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. Author manuscript; available in PMC: 2013 Dec 5.
Published in final edited form as: J Thorac Oncol. 2012 Jun;7(6):10.1097/JTO.0b013e31824de2fa. doi: 10.1097/JTO.0b013e31824de2fa

Phase I/II study of bortezomib in combination with carboplatin and bevacizumab as first line therapy in patients with advanced non-small cell lung cancer (NSCLC)

Bilal Piperdi 1, William V Walsh 2, Kendra Bradley 2, Zheng Zhou 2, Venu Bathini 2, Meredith Hanrahan-Boshes 2, Lloyd Hutchinson 3, Roman Perez-Soler 1
PMCID: PMC3852685  NIHMSID: NIHMS530556  PMID: 22534815

Abstract

Purpose

To establish the maximum tolerated dose (MTD) of weekly bortezomib in combination with fixed standard doses of carboplatin and bevacizumab and to estimate the efficacy (response rate and progression free survival) and safety of combination therapy with carboplatin, bortezomib and bevacizumab as first line therapy in patients with advanced NSCLC.

Experimental Design

Patients were assigned to three dose levels of weekly bortezomib with the fixed standard doses of carboplatin (AUC 6) and bevacizumab (15 mg/kg) q 3 wks using a standard phase I design. Bortezomib doses were 1.3 mg/m21.6 mg/m2 and 1.8 mg/m2 weekly on D1 and D8 of q 3wk cycle. A maximum of six cycles was administered. Patients with complete, partial response (PR) or stable disease were continued on single agent bevacizumab (15 mg/kg q 3 wks) as maintenance therapy. In phase II, either level III or MTD was administered to evaluate the efficacy and safety of the combination in first line treatment of advanced NSCLC.

Results

16 patients were enrolled (3, 4 and 9 pts in dose level I, II and III respectively). There was no pre-defined dose limiting toxicity in cycle 1 in all 16 patients. The recommended phase II dose is bortezomib 1.8 mg/m2 weekly on day 1 and day 8 in combination with carboplatin AUC of 6 and bevacizumab 15 mg/kg on every 21 day cycle. Total of 9 patients were treated at the recommended phase II dose level. The most common treatment related grade 3/4 toxicities during the subsequent cycles were thrombocytopenia (58%), lymphopenia (25%), neutropenia (12%) and diarrhea (25%). The grade 1/2 neuropathy was seen in 7 out of 16 pts (44%). The response rate, PFS and OS in all patients were 37.5% (95%CI 13.8% - 61.2%), 5.0 months (m) (95%CI: 3.1-8.4), 9.9 m (95% CI: 8.2-14.1) and the 9 patients in phase II portion are 44% (95%CI 15.3% - 77.3%), 5.5 m (95%CI: 3.1-12.2) and 10.9 months (95%CI: 8.0-14.1).

Conclusion

The recommended phase II dose for this combination is: carboplatin AUC 6, bevacizumab 15 mg/kg on D1 and bortezomib 1.8 mg/m2 on D1 and D8 on q 21 day cycle. The regimen was very well tolerated with interesting clinical activity in first line treatment of NSCLC.

Keywords: bortezomib, bevacizumab, metastatic non-small cell lung cancer

Introduction

Non-small cell lung cancer remains the leading cause of cancer deaths in United States (1). The platinum based palliative, systemic chemotherapy remains the cornerstone for the management of patients with metastatic disease or non-operable, locally advanced disease. It reduces cancer-related symptoms and improves quality of life and survival in patients with advanced NSCLC (2, 3). Although the first-line treatment of advanced NSCLC has evolved significantly over the past decade, currently doublets of second- and third-generation of chemotherapy regimens seem to reach a plateau with response rate of 30–40%, median survival of 8–9 months and 1-year survival rate of 35–40% (2–4). There is a need to identify novel targets and treatment strategies to improve the therapy for NSCLC patients.

Angiogenesis or new blood vessel formation is an important hallmark of many tumors. The vascular endothelial growth factor (VEGF) plays an important role in the growth and metastasis of many cancers, including non-small cell lung cancer (5). Bevacizumab (Genenetech, South San Francisco, CA) is a recombinant humanized anti-VEGF monoclonal antibody that is approved for treatment of metastatic colon and lung cancers. In randomized phase III studies, the addition of bevacizumab to standard chemotherapy improved both the response rate and survival in patients with advanced non-squamous NSCLC (6, 7). Based on the ECOG 4599 trial, the combination of carboplatin, paclitaxel and bevacizumab is a widely adopted front line regimen in patients with advanced NSCLC (5, 8).

The ubiquitin-proteasome pathway plays a pivotal role in the degradation of most intracellular proteins in eukaryotic cells, including those regulating apoptosis, cell cycle progression, transcription factor activation, and angiogenesis (9, 10). Bortezomib (VELCADE; Millennium Pharmaceuticals, Inc.), a dipeptide proteasome inhibitor, is a novel antineoplastic agent presently approved for the treatment of patients with multiple myeloma and relapsed mantle-cell lymphoma (11, 12). Bortezomib has been shown to have significant cytotoxic activity in human NSCLC cell lines in vitro. Bortezomib induced concentration and time-dependent G2/M cell cycle arrest of NSCLC cells (13–15). This G2/M arrest was unique and different from taxanes and vinca alkaloids and does not involve the tubulin (13, 14). Bortezomib also induces apoptosis in cells that over express bcl-2, a genetic trait that confers unregulated growth and resistance to conventional chemotherapeutics (16). Bortezomib has also been shown to have significant anti-angiogenic activity (25, 26).

In vivo activity was also observed in NSCLC xenografts as well as human NSCLC heterotransplant model (unpublished data). Clinical activity was also observed in patients with refractory NSCLC in phase I and II trials (17, 20). Due to its unique properties as well as single agent activity, we planned to do a pilot trial of combination of carboplatin, bortezomib and bevacizumab as first line therapy in patients with metastatic NSCLC. The rationale at the inception of the study was to substitute paclitaxel in the ECOG 4599 regimen by bortezomib due to its unique G2/M cell cycle arrest seen in pre-clinical studies.

The present study was designed as a phase I/II study. The primary objective of the phase I study was to define the maximum tolerated dose (MTD) of weekly bortezomib in combination with fixed standard doses of carboplatin and bevacizumab. The primary objective of the phase II study was to estimate the efficacy (response rate and progression free survival) and safety of combination therapy with carboplatin, bortezomib and bevacizumab as first line therapy in patients with advanced NSCLC. The study was closed early due to poor accrual. We will present the final safety and efficacy data from all 16 patients that were treated on the phase I and II portion of the study.

Patients and Methods

Patients

Adult patients with inoperable, locally advanced, or metastatic NSCLC (stage III B with pleural effusion or Stage IV disease based on AJCC 6th edition) were eligible for this study. Tumors were cytologically or histologically confirmed. Due to safety concerns with bevacizumab, the study initially included only non-squamous NSCLC patients. However, the study was subsequently amended to include patients with squamous cell histology if they meet the following criteria: a) extra-thoracic disease only; b) peripheral lung lesions only (peripheral lesion is defined as a lesion in which epicenter of tumor is < 2 cm from costal or diaphragmatic pleura and is > 2 cm from trachea, main and lobar bronchi). No squamous cell patients were accrued on the study at the time of study closure.

Other key eligibility criteria included measurable disease, age ≥ 18 years, ECOG performance status < or = 2, no gross hemoptysis (defined as bright red blood of ½ teaspoon or more), no history of thrombotic, hemorrhagic, or coagulopathy disorders, be free of active infection and be available for active follow up. Patients should also meet the key laboratory criteria including WBC > 3500/mm3 and ANC > 1500/mm3 and platelet count > 100 000/ mm3; total bilirubin < 1.5 mg/dl; creatinine < 1.5 mg/dl. or calculated creatinine clearance > 45 ml/min; INR < 1.5 and a PTT no greater than upper limits of normal within 1 week prior to registration. Patients with lung cancer placed on anticoagulant therapy for a thrombotic event are not eligible for this study.

Patients must have received no prior chemotherapy for metastatic disease. Patient may have received chemotherapy in adjuvant setting or with radiation for stage III disease. However, the disease-free interval from last chemotherapy had to be more than six months. Patients must be disease free for > 5 years if they had a prior second malignancy and agreed to use an acceptable method of contraception for the duration of the study.

Patients meeting any of the following exclusion criteria were not enrolled in the study: untreated CNS disease with the exception of treated brain metastasis; a history of ≥Grade 2 peripheral neuropathy within 14 days before enrollment; known previous sensitivity reactions to boron, or mannitol; known HIV positivity; myocardial infarction within 6 months prior to enrollment or New York Hospital Association (NYHA) Class III or IV heart failure, uncontrolled angina, severe uncontrolled ventricular arrhythmias, or electrocardiographic evidence of acute ischemia or active conduction system abnormalities; inadequately controlled hypertension (defined as systolic blood pressure > 150 mmHg and/or diastolic blood pressure of > 100 mmHg); history of stroke or transient ischemia attack or significant vascular disease (e.g. aortic aneurysm requiring surgical repair or recent peripheral arterial thrombosis) within 6 months prior to day 1; major surgical procedure, open biopsy, or significant traumatic injury within 28 days prior to day 1, anticipation of need for major surgical procedure during the course of the study; minor surgical procedure such as fine needle aspirations or core biopsies (excluding the placement of vascular access device) within 7 days prior to day 1; proteinuria as demonstrated by urine protein: creatinine ratio > 1.0 at screening; history of abdominal fistula, gastrointestinal perforation, or intra-abdominal abscess within 6 months prior to day 1; serious, non-healing wound, ulcer, or bone fracture; patient has received other investigational drugs with 14 days before enrollment or is expected to participate in an experimental drug study during this study treatment or serious medical or psychiatric illness likely to interfere with participation in this clinical study.

The study was reviewed and approved by the institutional review board of the University of Massachusetts and voluntary written informed consent was obtained before performance of any study-related procedure not part of normal medical care.

Study Design

The study was designed as a phase I/II study to evaluate the efficacy and safety of combination chemotherapy with carboplatin, bortezomib and bevacizumab in patients with advanced NSCLC.

During the phase I portion of the study, three dose levels of weekly bortezomib were studied in conjunction with fixed dose carboplatin and bevacizumab on a 21 day cycle to define the maximum tolerated dose (MTD). A maximum of six cycles was administered. Patients with complete response, partial response or stable disease after six cycles were allowed to continue on single agent bevacizumab every 3 weeks as maintenance therapy until disease progression.

Dose limiting toxicity (DLT) was defined as: grade 4 neutropenia or thrombocytopenia, any non-hematological toxicity >grade 3 except alopecia or inadequately treated nausea or vomiting, neurosensory toxicity of grade 2 with pain or any neurotoxicity > grade 2 during the first cycle of treatment.

During phase I, a traditional dose escalation model was used. If no dose limiting toxicity (DLT) was observed in 3 patients during the first cycle, the next dose level was accrued. If 1 DLT was observed, 3 additional patients were accrued to the dose level. If no additional DLTs were observed, the next dose level was accrued. However, if 2 or more DLTs were observed in a given dose level, one dose level below was to be defined as the maximum tolerated dose (MTD).

The following three pre-planned three dose levels of weekly bortezomib (day 1 and day 8) were studied with fixed doses of carboplatin AUC 6, bevacizumab 15 mg/kg on day 1 of every 21 day cycles: level I : 1.3 mg/m2; level II: 1.6 mg/m2 and level III 1.8 mg/m2. In phase II, either level III or MTD was administered in the same q21day cycle to evaluate the efficacy and safety of the regimen in first-line treatment of advanced NSCLC.

Drug administration

Bortezomib was supplied as a sterile lyophilized powder for reconstitution in vials containing bortezomib and mannitol at a 1:10 ratio. Bortezomib was administered as an intravenous push over 3 to 5 seconds followed by a standard saline flush or through a running IV line after carboplatin infusion on day 1 and bortezomib alone on day 8 of each cycle. The dose of bortezomib was determined based on body surface area.

Bevacizumab was supplied as a clear to slightly opalescent, colorless to pale brown, sterile liquid concentrate for solution for intravenous infusion. Bevacizumab was administered as an intravenous infusion on day 1 of each cycle. The dose of bevacizumab was based on actual body weight on day 1, cycle 1 of treatment. Unless there is a significant change (e.g., loss or gain of > 10% of the body weight), the dose remained the same in subsequent cycles. The initial dose was delivered over 90±15 minutes. If the first infusion is tolerated, the second infusion was delivered over 60±10 minutes and all subsequent infusions were delivered over 30±10 minutes.

Carboplatin is commercially available and supplied as a sterile lyophilized powder in single dose vials and was administered as an intravenous infusion over 30 minutes following bevacizumab infusion on day 1 of each cycle. Carboplatin dosage was calculated, for a target AUC 6 using the Calvert formula. The drugs were administered in the following order: bevacizumab followed by carboplatin followed by bortezomib.

Before each drug dose, the patients were evaluated for possible toxicities that may have occurred after the previous dose(s). Toxicities were assessed according to the NCI Common Terminology Criteria for Adverse Events (CTCAE), Version 3.0. Patients were required to have ANC > 1500 and platelets > 100, 000 prior to day one of each cycle. For non-hematologic toxicities grade 3 or greater, study drugs were held for up to 2 weeks until the toxicity returned to Grade 1 or better. On day 8, bortezomib was held if ANC < 1000 or platelet count < 100,000. The patient received one dose level lower of bortezomib if ANC 1000–1500 and platelet count > 100,000 and full dose if ANC > 1000 and platelet count > 100,000. If, after the study drugs were held, the toxicity does not resolve, as defined above, then the patient was discontinued from the protocol therapy. A treatment cycle delay or interruption of > 4 weeks because of toxicity resulted in removal of the patient from the study. If a treatment cycle was delayed due to hematological toxicity, all three agents including bevacizumab were held. If dose reduction is required for bortezomib on D8 of a particular cycle, the lowest prior dose level was used in both D1 and D8 of the subsequent cycles.

The anti-emetic regimen consisted of dexamethasone 20 mg intravenous on day 1, and ondansetron 8 mg intravenous on day 1 and ondansteron 8mg oral or intravenous on day 8. Patients were allowed to receive full supportive care, including transfusions of blood and blood products, erythropoietin, antibiotics and antiemetics when appropriate.

Patient evaluation

Patients enrolled in this study were evaluated clinically and with standard laboratory tests before and at regular intervals during their participation in this study. Safety evaluations consisted of medical interviews, recording of adverse events, physical examinations, blood pressure, and laboratory measurements. Patients were evaluated for adverse events (all grades), serious adverse events, and adverse events requiring study drug interruption or discontinuation at each study visit for the duration of their participation in the study. During cycle 1, all patients were monitored weekly with physical examination, blood count, vital signs and toxicity evaluation.

The patient’s target and non-target lesions were measured by conventional techniques and the overall disease response was assessed according to the Response Evaluation Criteria In Solid Tumors (RECIST) criteria every two cycles starting with cycle 2 (32).

An end of therapy visit was conducted 10 days after administration of the last dose of study drug and an end of study visit was conducted six weeks after the end of therapy visit or last chemotherapy cycle. Safety and efficacy evaluations were performed during these visits. Patients were followed at least every 3 months after the last dose of protocol therapy visit for two years to evaluate the long-term toxicity as well as the survival data.

Biomarker evaluation

Recent studies have identified the driver mutations in a subset of patient with NSCLC with adenocarcinoma histology (19). The mutations in EGFR and k-ras oncogene, as well as the translocation in EML/alk fusion protein can have a significant impact on the outcome of patients in the lung cancer therapeutic trials. The molecular mutation data for k-ras, EGFR and eml/alk translocation was collected in our patients. The purpose is to correlate with the clinical outcome including response rate and survival and to see whether there will be any hint of activity of this regimen in any particular subgroup. The tests were performed as part of routine clinical practice at our institution. The test were performed on formalin fixed, paraffin-embedded blocks or recut sections from the block prepared on slides including one stained with hematoxylin and eosin. EGFR mutation and k-ras mutation analysis was done by the molecular pathology department at University of Massachusetts.

Statistical Method and Analysis

The study was designed as a phaseI/II study. In the phase I portion, the traditional dose escalation model was used and it was anticipated that 9–18 patients would be enrolled. The phase II portion was designed based on the optimal two-stage phase II study by Simons et al (31). The efficacy of the combination chemotherapy was designed to be evaluated by response rate.

The study was designed to test the null hypothesis that the response rate is 15% or less versus the alternative hypothesis that the response rate is at least 30%. The response rate from the carboplatin, paclitaxel and bevacizumab in the E4599 study was 27%. Our hypothesis was that if we could achieve a similar response rate with a better tolerated regimen, it would be worth while to pursue this combination in a larger randomized study. Nineteen patients were planned to be accrued in the first stage of phase II. If 3 or fewer responses were observed during the first stage then the treatment was to be concluded early for lack of efficacy. Otherwise, 36 additional patients will be accrued for the second stage. If 12 or fewer responses among 55 patients are observed by the end of this phase II study, no further investigation of this treatment regimen is warranted. The power under this design is 80% and the type I error is 0.05. The probability of early stopping at the first stage is 68.4% if the true response rate is 15%. If the true response rate is 30%, the probability of early stopping at the first stage is 13.3%. The study was stopped early due to poor accrual. There were only 9 patients in the phase II portion of the study at the time of study closure.

Descriptive statistics were used for baseline characteristics and safety assessment. The response rate was calculated for all evaluable patients along with the 95% confidence interval (95% CI). The progression-free survival and overall survival was calculated using the Kaplan-Meier method, along with their 95% CI.

Results

Patients Population

A total of 16 patients with advanced NSCLC were enrolled between February 2007 and October 2010. All patients were included in the safety and efficacy analyses and received at least one dose of the study drug. Table 1 shows the baseline characteristics of all patients. The majority of the patients (n=11; 69%) had adenocarcinoma as histology; four patients with poorly differentiated NSCLC and one patient with adenocarcinoma with bronchoalveolar features. 14 patients (88%) were current or former smokers and none of the patients had prior adjuvant chemotherapy. 13 patients (81%) had metastatic disease and the rest has stage IIIB with pleural effusion.

Table 1.

Patient Demographics and Clinical Characteristics (N=16)

Characteristic No. of
Patients
(N=16)		 %
Age, yrs
Median (range)		 63 (47–73)
Sex
Female		 7 44
ECOG PS
0
1
2
unknown		 2
10
2
2		 12.5
62.5
12.5
12.5
Histology
Adenocarcinoma
Adenocarcinoma with BAC feature
Poorly differentiated NSCLC		 11
1
4		 69
6
25
Smoking Hx
Smoker
Non-smoker		 14
2		 88
12
Stage
IIIB(effusion)
IV		 3
13		 19
81
Prior adjuvant chemotherapy 0 0
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Treatment and follow up

16 patients received a total of 74 cycles of combination of bortezomib, bevacizumab and carboplatin. The number patients treated at dose level I, II and III are 3, 4 and 9 patients respectively. One patient at dose level II was found to be ineligible due to pre-existing peripheral vascular disease on retrospective review and one additional patient (hence total of 4 patients) were treated at level II. There was no predefined DLT in all 16 patients in the cycle 1 of treatment. The recommended phase II dose is bortezomib 1.8 mg/m2 weekly on day 1 and day 8 in combination with carboplatin AUC of 6 and bevacizumab 15 mg/kg on every 21 day cycle. Overall, 9 patients were treated at the recommended phase II dose level till the study was decided to close due to poor accrual.

The median follow up was 9.1 months. 8 patients (50%) completed all six cycles of combination therapy and six patients continue on maintenance bevacizumab therapy. The main reason for discontinuation of therapy was disease progression (n=11 patients; 69%). One patient at dose level II come off the study due worsening of preexisting peripheral vascular disease; one patient due to intolerable toxicity (peripheral neuropathy and unexplained pain syndrome in sacral area) after cycle 5 and one patient died on study due to rapid progression of the disease and general deterioration of clinical condition after cycle 3. Two patients are ongoing treatment on maintenance bevacizumab at the time of the report.

All the patients received planned full doses of carboplatin and bevacizumab. The dose reduction/ delay occurred in 50% of the patients. Total of 12 cycles of combination therapy was administered in 3 patients at cohort level I (bortezomib 1.3 mg/m2). There was no dose delay or reduction in any patients. At dose level 2 (bortezomib 1.6 mg/m2), 4 patients received total of 20 cycles of combination therapy. Two patients required dose reduction in bortezomib at cycle 4 due to thrombocytopenia. Total of 9 patients in cohort 3 (bortezomib 1.8mg/m2) received 42 cycles of combination of bortezomib, carboplatin and bevacizumab. 5 patients require dose delay/reduction in dose of bortezomib. The major cause of dose/delay or reduction is due to thrombocytopenia and usually in cycle 5 or 6.

Toxicity

Toxicity was defined as an adverse event considered possibly, probably or definitely related to treatment. Toxicity was assessed according to the NCI Common Terminology Criteria for Adverse Events (CTCAE), Version 3.0. The combination of bortezomib, carboplatin and bevacizumab was generally very well tolerated.

Common treatment-related adverse events occurring throughout the trial are summarized in table 2. There was no dose limiting toxicity in cycle one in all 16 patients treated. The predominant hematological toxicity included thrombocytopenia and neutropenia. Grade 3 thrombocytopenia and neutropenia were common at recommended phase II dose of bortezomib1.8 mg/m2 and usually occur after cycle 4. None of the patients had neutropenic fever. One patient each required platelet and packed red cell infusion. None of the patient required growth factor support (G-CSF or erythropoietin).

Table 2.

Treatment Related Adverse Events (N=16) (CTC AE version 3.0)

Toxicity Cycle 1
Gr1/2
N (%)		 Cycle 1
Gr3/4
N (%)		 All cycles
Gr1/2
N (%)		 All cycles
Gr3/4
N (%)
Hematological
Anemia 7 (44) 0 14 (88) 1 (6)
thrombocytopenia 4 (25) 1 (8) 10 (62) 7 (58)
neutropenia 2 (12) 0 8 (50) 2 (12)
lymphopenia 1 (6) 0 9 (56) 4 (25)
Non-hematological
Fatigue 5 (31) 0 10 (62) 2 (12)
Anorexia 4 (25) 0 7 (44) 2 (12)
Nausea 9 (56) 0 11 (69) 3 (19)
Vomiting 1 (6) 0 5 (31) 3 (19)
Diarrhea 5 (31) 0 9 (56) 4 (25)
constipation 3 (19) 0 7 (44) 0
Pain 5 (31) 1 (6) 6 (38) 1 (6)
Dehydration 1 (6) 0 5 (31) 2 (12)
Hypertension 1 (6) 2 (12) 2 (12) 2 (12)
Hemorrhage 3 (25) 0 9 (56) 0
Hemorrhage (pulmonary) 0 0 0 1 (6)
Thrombosis 0 0 1 (6) 1 (6)
hyponatremia 10 (62) 1 (8) 12 (75) 0
hyperglycemia 5 (32) 0 9 (56) 0
hypomagnesemia 0 0 5 (31) 0
neuropathy 0 0 7 (44) 0
infections 2 (12) 0 4 (25) 2 (12)
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The common non-hematological toxicities involving > 30% of the patients include fatigue, anorexia, nausea, vomiting, diarrhea and hyponatremia. The majority of these toxicities were grade 1 or 2. At the dose level 3, one patient developed grade 3 diarrhea and dehydration, requiring a dose reduction. Another patient developed herpes zoster reactivation involving face and cornea, most probably related to bortezomib. She was treated with acyclovir and continued on acyclovir prophylaxis throughout the rest of the treatment.

Grade 1 or 2 peripheral neuropathy was observed in 7 out of 16 patients (43%). Only one patient discontinued treatment due to peripheral neuropathy and unexplained pain syndrome.

Two patients discontinued the treatment due to toxicity. One patient had worsening of existing peripheral vascular disease and in retrospect did not meet the eligibility criteria. Another patient at dose level 3 discontinued the treatment due to peripheral neuropathy and unexplained pain syndrome as mentioned previously.

Tumor response and survival

All 16 patients were evaluable for tumor response. There was no complete response; 6 patients had partial response making the overall response rate of 37.5 % (95% CI: 13.8-61.2). Out of 9 patients at dose level III, 4 patients (44%; 95% CI: 15.3-77.3) had partial response. The disease control rate (PR+SD) at four month was 50%.

The median follow up was 9.1months. The progression free and overall survival of all the patients were 5.0 months (95% CI: 3.1-8.4) and 9.9 months (95% CI: 8.2-14.1) respectively. The study included patients with performance status of 2. The median PFS and OS in 13 patients with PS of 0-1 were 5.5 months (95% CI: 3.2-12.2) and 10.9 months (95% CI: 8.2-34.1).

There were total of 9 patients treated at dose level 3 (the recommended phase II dose). The study was close prematurely due to poor accrual. The overall response rate in this group is 44% (95% CI: 15.3- 77.3). This cohort of patients had the median PFS of 5.5 months (95% CI: 3.1, 12.2) and OS of 10.9 months (95% CI: 8.0-14.1)

Biomarker data

Recent studies have shown that a subset of patients with NSCLC with adenocarcinoma histology carries driver mutations that can have a significant impact on prognosis and response to different targeted therapies (19). EGFR mutation is present in 15–20% of adenocarinoma and is associated with better prognosis and the response to EGFR tyrosine kinase inhibitors. The k-ras mutation is reported in 20–30% of adenocarcinoma and is associated with poor prognosis. EML/alk translocation is present in 4% of adenocarcinoma and has been associated with response to crizotinib (19).

We collected the mutation data from our patient in the current study. 13 patients had k-ras mutation testing and 4 (30%) have mutations in k-ras oncogene. Out of six patients with partial response to the combination regimen: 5 patients had wild type k-ras, one patient had k-ras mutation.

EGFR mutation status was available in 13 patients and all the patients had EGFR wild type. One patient on the study was subsequently found out to have eml/alk translocation. This patient had partial response to the combination of carboplatin, bortezomib and bevacizumab and had the progression free survival of 12 months. He subsequently was treated with crizotinib on a clinical trial and had prolonged disease control. Although this is clearly anecdotal, we found this interesting to have a very good partial response and prolong progression free survival to combination of carboplatin, bortezomib and bevacizumab in this particular patient.

Discussion

Bortezomib is a novel proteosome inhibitor that has shown interesting pre-clinical and clinical activity in NSCLC (13–17). In this study, we examined the combination of bortezomib with carboplatin and bevacizumab. We used weekly dosing of bortezomib in combination with standard doses of carboplatin and bevacizumab. The study was designed as a phase I/II study with the primary aim of phase I portion of the study is to determine the maximum tolerated dose of bortezomib that can be administered with standard doses of carboplatin and bevacizumab.

The combination of bortezomib, carboplatin and bevacizumab was generally very well tolerated. We did not notice any dose limiting toxicity during the first cycle in all 16 patients treated. Our recommended phase II dose level is bortezomib 1.8 mg/m2 on day 1 and day 8 with standard doses of carboplatin (AUC of 6) and bevacizumab (15 mg/kg). At this dose level, increased hematological toxicity, particularly neutropenia and thrombocytopenia, was seen in patients receiving more than 4 cycles. This required dose delays but 4 out of 9 patients completed all planned six cycles of chemotherapy. The major reason for discontinuation of treatment was disease progression and only one patient come of the study due to intolerable toxicity and this was not due to hematological toxicity.

The peripheral neuropathy was one of the known side effects of bortezomib (11, 12). 43% of patients in our study developed grade I or II peripheral neuropathy. One patient at dose level three had an unusual painful syndrome and stopped treatment after 5 cycles. Painful grade II sensory neuropathy or grade III peripheral neuropathy was not observed. This may be due to the fact that we used weekly regimen and the bortezomib was limited to six cycles with chemotherapy.

The primary end point of the phase II portion of the study was to estimate the response rate and progression free survival. The study was closed prematurely due to poor accrual and is under powered to definitely estimate the response rate and progression free survival. 4 out of 9 patients at the recommended phase II dose level received partial remission with the median PFS and OS of 5.5 months and 10.9 months. This exceeded our preplanned response rate of 30% or 3 out of initial planned 19 patients in the first stage of phase II portion of the study. Our response rate from bortezomib, carboplatin and bevacizumab seems to be comparable with bevacizumab in combination with other standard doublet chemotherapies (carboplatin/paclitaxel or gemcitabine/cisplatin or carboplatin/permetrexed) (6, 7). The number of patients in the phase II portion of this study is very small and the study is underpowered to draw any definite conclusion regarding the survival data and efficacy of this regimen. Regardless, the regimen was very well tolerated with interesting activity. A larger randomized phase II study with the standard doublet with bevacizumab as control arm will further define the activity of this regimen.

Bortezomib has been extensively studied in the treatment of metastatic NSCLC with mixed results (17–24). The standard dose of bortezomib 1.3 mg/m2 twice a week had limited single agent activity in patients with advanced NSCLC (18). In a multicenter phase II study, the addition of bortezomib to gemcitabine and carboplatin had shown interesting activity and survival (21). However, in other randomized phase II studies, the addition of bortezomib to single agent docetaxel, pemetrexed or erlotinib offered no statistically significant response or survival advantage (22–24). Our study is the first to evaluate the combination of bortezomib with chemotherapy and bevacizumab based regimen.

There is considerable interest to combine different targeted therapies. In pre-clinical models, bortezomib has been shown to have anti-angiogenic effects via direct or indirect effects on endothelial cell (25). In multiple myeloma derived endothelial cells, bortezomib triggered a dose dependent inhibition of vascular endothelial growth factor (VEGF) and interleukin-6 (IL-6) secretion, and induced down regulation of VEGF, IL-6, insulin-like growth factor-I, Angiopoietin 1 (Ang1), and Ang2 transcription (25). Bortezomib has also been shown to induce the autophagic death of proliferating endothelial cell (26).

Recent studies have shown increased level of VEGF and other proangiogenic factors in patients treated with bevacizumab (27, 28). VEGF feedback production in response to bevacizumab therapy was shown to be entirely host derived and tumor independent (29). High level of post-treatment plasma VEGF level has been associated with poorer outcome in NSCLC patients treated with bevacizumab (28). Bortezomib has been shown to augment the activity of bevacizumab by modulating hypoxia-inducible factor-1α (HIF-1α) transcription and subsequent pro-angiogenic factor production. In a phase I study, the combination of bortezomib and bevacizumab was well tolerated and the pharmacodynamic studies showed significant decrease in levels of pro-angiogenic factors including VEGF levels (30). When we designed this trial, we decided only to continue single agent bevacizumab as maintenance therapy after six cycles of the combination therapy based on initial ECOG study of bevacizumab in combination with carboplatin and paclitaxel. The majority of the patients who finished the planned six cycles of chemotherapy and received single agent bevacizumab progressed on single agent bevacizumab. It will be interesting in future trials to examine the role of bortezomib and bevacizumab maintenance therapy after initial induction chemotherapy with a platinum based regimen or the current combination regimen.

In summary, the combination of weekly bortezomib with standard doses of carboplatin and bevacizumab is very well tolerated in patients with advanced NSCLC. The recommended phase II dose is bortezomib 1.8 mg/m2 on day 1 and day 8 in combination with carboplatin AUC of 6 and bevacizumab 15 mg/kg on D1 every three week. Interesting clinical activity was observed in patients with advanced NSCLC and this trial is underpowered to determine the efficacy.

Figure 1.

Figure 1

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CT scans showing partial responses : a) lung nodule in patient #8 pre-chemotherapy; b) post 2 cycles; c) hilar mass and malignant pleural effusion in patient #11 pre-chemotherapy and d) post 4 cycles, marked reduction in hilar mass and complete resolution of malignant pleural effusion.

Figure 2.

Figure 2

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Kaplan-Meier plots for a) progression-free survival (PFS) b) overall survival (OS) of all patients and (c) PFS and OS of patient treated in cohort 3 (recommended phase II dose).

Table 3.

Results for known genetic mutations, response, PFS and OS (N=16)

Patient
ID		 Dose
level		 EGFR
mutation		 k-ras
mutation		 Eml/alk
transloc		 # of
cycles		 response PFS
(months)		 OS
(months)
01 1 n/a n/a n/a 6 SD 4.6 8.4
02 1 n/a n/a n/a 4 SD 2.4 7.6
03 1 wild pos n/a 2 PD 1.2 3.7
04 2 wild wild pos 15* PR 12.2 33.6
05 2 wild wild n/a 2 SD 1.5 9.5
06 2 wild pos n/a 6 SD 3.8 8.1
07 2 wild wild n/a 8* PR 8.3 26.7+
08 3 wild pos n/a 5 PR 9.2 12
09 3 wild wild n/a 3 SD 2.1 2.1
10 3 n/a n/a n/a 2 PD 1.1 9.8
11 3 wild wild n/a 8* PR 6.5 21.1+
12 3 wild wild n/a 4 SD 2.7 13.9
13 3 wild wild n/a 10 PR 5.4 10.8
14 3 wild wild n/a 4 SD 3.2 7.8
15 3 wild wild n/a 11*# PR 9.0+ 9.0+
16 3 wild pos n/a 9*# SD 7.0+ 7.0+
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*

patients continued on single agent bevacizumab after 6 cycles

#

ongoing active treatment

+

alive in follow up

Acknowledgement

Supported by Millennium Pharmaceuticals, Inc.

Footnotes

Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

Presented in part as an abstract at the 101st Annual Meeting of the American Association for Cancer Research, April 17–21, 2010.

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