weekly evaluations during the first treatment cycle included history, physical examination, toxicity assessment, CBC, prothrombin time, aPTT, and serum biochemistry. The same evaluations were also performed after 2 months and every 3 months thereafter. Additional visits were allowed at the discretion of the responsible physicians. Tumor measurements were performed after two cycles and then after every three cycles thereafter. Tumor response was assessed using WHO criteria.9 Patients were allowed to continue treatment in the absence of progressive disease or unacceptable toxicity.
Pharmacokinetic Sampling and Assays
Blood samples (5 mL) for pharmacokinetic analysis were collected from the continuous infusion patients using an indwelling intravenous canula before switching the reservoir and 0.5, 1, 2, 4, 6, 8, and 10 hours after the reservoir changes on days 1 and 22 of the first cycle. For the bolus injection patients, blood samples were collected before dosing and 5, 15, and 30 minutes and 1, 2, 4, 6, 8, and 10 hours after the morning dose on days 1 and 22 of the first cycle. Samples were collected in EDTA-containing tubes and then placed on ice and centrifuged at 2,000 x g for 10 minutes within 1 hour using a refrigerated (4°C) centrifuge. Plasma was stored in polypropylene tubes at –20°C until analysis. Plasma concentrations of ABT-510 and its major metabolite M-1 were determined using a validated liquid chromatography with mass spectrometry method. Sample preparation involved a plasma aliquot supplemented with internal standard (ABT-818) and solid-phase extraction with SPEC C18AR discs in a 96-well plate format (Varian, Lake Forest, CA). Extracts were injected via autosampler into a BHK ODS-W/S C18 5 μm 100 x 3.0 mm analytic column (BHK Laboratories Inc, Naperville, IL), with mobile phase of acetonitrile/0.1% formic acid in water (pH 4.0) at a flow rate of 0.3 mL/min. Analysis was performed on an API III+ mass spectrometer (Perkin Elmer Sciex, Concord, Ontario, Canada), with positive ionization using the Turbo IonSpray source. Multiple reaction monitoring with the following nominal transitions, in chromatographic elution order, was m/z (mass-charge ratio) 502 72 for ABT-388 (M-1 metabolite), m/z 1210 249 for internal standard, and m/z 995 270 for ABT-510. Peak areas were integrated using MacQuan v1.6 (Perkin Elmer Sciex). Watson v6.2.0.02 LIMS (PSS, Inc, Wayne, PA) was used for regression (weighted 1/x2) and quantitation. The lower limit of quantitation in plasma was approximately 0.5 ng/mL for ABT-510 and 3 ng/mL for the M-1 metabolite.
Urine was collected on days 1 and 22 of the first cycle in the bolus injection cohorts only. Two samples of at least 15 mL were collected immediately before dosing on day 1 for the baseline drug assay. Urine was collected over 24 hours after dosing on days 1 and 22 for the patients dosed once daily and from 0 to 12 and 12 to 24 hours after dosing on days 1 and 22 for the patients dosed twice daily. The urine samples were refrigerated during the collection period, and total volumes were measured. Two 15-mL samples from each sampling period were stored at –20°C until analysis. For measurement of urine concentrations of ABT-510 and its major metabolite M-1, 200 μL of urine was combined with internal standard and diluted. In addition, a trapping column (20 x 2.0 mm, 5 μm; ODS-W/S C18; BHK Laboratories) was used on line for cleanup of the diluted urine samples. The analytes were eluted from the trapping column and onto the analytic column for chromatographic separation and introduction to the source of the mass spectrometer. The mass spectrometer detection and quantification were the same as for the plasma assay. The lower limits of quantification were approximately 6 ng/mL for ABT-510 and 99 ng/mL for the M-1 metabolite.
Pharmacodynamic Sampling and Assays
Blood samples (5 mL) for determination of VEGF, bFGF, and IL-8 levels were drawn at baseline, on day 22, at the end of cycle 2, after every three cycles thereafter, and at the final study visit. Samples were preferably drawn in the morning before the injection of ABT-510 or change of the reservoir. Samples were collected in Becton Dickinson Vacutainer serum separation tubes (Becton Dickinson, Mountain View, CA). After collection, samples were allowed to clot and were centrifuged within 30 minutes at 800 x g for 15 minutes. Serum was stored in polypropylene tubes at –20°C until analysis. At the same collection times, urine samples of at least 10 mL were collected. After collection, urine was stored in the refrigerator and, within 4 hours, the following protease inhibitors were added: aminoethyl benzenesulfonic acid 80 μg (Pefabloc SC; Pentafarm, AG, Basel, Switzerland), EDTA-sodium 200 μg, leupeptin 0.2 μg (Roche Molecular Biochemicals, Basel, Switzerland), and pepstatin 0.2 μg (Roche). Thereafter, the urine specimen was centrifuged at 4°C at 3,000 x g for 10 minutes. The supernatant was stored in polypropylene tubes at –20°C until analysis. Serum and urine VEGF, bFGF, and IL-8 were measured using commercially available kits (Human VEGF Quantikine Immunoassay Kit, Human FGF Basic Quantikine Immunoassay Kit, and Human IL-8/CXLX8 Quantikine ELISA Kit; R&D Systems Inc, Minneapolis, MN). The lower limits of quantitation in serum were 0.9, 0.22, and 0.5 pg/mL for VEGF, bFGF, and IL-8, respectively. The lower limits of quantitation in urine were 1.0 and 0.1 ng/g creatinine for VEFG and bFGF, respectively.
Pharmacokinetic Analysis
Noncompartmental methods were used to determine values of pharmacokinetic variables of ABT-510 after administration by continuous infusion or subcutaneous bolus injection using WINNonlin (Scientific Consultant, Apex, NC) -Pro, version 4.1 (Pharsight Corporation, Cary, NC). Maximum measured concentration, minimum measured concentration, and time of maximum observed concentration were calculated from the concentration-time curves. Additional parameters estimated were terminal elimination rate constant (?), the corresponding half-life (t1/2), apparent clearance, and apparent volume of distribution. The trapezoidal rule was used to calculate area under the blood concentration–time curve (AUC) to infinity on day 1 and AUC for dosing interval for day 22. The percentage of dose recovered in urine as ABT-510 and M-1 was calculated as the amount recovered in urine divided by the dose and multiplied by 100. The amount of M-1 recovered in urine was converted to the equivalent ABT-510 amount by multiplying by the ratio of molecular weights (994:501).
Statistics
The sample size was based on clinical justification and patient numbers historically used for testing of new antineoplastic compounds. In models for the analysis of safety data, dose was treated as a factor with discrete levels or as a continuous variable. The nonparametric Friedman's test was performed to compare pharmacodynamic data on days 1, 22, and 56. Two-sided P < .05 was considered significant. For the pharmacokinetic analysis, descriptive statistics of parameters were determined with a breakdown by regimen and dose level on days 1 and 22.
RESULTS
A total of 39 patients were enrolled onto six dosing cohorts. The patient characteristics are listed in Table 1. Four patients received ABT-510 by continuous infusion at a dose of 100 mg/24 h. The remaining 35 patients received ABT-510 by bolus injection (50 and 100 mg twice daily and 100, 200, and 260 mg once daily). A total of 144 cycles of ABT-510 were administered, with a median of two cycles per patient (range, one to 19 cycles; Table 2).
Toxicity
The incidence of the observed side effects, possibly or probably related to ABT-510, as a function of the dose and schedule are listed in Table 3. All patients in the continuous infusion cohort developed grade 2 skin infiltrates at the site of the ABT-510 infusion after 48 hours. These infiltrates consisted of erythema and edema of the skin with a maximum diameter of 5 cm. The infiltrates were sometimes painful and persisted for 7 to 21 days after discontinuation of therapy. A skin biopsy of one of these infiltrates revealed an influx of neutrophils around the smooth vascular endothelium without any signs of vasculitis. These skin infiltrates did not reoccur when the infusion site was changed daily instead of once every 3 days. On the basis of the observed skin reactions and the inconvenience of daily changing of infusion site, dosing by continuous infusion was discontinued. In the bolus injection cohorts, only mild to moderate (grade 1 and 2) skin reactions were observed, consisting of redness, slight edema, and sporadic pain at the injection site. These symptoms were of short duration and disappeared within a few minutes to 1 hour after the injection. Mild to moderate skin reactions and fatigue were the most common side effects observed with ABT-510 administration. There was no correlation between side effects observed and ABT-510 dose. MTD could not be defined in the bolus injection schedules. A total of 17 severe adverse events were reported in 10 different patients. Only three severe adverse events were considered to be possibly related to ABT-510 (a fatal intracranial hemorrhage, a transient ischemic attack, and new-onset diabetes mellitus).
The intracranial hemorrhage occurred in a 57-year-old male patient with non–small-cell lung cancer. He was hospitalized on day 4 of cycle 2 of treatment with ABT-510 100 mg once daily because of progressive headache, accompanied by dizziness and nausea. Neurologic examination revealed cerebellar ataxia, and magnetic resonance imaging evaluation showed cerebellar bleeding in a previously undetected cerebellar metastasis. The patient's medical history was positive for hypertension but negative for cardiovascular events, diabetes mellitus, hyperlipidemia, hemorrhagic diatheses, and anticoagulant medication. Laboratory evaluation did not indicate a coagulopathy. This patient died the next day. Although a causal relationship with ABT-510 cannot be ruled out, hemorrhage in brain metastases is considered not uncommon in non–small-cell lung cancer.10,11
The transient ischemic attack occurred in a 53-year-old woman with an advanced leiomyosarcoma receiving ABT-510 260 mg once daily. The patient had received extensive prior therapy. She experienced motor aphasia and facial nerve palsy at the clinic before dosing for pharmacokinetic sampling on day 22 of cycle 1. She had no history of cardiovascular events, hypertension, diabetes mellitus, or hyperlipidemia, but she used oral contraceptives. The brain magnetic resonance imaging scan and ECG were normal. Additional laboratory investigations revealed a platelet count of 709 x 109/L, slightly elevated D-dimers, and normal antithrombin III and aPTT. The elevated platelet count was already present at baseline. The patient was treated with an oral platelet aggregation-inhibiting agent and subcutaneous low molecular weight heparin. Because a relationship with ABT-510 could not be ruled out, this drug was discontinued. In the following months, no new cardiovascular events were observed. Both platelets and D-dimers remained elevated after discontinuation of ABT-510 and were considered related to the advanced malignancy.
A 54-year-old male patient with an advanced liposarcoma receiving ABT-510 100 mg twice daily was diagnosed with diabetes mellitus and grade 4 hyperglycemia on day 7 of cycle 1. The patient was known to have lung and bone metastases and an infiltrating mass in the pancreatic region. No other risk factors for diabetes mellitus were present. The patient was treated with subcutaneous insulin twice daily and continued receiving ABT-510 injections for two cycles. The diabetes was most likely caused by an infiltrating mass in the pancreatic region, although a relationship with ABT-510 could not be ruled out.
Pharmacokinetics
Plasma sampling for pharmacokinetic studies was performed on day 1 of the first cycle in 38 patients and on day 22 in 36 patients. Because plasma concentration data were not available at all time points from some subjects, a complete pharmacokinetic analysis could be performed using only 28 subjects on day 1 and 27 subjects on day 22. The values for the pharmacokinetic parameters after dosing with continuous infusion or subcutaneous bolus injections are listed in Table 4. The ABT-510 plasma concentration versus time curves on days 1 and 22 for all dose levels studied are shown in Figures 2 and 3, respectively.
The continuous infusion cohort achieved a steady-state ABT-510 concentration of 242 ± 19 ng/mL on day 22 (n = 4). After subcutaneous bolus injection, ABT-510 was rapidly absorbed and eliminated with overall (n = 47) values of 0.7 ± 0.3 hour for time of maximum observed concentration, 22.5 ± 6.5 L/h for apparent clearance, 36.8 ± 10.8 L for apparent volume of distribution, and 1.1 ± 0.2 hour for t1/2; these values appeared to be similar across all the bolus injection regimens and across days 1 and 22. ABT-510 plasma concentrations did not accumulate with the once-daily or twice-daily dosing regimens.
Pharmacokinetic evaluation of the major metabolite M-1 showed that the continuous infusion regimen produced a steady-state concentration of 273 ± 66 ng/mL on day 22 (n = 4). After subcutaneous bolus injections, the overall mean t1/2 of M-1 was 2.9 hours; this was similar across all dosing groups for both days 1 and 22. On average, 58% ± 19% (n = 10) of the ABT-510 dose was recovered in the urine as M-1, and approximately 1% was recovered as unchanged drug. The percentage of dose excreted in the urine as M-1 was similar after the morning and evening doses in the groups dosed twice daily. There was no consistent trend across doses or days in the percentage of dose excreted as M-1.
Pharmacodynamics
Serum bFGF, VEGF, and IL-8 results are listed in Table 5 and presented in Figures 4, 5, and 6. Median serum bFGF concentrations decreased between days 1 and 56 (P = .003), whereas median serum VEGF and IL-8 concentrations did not change significantly. The duration of treatment was not correlated with the change in serum concentrations of bFGF. Changes in serum concentrations of bFGF did not correlate with changes in serum concentrations of VEGF and IL-8, and the changes in serum concentrations of bFGF, VEGF, or IL-8 over time were not correlated across individuals with ABT-510 dose, maximum measured concentration, or AUC values.
Urine concentrations of bFGF and VEGF did not change significantly (n = 23) and did not correlate with changes in serum concentrations of bFGF. Median urine concentrations of bFGF on days 1 and 56 were 0.1 ng/g creatinine (range, 0.1 to 0.7 ng/g creatinine) and 0.2 ng/g creatinine (range, 0.1 to 0.8 ng/g creatinine; P = .99), respectively; and urine concentrations of VEGF on days 1 and 56 were 41 ng/g creatinine (range, 4 to 412 ng/g creatinine) and 48 ng/g creatinine (range, 1 to 222 ng/g creatinine; P = .32), respectively.
Antitumor Activity
There were no partial or complete responses observed. Stable disease lasting more than two cycles was observed in 13 patients, with six of the patients experiencing stable disease lasting six cycles. One female patient with a recurrent angiosarcoma on the upper left leg, who developed new metastatic skin lesions every week before the start of ABT-510, had a period of stable disease without development of new lesions for 10 cycles. One male patient with renal cell cancer, who had bone metastases and three new primary tumors and/or metastases in the other kidney, experienced stable disease for 17 cycles. One male patient with a myxoid chondrosarcoma and multiple lymph node and pulmonary metastases has been treated for 19 cycles without clinical and radiologic signs of progression. There was no apparent relationship between the occurrence of prolonged stable disease and the ABT-510 dose or treatment schedule.
DISCUSSION
This phase I, two-center, open-label, multiple dose–escalation study of ABT-510 is the first clinical study in cancer patients with an agent that mimics the naturally occurring angiogenesis inhibitor TSP-1. This study demonstrates that ABT-510 has linear, time-independent pharmacokinetics and a favorable toxicity profile.
In preclinical mouse tumor models, ABT-510 inhibits tumor growth and metastasis formation but does not induce tumor regression. Efficacious doses range from 0.1 to 200 mg/kg/d, depending on tumor model and mode of administration (intraperitoneal, subcutaneous bolus, or continuous infusion). A limited number of tumor regressions, along with prolonged disease stabilization, were observed in a study involving tumor-bearing companion dogs evaluating ABT-510 at doses of 0.5 mg/kg/d twice daily. Regressions were observed in dogs with soft tissue sarcomas, epithelial tumors, and lymphomas.12 These composite preclinical data suggest that ABT-510 would primarily exhibit cytostatic activity with occasional tumor responses. In addition, ABT-510 exhibited a favorable safety profile in preclinical efficacy and toxicology studies, with renal DLT being observed at a dose of 75 mg/kg/d in rats. Given the low likelihood of observing tumor responses with ABT-510, particularly in a phase I study population, and the potential that an MTD would not be defined or that the efficacious dose would be significantly lower than the MTD, potential biomarkers for antitumor activity were being looked for to assist in the selection of an optimal biologically active phase II dose.
A potential pharmacokinetic end point was defined as a plasma concentration exceeding 100 ng/mL for at least 3 hours per day because preclinical models have shown that this time over threshold produced 75% of maximally observed efficacy in 11 different models.8 This finding is consistent with in vitro studies that demonstrated that an exposure of more than 20 nmol/L (approximately 20 ng/mL) for approximately 4 hours is required to induce Fas ligand expression in endothelial cells (unpublished data). Increases in time over threshold beyond 3 hours and further increases in overall exposure (AUC) correlated with increased efficacy in some models. With all doses of ABT-510 administered by bolus subcutaneous injection, the pharmacokinetic target of 100 ng/mL for at least 3 hours per day was achieved (Table 4). On the basis of the concept of time over threshold, it is obvious that increasing the frequency of injections from once daily to twice daily results in more prolonged time above threshold than doubling the dose. The 100-mg twice-daily dose regimen, for example, results in approximately 11 hours above the threshold of 100 ng/mL compared with approximately 7 hours for the 200-mg once-daily dose regimen. For this reason, twice-daily administration is being recommended for evaluation in phase II studies. However, it remains to be seen whether time above threshold will correlate with clinical outcome in humans and can be used as a surrogate end point.
Dose escalation was halted at 260 mg/d once daily because this involved two injections of 1.3 mL, which was the predefined maximum volume and number of injections per day. Also, the incremental exposure over the target threshold projected for increased doses was determined to be of modest relative value. This defined maximum dose is relative because, even at this volume of injection, the toxicities generally were mild, and the local skin reactions were acceptable. On the basis of this study, the recommended phase II dose of ABT-510 is 100 mg twice daily subcutaneously, although additional evaluation of the dose-response effect (dose ranging) with ABT-510 might be of value.
Dose escalation in the 24-hour continuous subcutaneous infusion arm was discontinued at 100 mg/d because of the skin toxicity observed at the injection site in the four subjects treated with this regimen. Although the skin infiltrates were not dose limiting according to the defined criteria, further dose escalation was not considered feasible. The pharmacokinetic data from this dosing cohort also predicted drug exposure less than 100 ng/mL with a 50% dose reduction; therefore, no further administration using this dosing method was pursued.
The evaluation of ABT-510 effects on serum markers of angiogenesis, such as VEGF, bFGF, and IL-8, was performed in this phase I study strictly as an exploratory exercise to investigate the value of the serum markers as potential biomarkers. The evaluation of serum angiogenesis markers identified a significant decrease in median serum bFGF levels on day 56 compared with day 1. VEGF and IL-8 levels did not show significant changes. The cause of the observed bFGF decrease remains speculative. It could be related to direct effects of ABT-510 and/or changes in tumor status. However, the clinical significance of circulating bFGF in relation to tumor status is still controversial.13 The fact that no changes in VEGF levels were observed during treatment with ABT-510 is consistent with preclinical studies showing that inhibition of carcinogenesis and angiogenesis by TSP-1 was not caused by changes in VEGF expression, receptor binding, or receptor activation.14 Associations between bFGF, VEGF, and IL-8 and the angiogenic state in cancer patients are complex because this state is controlled by the angiogenic switch in such a way that predominance of inducers results in angiogenesis and predominance of inhibitors results in vascular quiescence.15 It is still far from clear whether serial measurements of these pro-angiogenic factors during treatment with an angiogenesis inhibitor will be useful as a marker of their activity. In previous studies with other angiogenesis inhibitors, monitoring of urinary or plasma VEGF and bFGF provided no significant information.16-19
No tumor regressions were observed among the patients treated with ABT-510 in this study; this was not unexpected given the preclinical efficacy profile. However, in a number of patients, prolonged stable disease was observed, and of these patients, six patients had stable disease for more than six cycles; tumor types included sarcoma (n = 2), renal cell carcinoma, cervix carcinoma, colorectal carcinoma, and germ cell tumor. Although from this uncontrolled trial setting it cannot be concluded whether this is a drug effect or a result of indolent growth patterns, these data potentially add to the fact that relevant plasma concentrations were achieved.
Currently, ABT-510 is being tested in phase II studies, either as single agent or in combination with cytotoxic chemotherapy, in patients with soft tissue sarcoma, renal cell cancer, lymphoma, and non–small-cell lung cancer. Several of these trials are randomized, comparing different doses of ABT-510. The absence of cytochrome P-450 interactions and the favorable safety profile make ABT-510 well suited for combination therapy, both with chemotherapeutic agents and with other antiangiogenic agents. Several preclinical models have demonstrated that both elevated VEGF expression and downregulation of TSP-1 are necessary for tumor angiogenesis.20,21 Blockade of pro-angiogenic signaling accompanied by simultaneous augmentation of a suppressed inhibitory signal may be an attractive approach to inhibit tumor angiogenesis. Preclinical studies evaluating this approach have been initiated. Phase I studies combining ABT-510 with several standard chemotherapy regimens (fluorouracil/leucovorin and cisplatin/gemcitabine) have recently been completed. The combinations seemed feasible, without pharmacokinetic interactions and without additional toxicity.22
Authors' Disclosures of Potential Conflicts of Interest
Althoguh all authors have completed the disclosure declaration, the following authors or their immediate family members have indicated a financial interest. No conflict exists for drugs or devices used in a study if they are not being evaluated as part of the investigation. For a detailed description of the disclosure categories, or for more information about ASCO's conflict of interest policy, please refer to the Author Disclosure Declaration and the Disclosures of Potential Conflicts of Interest section in Information for Contributors.
Acknowledgment
The authors would like to thank the following individuals for their participation in this clinical trial: Dan Daszkowski, Chetan Karyekar, Susan Glad-Anderson, and Leny van Doorn.
NOTES
Supported by Abbott Laboratories, Chicago, IL.
Presented in part at the 38th Annual Meeting of the American Society of Clinical Oncology, Orlando, FL, May 18-21, 2002.
Authors' disclosures of potential conflicts of interest are found at the end of this article.
REFERENCES
Lawler J: The structural and functional properties of thrombospondin. Blood 67:1197-1209, 1986
Dameron KM, Volpert OV, Tainsky MA, et al: Control of angiogenesis of fibroblasts by p53 regulation of thrombospondin-1. Science 265:1582-1584, 1994
Volpert OV, Tolsma SS, Pellerin S, et al: Inhibition of angiogenesis by thrombospondin-2. Biochem Biophys Res Commun 217:326-332, 1995
Jimenez B, Volpert OV, Crawford SE, et al: Signals leading to apoptosis dependent of neovascularization by thrombospondin-1. Nat Med 6:41-48, 2000
Crawford SE, Stellmach V, Murphey-Ullrich JE, et al: Thrombospondin-1 is a major activator of TGF-B1 in vivo. Cell 93:1159-1170, 1998
Volpert OV, Zaichuk T, Zhou W, et al: Inducer-stimulated Fas targets activated endothelium for destruction by anti-angiogenic thrombospondin-1 and pigment epithelium-derived factor. Nat Med 8:349-357, 2002
Abbott Laboratories: Information for Clinical Investigators ABT-510. Global Pharmaceutical Research and Development of Abbott Laboratories, Abbott Park, Illinois 60064-3500, USA (ed 4). Abbott Park, IL, Abbott Laboratories, 2002
Carr R, Marsh K, Schneider A, et al: Pharmacokinetic/pharmacodynamic relationships for the angiogenesis inhibitor ABT-510 in preclinical efficacy models. Eur J Cancer 38:S79, 2002 (abstr 250)
World Health Organization. WHO Handbook for Reporting Results of Cancer Treatment. WHO Offset Publication No. 4, Geneva, Switzerland, World Health Organization, 1979
Bitoh S, Hasegawa H, Ohtsuki H, et al: Cerebral neoplasms initially presenting with massive intracerebral hemorrhage. Surg Neurol 22:57-62, 1984
Satoh H, Yamashita YT, Isjikawa H, et al: Intracranial hemorrhage due to cerebral metastases of lung-cancer: A case report. Oncol Rep 6:371-372, 1999
Khanna C, Rusk T, Haviv F et al: Antiangiogenic thrombospondin-1 peptides result in regression of naturally occurring cancers in pet dogs. Proc Am Soc Clin Oncol 21:22a, 2002 (abstr 85)
Poon RTT, Fan ST, Wong J, et al: Clinical implications of circulating angiogenic factors in cancer patients. J Clin Oncol 19:1207-1225, 2001
Hawighorst T, Oura H, Streit M, et al: Thrombospondin-1 selectively inhibits early-stage carcinogenesis and angiogenesis but not tumor lymphangiogenesis and lymphatic metastasis in transgenic mice. Oncogene 21:7945-7956, 2002
Volpert OV: Modulation of endothelial cell survival by an inhibitor of angiogenesis thrombospondin-1: A dynamic balance. Cancer Metastasis Rev 19:87-92, 2000
Voest EE, Beerepoot LV, Groenewegen G, et al: Phase I trial of recombinant human angiostatin by twice-daily subcutaneous injection in patients with advanced cancer. Proc Am Soc Clin Oncol 21:81a, 2002 (abstr 322)
Eder JP, Supko JG, Clark JW, et al: Phase I clinical trial of recombinant human endostatin administered as a short intravenous infusion repeated daily. J Clin Oncol 20:3772-3784, 2002
Herbst RS, Hess KR, Tran HT, et al: Phase I study of recombinant human endostatin in patients with advanced solid tumors. J Clin Oncol 20:3792-3803, 2002
Thomas JP, Arzoomanian RZ, Alberti D, et al: Phase I pharmacokinetic and pharmacodynamic study of recombinant human endostatin in patients with advanced solid tumors. J Clin Oncol 21:223-231, 2003
Watnick R, Cheng Y, Rangarajan A, et al: Ras modulates Myc activity to repress thrombospondin-1 expression and increase tumor angiogenesis. Cancer Cell 3:219-321, 2003
Filleur S, Courtin A, Ait-Si-Ali S, et al: SiRNA-mediated inhibition of vascular endothelial growth factor severely limits tumor resistance to antiangiogenic thrombospondin-1 and slows tumor vascularization and growth. Cancer Res 63:3919-3922, 2003
Vos de FY, Hoekstra R, Eskens FALM et al: Dose-finding and pharmacokinetic study of ABT-510 with gemcitabine and cisplatin in patients with advanced cancer. Proc Am Soc Clin Oncol 22:214s, 2004 (abstr 3077)(Ronald Hoekstra, Filip Y.)