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Expanding the Clinical Development of Bevacizumab

Cancer Therapy Evaluation Program, National Cancer Institute, Bethesda, Maryland, USA

Correspondence: Helen X. Chen, M.D., Cancer Therapy Evaluation Program, National Cancer Institute, 6130 Executive Boulevard, EPN 7131, Bethesda, Maryland 20892, USA. Telephone: 301-496-8798; Fax: 301-402-0428; e-mail: chenh{at}ctep.nci.nih.gov

	LEARNING OBJECTIVES

Top Learning Objectives Abstract Introduction Clinical Development of... Conclusions and Future... References

 
After completing this course, the reader will be able to:

1. Describe the clinical development of bevacizumab.
   
2. Summarize the clinical trials conducted using bevacizumab.
   
3. Discuss the anticipated role of bevacizumab in cancer treatment and future directions for research.
   

	ABSTRACT

Top Learning Objectives Abstract Introduction Clinical Development of... Conclusions and Future... References

 
Bevacizumab (Avastin^TM; Genentech, Inc.; South San Francisco, CA) is a recombinant, humanized monoclonal antibody to vascular endothelial growth factor, a key regulator of tumor angiogenesis. Bevacizumab demonstrated potent antitumor activity in preclinical models and has also shown biologic activity and clinical benefit in clinical studies. Notably, a randomized, placebo-controlled phase II trial in renal cell carcinoma demonstrated a significantly longer time to tumor progression with bevacizumab monotherapy. Furthermore, in a phase III trial for untreated advanced colorectal cancer, the addition of bevacizumab to chemotherapy led to significantly longer overall survival and progression-free survival times than chemotherapy alone. The clinical development of bevacizumab has been expanded to include confirmatory phase III trials and exploratory phase II trials in a variety of solid tumors and hematologic malignancies. Treatment regimens being examined include bevacizumab alone and in combination with conventional chemotherapy, radiation, immune therapy, and biologically targeted agents.

Key Words. Vascular endothelial growth factor • Clinical trial • Cancer • Monoclonal antibody • Bevacizumab

	INTRODUCTION

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The switch to an angiogenic phenotype represents a pivotal step in the multistage process toward malignancy [1] and is regulated by a diverse group of endogenous proangiogenic and antiangiogenic factors [2]. Vascular endothelial growth factor (VEGF) is one of the most potent promoters of angiogenesis and has been identified as a fundamental regulator of tumor neovascularization. VEGF is, thus, considered to be an important therapeutic target in antiangiogenic cancer therapy strategies [3]. Various approaches aimed at the VEGF pathway are currently under clinical investigation, including agents that target VEGF receptors and agents that neutralize VEGF.

Bevacizumab (Avastin^TM; Genentech, Inc.; South San Francisco, CA), a recombinant humanized monoclonal antibody (mAb), binds to all isoforms of human VEGF with high affinity [4]. In preclinical models, VEGF-neutralizing antibodies led to potent tumor growth inhibition in a number of human cancer xenograft and metastatic models [5, 6]. The antitumor effect of VEGF-neutralizing antibodies is enhanced by their combination with chemotherapy [7], radiation [8, 9], and other antiangiogenic agents [10].

Clinical experience with bevacizumab has demonstrated that the agent is biologically active. In a randomized phase II study in patients with renal cell carcinoma (RCC), bevacizumab monotherapy significantly prolonged the time to disease progression (TTP) [11]. In patients with breast cancer, bevacizumab was shown to induce objective tumor response with monotherapy [12], and enhance the response rate (RR) when combined with capecitabine, although no difference in TTP was observed in a phase III trial comparing the combination with capecitabine alone [13]. Phase II studies have also demonstrated promising clinical efficacy for bevacizumab in combination with chemotherapy in colorectal cancer and non-small cell lung cancer (NSCLC) [14, 15]. More recently, results from a phase III study in patients with untreated colorectal cancer have become available, which demonstrated a significant survival benefit conferred by the addition of bevacizumab to irinotecan/5-fluorouracil (5-FU)/leucovorin chemotherapy [16]. Adverse events associated with bevacizumab therapy mainly relate to the vascular system and include thrombosis, bleeding, hypertension, and proteinuria [17].

The demonstration of the biologic activity of bevacizumab, together with the pivotal role of VEGF in the pathophysiology of cancer, has provided the rationale for evaluating the role of bevacizumab in different tumor types and for exploring strategies to optimize its therapeutic potential. To further the development of bevacizumab, a collaborative research and development agreement has been established between the National Cancer Institute (NCI) and Genentech, Inc. Confirmatory phase III trials of bevacizumab are being conducted in breast and colon cancers, NSCLC, and RCC, and exploratory phase I and II trials in a variety of solid tumors and hematologic malignancies are ongoing (Table 1 and Table 2). Studies in breast, colorectal, and lung cancers are covered elsewhere in this supplement [14, 15, 18]. This review covers studies in other tumor types.

	CLINICAL DEVELOPMENT OF BEVACIZUMAB IN SOLID TUMORS AND HEMATOLOGIC MALIGNANCIES

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More than 30 clinical trials with bevacizumab in solid tumors and hematologic malignancies are planned, ongoing, or recently completed (Table 1 and Table 2). In general, the therapeutic strategies being evaluated can be divided into the following categories:

• bevacizumab monotherapy
  
• bevacizumab in combination with chemotherapy or radiotherapy
  
• bevacizumab in combination with other putative antiangiogenic strategies (e.g., low-dose interferon, thalidomide, low-dose, orally administered chemotherapy)
  
• bevacizumab in combination with tumor-targeted therapies (e.g., erlotinib, trastuzumab, rituximab, imatinib)
  
• bevacizumab in combination with immunotherapy (e.g., dendritic cells pulsed with tumor antigen).
  

Renal Cell Cancer
Standard therapy for advanced RCC is extremely limited. High-dose IL-2, the only agent approved for this disease, is associated with long-term remission, but only in a small subset of patients with stage IV disease [20, 21]. Interferon alpha (IFN-) is commonly used but is associated with only a modest, albeit statistically significant, survival benefit [22, 23]. New treatment options are clearly needed for this patient population.

Compared with many other tumor types, the pathophysiology of clear-cell renal carcinoma has distinctive features, with disruption of the von Hippel-Lindau (VHL) tumor-suppressor gene documented in the majority of hereditary or sporadic clear-cell renal carcinomas [24]. Loss of VHL function results in constitutive overexpression of hypoxia-inducible proteins, including VEGF, platelet-derived growth factor (PDGF)-ß, TGF-ß, and Glu-1 [25]. All RCCs are highly vascularized, and the vast majority of hypervascular RCCs demonstrate increased expression of VEGF, regardless of tumor size, stage, or grade [26]. In addition, increased levels of VEGF in serum are noted in patients with RCC, and these correlate with advanced disease and poor prognosis [27, 28]. In a xenograft model study, transfection of renal cancer cells with VEGF enhanced angiogenesis, tumor growth, and production of ascites [29].

The high frequency of VHL mutation and its close association with increased VEGF expression identified RCC as one of the most promising target diseases for anti-VEGF therapy. Accordingly, a prospective three-arm, double-blind, placebo-controlled, phase II trial of bevacizumab monotherapy was conducted at the NCI in patients with metastatic RCC [11]. In that trial, patients were randomized to high-dose (HD) bevacizumab (10 mg/kg every 2 weeks), low-dose (LD) bevacizumab (3 mg/kg every 2 weeks), or placebo (n = 39, n = 37, and n = 40, respectively). Primary end points were TTP and RR.

The majority of patients included in the trial had previously received IL-2, and approximately 25% of patients had received prior chemotherapy or IFN-. Prior therapies, baseline characteristics, and prognostic factors were well balanced in the three trial arms. After 110 patients were randomized (38 placebo, 35 LD, and 37 HD), an interim analysis showed a highly significant prolongation of TTP in the HD bevacizumab group compared with the placebo group (hazard ratio = 2.55, p < 0.001) (Fig. 1) and a small difference of borderline significance between the LD bevacizumab and placebo arms. These efficacy data satisfied the early stopping criteria. The difference in TTP was also confirmed by an independent panel of radiologists. Tumor response evaluation identified four partial responses in the HD bevacizumab treatment arm (RR = 10%). Although the trial failed to demonstrate a survival benefit, this is attributable, at least in part, to the small sample size and the protocol-defined crossover from the placebo arm after disease progression. Bevacizumab was well tolerated in the trial, with hypertension and asymptomatic proteinuria being the most common adverse events.

View larger version (15K): [in this window] [in a new window]   Figure 1. Time to disease progression in patients with renal cell carcinoma treated with high-dose (A) or low-dose (B) bevacizumab or placebo (reproduced with permission from Yang et al., 2003 [11]).

Gynecologic Cancers
Angiogenesis is crucial in the development of ovarian cancer. VEGF is closely linked to normal ovarian function and is required for the development of the corpus luteum and the maturation of the endometrium [30]. Elevated VEGF expression occurs in all stages of ovarian cancer and is associated with poor prognosis, including shorter survival [30–33]. In addition to its role in ovarian-cancer-associated angiogenesis [34, 35], VEGF overexpression is directly associated with the production of ascitic fluid [36], a feature probably related to the ability of VEGF to induce endothelial hyperpermeability [37]. The parent murine antibody of bevacizumab, A.4.6.1, demonstrated promising antitumor activity in a subcutaneous SKOV-3 human ovarian cancer xenograft model [38]. Interestingly, in the intraperitoneal model of the same cell line, A.4.6.1 produced only partial inhibition of tumor growth, but was associated with almost complete inhibition of ascites production.

These data provided the rationale for evaluation of bevacizumab in ovarian cancer. A Gynecologic Oncology Group (GOG) phase II study of bevacizumab monotherapy is ongoing, and a second trial will examine the combination of bevacizumab with daily, low-dose, oral cyclophosphamide. The concept of the second study is based on preclinical evidence that repetitive, low-dose (metronomic) chemotherapy, such as cyclophosphamide, decreases angiogenesis and retards the growth of cancer cells that have demonstrated in vitro resistance to the same chemotherapy [39, 40]. Furthermore, the combination of a metronomic chemotherapy regimen (vincristine) with specific VEGF receptor blockage sustains tumor regression, while the effect of either agent alone is transient [41].

VEGF is also an important regulator of angiogenesis in cervical cancer; VEGF expression is increased in cervical intraepithelial neoplasia and invasive cervical cancer [42, 43]. Therefore, a study by the GOG is evaluating bevacizumab monotherapy in patients with persistent/recurrent squamous cell cervical cancer.

Prostate Cancer
VEGF expression is increased in both localized and metastatic prostate tumors [44], and this VEGF overexpression correlates with greater MVD, worse tumor grade, and poor prognosis [45]. Furthermore, expression of the VEGF receptor (VEGFR)-1 and, to a lesser extent the VEGFR-2, has also been demonstrated for prostate tumor cells [46, 47].

Based on these observations, a phase II trial was conducted to evaluate bevacizumab monotherapy in androgen-independent prostate cancer [48]. In that trial, no objective partial or complete responses were observed, and no patient achieved a >50% decline in prostate-specific antigen. A phase II CALGB trial of bevacizumab in combination with docetaxel and estramustine for hormone-refractory prostate cancer has just completed accrual, and the results are awaited with interest.

In addition, the VEGF-associated inhibition of dendritic-cell maturation [49] and the augmented antitumor effect of peptide-pulsed dendritic cells by an anti-VEGF mAb [50], provide the rationale for clinical evaluation of VEGF antibodies in cancer immunotherapy. The first study to test this concept is ongoing, using a combined regimen of bevacizumab and autologous dendritic cells pulsed with prostatic acid phosphatase (PAP) and GM-CSF fusion protein (Provenge^®; Dendreon Corporation; Seattle, WA).

Head and Neck Cancer
As with other solid tumors, increased VEGF expression is documented in many forms of head and neck cancer, including esophageal [51, 52], oral and oropharyngeal [53], nasopharyngeal [54], and thyroid [55] cancers. In some tumor types, clinical responses to chemoradiotherapy and 5-year survival rates were significantly poorer in patients with high-serum VEGF concentrations [52].

Radiotherapy is an important treatment modality for head and neck cancer, but treatment failure is common [56]. In preclinical studies, VEGF production by tumor cells could be induced upon ionizing irradiation [57], and VEGF, in turn, protected the endothelial cells from the cell-killing effect of radiation [58]. Furthermore, in vivo administration of anti-VEGF antibodies prior to radiotherapy resulted in a greater than additive antitumor effect on human tumor xenografts [8, 9]. These findings support the possibility that disruption of the paracrine relationship between tumor and endothelium may enhance the efficacy of radiotherapy. A phase I, dose-escalation study was initiated to evaluate the safety and feasibility of bevacizumab in combination with chemoradiation in patients with locally advanced or recurrent head and neck cancer. In addition, a phase I/II study of the combination of bevacizumab with erlotinib has recently opened. Given the feasibility of tumor biopsy in this disease, tumor specimens will be examined for molecular effects of a dual EGFR and VEGF blockage on tumor cells and endothelial cells.

Pancreatic Cancer
Pancreatic cancer is a devastating disease with a poor prognosis, including an overall 5-year survival rate of less than 5% and a median survival time of 5–6 months after diagnosis [59]. Investigation of various chemotherapeutic regimens in the past few decades has not resulted in a significant improvement in survival. Testing of new agents with novel mechanisms of action is highly desirable in the effort to explore effective therapies for this disease.

Overexpression of VEGF is demonstrated in most clinical pancreatic cancer specimens [60, 61]. Although no association between MVD and prognosis was evident in some studies [61], analysis of a larger patient population showed positive correlations between strong cellular expression of VEGF and greater MVD, liver metastasis, and poor survival [60]. Expression of VEGF receptors (VEGFR-1 and VEGFR-2) has also been shown in pancreatic cancer cells, as well as in surrounding microvessels [62]. Moreover, some pancreatic cancer cell lines may be growth stimulated in vitro by recombinant VEGF [62–64]. Although several other angiogenic promoters are overexpressed in pancreatic cancer, VEGF-targeted agents, including the anti-VEGF mAb A.4.6.1, were effective in retarding the growth and spread of pancreatic cancer in animal models. These findings underscore the importance of VEGF in the growth and progression of pancreatic cancer. Three studies investigating bevacizumab combination therapy are planned or already in progress. The phase II trial with bevacizumab and gemcitabine has shown promising preliminary results in terms of RR (24% in the first 33 evaluable patients) [65]. Accrual is ongoing in this trial, and progression and overall survival data are pending.

Other Solid Tumor Types
The importance of VEGF is recognized in a number of less common tumor types. Notably, the potential for autocrine stimulation of the VEGF receptor has been identified in malignant melanoma and Kaposi’s sarcoma (KS) [66, 67]. These tumor types, therefore, represent promising targets for bevacizumab therapy.

Evaluations of single-agent bevacizumab are being undertaken in KS, melanoma, and hepatocellular carcinoma. Combination therapy with low-dose IFN- (either in the conventional form or pegelated) is being studied in malignant melanoma and carcinoid tumors. Other ongoing trials include a double-blind, placebo-controlled phase II trial in mesothelioma comparing progression-free survival in patients receiving chemotherapy with that of patients receiving chemotherapy plus bevacizumab. A phase II trial of bevacizumab/doxorubicin is also ongoing in advanced soft-tissue sarcoma.

Bevacizumab in Hematologic Malignancies and Lymphomas
While the role of the bone marrow microenvironment in hematopoiesis has long been recognized [68], the importance of endothelial cells and angiogenic factors in hematopoietic malignancies has only recently become apparent [69–71]. Although the precise mechanisms of the interaction are not fully understood, both autocrine and paracrine loops have been shown to mediate growth and survival signals for hematopoietic malignant cells [72]. Coexpression of VEGF and its receptors (Flt-1 and KDR) has been demonstrated in myeloblasts and immature myeloid elements in acute myeloid leukemia (AML), multiple myeloma (MM) [71], and myelodysplastic syndrome (MDS) [73], suggesting the potential for autocrine action of VEGF on these malignant cells. Indeed, VEGF has been shown to stimulate in vitro colony formation of leukemic cells derived from patients with chronic myelomonocytic leukemia (CMML) and MDS [73], and antibody neutralization of VEGF inhibited such effects. In addition, endothelial cell activation by VEGF results in the production of additional cytokines and hematopoietic growth factors (e.g., GM-CSF, IL-6, and IL-8), which, in turn, support leukemic cell growth through a paracrine mechanism [73, 74].

Increase in marrow vascularity and overexpression of proangiogenic factors, including VEGF, have been observed in patients with various hematologic malignancies, such as CMML, AML [75], MDS [76], and MM [77]. Furthermore, increased VEGF expression was shown to be an independent negative prognostic factor in AML [76].

The body of evidence, therefore, substantiates a role of VEGF and other proangiogenic factors in the pathophysiology of hematologic malignancies and provides the rationale for VEGF-targeted therapy in these indications [78]. Currently, the single-agent activity of bevacizumab is being evaluated in MDS and non-Hodgkin’s lymphoma (NHL), and bevacizumab in combination with chemotherapy is being assessed in patients with AML and chronic myeloid leukemia (CML) in blast crisis. A comparison of single-agent bevacizumab with bevacizumab/thalidomide combination therapy in patients with MM is ongoing. The feasibility and efficacy results for bevacizumab in hematologic malignancies are eagerly awaited.

	CONCLUSIONS AND FUTURE DIRECTIONS

Top Learning Objectives Abstract Introduction Clinical Development of... Conclusions and Future... References

 
The demonstration of a delay in RCC progression with bevacizumab therapy and the survival benefit conferred by the addition of bevacizumab to chemotherapy in colorectal cancer have provided proof-of-principle evidence that bevacizumab is biologically active and that tumor-expressed VEGF is a valid target for anticancer therapy. Ongoing phase III and phase I/II studies will determine the clinical utility of bevacizumab in specific tumor types and stages and provide insight into further clinical development of this agent.

It is well recognized that, in any given tumor, a multitude of angiogenic factors can be demonstrated. Furthermore, the profiles and relative contributions of individual factors not only vary among tumor types, but may also evolve within the same patients, owing to inherent tumor biology or driven by therapeutic interventions [2]. Such complexity of tumor growth and angiogenesis pathways suggests that, although VEGF is one of the most important angiogenic promoters, suppression of VEGF alone will not be sufficient for all tumor types and during all stages of tumor progression. Indeed, even in RCC, where the role of VEGF is assumed to be predominant, mixed tumor responses were not uncommon with bevacizumab monotherapy, and tumor progression, although significantly delayed, eventually occurred after a median of 5–6 months [11]. In the phase III trial in pretreated metastatic breast cancer, the addition of bevacizumab to capecitabine was not associated with a significant increase in TTP, despite its ability to increase the RR [13].

The challenges in the clinical development of bevacizumab are common to other single targeted agents, particularly those directed at growth factors. To achieve the full potential of molecularly targeted therapies, strategies should be developed to ensure appropriate patient selection and rational combination with cytotoxic and multiple targeted agents. Critical to these aims are a better understanding of tumor biology and more in-depth preclinical and clinical correlative studies. Specific tasks of ongoing and future studies should include understanding the mechanisms of resistance, identifying predictive markers of response or resistance, establishing reliable assays for clinical use, and evaluation of proof-of-principle combination regimens in appropriate tumor models.

Other issues to be addressed include selection of optimal bevacizumab doses and schedules, as well as the identification of risk factors and management of adverse effects. These are discussed in more detail elsewhere in this supplement [17].

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