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Angiogenesis is critical for growth, metastasis and invasion of tumours (Kerbel, 2008). Vascular endothelial growth factor (VEGF) is the major mediator of angiogenesis, although multiple other angiogenic factors such as fibroblast growth factor-1 (FGF-1, aFGF) and -2 (FGF-2, bFGF) and platelet-derived growth factor are involved in the initiation and maintenance of angiogenesis and tumourigenesis (Kerbel, 2008). Bevacizumab, which inhibits VEGF receptor-2 (VEGFR-2) signalling through inhibition of VEGF-A ligand binding, prolongs survival in patients with metastatic colorectal cancer in both the first- and second-line settings when used in combination with cytotoxic chemotherapy; however, observed responses are transient, with a median overall survival of 12.9 months for patients receiving second-line treatment and 23 months for those receiving first-line treatment (Kabbinavar et al, 2003; Hurwitz et al, 2004; Giantonio et al, 2007).

Cetuximab, a chimeric monoclonal antibody targeting the epidermal growth factor receptor (EGFR), has demonstrated efficacy and overall survival benefits in patients with K-Ras wild-type metastatic colorectal cancer (Cunningham et al, 2004; Sobrero et al, 2008). Preclinical studies suggest that VEGF overexpression has a role in acquired resistance to anti-EGFR therapy (Bianco et al, 2008) and that dual inhibition of VEGF and EGF signalling may overcome such resistance (Naumov et al, 2009). It is unclear whether these observations translate into prolonged clinical benefit.

Upregulation of alternate proangiogenic signals, such as the FGF signalling pathway, is one of the proposed mechanisms in the development of resistance to VEGF-directed antiangiogenic therapy (Bergers and Hanahan, 2008). Serum bFGF levels have been shown to be increased following first-line treatment of patients with metastatic colorectal cancer treated with combination chemotherapy and monoclonal anti-VEGF antibody therapy (Kopetz et al, 2009). Targeting FGF, VEGF and EGF signalling pathways simultaneously may provide additional clinical benefit to patients with advanced gastrointestinal malignancies. Brivanib is the first orally bioavailable selective dual inhibitor of FGF and VEGF signalling and is administered as brivanib alaninate, the L-alanine ester prodrug of the active moiety brivanib. In preclinical studies, brivanib has shown strong antiangiogenic and antitumour effects on tumour cells across a range of tumour types, including colon, breast, liver and lung (Bhide et al, 2006, 2010; Huynh et al, 2008), and no evidence of evasive resistance when administered to RIP-Tag2/Bl6 mice (Allen et al, 2009). In a recent phase II study in advanced/metastatic hepatocellular carcinoma, brivanib demonstrated encouraging efficacy, with an overall survival of 10 months, and was generally well tolerated (Park et al, 2011). Cetuximab has been combined with anti-VEGF monoclonal antibody therapy (bevacizumab) safely with preliminary suggestion of improved efficacy (Saltz et al, 2007); however, large, randomised phase III studies failed to show a survival benefit associated with cetuximab or panitumumab when added to bevacizumab plus cytotoxic chemotherapy (Hecht et al, 2009; Tol et al, 2009). The FGF pathway is a known alternate angiogenesis pathway to the VEGF pathway and may account for the resistance to anti-VEGF therapy (Kopetz et al, 2009); thus a combined VEGF and FGF receptor inhibitor holds the promise of superior efficacy when used in combination with an EGFR pathway inhibitor.

The primary goals of the current study were to (a) define a dose for combination treatment with brivanib alaninate and cetuximab for further evaluation in phase II/III studies, (b) develop knowledge about dose-limiting toxicity (DLT) and (c) assess the preliminary efficacy and safety of this combination therapy in patients with advanced gastrointestinal malignancies.

Materials and methods

Patients

Patients were required to have the following characteristics: age ⩾18 years; Eastern Cooperative Oncology Group performance status score 0 or 1; histologic or cytologic diagnosis of gastrointestinal malignancy (if the patient entered during the dose-expansion phase, they had to have tissue-verified colorectal cancer); tumour biopsy tissue available for correlative biomarker analysis (K-Ras testing on tissue was performed post hoc after trial completion); radiographic or tissue confirmation that the disease was locally advanced/metastatic; measurable disease; adequate bone marrow, hepatic and renal function; toxicity related to prior therapy had to be resolved to baseline or deemed irreversible; at least 4 weeks had to pass since last chemotherapy, immunotherapy, radiotherapy, anticancer hormonal therapy or targeted therapy, and at least 6 weeks since last therapy with bevacizumab, nitrosoureas, mitomycin C and/or liposomal doxorubicin; and women of child-bearing age had to have a negative pregnancy test. Prior anti-EGFR therapy and anti-VEGF monoclonal antibody therapy were allowed. Patients who had prior treatment with VEGFR-tyrosine kinase inhibitors were ineligible. A DLT was defined, for the purposes of this study, as any of the following events occurring in the first 4 weeks of study treatment: grade 4 neutropenia (i.e. absolute neutrophil count (ANC) <500 cells mm–3 for 5 or more consecutive days) or febrile neutropenia (i.e. fever >38°C with an ANC <500 cells mm–3 requiring hospitalisation); grade 4 thrombocytopenia or bleeding episode requiring platelet transfusion; grade 3 nausea and/or emesis despite the use of maximal medical intervention; grade 2 or greater cardiovascular toxic effect; any grade 3 or greater nonhaematologic toxic effect; or delayed recovery (2 weeks or more) after scheduled re-treatment from a delayed toxic effect related to treatment with cetuximab and brivanib.

Study design

This was an open-label, phase I study of brivanib alaninate administered orally in combination with intravenous cetuximab to patients with advanced gastrointestinal malignancy. This study was conducted in accordance with good clinical practice, as defined by the International Conference on Harmonization and in accordance with the ethical principles underlying European Union Directive 2001/20/EC and the United States Code of Federal Regulations, Title 21, Part 50 (21CFR50). The protocol, amendments and patient-informed consent received appropriate approval by the respective Institutional Review Board/Independent Ethics Committees prior to study initiation. Informed consent was obtained from each patient prior to study participation.

The primary objective was to assess the DLT of brivanib alaninate in combination with cetuximab and to define the maximum tolerated dose (MTD) in patients with advanced gastrointestinal malignancy who had failed prior therapy.

Secondary objectives included assessment of radiographic evidence of antitumour activity, evaluation of changes by 2[18F]fluoro-2-deoxyglucose positron-emitting tomography (FDG-PET) scan and/or radiologic response as defined by the modified World Health Organisation (WHO) criteria, duration of response, duration of disease control and time to progression at doses other than the MTD. Additional FDG-PET-specific objectives were to assess the tumour metabolic response and the association of tumour metabolic changes with clinical outcome (progression-free survival; PFS) in this study population and to assess the reproducibility of FDG-PET measurements of standardised uptake value (SUV) parameters in this multicentre trial. Additional secondary objectives were to determine the disease control rates, duration of response, duration of disease control and PFS based on the modified WHO criteria in response-evaluable patients at the MTD, and to assess the pharmacokinetics (PK) of brivanib alaninate when administered in combination with cetuximab.

A post hoc additional exploratory biomarker analysis to evaluate the relationships between K-Ras mutation status and efficacy end points in patients with colorectal cancer was performed.

Treatment

On cycle 1, day 1 of a 28-day treatment cycle, a single dose of brivanib alaninate was administered, followed by a 6-day washout period. On cycle 1, day 8, continuous daily oral dosing of brivanib alaninate was started together with a single loading dose of intravenous cetuximab 400 mg m–2 infused over 120 min. Beginning on cycle 1, day 15, cetuximab was administered weekly at 250 mg m–2, infused over 60 min. For the remainder of the study, patients received oral brivanib alaninate on a daily continuous schedule and intravenous cetuximab on a weekly basis. Dose escalation of brivanib alaninate starting at 320 mg with two additional escalations of 600 and 800 mg was explored (see Appendix A for further treatment details).

Assessments

Safety

Adverse events (AEs) were evaluated according to the National Cancer Institute Common Terminology Criteria for Adverse Events (v 3.0) on a continuous basis, while the patient was on study and until ⩽30 days after the last dose of study drug or until all treatment-related AEs had recovered to baseline or were deemed irreversible. Once a subject had been off treatment due to toxicity, assessments were to be made every 28 days until all study-related toxicities resolved to baseline, stabilised or were deemed irreversible.

Pharmacokinetics

Brivanib alaninate is rapidly converted to brivanib (the active moiety) in vivo. Hence, only the active moiety was measured in plasma. Plasma samples for determination of brivanib concentrations were drawn pre-dose and serially (15, 30 and 45 min, and 1, 2, 4, 6, 8 and 24 h after dosing) after dosing on cycle 1, day 1 and cycle 1, day 8. Serum samples for determination of cetuximab concentrations were drawn pre-dose and serially (2, 4, 6 and 24 h after dosing) after dosing on cycle 1, day 8. Serum samples for both brivanib and cetuximab were also drawn pre-dose on cycle 1, day 15 and cycle 2, day 1. For analyses of brivanib and cetuximab concentrations in serum and plasma samples, see Appendix B.

Efficacy

Tumour response was determined using modified WHO criteria (see Appendix C).

FDG-PET and K-Ras mutations analyses

See Appendix D.

Statistical methods

Cohorts of 3–10 patients were to be treated at each dose level until the MTD was defined based on observance of DLTs; however, the MTD was not reached due to lack of DLT. Thus, at the maximum dose level (MDL) of 800 mg, a total of 40 response-evaluable patients were accrued in two stages, with the number of patients at each stage based on a modified Gehan two-stage approach (see Appendix E for further details on statistical analyses).

Results

Patient disposition and demographics

Eighty-five patients were enrolled on this study, 62 of whom were treated with brivanib alaninate (Table 1) and 61 of whom received brivanib alaninate plus cetuximab (24 of the 85 patients failed screening). The majority of patients were males (61% vs 39%); median age was 60 years. Fifty-nine patients had colorectal cancer, two had oesophageal cancer and one had fibrolamellar hepatocellular carcinoma. The most common metastatic sites were liver (53 patients), lung (51 patients) and lymph node (19 patients).

Table 1 (a) Baseline demographics and characteristics and (b) prior therapy
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Safety

The MTD was not formally reached and the expansion cohort was opened at the predefined MDL based on the MTD (800 mg) reached in the phase I brivanib monotherapy study (CA182002) (Jonker et al, 2010). One patient had a DLT (bilateral pulmonary embolism) considered possibly related to treatment with brivanib alaninate at a dose of 320 mg combined with cetuximab. The event did not lead to any further complications, but the patient died 3 weeks later due to disease progression. No further DLTs occurred at any dose.

Overall, brivanib treatment-related AEs were acceptable when administered daily at doses of 320, 600 and 800 mg in combination with cetuximab (Table 2). Six patients (9.7%), all of whom were in the 800-mg cohort, had grade 1/2 palmar–plantar dysesthesias.

Table 2 Most common treatment-related (investigator determined) AEs and serious AEs
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The majority of treatment-related AEs were grade 1/2 in severity. The most frequently reported treatment-related grade 3/4 AEs (>5% of patients) were fatigue and increase in hepatic transaminases. The incidence of grade 3/4 palmar–plantar dysesthesias was infrequent, with only one reported case in the 800-mg cohort (1.6% of patients). In total, 14 patients (22.6%) reported serious (grade 3/4) treatment-related AEs during the study, including vomiting, dehydration and pyrexia. Seven patients discontinued due to study drug-unrelated AEs, including small intestinal obstruction, vomiting, pneumonia, left ventricular dysfunction, hyperbilirubinaemia, drug hypersensitivity to cetuximab, disease progression, angio-oedema and hypertension in one patient each. In addition, pneumonia and malignant neoplasm progression occurred in the same patient. Four patients discontinued due to drug-related toxicities, which included sepsis, aspartate aminotransferase elevation, dehydration and angio-oedema. All AEs leading to study discontinuation occurred in the 800-mg cohort.

Twenty patients had at least one dose reduction, including two in the 600-mg cohort and 18 in the 800-mg cohort. No patient in the 320-mg cohort had a dose reduction. Dose reductions due to cetuximab-related AEs occurred across all cohorts in 16 patients (26%), two in the 600-mg cohort and 14 in the 800-mg cohort. Reasons for dose reduction were delayed nonhaematologic recovery (five patients, 8%), skin toxicity (four patients, 6%) and delayed haematologic recovery (one patient, 2%).

In all, 15 patients died during the study, 14 due to disease progression and 1 due to study drug toxicity. One patient, who was treated with brivanib alaninate 800 mg in combination with cetuximab, died on day 47 due to sepsis related to rectal perforation. Both events were considered possibly related to treatment.

Pharmacokinetics

At the MDL, cetuximab had no effect on brivanib maximum plasma concentration (Cmax) or area under the plasma concentration–time curve extrapolated to infinity (AUC(INF)): 90% CIs for the Cmax and AUC(INF) geometric mean ratios with and without cetuximab were within the 80–125% interval (Table 3).

Table 3 Effect of cetuximab on brivanib pharmacokinetics following single 800 mg doses of brivanib alaninate with cetuximab 400 mg m–2 on day 8 and without cetuximab 400 mg m–2 on day 1
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Efficacy

Six patients (9.7%) achieved a partial response, all of whom were in the brivanib alaninate 800 mg dose group, resulting in an objective response rate of 10%. Twenty-eight patients (45%) achieved stable disease (three in the brivanib alaninate 320 mg dose group, three in the brivanib alaninate 600 mg dose group and 22 in the brivanib alaninate 800 mg dose group), 15 patients (24.2%) had disease progression (two in the brivanib alaninate 320 mg dose group, two in the brivanib alaninate 600 mg dose group and 11 in the brivanib alaninate 800 mg dose group). Radiographic response was not evaluable in 13 patients (21%). The median duration of response was 9.2 months (95% CI: 7.20, 16.4 months; n=6), the median duration of disease control was 5.5 months (95% CI: 3.84, 7.32 months; n=28) and the median duration of stable disease was 5.3 months (95% CI: 3.68, 5.55; n=21). The median PFS was 3.9 months (95% CI: 3.38, 5.42 months; n=51).

FDG-PET and K-Ras analyses

Results of FGD-PET and K-Ras analyses are reported in Appendix F.

Discussion

This phase I study evaluated the safety, PK, pharmacodynamics and efficacy of brivanib combined with full-dose cetuximab in patients with advanced gastrointestinal malignancies. The MTD of the combination of cetuximab and brivanib was not established at the prespecified MDL of 800 mg, although this dose was determined to be the MTD for monotherapy in a previous phase I study (Jonker et al, 2010). One DLT occurred at the 320-mg dose level in one patient who experienced bilateral pulmonary artery emboli. The toxicity profile of brivanib alaninate in combination with cetuximab observed in this study was manageable.

The population in the current study was heavily pretreated, having advanced-stage disease; the majority of patients had received three or more previous lines of treatment and ∼40% of patients had prior EGFR inhibitor therapy. Despite this, brivanib alaninate in combination with cetuximab demonstrated promising antitumour activity, with an objective response rate of 10% at the MDL and a PFS of 3.9 months. Objective responses were not noted in the 11 patients treated with low-dose brivanib group, despite concomitant cetuximab therapy, possibly because of the low number of patients in this subgroup and the inclusion of patients with gastrointestinal malignancies other than colorectal cancer (where cetuximab does not have a proven efficacy as monotherapy), patients with unknown K-Ras status and patients who had received prior anti-EGFR therapies. Consistent with our findings, results from a phase III study of cetuximab monotherapy vs cetuximab plus irinotecan combination therapy in irinotecan-refractory patients showed a median PFS of 1.5 months with cetuximab monotherapy (Cunningham et al, 2004), and a separate study of cetuximab monotherapy in patients who were refractory or intolerant to both irinotecan and oxaliplatin by Jonker et al (2007) reported a median PFS of 1.9 months, with an overall survival of 6.1 months. Notably, the K-Ras wild-type subgroup in the study by Jonker's group had a response rate of 12.8% and overall survival of 9.5 months (Karapetis et al, 2008). In contrast, in the small subgroup of patients in the current study who had K-Ras wild-type status and did not receive prior anti-EGFR-targeted therapy, 6 of 11 patients had a partial response. However, it should be noted that these K-Ras analyses were unplanned and, therefore, limited. In a first-line setting, the combination of cetuximab with the VEGF-A-binding monoclonal antibody bevacizumab and oxaliplatin-based chemotherapy did not show a benefit for adding cetuximab in patients with K-Ras wild-type tumours. In patients with K-Ras mutant tumours, there was a detrimental effect (median PFS decreased from 12.5 to 8.1 months and overall survival decreased from 24.9 to 17.2 months) (Tol et al, 2009).

2[18F]Fluoro-2-deoxyglucose positron-emitting tomography is increasingly becoming an integral part of multicentre trials for tumour detection, staging and follow-up studies. In particular, changes in SUV parameters over the course of treatment can serve as an early surrogate marker for clinical benefit (Kelloff et al, 2005). The FDG-PET repeatability coefficients reported here indicate that 95% of patients had an SUVmax per cent difference in the two repeat baseline scans within −34% and 51%. Notably, the lower repeatability coefficient value of −34% rather than the −25%, as suggested by European Organisation for Research and Treatment of Cancer (EORTC) guidance, indicates a metabolic response by SUVmax, while individual changes within −34% and 51% may not represent a metabolic response to treatment. The metabolic response rate, as determined by FDG-PET, was similar across all SUV parameters when assessed by the EORTC and lower repeatability coefficient threshold. At the 800-mg dose, metabolic response rate on day 15 (after 2 weekly doses of cetuximab and 1 week of continuous dosing of brivanib alaninate) was 53% by EORTC criteria and 43% by repeatability threshold. In addition, the metabolic response rate on day 56 was 39% by EORTC and 26% by repeatability threshold. Association of metabolic response and clinical outcome was assessed by SUVmax, with results suggesting that metabolic response at days 15 and 56, based on the SUVmax parameter, may be associated with longer PFS in this population. In addition, greater separation of the Kaplan–Meier curves at day 56 than at day 15 may suggest that metabolic response at day 56 is a better predictor of PFS, a hypothesis that could be tested in future trials. Overall, these data support the concept that metabolic response may represent a predictive marker of clinical outcomes.

The promising efficacy observed in this study in patients with advanced gastrointestinal cancer and the manageable toxicity profile of brivanib justifies further study of brivanib in combination with cetuximab. A prospective, randomised, phase III study of brivanib alaninate plus cetuximab compared with placebo plus cetuximab in patients with advanced colorectal cancer whose tumours are K-Ras wild type and who have been previously treated with combination chemotherapy but are treatment naive for EGFR-targeted therapy has completed accrual February 2011 (the National Cancer Institute of Canada trial, clinicaltrials.gov identifier: NCT00640471); this study should help us better understand what additional benefit (if any) brivanib adds in combination with cetuximab in patients with advanced colorectal cancer.