Phase I Trial of the Oral Antiangiogenesis Agent AG-013736 in Patients With Advanced Solid Tumors: Pharmacokinetic and Clinical Results
http://www.100md.com
《临床肿瘤学》
the University of California, San Francisco Comprehensive Cancer Center, San Francisco
Pfizer Global Research and Development, San Diego, CA
University of Wisconsin Comprehensive Cancer Center, Madison, WI
The University of Texas M.D. Anderson Cancer Center, Houston, TX
ABSTRACT
PURPOSE: We studied the safety, clinical activity, and pharmacokinetics (PK) of AG-013736, an oral receptor tyrosine kinase inhibitor of vascular endothelial cell growth factor, platelet-derived growth factor, and c-Kit, in patients with advanced cancer.
PATIENTS AND METHODS: Patients received fixed doses of AG-013736 orally in 28-day cycles. In the first cohort, patients initially received two single test doses of AG-013736 (10 and 30 mg); subsequent dosing was determined by individual PK parameters. Doses in subsequent cohorts were assigned by using a traditional dose-escalation/de-escalation rule based on observed toxicities in the current and previous cohorts. PK analysis included evaluation of the effect of food and antacid.
RESULTS: Thirty-six patients received AG-013736 at doses ranging from 5 to 30 mg by mouth twice daily. The dose-limiting toxicities observed included hypertension, hemoptysis, and stomatitis and were seen primarily at the higher dose levels. The observed hypertension was manageable with medication. Stomatitis was generally tolerable and managed by dose reduction or drug holidays. AG-013736 was absorbed rapidly, with peak plasma concentrations observed within 2 to 6 hours after dosing. The maximum-tolerated dose and recommended phase II dose of AG-013736 is 5 mg, twice daily, administered in the fasted state. No significant drug interaction with antacid was seen. There were three confirmed partial responses and other evidence of clinical activity.
CONCLUSION: In this study, we have demonstrated clinical activity and safety of AG-013736 in patients with advanced solid tumors and identified the dose for phase II testing. The unique phase I study design allowed early identification of important absorption and metabolic issues critical to phase II testing of this agent.
INTRODUCTION
Angiogenesis is necessary for the progression from benign to malignant tumors, as well as growth and metastases of malignant cells.1-7 Disruption of endothelial cell responses responsible for abnormal blood vessel formation may be used to stop tumor growth.8-11
A strong correlation has been observed between vascular endothelial cell growth factor (VEGF) expression and tumor microvessel density in multiple tumor types,12-14 supporting the role of VEGF in facilitating tumor angiogenesis15,16 through its receptors. Abrogation of signaling through both the VEGF receptor (VEGFR) and platelet-derived growth factor receptor-? (PDGFR-?) has been demonstrated to inhibit angiogenesis and tumor growth in preclinical models, with early supportive clinical results.17-20
AG-013736 is a substituted indazole derivative that was discovered by using a structure-based drug design. Mechanistically, AG-013736 is a potent small molecule tyrosine kinase inhibitor of all known VEGFRs at subnanomolar concentrations and PDGFR-? and c-Kit in low nanomolar concentrations. In vitro, AG-013736 selectively blocks VEGF-stimulated receptor autophosphorylation leading to inhibition of endothelial cell proliferation and survival. In mice, AG-013736 inhibited tumor vascular angiogenesis and the growth of human colorectal and murine lung tumors. This antitumor effect was associated with a significant decrease in microvessel density and increased tumor necrosis. In a transgenic mouse model of spontaneous islet cell tumors, treatment with AG-013736 eliminated endothelial fenestrations and suppressed vascular sprouting within 24 hours. At 7 days, vascular density decreased more than 70%, and significant tumor shrinkage was seen at 21 days.21
Given strong preclinical evidence for antitumor/antimetastatic activity, a first-in-human phase I trial was conducted to test AG-013736 in patients with advanced solid malignancies. The primary objective was to determine dose-limiting toxicity (DLT) and the maximum-tolerated dose (MTD) of AG-013736. Secondary objectives were to (1) evaluate the pharmacokinetics (PK) of oral AG-013736, (2) conduct a pilot evaluation of the effect of food on AG-013736 PK, (3) conduct a pilot evaluation of the effect of an antacid on the PK of AG-013736, and (4) document preliminary evidence of antitumor activity. Dynamic contrast-enhanced (DCE) imaging was performed to evaluate the association between PK and vascular response; these data are reported in a separate publication.22
PATIENTS AND METHODS
Patients were enrolled at the University of Wisconsin Comprehensive Cancer Center, University of Texas M.D. Anderson Cancer Center, and University of California, San Francisco Comprehensive Cancer Center between May 2002 and January 2004. All patients were 18 years of age with histologically confirmed advanced nonhematologic cancer refractory to standard therapy or for which no effective therapy was available. An Eastern Cooperative Group performance status of 0 to 2 and a life expectancy of 12 weeks were required. All patients had completed prior therapies with adequate time allowed for recovery from toxicity. Additional inclusion criteria included: adequate bone marrow, renal, and liver function (bilirubin 1.5 mg/dL, transaminases 2.5 x upper limit of normal or 5 x upper limit of normal with documented liver metastasis). No prior history of coronary artery disease, significant gastrointestinal abnormalities, or uncontrolled brain metastases was permitted. Patients with a positive stool guaiac test without evidence of active bleeding were included provided monitoring of the hemoglobin was performed during the trial. In addition, patients requiring the use of potent CYP3A4 inhibitors or CYP3A4/CYP1A2 inducers, chronic H2 antagonists or proton-pump inhibitors, or therapeutic anticoagulant therapy were not eligible. By amendment, patients with proteinuria 500 mg/24 hours, squamous cell lung cancer, centrally located lung lesions (of any histology), and uncontrolled hypertension (systolic > 140 mmHg, diastolic > 90 mmHg) were excluded because of observed toxicities.
Drug Administration and Study Design
AG-013736 was supplied as 1- and 10-mg film-coated tablets. AG-013736 was administered either once daily (QD) or BID, with no scheduled breaks. Each cycle of therapy was defined as 28 days.
Calculation of Starting Dose and Continuous Drug Dosing
Initial dosing was chosen based on a modification of the recommendations of DeGeorge et al23: one sixth of the body-surface area–adjusted minimal effect dose (MED; 10 mg/kg/d) identified in a 28-day dog study of AG-013736. This resulted in a calculated safe human starting dose of 30 mg BID. Cohorts of 6 patients were planned at each dose level. To obtain an early evaluation of in vivo PK and safety, patients in cohort 1 were initially treated with a single test dose of 10 mg, followed by a single test dose of 30 mg at least 48 hours later. The two single doses were selected to not exceed the projected starting dose and to provide an early assessment of PK dose proportionality.
Each patient in the first cohort was then assigned a continuous dose based on their single-dose PK to obtain steady-state plasma exposures similar to that associated with the MED in dogs. The specific continuous dose was estimated as the product of observed apparent plasma clearance and target MED exposure in dogs. The six patients in cohort one received doses between 10 mg QD and 30 mg BID.
Based on the overall safety profile of the first cohort, the average dose evaluated in the first cohort (20 mg BID) was selected as the continuous dose for the second cohort. In subsequent cohorts, escalation of doses by 100% or 40% was planned. However, because of the significant toxicity seen in the first two cohorts, a dose de-escalation schema was used.
Dose Reductions and Modifications
Dose reductions or interruptions of AG-013736 treatment were added by amendment for hypertension, proteinuria, and hemoptysis. Dose modifications for other toxicities were decided between the investigator and sponsor. Intrapatient dose escalation was not allowed. Treatment was discontinued for disease progression, unacceptable toxicity, or withdrawal of consent. Patients were considered to have completed the study after receiving eight cycles of treatment; however, they could continue to receive AG-013736 on a separate follow-on protocol for as long as they derived benefit.
Definition of DLT/MTD
DLT was defined as any of the following during the first cycle when considered related to AG-013736 treatment: select grade 2 or higher gastrointestinal toxicity despite the use of medical intervention and/or prophylaxis, grade 3 anemia, grade 4 thrombocytopenia, or grade 3 nonhematologic toxicities except nausea, vomiting, diarrhea, constipation, pain, and hypertension controlled with medication. The MTD was defined as the dose at which no more than one of six patients in a single cohort experienced DLT in the first cycle.
PK
Plasma samples were collected over 12 hours after dosing in all patients for evaluation of multiple-dose PK on days 1 and 15 (cycle 1) and day 29 (cycle 2, day 1). Additionally, predose trough samples were collected on days 43 (cycle 2, day 15) and 57 (cycle 3, day 1). Whole blood (5 mL) was collected in K3-EDTA vacutainer tubes at predose, 0.5, 1, 2, 4, 8, and 12 hours after dosing and centrifuged at 1,000 x g for 15 minutes, and plasma was frozen at –20°C. Analysis was conducted at Charles River Discovery and Development Services (Worcester, MA). Adequate precautions were taken to minimize degradation of AG-013736 in plasma/whole blood when exposed to visible light.
Assay for Measurement of AG-013736 in Human Plasma
Plasma concentrations of AG-013736 were measured by using a validated liquid chromatography-tandem mass spectrometric method (lower limit of quantitation, 0.1 ng/mL). Briefly, AG-013736 and the deuterated internal standard were extracted from plasma with an ethyl acetate/hexanes mixture. The extracts were evaporated to dryness, reconstituted in aqueous methanol mobile phase, and eluted from a reversed-phase column onto a triple-quadrupole mass spectrometer for detection.
Standard AG-013736 plasma PK parameters were estimated by using noncompartmental methods (using the WINNonlin Professional software; Scientific Consultant, Apex, NC), version 3.2. AG-013736 levels were also measured in the urine of patients.
Evaluation of Food Effect
Patients in cohorts two through four participated in a pilot food-effect study. On day 29, patients received their morning AG-013736 dose within 30 minutes of completion of a 1,000-calorie meal (50% caloric content from fat). On day 30, patients received their morning dose after an overnight fast of at least 6 hours. PK samples were collected for 12 hours on both days. Based on this evaluation, patients in the fifth and sixth cohorts were asked to fast for 2 hours before and after each AG-013736 dose.
Evaluation of Antacid Effect
Patients in cohort six participated in a pilot antacid-effect study to evaluate the effect of the proton-pump inhibitor rabeprazole (Aciphex) on AG-013736 PK. On days 30 to 34, patients received 20 mg of rabeprazole daily 3 hours before their morning AG-013736 dose. On day 34, PK samples were collected for 12 hours after the morning AG-013736 dose.
Pretreatment and Follow-Up Evaluations
Complete medical history, physical examination including vital signs, Eastern Cooperative Oncology Group performance status, full blood count, and serum chemistry and urinalysis were performed at baseline, weekly during the first cycle, and monthly thereafter. A 12-lead ECG and stool guaiac test were performed at baseline. A serum or urine pregnancy test was performed in women of childbearing potential. Toxicity was evaluated by clinical and laboratory examination and graded by using the National Cancer Institute Common Toxicity Criteria, version 2.0.24 After hypertension was identified as a DLT, all patients were required to monitor and record their blood pressures at least once daily. Radiological or physical assessment of tumors was done within 28 days before the first dose and then repeated every two cycles (every 8 weeks). Patients were evaluated for response by using standard Response Evaluation Criteria in Solid Tumors (RECIST).25
Additional Assessments
Plasma samples were collected to evaluate exploratory biomarkers related to the VEGFR pathway. In an effort to evaluate a possible renal etiology for the hypertension seen in patients treated with AG-013736, renin, angiotensin II, and aldosterone levels were measured in the plasma of patients from cohorts five and six receiving 5 mg of AG-013736 BID in the fasted state. Blood samples were collected before dosing on day 1 and then 4 hours after the morning AG-013736 dose on days 1, 15, and 29 (cycle 2, day 1).
RESULTS
Patient Characteristics
A total of 36 patients with advanced solid malignancies were treated with AG-013736 in this phase I trial. The patients received oral doses ranging from 5 mg BID to 30 mg BID. The first four cohorts were dosed in the fed state, and the last two cohorts were dosed in the fasted state (defined as no food or drink other than water in the 2 hours before and after AG-013736 dosing; Table 1) after PK data demonstrated improved absorption in the fasted state (see PK). Selected patient characteristics are summarized in Table 2. Twenty-six patients (72%) had received prior chemotherapy, and 18 patients (50%) had prior radiotherapy.
PK
AG-013736 administered in the fed state was absorbed rapidly, with peak plasma concentrations occurring within 2 to 6 hours after dosing (Table 3, Fig 1). Plasma concentrations declined with a terminal plasma half-life between 2 and 5 hours. AG-013736 plasma PK reached steady state within 15 days, with no unexpected accumulation. Patients in cohorts two through four demonstrated generally linear PK, as evidenced by dose-proportional increases in maximum plasma concentration (Cmax) and area under the curve (AUC; Fig 2).
At the phase II dose of 5 mg BID given in the fasted state, the between-patient coefficient of variation for AUC0-24 on cycle 1, day 15 was 90% (Table 3); the corresponding coefficient of variation for Cmax was 63%. A total of 9 of 16 patients in cohorts two through four completed the pilot food-effect study. Both the rate and extent of absorption were higher in the fasted state, as evidenced by higher Cmax, higher AUC0-24, and a shorter Tmax in the majority of patients. Peak concentrations occurred 1 to 2 hours after dosing in the fasted state, and there was a median 49% increase in plasma exposures compared with the fed state. Plasma half-life was not changed appreciably in fed and fasted states (Table 3, Fig 3). There was no difference in the effect of food across dose groups.
In the presence of the potent proton-pump inhibitor rabeprazole, the rate of absorption of AG-013736 was decreased (reduction in Cmax), but the extent of absorption was unaffected (marginal to negligible changes in the AUC; Fig 4). Rabeprazole was administered in this pilot evaluation for 5 days to maximize its gastric pH-lowering effect. Because of the minimal change in plasma exposure in the presence of rabeprazole, the effect of antacids on AG-013736 absorption was not considered to be clinically significant.
Twelve-hour urinary collections obtained on days 1 and 29 (cycle 2, day 1) demonstrated that < 1% of the administered dose appeared as unchanged drug in the urine regardless of dose, which indicates that the majority of drug elimination was through systemic metabolism.
Soluble plasma proteins (VEGF, fibroblast growth factor, tumor necrosis factor-alpha, and matrix metalloproteinases 2 and 9) evaluated in this study as exploratory markers related to VEGFR signal transduction pathways did not show any variation with treatment.
DLT and MTD
The primary DLT was hypertension. All DLTs are summarized in Table 4. Based on the experience of the first two cohorts, a dose of 20 mg BID exceeded the MTD. DLTs also occurred in patients who were treated in the first two cohorts with 10 mg BID; consequently, this dose was considered to be above MTD and was never tested in a separate cohort.
The MTD and recommended phase II dose of AG-013736 is 5 mg BID in the fasted state. Of 14 patients treated in two cohorts with this dose, two DLTs (grade 2 stomatitis and grade 3 diarrhea) were observed (Table 4).
Safety
Toxicities reported in at least 10% of patients and all grade 3 and 4 toxicities are summarized in Table 5. The principal toxicities were hypertension, fatigue, diarrhea, stomatitis, nausea, and vomiting. Of these, hypertension was reported the most frequently, occurring in 22 patients (61%). The majority of cases (18 patients) were controlled easily with antihypertensive medications. Of the remaining four patients, two developed grade 3 or 4 hypertension and withdrew from the study without attempting medical management. The other two patients developed grade 1/2 hypertension not requiring medication and withdrew from the study early because of disease progression.
The incidence and severity of hypertension was dose dependent. In the first two cohorts in which doses ranged from 10 mg QD to 30 mg BID, hypertension was observed in all 10 patients and was grade 3 or 4 in severity in five patients. Seizures occurred in two patients in the first cohort (before hypertension was monitored and treatment guidelines were established): one patient at 20 mg BID and one at 10 mg BID. Neither of these patients had brain metastases or a prior history of seizures; both patients recovered without sequelae. In subsequent cohorts of patients receiving doses < 10 mg BID (n = 26), hypertension was a DLT in one patient receiving 15 mg QD. This patient continued on treatment at a lower dose with concurrent antihypertensive medication with good blood pressure control. Eleven other patients developed hypertension (including six of the 14 patients treated at the recommended phase II dose) that was not dose limiting and was managed by standard antihypertensive medications. No consistent shifts in the levels of renin, angiotensin II, and aldosterone were seen with drug treatment, suggesting that AG-013736–related hypertension is unlikely to be mediated through alteration of the renin-angiotensin-aldosterone pathway.
A variety of grade 1 to 2 mouth-pain events (eg, stomatitis, glossodynia) were reported as related to AG-013736 in 11 patients. Two patients experienced grade 3 stomatitis, accompanied by grade 3 throat pain in one patient. Dose reductions or drug holidays helped to manage this toxicity. Grade 1 hoarseness or voice changes were reported as a drug-related event in five patients.
Drug-related diarrhea was generally grade 1 to 2: one patient in cohort six experienced intermittent grade 3 diarrhea and required dose reduction. Loperamide was used successfully to manage diarrhea in most patients. Nausea and vomiting were grade 1 to 2 in severity and manageable with medications.
Five patients died during the study; three of the deaths were related to disease progression. Two patients with adenocarcinoma of the lung (non–small-cell lung cancer) in the second cohort (20 mg BID, reduced to 10 mg BID) developed fatal hemoptysis. One patient had a centrally located lung lesion and died acutely with grade 4 hemoptysis while on AG-013736. Treatment with high-dose AG-013736 likely led to the rapid breakdown of a vessel wall with tumor infiltration, followed by fatal hemoptysis. The second patient with a peripherally located lung lesion developed grade 1 hemoptysis while on AG-013736; consequently, treatment with the study drug was discontinued. Two weeks later, the patient died of lung complications including grade 4 hemoptysis, which was ultimately determined by the investigator to be related to disease and concurrent infection. Based on these episodes of serious hemoptysis as well as reported similar bleeding events seen with other antiangiogenic agents, the protocol was amended to exclude patients with centrally located lung lesions of any type as well as patients with squamous cell carcinoma of the lung. The only other bleeding event reported as related to AG-013736 was one case of grade 1 rectal bleeding.
Abnormalities in laboratory parameters were reported as adverse events in Table 3 if they resulted in dose reduction, treatment interruption, or clinical sequelae or were otherwise considered important by the investigator. Findings from review of the laboratory data are summarized below.
Asymptomatic proteinuria was observed in 7 of the 10 patients enrolled in the first two cohorts, including two patients with grade 3 proteinuria by dipstick. The protocol was subsequently amended to exclude patients with proteinuria > 500 mg over 24 hours; dose adjustment or suspension of AG-013736 dosing was required for patients with proteinuria 1 g over 24 hours. In subsequent cohorts treated at lower doses of AG-013736, proteinuria was less frequent and less severe. Of these 26 patients, only six developed grade 1 or 2 proteinuria after starting treatment.
Three patients developed grade 3 transaminase elevations while receiving AG-013736. One patient with metastatic breast cancer developed pancreatitis and ischemic bowel associated with superior mesenteric vein thrombosis on15 mg QD. The cause of this constellation of events was not determined, although tumor infiltration was suspected. In one patient, the change in liver-function tests was reported to be related to a concomitant medication (enalapril). The third case occurred in a patient receiving 20 mg BID.
There was only one notable hematologic toxicity at any dose level: grade 2 thrombocytopenia resulting in treatment discontinuation in a patient taking 20 mg BID.
Efficacy
The primary purpose of this study was to establish the safety and MTD of AG-013736. In addition, patients were evaluated for response according to RECIST. There were three confirmed partial responses and other indications of antitumor activity. One patient with adenoid cystic carcinoma enrolled in the 15-mg-QD cohort was identified as a responder after completing three cycles. The response was ongoing 4 months later, when the patient stopped treatment because of subjectively intolerable grade 2 stomatitis accompanied by weight loss. Partial response was also observed in two of six patients with renal cell carcinoma (RCC). In one of these patients, response was identified after 10 weeks of treatment with 5 mg BID in the fasted state. Disease progression was documented 18 weeks later. The second patient with RCC was identified as a responder after two cycles. One year later, the patient stopped treatment with AG-013736 to be evaluated for surgical excision of his remaining lung metastases. Four weeks later, his disease progressed such that surgery was no longer an option. AG-013736 treatment was restarted, resulting in renewed tumor shrinkage (Fig 5).
Cavitation of lung lesions was observed in the two patients with non–small-cell lung cancer (Fig 6). Although both patients subsequently died with hemoptysis (only one of the deaths was considered related to AG-013736), this may be evidence of a significant antiangiogenic effect of AG-013736. Three additional patients had decreases in tumor burden that did not qualify by RECIST criteria for response, including one patient each with mesothelioma (on treatment for 16 months), thyroid cancer (9 months), and RCC (4 months). Finally, a patient with refractory breast cancer experienced healing of skin lesions before starting treatment with the CYP3A inducer phenytoin (see Discussion for details).
DISCUSSION
This phase I study has defined the MTD for AG-013736 and identified important issues regarding absorption, drug interactions, and toxicities. In addition, data from this trial support the clinical activity of AG-013736 in advanced and refractory cancer, providing the information necessary to proceed with phase II testing in specific malignancies.
As is commonly seen with orally administered drugs, PK was variable; the major determinant for variability is not clear at this time. In vitro studies with human liver indicate that AG-013736 is metabolized by conversion to glucuronide metabolites as well as oxidation by the CYP3A4 and CYP1A2 isozymes, two P450 enzymes known to be inducible and thus likely to exhibit variability in patients. Additionally, the CYP3A4 isozyme content in human intestine and liver is known to be highly variable even in healthy (noninduced) individuals.
Another contributor toward the PK variability could be differences in AG-013736 gastrointestinal absorption. PK testing in this trial revealed that dosing in the fasted state resulted in the best absorption; this recommendation has been incorporated into phase II trials. It is interesting to note that although the aqueous solubility of AG-013736 is pH dependent with low (acidic) pH, resulting in the highest solubility, drug concentrations in this study were not significantly affected by the proton-pump inhibitor rabeprazole. Several clinical evaluations in healthy volunteers are ongoing or planned to better understand sources of variability in the oral PK of AG-013736.
Data from a single patient in cohort one indicated a likely drug interaction with phenytoin. After phenytoin use, the AUC0-24 and Cmax of AG-013736 were reduced approximately 10-fold. Phenytoin is a potent inducer of multiple CYP450 enzymes, and it is likely that it induced AG-013736 clearance. Although there had been clinical evidence of healing of malignant and ulcerating skin lesions in this patient before the start of phenytoin, she subsequently experienced disease progression that resulted in discontinuation from study treatment. Concomitant use of potent inducers of the CYP3A4 and CYP1A2 isozymes was subsequently excluded during treatment with AG-013736.
The initial doses used in this trial were clearly above the MTD for AG-013736. Nonclinical safety studies conducted to support the phase I starting dose did not identify the hypertensive effect of AG-013736, which resulted in the clinical administration of the maximum administrable dose within the first two cohorts. In addition, DLT was not identified in cohort one (partly because the need for daily blood pressure monitoring had not been recognized yet) before enrollment was permitted by protocol into cohort two. This led to the selection of the cohort-two dose (20 mg BID) based on the relatively unremarkable early safety profile of patients in cohort one, who were receiving doses between 10 mg QD and 30 mg BID. The subsequent dose de-escalation and study amendment to monitor and treat hypertension adequately avoided additional significant toxicity. The experiences described in this trial highlight the importance of well-designed phase I studies with detailed PK sampling to better understand human-specific end-organ toxicities, potential drug interactions affecting therapeutic blood concentrations, and therapeutic dosing.
The observed toxicities are similar to those observed for other antiangiogenic agents,26-30 with some variability in DLT, and are dose related. Diarrhea and nonulcerating stomatitis (glossodynia) were generally tolerable. Hypertension, which was easily controlled with antihypertensive medications in patients receiving the recommended phase II dose of AG-013736, has also been seen with the antibody to VEGF, bevacizumab,31,32 as well as with oral angiogenesis inhibitors.27,33 The development of hypertension is likely to be related to a mechanistic effect of antiangiogenic agents and may be a marker of VEGFR inhibition. However, increases in indices of vascular stiffness seen with the angiogenesis inhibitor BAY 43-9006 do not correlate with levels of circulating VEGF.33 In addition to its role in triggering angiogenesis, VEGF exerts a stimulatory effect on endothelial nitric oxide (NO) production by upregulating endothelial nitric oxide synthase expression. NO in turn has a vasodilatory effect.34-36 Inhibition of VEGFR signaling by AG-013736 may cause a decrease in NO production, leading to vasoconstriction and hypertension in a subset of patients. Preclinical data support this hypothesis; in vitro evaluation of AG-013736 in human umbilical vein endothelial cells revealed inhibition of endothelial nitric oxide synthase activity at the median infectious dose that inhibited VEGFR-2 phosphorylation.37
AG-013736 has clear clinical activity in advanced and refractory malignancies, as shown in this phase I trial. Sustained tumor response was seen in renal cell cancer as well as in adenoid cystic cancer, and antitumor activity was demonstrated in patients with lung cancer. Tumor necrosis with hemoptysis in centrally located lung lesions has been seen also with bevacizumab and may be a specific marker of antiangiogenic activity. Earlier treatment of lung carcinomas and careful selection of patients could avoid this potentially life-threatening toxicity. The evaluation of pharmacodynamic response to AG-013736 measured by DCE magnetic resonance imaging, reported separately, demonstrated a decrease in tumor vascular parameters at the selected phase II dose.
Based on the encouraging clinical results from this trial, evidence of antiangiogenic activity on DCE magnetic resonance imaging, and the tolerability of AG-013736 at a dose of 5 mg BID in the fasted state, current and planned phase II trials are evaluating efficacy as a single agent or in combination with chemotherapy in a variety of malignancies. A variety of anti-VEGFR small molecules, differentiated by target and specificity, are in development.26,30,38-40 The multitargeted nature of these molecules, for example combining VEGF and PDGFR inhibition, may be critical to antitumor effects19,41 and may be a specific advantage over antibodies that target the ligand alone. As we continue to explore the use of potent antiangiogenic agents in the treatment of cancer, it will be important to include appropriate PK evaluations, as well as surrogate markers to evaluate target inhibition.22,42-45
Authors' Disclosures of Potential Conflicts of Interest
Although 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
We gratefully acknowledge the help of Marina Kenzer, Tracey Borst, Donna Alberti, Kimberly Binger, Cathy Henceroth, Kavitha Krishnan, Michelle Purdom, Debra Chicks, and Krista McAlee for patient recruitment, data management, and patient care during the conduct of this phase I trial. We also thank Brian Hee for assistance with bioanalysis of the pharmacokinetics samples. Finally, we appreciate the efforts of Anton Mulvaney, who was responsible for study management.
NOTES
Research support provided by Pfizer Global Research and Development, San Diego, CA, and Groton, CT.
Presented at the American Association for Cancer Research-National Cancer Institute-European Organisation for Research and Treatment of Cancer International Conference on Molecular Targets and Cancer Therapeutics, 2003, Boston, MA; 6th International Symposium on Antiangiogenesis Agents, 2004, San Diego, CA; and American Society of Clinical Oncology Annual Meeting, 2004, New Orleans, LA.
Authors' disclosures of potential conflicts of interest are found at the end of this article.
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Rugo HS: Bevacizumab in the treatment of breast cancer: Rationale and current data. Oncologist 9:43-49, 2004 (suppl 1)
Veronese ML, Flaherty R, Townsend A, et al: Pharmacodynamic study of the raf kinase inhibitor BAY 43-9006: Mechanisms of hypertension. Proc Am Soc Clin Oncol 22:135s, 2004 (abstr 2035)
Bouloumie A, Schini-Kerth VB, Busse R: Vascular endothelial growth factor up-regulates nitric oxide synthase expression in endothelial cells. Cardiovasc Res 41:773-780, 1999
Kroll J, Waltenberger J: A novel function of VEGF receptor-2 (KDR): rapid release of nitric oxide in response to VEGF-A stimulation in endothelial cells. Biochem Biophys Res Commun 265:636-639, 1999
Shiojima I, Walsh K: Role of Akt signaling in vascular homeostasis and angiogenesis. Circ Res 90:1243-1250, 2002
Wickman G, Hallin M, Salansky KA, et al: Further characterization of the potent VEGF/PDGF receptor tyrosine kinase inhibitor AG-013736 in preclinical tumor models for its antiangiogenesis and antitumor activity. Proc Am Assoc Cancer Res 44:865, 2003 (abstr 3780)
Sun L, McMahon G: Inhibition of tumor angiogenesis by synthetic receptor tyrosine kinase inhibitors. Drug Discov Today 5:344-353, 2000
Kerbel R, Folkman J: Clinical translation of angiogenesis inhibitors. Nat Rev Cancer 2:727-739, 2002
Ciardiello F, Bianco R, Caputo R, et al: Antitumor activity of ZD6474, a vascular endothelial growth factor receptor tyrosine kinase inhibitor, in human cancer cells with acquired resistance to antiepidermal growth factor receptor therapy. Clin Cancer Res 10:784-793, 2004
Bergers G, Javaherian K, Lo KM, et al: Effects of angiogenesis inhibitors on multistage carcinogenesis in mice. Science 284:808-812, 1999
Mundhenke C, Thomas JP, Wilding G, et al: Tissue examination to monitor antiangiogenic therapy: A phase I clinical trial with endostatin. Clin Cancer Res 7:3366-3374, 2001
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Willett CG, Boucher Y, di Tomaso E, et al: Direct evidence that the VEGF-specific antibody bevacizumab has antivascular effects in human rectal cancer. Nat Med 10:145-147, 2004
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Pfizer Global Research and Development, San Diego, CA
University of Wisconsin Comprehensive Cancer Center, Madison, WI
The University of Texas M.D. Anderson Cancer Center, Houston, TX
ABSTRACT
PURPOSE: We studied the safety, clinical activity, and pharmacokinetics (PK) of AG-013736, an oral receptor tyrosine kinase inhibitor of vascular endothelial cell growth factor, platelet-derived growth factor, and c-Kit, in patients with advanced cancer.
PATIENTS AND METHODS: Patients received fixed doses of AG-013736 orally in 28-day cycles. In the first cohort, patients initially received two single test doses of AG-013736 (10 and 30 mg); subsequent dosing was determined by individual PK parameters. Doses in subsequent cohorts were assigned by using a traditional dose-escalation/de-escalation rule based on observed toxicities in the current and previous cohorts. PK analysis included evaluation of the effect of food and antacid.
RESULTS: Thirty-six patients received AG-013736 at doses ranging from 5 to 30 mg by mouth twice daily. The dose-limiting toxicities observed included hypertension, hemoptysis, and stomatitis and were seen primarily at the higher dose levels. The observed hypertension was manageable with medication. Stomatitis was generally tolerable and managed by dose reduction or drug holidays. AG-013736 was absorbed rapidly, with peak plasma concentrations observed within 2 to 6 hours after dosing. The maximum-tolerated dose and recommended phase II dose of AG-013736 is 5 mg, twice daily, administered in the fasted state. No significant drug interaction with antacid was seen. There were three confirmed partial responses and other evidence of clinical activity.
CONCLUSION: In this study, we have demonstrated clinical activity and safety of AG-013736 in patients with advanced solid tumors and identified the dose for phase II testing. The unique phase I study design allowed early identification of important absorption and metabolic issues critical to phase II testing of this agent.
INTRODUCTION
Angiogenesis is necessary for the progression from benign to malignant tumors, as well as growth and metastases of malignant cells.1-7 Disruption of endothelial cell responses responsible for abnormal blood vessel formation may be used to stop tumor growth.8-11
A strong correlation has been observed between vascular endothelial cell growth factor (VEGF) expression and tumor microvessel density in multiple tumor types,12-14 supporting the role of VEGF in facilitating tumor angiogenesis15,16 through its receptors. Abrogation of signaling through both the VEGF receptor (VEGFR) and platelet-derived growth factor receptor-? (PDGFR-?) has been demonstrated to inhibit angiogenesis and tumor growth in preclinical models, with early supportive clinical results.17-20
AG-013736 is a substituted indazole derivative that was discovered by using a structure-based drug design. Mechanistically, AG-013736 is a potent small molecule tyrosine kinase inhibitor of all known VEGFRs at subnanomolar concentrations and PDGFR-? and c-Kit in low nanomolar concentrations. In vitro, AG-013736 selectively blocks VEGF-stimulated receptor autophosphorylation leading to inhibition of endothelial cell proliferation and survival. In mice, AG-013736 inhibited tumor vascular angiogenesis and the growth of human colorectal and murine lung tumors. This antitumor effect was associated with a significant decrease in microvessel density and increased tumor necrosis. In a transgenic mouse model of spontaneous islet cell tumors, treatment with AG-013736 eliminated endothelial fenestrations and suppressed vascular sprouting within 24 hours. At 7 days, vascular density decreased more than 70%, and significant tumor shrinkage was seen at 21 days.21
Given strong preclinical evidence for antitumor/antimetastatic activity, a first-in-human phase I trial was conducted to test AG-013736 in patients with advanced solid malignancies. The primary objective was to determine dose-limiting toxicity (DLT) and the maximum-tolerated dose (MTD) of AG-013736. Secondary objectives were to (1) evaluate the pharmacokinetics (PK) of oral AG-013736, (2) conduct a pilot evaluation of the effect of food on AG-013736 PK, (3) conduct a pilot evaluation of the effect of an antacid on the PK of AG-013736, and (4) document preliminary evidence of antitumor activity. Dynamic contrast-enhanced (DCE) imaging was performed to evaluate the association between PK and vascular response; these data are reported in a separate publication.22
PATIENTS AND METHODS
Patients were enrolled at the University of Wisconsin Comprehensive Cancer Center, University of Texas M.D. Anderson Cancer Center, and University of California, San Francisco Comprehensive Cancer Center between May 2002 and January 2004. All patients were 18 years of age with histologically confirmed advanced nonhematologic cancer refractory to standard therapy or for which no effective therapy was available. An Eastern Cooperative Group performance status of 0 to 2 and a life expectancy of 12 weeks were required. All patients had completed prior therapies with adequate time allowed for recovery from toxicity. Additional inclusion criteria included: adequate bone marrow, renal, and liver function (bilirubin 1.5 mg/dL, transaminases 2.5 x upper limit of normal or 5 x upper limit of normal with documented liver metastasis). No prior history of coronary artery disease, significant gastrointestinal abnormalities, or uncontrolled brain metastases was permitted. Patients with a positive stool guaiac test without evidence of active bleeding were included provided monitoring of the hemoglobin was performed during the trial. In addition, patients requiring the use of potent CYP3A4 inhibitors or CYP3A4/CYP1A2 inducers, chronic H2 antagonists or proton-pump inhibitors, or therapeutic anticoagulant therapy were not eligible. By amendment, patients with proteinuria 500 mg/24 hours, squamous cell lung cancer, centrally located lung lesions (of any histology), and uncontrolled hypertension (systolic > 140 mmHg, diastolic > 90 mmHg) were excluded because of observed toxicities.
Drug Administration and Study Design
AG-013736 was supplied as 1- and 10-mg film-coated tablets. AG-013736 was administered either once daily (QD) or BID, with no scheduled breaks. Each cycle of therapy was defined as 28 days.
Calculation of Starting Dose and Continuous Drug Dosing
Initial dosing was chosen based on a modification of the recommendations of DeGeorge et al23: one sixth of the body-surface area–adjusted minimal effect dose (MED; 10 mg/kg/d) identified in a 28-day dog study of AG-013736. This resulted in a calculated safe human starting dose of 30 mg BID. Cohorts of 6 patients were planned at each dose level. To obtain an early evaluation of in vivo PK and safety, patients in cohort 1 were initially treated with a single test dose of 10 mg, followed by a single test dose of 30 mg at least 48 hours later. The two single doses were selected to not exceed the projected starting dose and to provide an early assessment of PK dose proportionality.
Each patient in the first cohort was then assigned a continuous dose based on their single-dose PK to obtain steady-state plasma exposures similar to that associated with the MED in dogs. The specific continuous dose was estimated as the product of observed apparent plasma clearance and target MED exposure in dogs. The six patients in cohort one received doses between 10 mg QD and 30 mg BID.
Based on the overall safety profile of the first cohort, the average dose evaluated in the first cohort (20 mg BID) was selected as the continuous dose for the second cohort. In subsequent cohorts, escalation of doses by 100% or 40% was planned. However, because of the significant toxicity seen in the first two cohorts, a dose de-escalation schema was used.
Dose Reductions and Modifications
Dose reductions or interruptions of AG-013736 treatment were added by amendment for hypertension, proteinuria, and hemoptysis. Dose modifications for other toxicities were decided between the investigator and sponsor. Intrapatient dose escalation was not allowed. Treatment was discontinued for disease progression, unacceptable toxicity, or withdrawal of consent. Patients were considered to have completed the study after receiving eight cycles of treatment; however, they could continue to receive AG-013736 on a separate follow-on protocol for as long as they derived benefit.
Definition of DLT/MTD
DLT was defined as any of the following during the first cycle when considered related to AG-013736 treatment: select grade 2 or higher gastrointestinal toxicity despite the use of medical intervention and/or prophylaxis, grade 3 anemia, grade 4 thrombocytopenia, or grade 3 nonhematologic toxicities except nausea, vomiting, diarrhea, constipation, pain, and hypertension controlled with medication. The MTD was defined as the dose at which no more than one of six patients in a single cohort experienced DLT in the first cycle.
PK
Plasma samples were collected over 12 hours after dosing in all patients for evaluation of multiple-dose PK on days 1 and 15 (cycle 1) and day 29 (cycle 2, day 1). Additionally, predose trough samples were collected on days 43 (cycle 2, day 15) and 57 (cycle 3, day 1). Whole blood (5 mL) was collected in K3-EDTA vacutainer tubes at predose, 0.5, 1, 2, 4, 8, and 12 hours after dosing and centrifuged at 1,000 x g for 15 minutes, and plasma was frozen at –20°C. Analysis was conducted at Charles River Discovery and Development Services (Worcester, MA). Adequate precautions were taken to minimize degradation of AG-013736 in plasma/whole blood when exposed to visible light.
Assay for Measurement of AG-013736 in Human Plasma
Plasma concentrations of AG-013736 were measured by using a validated liquid chromatography-tandem mass spectrometric method (lower limit of quantitation, 0.1 ng/mL). Briefly, AG-013736 and the deuterated internal standard were extracted from plasma with an ethyl acetate/hexanes mixture. The extracts were evaporated to dryness, reconstituted in aqueous methanol mobile phase, and eluted from a reversed-phase column onto a triple-quadrupole mass spectrometer for detection.
Standard AG-013736 plasma PK parameters were estimated by using noncompartmental methods (using the WINNonlin Professional software; Scientific Consultant, Apex, NC), version 3.2. AG-013736 levels were also measured in the urine of patients.
Evaluation of Food Effect
Patients in cohorts two through four participated in a pilot food-effect study. On day 29, patients received their morning AG-013736 dose within 30 minutes of completion of a 1,000-calorie meal (50% caloric content from fat). On day 30, patients received their morning dose after an overnight fast of at least 6 hours. PK samples were collected for 12 hours on both days. Based on this evaluation, patients in the fifth and sixth cohorts were asked to fast for 2 hours before and after each AG-013736 dose.
Evaluation of Antacid Effect
Patients in cohort six participated in a pilot antacid-effect study to evaluate the effect of the proton-pump inhibitor rabeprazole (Aciphex) on AG-013736 PK. On days 30 to 34, patients received 20 mg of rabeprazole daily 3 hours before their morning AG-013736 dose. On day 34, PK samples were collected for 12 hours after the morning AG-013736 dose.
Pretreatment and Follow-Up Evaluations
Complete medical history, physical examination including vital signs, Eastern Cooperative Oncology Group performance status, full blood count, and serum chemistry and urinalysis were performed at baseline, weekly during the first cycle, and monthly thereafter. A 12-lead ECG and stool guaiac test were performed at baseline. A serum or urine pregnancy test was performed in women of childbearing potential. Toxicity was evaluated by clinical and laboratory examination and graded by using the National Cancer Institute Common Toxicity Criteria, version 2.0.24 After hypertension was identified as a DLT, all patients were required to monitor and record their blood pressures at least once daily. Radiological or physical assessment of tumors was done within 28 days before the first dose and then repeated every two cycles (every 8 weeks). Patients were evaluated for response by using standard Response Evaluation Criteria in Solid Tumors (RECIST).25
Additional Assessments
Plasma samples were collected to evaluate exploratory biomarkers related to the VEGFR pathway. In an effort to evaluate a possible renal etiology for the hypertension seen in patients treated with AG-013736, renin, angiotensin II, and aldosterone levels were measured in the plasma of patients from cohorts five and six receiving 5 mg of AG-013736 BID in the fasted state. Blood samples were collected before dosing on day 1 and then 4 hours after the morning AG-013736 dose on days 1, 15, and 29 (cycle 2, day 1).
RESULTS
Patient Characteristics
A total of 36 patients with advanced solid malignancies were treated with AG-013736 in this phase I trial. The patients received oral doses ranging from 5 mg BID to 30 mg BID. The first four cohorts were dosed in the fed state, and the last two cohorts were dosed in the fasted state (defined as no food or drink other than water in the 2 hours before and after AG-013736 dosing; Table 1) after PK data demonstrated improved absorption in the fasted state (see PK). Selected patient characteristics are summarized in Table 2. Twenty-six patients (72%) had received prior chemotherapy, and 18 patients (50%) had prior radiotherapy.
PK
AG-013736 administered in the fed state was absorbed rapidly, with peak plasma concentrations occurring within 2 to 6 hours after dosing (Table 3, Fig 1). Plasma concentrations declined with a terminal plasma half-life between 2 and 5 hours. AG-013736 plasma PK reached steady state within 15 days, with no unexpected accumulation. Patients in cohorts two through four demonstrated generally linear PK, as evidenced by dose-proportional increases in maximum plasma concentration (Cmax) and area under the curve (AUC; Fig 2).
At the phase II dose of 5 mg BID given in the fasted state, the between-patient coefficient of variation for AUC0-24 on cycle 1, day 15 was 90% (Table 3); the corresponding coefficient of variation for Cmax was 63%. A total of 9 of 16 patients in cohorts two through four completed the pilot food-effect study. Both the rate and extent of absorption were higher in the fasted state, as evidenced by higher Cmax, higher AUC0-24, and a shorter Tmax in the majority of patients. Peak concentrations occurred 1 to 2 hours after dosing in the fasted state, and there was a median 49% increase in plasma exposures compared with the fed state. Plasma half-life was not changed appreciably in fed and fasted states (Table 3, Fig 3). There was no difference in the effect of food across dose groups.
In the presence of the potent proton-pump inhibitor rabeprazole, the rate of absorption of AG-013736 was decreased (reduction in Cmax), but the extent of absorption was unaffected (marginal to negligible changes in the AUC; Fig 4). Rabeprazole was administered in this pilot evaluation for 5 days to maximize its gastric pH-lowering effect. Because of the minimal change in plasma exposure in the presence of rabeprazole, the effect of antacids on AG-013736 absorption was not considered to be clinically significant.
Twelve-hour urinary collections obtained on days 1 and 29 (cycle 2, day 1) demonstrated that < 1% of the administered dose appeared as unchanged drug in the urine regardless of dose, which indicates that the majority of drug elimination was through systemic metabolism.
Soluble plasma proteins (VEGF, fibroblast growth factor, tumor necrosis factor-alpha, and matrix metalloproteinases 2 and 9) evaluated in this study as exploratory markers related to VEGFR signal transduction pathways did not show any variation with treatment.
DLT and MTD
The primary DLT was hypertension. All DLTs are summarized in Table 4. Based on the experience of the first two cohorts, a dose of 20 mg BID exceeded the MTD. DLTs also occurred in patients who were treated in the first two cohorts with 10 mg BID; consequently, this dose was considered to be above MTD and was never tested in a separate cohort.
The MTD and recommended phase II dose of AG-013736 is 5 mg BID in the fasted state. Of 14 patients treated in two cohorts with this dose, two DLTs (grade 2 stomatitis and grade 3 diarrhea) were observed (Table 4).
Safety
Toxicities reported in at least 10% of patients and all grade 3 and 4 toxicities are summarized in Table 5. The principal toxicities were hypertension, fatigue, diarrhea, stomatitis, nausea, and vomiting. Of these, hypertension was reported the most frequently, occurring in 22 patients (61%). The majority of cases (18 patients) were controlled easily with antihypertensive medications. Of the remaining four patients, two developed grade 3 or 4 hypertension and withdrew from the study without attempting medical management. The other two patients developed grade 1/2 hypertension not requiring medication and withdrew from the study early because of disease progression.
The incidence and severity of hypertension was dose dependent. In the first two cohorts in which doses ranged from 10 mg QD to 30 mg BID, hypertension was observed in all 10 patients and was grade 3 or 4 in severity in five patients. Seizures occurred in two patients in the first cohort (before hypertension was monitored and treatment guidelines were established): one patient at 20 mg BID and one at 10 mg BID. Neither of these patients had brain metastases or a prior history of seizures; both patients recovered without sequelae. In subsequent cohorts of patients receiving doses < 10 mg BID (n = 26), hypertension was a DLT in one patient receiving 15 mg QD. This patient continued on treatment at a lower dose with concurrent antihypertensive medication with good blood pressure control. Eleven other patients developed hypertension (including six of the 14 patients treated at the recommended phase II dose) that was not dose limiting and was managed by standard antihypertensive medications. No consistent shifts in the levels of renin, angiotensin II, and aldosterone were seen with drug treatment, suggesting that AG-013736–related hypertension is unlikely to be mediated through alteration of the renin-angiotensin-aldosterone pathway.
A variety of grade 1 to 2 mouth-pain events (eg, stomatitis, glossodynia) were reported as related to AG-013736 in 11 patients. Two patients experienced grade 3 stomatitis, accompanied by grade 3 throat pain in one patient. Dose reductions or drug holidays helped to manage this toxicity. Grade 1 hoarseness or voice changes were reported as a drug-related event in five patients.
Drug-related diarrhea was generally grade 1 to 2: one patient in cohort six experienced intermittent grade 3 diarrhea and required dose reduction. Loperamide was used successfully to manage diarrhea in most patients. Nausea and vomiting were grade 1 to 2 in severity and manageable with medications.
Five patients died during the study; three of the deaths were related to disease progression. Two patients with adenocarcinoma of the lung (non–small-cell lung cancer) in the second cohort (20 mg BID, reduced to 10 mg BID) developed fatal hemoptysis. One patient had a centrally located lung lesion and died acutely with grade 4 hemoptysis while on AG-013736. Treatment with high-dose AG-013736 likely led to the rapid breakdown of a vessel wall with tumor infiltration, followed by fatal hemoptysis. The second patient with a peripherally located lung lesion developed grade 1 hemoptysis while on AG-013736; consequently, treatment with the study drug was discontinued. Two weeks later, the patient died of lung complications including grade 4 hemoptysis, which was ultimately determined by the investigator to be related to disease and concurrent infection. Based on these episodes of serious hemoptysis as well as reported similar bleeding events seen with other antiangiogenic agents, the protocol was amended to exclude patients with centrally located lung lesions of any type as well as patients with squamous cell carcinoma of the lung. The only other bleeding event reported as related to AG-013736 was one case of grade 1 rectal bleeding.
Abnormalities in laboratory parameters were reported as adverse events in Table 3 if they resulted in dose reduction, treatment interruption, or clinical sequelae or were otherwise considered important by the investigator. Findings from review of the laboratory data are summarized below.
Asymptomatic proteinuria was observed in 7 of the 10 patients enrolled in the first two cohorts, including two patients with grade 3 proteinuria by dipstick. The protocol was subsequently amended to exclude patients with proteinuria > 500 mg over 24 hours; dose adjustment or suspension of AG-013736 dosing was required for patients with proteinuria 1 g over 24 hours. In subsequent cohorts treated at lower doses of AG-013736, proteinuria was less frequent and less severe. Of these 26 patients, only six developed grade 1 or 2 proteinuria after starting treatment.
Three patients developed grade 3 transaminase elevations while receiving AG-013736. One patient with metastatic breast cancer developed pancreatitis and ischemic bowel associated with superior mesenteric vein thrombosis on15 mg QD. The cause of this constellation of events was not determined, although tumor infiltration was suspected. In one patient, the change in liver-function tests was reported to be related to a concomitant medication (enalapril). The third case occurred in a patient receiving 20 mg BID.
There was only one notable hematologic toxicity at any dose level: grade 2 thrombocytopenia resulting in treatment discontinuation in a patient taking 20 mg BID.
Efficacy
The primary purpose of this study was to establish the safety and MTD of AG-013736. In addition, patients were evaluated for response according to RECIST. There were three confirmed partial responses and other indications of antitumor activity. One patient with adenoid cystic carcinoma enrolled in the 15-mg-QD cohort was identified as a responder after completing three cycles. The response was ongoing 4 months later, when the patient stopped treatment because of subjectively intolerable grade 2 stomatitis accompanied by weight loss. Partial response was also observed in two of six patients with renal cell carcinoma (RCC). In one of these patients, response was identified after 10 weeks of treatment with 5 mg BID in the fasted state. Disease progression was documented 18 weeks later. The second patient with RCC was identified as a responder after two cycles. One year later, the patient stopped treatment with AG-013736 to be evaluated for surgical excision of his remaining lung metastases. Four weeks later, his disease progressed such that surgery was no longer an option. AG-013736 treatment was restarted, resulting in renewed tumor shrinkage (Fig 5).
Cavitation of lung lesions was observed in the two patients with non–small-cell lung cancer (Fig 6). Although both patients subsequently died with hemoptysis (only one of the deaths was considered related to AG-013736), this may be evidence of a significant antiangiogenic effect of AG-013736. Three additional patients had decreases in tumor burden that did not qualify by RECIST criteria for response, including one patient each with mesothelioma (on treatment for 16 months), thyroid cancer (9 months), and RCC (4 months). Finally, a patient with refractory breast cancer experienced healing of skin lesions before starting treatment with the CYP3A inducer phenytoin (see Discussion for details).
DISCUSSION
This phase I study has defined the MTD for AG-013736 and identified important issues regarding absorption, drug interactions, and toxicities. In addition, data from this trial support the clinical activity of AG-013736 in advanced and refractory cancer, providing the information necessary to proceed with phase II testing in specific malignancies.
As is commonly seen with orally administered drugs, PK was variable; the major determinant for variability is not clear at this time. In vitro studies with human liver indicate that AG-013736 is metabolized by conversion to glucuronide metabolites as well as oxidation by the CYP3A4 and CYP1A2 isozymes, two P450 enzymes known to be inducible and thus likely to exhibit variability in patients. Additionally, the CYP3A4 isozyme content in human intestine and liver is known to be highly variable even in healthy (noninduced) individuals.
Another contributor toward the PK variability could be differences in AG-013736 gastrointestinal absorption. PK testing in this trial revealed that dosing in the fasted state resulted in the best absorption; this recommendation has been incorporated into phase II trials. It is interesting to note that although the aqueous solubility of AG-013736 is pH dependent with low (acidic) pH, resulting in the highest solubility, drug concentrations in this study were not significantly affected by the proton-pump inhibitor rabeprazole. Several clinical evaluations in healthy volunteers are ongoing or planned to better understand sources of variability in the oral PK of AG-013736.
Data from a single patient in cohort one indicated a likely drug interaction with phenytoin. After phenytoin use, the AUC0-24 and Cmax of AG-013736 were reduced approximately 10-fold. Phenytoin is a potent inducer of multiple CYP450 enzymes, and it is likely that it induced AG-013736 clearance. Although there had been clinical evidence of healing of malignant and ulcerating skin lesions in this patient before the start of phenytoin, she subsequently experienced disease progression that resulted in discontinuation from study treatment. Concomitant use of potent inducers of the CYP3A4 and CYP1A2 isozymes was subsequently excluded during treatment with AG-013736.
The initial doses used in this trial were clearly above the MTD for AG-013736. Nonclinical safety studies conducted to support the phase I starting dose did not identify the hypertensive effect of AG-013736, which resulted in the clinical administration of the maximum administrable dose within the first two cohorts. In addition, DLT was not identified in cohort one (partly because the need for daily blood pressure monitoring had not been recognized yet) before enrollment was permitted by protocol into cohort two. This led to the selection of the cohort-two dose (20 mg BID) based on the relatively unremarkable early safety profile of patients in cohort one, who were receiving doses between 10 mg QD and 30 mg BID. The subsequent dose de-escalation and study amendment to monitor and treat hypertension adequately avoided additional significant toxicity. The experiences described in this trial highlight the importance of well-designed phase I studies with detailed PK sampling to better understand human-specific end-organ toxicities, potential drug interactions affecting therapeutic blood concentrations, and therapeutic dosing.
The observed toxicities are similar to those observed for other antiangiogenic agents,26-30 with some variability in DLT, and are dose related. Diarrhea and nonulcerating stomatitis (glossodynia) were generally tolerable. Hypertension, which was easily controlled with antihypertensive medications in patients receiving the recommended phase II dose of AG-013736, has also been seen with the antibody to VEGF, bevacizumab,31,32 as well as with oral angiogenesis inhibitors.27,33 The development of hypertension is likely to be related to a mechanistic effect of antiangiogenic agents and may be a marker of VEGFR inhibition. However, increases in indices of vascular stiffness seen with the angiogenesis inhibitor BAY 43-9006 do not correlate with levels of circulating VEGF.33 In addition to its role in triggering angiogenesis, VEGF exerts a stimulatory effect on endothelial nitric oxide (NO) production by upregulating endothelial nitric oxide synthase expression. NO in turn has a vasodilatory effect.34-36 Inhibition of VEGFR signaling by AG-013736 may cause a decrease in NO production, leading to vasoconstriction and hypertension in a subset of patients. Preclinical data support this hypothesis; in vitro evaluation of AG-013736 in human umbilical vein endothelial cells revealed inhibition of endothelial nitric oxide synthase activity at the median infectious dose that inhibited VEGFR-2 phosphorylation.37
AG-013736 has clear clinical activity in advanced and refractory malignancies, as shown in this phase I trial. Sustained tumor response was seen in renal cell cancer as well as in adenoid cystic cancer, and antitumor activity was demonstrated in patients with lung cancer. Tumor necrosis with hemoptysis in centrally located lung lesions has been seen also with bevacizumab and may be a specific marker of antiangiogenic activity. Earlier treatment of lung carcinomas and careful selection of patients could avoid this potentially life-threatening toxicity. The evaluation of pharmacodynamic response to AG-013736 measured by DCE magnetic resonance imaging, reported separately, demonstrated a decrease in tumor vascular parameters at the selected phase II dose.
Based on the encouraging clinical results from this trial, evidence of antiangiogenic activity on DCE magnetic resonance imaging, and the tolerability of AG-013736 at a dose of 5 mg BID in the fasted state, current and planned phase II trials are evaluating efficacy as a single agent or in combination with chemotherapy in a variety of malignancies. A variety of anti-VEGFR small molecules, differentiated by target and specificity, are in development.26,30,38-40 The multitargeted nature of these molecules, for example combining VEGF and PDGFR inhibition, may be critical to antitumor effects19,41 and may be a specific advantage over antibodies that target the ligand alone. As we continue to explore the use of potent antiangiogenic agents in the treatment of cancer, it will be important to include appropriate PK evaluations, as well as surrogate markers to evaluate target inhibition.22,42-45
Authors' Disclosures of Potential Conflicts of Interest
Although 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
We gratefully acknowledge the help of Marina Kenzer, Tracey Borst, Donna Alberti, Kimberly Binger, Cathy Henceroth, Kavitha Krishnan, Michelle Purdom, Debra Chicks, and Krista McAlee for patient recruitment, data management, and patient care during the conduct of this phase I trial. We also thank Brian Hee for assistance with bioanalysis of the pharmacokinetics samples. Finally, we appreciate the efforts of Anton Mulvaney, who was responsible for study management.
NOTES
Research support provided by Pfizer Global Research and Development, San Diego, CA, and Groton, CT.
Presented at the American Association for Cancer Research-National Cancer Institute-European Organisation for Research and Treatment of Cancer International Conference on Molecular Targets and Cancer Therapeutics, 2003, Boston, MA; 6th International Symposium on Antiangiogenesis Agents, 2004, San Diego, CA; and American Society of Clinical Oncology Annual Meeting, 2004, New Orleans, LA.
Authors' disclosures of potential conflicts of interest are found at the end of this article.
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