William J. Gradishar
Cancer development is a complex process involving the conversion of a normal cell into a cancer cell. In addition to this malignant transformation, a tumor requires a vascular supply. Over 30 years ago, Judah Folkman proposed the theory that tumors are unable to grow beyond a size of 2 to 3 mm in the absence of a new vascular supply.1 Research has confirmed that a necessary step in the growth of both the primary tumor and metastases is the development of vasculature, termed angiogenesis.2, 3 Normally, this process is tightly regulated by activators and inhibitors of angiogenesis (Table 28.1). Tumors can produce some of these activator molecules or down-regulate expression of inhibitors, therefore altering the balance in favor of an “angiogenic switch.”4, 5 Due to the imbalance, these endothelial cells continue to proliferate, unlike normal endothelial cells, which mature and then become quiescent. In addition to an altered growth rate, the cytokine balance results in tumor-associated blood vessels that are structurally abnormal. They are not organized into an intact network of capillaries; the perivascular cells are more loosely associated with the endothelial cells, leading to a leaky basement membrane; and tumor cells can become integrated into the vessel wall.6 The importance of neoangiogenesis to tumor growth provides a rationale for exploring antiangiogenic therapies in the treatment of cancer.1
As understanding of the complex process of angiogenesis in tumors has advanced, many potential strategies for disrupting this process have been identified.7Drugs currently in development are grouped into five categories based on mechanism of action: (a) agents that block breakdown of the extracellular matrix, such as matrix metalloproteinase inhibitors (MMPIs); (b) drugs that inhibit endothelial cells; (c) agents that inhibit endothelial cell-specific integrin/survival signaling; (d) drugs that block activation of angiogenesis, such as anti–vascular endothelial growth factor (VEGF) agents; and (e) drugs with unknown mechanisms of action.8
Dozens of agents have been developed and are in various stages of clinical development. This chapter highlights promising agents and the results of clinical trials. To date, the only FDA-approved agent is bevacizumab (Avastin), a recombinant humanized anti-VEGF monoclonal antibody. There have been a number of disappointing clinical trials with a variety of single-agent antiangiogenesis therapies. Due to the complexity of tumorigenesis and angiogenesis, this is not altogether surprising. Future strategies should include combinations of agents that target various processes involved in tumor growth. In addition, correlative studies and measurement of potential targets, such as VEGF or MMP, should be done to better identify the population most likely to benefit or to elucidate whether the desired endpoint is being achieved with a given agent.
MATRIX METALLOPROTEINASE INHIBITORS
MMPs are a family of structurally related zinc-containing endopeptidases that are involved in the degradation of extracellular matrix components (ECM).9 MMP gene expression and enzymatic activity is an important physiologic process that plays an important role in embryogenesis, wound healing, and the female reproductive cycle.10 Because MMPs facilitate the breakdown of the basement membrane and the underlying stroma, they have been implicated in tumor invasion and metastasis formation.
Over 20 members of the MMP family have been identified. The MMPs are produced by a variety of different cells, including fibroblasts, epithelial cells, inflammatory cells, and endothelial cells.11 Several studies have demonstrated high levels of MMPs in tumors and in the plasma and urine of patients with malignancy.9, 12 MMP expression is also increased in metastatic tumors as compared with the primary tumor.13 Physiologic inhibitors known as tissue inhibitors of metalloproteinases (TIMPs) control MMP activity. The TIMP family currently consists of four members: TIMP-1, TIMP-2, TIMP-3, and TIMP-4.10
TABLE 28.1 ANGIOGENIC ACTIVATORS AND INHIBITORS
For a primary tumor to progress locally through adjacent normal tissue or for metastases to expand at a site distant from the primary tumor, the ECM must be digested.14 MMPs are also directly involved in the angiogenic response, as they mediate remodeling and invasion of the ECM by new vessels.15, 16 MMPs promote angiogenesis by regulating endothelial cell attachment, proliferation, and migration.15, 16 These observations suggest that matrix metalloproteinase inhibitors (MMPIs) could inhibit tumor progression at both the primary tumor site and sites of metastases. A concern with the long-term administration of MMPIs is the effect they may have on normal physiologic processes that require MMP activity, such as wound repair or reproduction.17, 18 Several MMPIs have been developed, but results in clinical trials of patients with advanced malignancy have been disappointing; therefore, ongoing studies are limited.
Marimastat is the most extensively tested MMPI. It is a synthetic MMPI that is orally bioavailable and is nonspecific, as it inhibits the activity of MMP-1, -2, -3, -7, and -9.19 Phase I clinical trials demonstrated that the drug was relatively well tolerated, and musculoskeletal complaints were the most common adverse effect.20, 21, 22 Marimastat has been evaluated in patients with pancreatic, breast, lung, ovarian, colorectal, gastric, and prostate cancers and glioblastoma. It was tested in a phase III randomized trial as first-line therapy in patients with unresectable pancreatic cancer. A total of 414 patients were randomized to receive 5, 10, or 25 mg bid of marimastat or 1,000 mg/m2 of gemcitabine.23 There was evidence of a dose-response, as patients treated with the 25-mg dose had better overall survival than patients treated with the lower doses of marimastat. There was no difference in 1-year survival between patients treated with gemcitabine (19%) and the 25-mg dose of marimastat (20%). Patients treated with gemcitabine had a statistically significant improvement in pain.
A number of phase III trials were performed to test the role of marimastat in patients with metastatic disease who had responded or had stable disease after first-line chemotherapy. These studies included patients with breast cancer, small cell lung cancer, or gastric cancer.24, 25, 26 There was no improvement in overall or disease-free survival in these cohorts. There are a variety of reasons why marimastat may not have been effective. First, only a minority of patients had trough levels of marimastat that were therapeutic. Interestingly, higher levels of marimastat were associated with a higher mortality.26, 27 Higher drug levels were also associated with a rise in the serum concentrations of MMP-9, one of the MMPs associated with tumor grade and metastases. Therefore, despite therapy with an MMPI, other mechanisms overcame inhibition and resulted in stimulation of MMP expression.
More recently marimastat has also been tested in combination with other antiangiogenic therapies. Fifty patients with a variety of advanced malignancies were treated with marimastat, captopril, and dalteparin. One patient with renal cell carcinoma had a prolonged partial remission, while three other patients with renal cell carcinoma had stable disease for 204 to 337 days.28 There were three major hemorrhagic events, one of which was fatal. In addition, 14% of the patients had grade 3 musculoskeletal toxicity that responded to a reduction in the dose of marimastat; this toxicity has been the most significant in other studies of marimastat.22, 29 It is possible that combination regimens with other antiangiogenic therapies will have more efficacy than single-agent MMPIs.
The disappointing results of phase III trials with marimastat demonstrate the complexities of this targeted therapy. Inhibition of MMP may not achieve the desired effect, and instead MMP expression may be stimulated. This phenomenon highlights the importance of correlative studies in clinical trials. It is necessary to measure the proposed target during therapy to ensure that the desired effect is achieved, for if it is not, this may explain negative results.
Prinomastat and BAY 12-9566 are two additional MMPIs that have shown disappointing results in clinical trials. BAY 12-9566 was found to result in significantly shorter survival times than gemcitabine in a randomized phase III trial in patients with advanced pancreatic cancer.30 In addition, prinomastat combined with chemotherapy for patients with advanced lung cancer demonstrated a significantly higher incidence of venous thromboembolism than either prinomastat or chemotherapy alone.31 Due to their lack of efficacy and their toxicity profile, these agents are no longer in development.
BMS-275291 is another MMPI currently in clinical trials. It is similar in structure to marimastat, is orally bioavailable, and inhibits a number of MMPs, including MMP-1, -2, -7, -9, and -14.32 It was designed to be specific for the MMPs and not to affect other metalloproteinases, such as sheddases, involved in the release of tumor necrosis factor-α (TNF-α), TNF-α receptor, and interleukin-6 receptor, which were thought to account for musculoskeletal toxicity.33 In a phase I study in patients with a variety of advanced malignancies, 44 patients were treated with escalating doses of BMS-275291, from 600 to 2,400 mg/day.33 The maximum tolerated dose was not achieved, as there were no dose-limiting toxicities at 900, 1,800, or 2,400 mg/day. The most common adverse events were grade 1 or 2 myalgias and arthralgias in 59% of patients. Unlike previous trials with MMPIs, the musculoskeletal toxicity did not necessitate discontinuation of the drug. A rash developed in 23% of patients, but in 90% of cases it resolved despite continuation of the drug. The other observed side effects included fatigue (32%), nausea (23%), and headache (16%). The median time on the study was 8 weeks, although 6 patients were treated for over 8 months, and 3 patients were treated for over 1 year. There were no objective partial or complete tumor responses; stable disease was seen in 27% of cases.33
The mechanism of action of these drugs suggests that they may be more effective in the setting of small-volume disease or micrometastatic disease. This provides the rationale for treatment in the adjuvant setting. A randomized phase II trial was performed in patients with stage I (T1c) to IIIa breast cancer who were to receive adjuvant therapy with tamoxifen alone, four cycles of doxorubicin and cyclophosphamide (AC), or AC followed by four cycles of paclitaxel.34Patients were randomly assigned to receive 1,200 mg of BMS-275291 once daily or placebo for 1 year. Therapy started concurrently with systemic chemotherapy or with tamoxifen if patients were not receiving chemotherapy. The primary objectives of the study were to evaluate rates of drug discontinuation due to intolerance and to ensure that adequate trough levels could be achieved. In the 71 patients treated in the study, the major toxicity was musculoskeletal, seen in 36% of the BMS-275291 group and characterized primarily by tendonitis and bursitis.34 The symptoms were reversible after drug discontinuation. Two patients in this group developed palpable nontender nodules on tendon surfaces. Musculoskeletal symptoms were seen in 21% of the placebo group, which was not statistically significantly different. Discontinuation rates at 1 year were similar, 33% in the treatment group and 21% in the placebo group. Only 19% of patients had trough levels that were higher than the IC90 for MMP-9 more than half the time. The musculoskeletal toxicity, discontinuation rate, and therapeutic trough levels in a minority of patients resulted in early termination of this study and the conclusion that this drug was not feasible in the adjuvant setting. Clinical trials continue with this agent in other settings, such as hormone-refractory prostate cancer.
DRUGS THAT INHIBIT ENDOTHELIAL CELLS
TNP-470, is a synthetic analog of fumagillin, which is secreted by the fungus Aspergillus fumigatus Fresenius and was isolated as a contaminant from an endothelial cell culture.35 Fumagillin inhibited in vitro endothelial cell proliferation and in vivo tumor growth and angiogenesis but was associated with significant weight loss in mice.35 TNP-470 is one of the compounds obtained when fumagillin undergoes alkaline hydrolysis and is more potent. TNP-470 blocks endothelial cell proliferation. The potential mechanisms of action include inhibition of the metalloprotease methionine animopeptidase (MetAP-2)36 and inhibition of protein kinases, including protein kinase C (PKC) and mitogen-activated protein kinase (MAPK).37 In addition, TNP-470 has been shown to be cytotoxic to cancer cell lines.38
Phase I trials of TNP-470 have been conducted in patients with AIDS-related Kaposi's sarcoma, cervical cancer, and other advanced malignancies. Treatment schedules have included drug administration once a week, every other day, or three times a week.39, 40, 41 Pharmacokinetic data from these studies demonstrated a very short half-life for TNP-470 and its metabolites, from 2 to 6 minutes.40 After the initial trials, the most commonly used dosing regimen has been 60 mg/m2 infused over 1 hour three times per week.42, 43 The most common toxicity, and the dose-limiting toxicity, in the phase I trials was neurotoxicity.39, 40 Neurotoxicity developed in 44% of patients; it included dizziness/vertigo (43%), ataxia (24%), short-term memory loss (24%), confusion (14%), anxiety/depression (14%), and insomnia (5%).40 At the highest dose level (235 mg/m2 once per week), ataxia was the dose-limiting toxicity, with 33% of patients developing grade 3 or 4 toxicity. Other nonneurologic toxicities include nausea (19%), anorexia (19%), and fatigue (19%).40 A phase II trial was performed in 33 patients with metastatic renal cell cancer; 60 mg/m2 TNP-470 was infused over 1 hour three times per week.42 Only 1 patient had a partial response, but 6 others had stable disease for over 6 months. Neurotoxicity was the most common adverse effect (67%); it included cerebellar symptoms, psychiatric changes, and confusion. Other toxicities included fatigue/asthenia (60%), anorexia (36%), and nausea (36%). Fifteen percent of patients discontinued therapy due to neurotoxicity.42
TNP-470 has also been tested in combination with traditional chemotherapy. A total of 32 patients with advanced solid tumors were enrolled in a dose-finding study with TNP-470 and paclitaxel.43 The regimen of paclitaxel 225 mg/m2 every 3 weeks and TNP-470 60 mg/m2 three times per week was well tolerated, and TNP-470 did not adversely affect the clearance of paclitaxel.43 Partial responses were observed in 25% of patients, and 53% had stable disease. The incidence of hematologic toxicities did not appear to be increased with the addition of TNP-470. Nonhematologic toxicities included fatigue (78%), peripheral neuropathy (66%), arthralgias (62%), nausea (59%), diarrhea (41%), dizziness (34%), abnormal vision (34%), and abnormal gait (22%). The neurotoxicity in this study was predominantly mild (grade 1 or 2) and reversible after discontinuation of treatment. Formal neuropsychologic testing was performed as part of this trial and demonstrated that the most commonly seen deficits included decline in executive function, memory, and motor dexterity. In only 9% of cases did function not return to baseline. The addition of carboplatin to a regimen of TNP-470 60 mg/m2 thrice weekly and paclitaxel 225 mg/m2 every 3 weeks was evaluated in a clinical trial.44 Carboplatin at an AUC of 6 was found to be well tolerated, with toxicity similar to that in the previous study by Herbst et al. Responses were also similar; 24% of patients had a partial response and 47% had stable disease.
The responses to date have been relatively modest with TNP-470. Given the short half-life of the drug and its metabolites, a different administration schedule may be more effective. Studies in animal models have demonstrated that daily administration was more effective than thrice weekly administration at inhibiting tumor growth, metastasis, and angiogenesis.45 Therefore, a phase I trial was designed to determine the feasibility of continuous infusion TNP-470 with or without paclitaxel and carboplatin in patients with advanced solid tumors. The most tolerable regimen when given with paclitaxel (200 mg/m2) and carboplatin (AUC 5 to 6) was TNP-470 at 2.5 mg/m2 continuous infusion for 5 days every week.46 Response data have not been published.
INHIBITION OF ENDOTHELIAL CELL–SPECIFIC INTEGRIN/SURVIVAL SIGNALING
EMD 121974 (Cilengitide)
The interaction between endothelial cells and the extracellular matrix is critical to neoangiogenesis. The integrin αVβ3 is expressed on a number of cells, including endothelial cells; is responsible for their binding to the extracellular matrix components during angiogenesis; and is up-regulated by basic fibroblast growth factor (bFGF).47 Apoptosis is induced in vascular cells when this interaction is inhibited.48 In patients with breast cancer, αVβ3 was higher in patients with metastatic tumors and was predictive of relapse-free survival.49 EMD 121974 is a peptide developed to inhibit the αVβ3 receptor.47 A total of 37 patients with a variety of advanced malignancies were treated in a phase I study.50 The drug was given intravenously twice a week and was tested at doses from 30 to 1,600 mg/m2. No hematologic toxicity was seen, and nonhematologic toxicities, including nausea, anorexia, fatigue, and malaise, were mild, with none higher than grade 2.50 In addition, the plasma half-life is short, 3 to 4 hours. Therefore, the optimal dose and administration schedule have not yet been determined. Further phase I and II studies are ongoing in patients with glioblastoma multiforme, melanoma, acute myeloid leukemia (AML), and other advanced malignancies.
INHIBITION OF ANGIOGENESIS ACTIVATORS
The VEGF signaling family is an important factor in both physiologic and pathologic angiogenesis. The complexities of this system are beyond the scope of this chapter; therefore, only a brief overview is provided, but detailed reviews are available.51, 52, 53 The VEGF family is made up of four main ligands and three tyrosine kinase receptors (see Table 28.2). VEGF-A, also referred to as VEGF, is a primary regulator of angiogenesis. It is important in early endothelial cell survival, but established blood vessels are not VEGF-dependent.51 Vascular permeability is also mediated by VEGF, which forms fenestrations in blood vessels.52, 53 VEGF expression can be induced by hypoxia and hypoxia-inducible factor 1 (HIF-1); a number of cytokines and growth factors (including fibroblast growth factor-4, platelet-derived growth factor [PDGF], TNF-α, and transforming growth factor-β [TGF-β]); UV-B radiation; and inactivation of the vHL (von Hippel Landau) tumor suppressor gene. Natural inhibitors of VEGF include cytokines, such as IL-10 and IL-13.
VEGF-A exerts most of its actions by binding to VEGFR-2, although it also binds to VEGFR-1(flt-1). VEGFR-2 (KDR in humans or flk-1 in the mouse) is expressed on both vascular and lymphatic endothelial cells and is the main regulator of angiogenesis, endothelial cell proliferation and survival, and vascular permeability.52 The scope of activity of VEGFR-1 varies; if VEGFR-2 is not present, activation of VEGFR-1 will not result in endothelial cell proliferation, yet cell migration is induced. VEGFR-1 may also have an inhibitory role by binding to VEGF-A, thus interfering with its ability to activate VEGFR-2.51 VEGFR-3 is found predominantly in lymphatics and likely has an important role in lymphangiogenesis, but can also be induced in tumor-associated endothelial cells.54
TABLE 28.2 VEGF (VASCULAR ENDOTHELIAL GROWTH FACTOR) SIGNALING FAMILY
VEGF and its receptors have been found to be overexpressed in a number of malignant cell lines and tumors, including colorectal, gastric, hepatocellular, lung, breast, and endometrial cancers, AIDS-associated Kaposi's sarcoma, and AML.55, 56, 57, 58, 59 The significance of this overexpression is not completely understood. A number of studies have demonstrated that increased expression of VEGF correlates with an increased vessel density, but the findings on the relationship between VEGFR-2 and vessel density have been mixed.56, 60, 61 Retrospective studies to determine whether VEGF or receptor expression correlates with prognosis have not been definitive. In one of the largest series performed, tumors from 259 patients with colorectal cancer were examined, and survival was found to be lower in patients whose tumors were positive for VEGF.61 Similar studies in patients with gastric and endometrial cancer, however, did not find VEGF expression to have prognostic significance.60, 62 Despite these uncertainties, animal studies have demonstrated that inhibiting VEGF signaling interrupts tumor growth and invasion.54, 63, 64, 65, 66 Therefore, based on evidence for the critical role of VEGF in angiogenesis and preclinical data suggesting that tumor-associated angiogenesis and growth could be altered by targeting this system, a number of agents aimed at the VEGF family have been developed and are presented here (Table 28.3).
TABLE 28.3 AGENTS TARGETING THE VEGF FAMILY
Bevacizumab is a recombinant humanized monoclonal antibody to VEGF. It binds to VEGF, including a number of its splice variants that are biologically active.67 In phase I trials, pharmacokinetic studies determined that the half-life of the drug is about 21 days at doses greater than 0.3 mg/kg.67 No patients developed antibodies to the recombinant antibody. Bevacizumab has been tested at doses from 3 to 20 mg/kg. Of interest, the dose-response relationship has not been consistent. In colorectal cancer, a dose of 5 mg/kg was more effective than 10 mg/kg.68 On the other hand, in renal cell cancer and non–small cell lung cancer, higher doses up to 15 mg/kg are more effective than lower doses.69,70 A number of clinical trials have been undertaken with bevacizumab either as a single agent or in combination with chemotherapy in a variety of malignancies.
Renal cell carcinoma was felt to be a potentially promising target for bevacizumab, since most clear cell renal carcinomas have a mutation in the vHL tumor suppressor gene, leading to HIF-1–mediated VEGF production.51, 69 A randomized phase II trial with single-agent bevacizumab was conducted in patients with metastatic clear cell renal carcinoma who had progressed after immunotherapy. A total of 116 patients were randomly assigned to either placebo, bevacizumab 3 mg/kg every 2 weeks, or bevacizumab 10 mg/kg every 2 weeks.69 There were no responses in the low-dose group, while a partial response was seen in 10% of patients in the higher dose group. Toxicities seen in this study included proteinuria (53%), hypertension (20%), and epistaxis (17%). There was a difference between progression-free survival in the high-dose group compared with the placebo group (4.8 vs. 2.5 months, P < .001), but no improvement in overall survival occurred in any of the cohorts.69 Single-agent therapy was not felt to be promising in renal cell cancer, and studies were undertaken with combination therapy. Thalidomide plus bevacizumab was also not effective.71 A phase II trial combined bevacizumab with erlotinib (Tarceva), an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor.72 All patients were treated with bevacizumab 10 mg/kg every 2 weeks and erlotinib 150 mg orally daily. Of 57 evaluable patients, a partial response was seen in 25% of patients, and 62% had either minor responses or stable disease.72 Grade 3 or 4 toxicities included hypertension (11%), diarrhea (9%), and rash (7%). Further studies are warranted to evaluate the role of this combination or others in the treatment of renal cell carcinoma. An ongoing phase III trial randomizes patients to either interferon-α alone or interferon-α plus bevacizumab.73
Bevacizumab has been approved by the FDA for use with 5-fluorouracil (5-FU)–based chemotherapy in first-line therapy of patients with metastatic colorectal cancer. This approval was based on two clinical trials in 917 patients with previously untreated metastatic disease.68, 74 In the phase II trial, patients were randomly assigned to one of three treatment regimens: 5-FU/leucovorin (LV), 5-FU/LV plus bevacizumab 5 mg/kg every 2 weeks, or 5FU/LV plus bevacizumab 10 mg/kg every 2 weeks.68 The 5-FU/LV in all groups was given weekly for the first 6 weeks of an 8-week cycle. The difference in overall median survival was not statistically significant. The time to progression was improved in the 5 mg/kg bevacizumab group when compared with the control group (9 months vs 5.2 months, P = .005), as was the overall response (40% vs. 17%, P = .029).68 The time to progression and overall response in the cohort that received the higher dose bevacizumab were no better than for the control group. The phase III trial randomly assigned 411 patients to irinotecan, bolus 5-FU, and LV (IFL) and 402 patients to IFL plus bevacizumab 5 mg/kg every 2 weeks.74 The group that received bevacizumab had a better overall response rate and improved progression-free and overall survival. The median survival was 15.6 months in the IFL cohort, and 20.3 months in the IFL plus bevacizumab group (P < .001).74 These significant results led to FDA approval of this agent in February 2004.
The toxicities associated with bevacizumab have been similar in these large studies; the most notable have been hypertension, proteinuria, minor and major hemorrhage, wound-healing complications, and gastrointestinal perforation. Hypertension is seen in up to 19% of cases, with grade 3 hypertension seen in 11% of bevacizumab-treated patients in the study of IFL with or without bevacizumab.74 Patients must be monitored carefully for development of hypertension, and antihypertensive therapy instituted promptly. If significant hypertension develops, the drug should be discontinued. Although rare, hypertensive encephalopathy did develop in 4 patients, of over 1,000 treated in clinical trials (bevacizumab [Avastin] package insert, Genentech, San Francisco, April 2004). Bleeding episodes have also been more frequent with bevacizumab treatment than with chemotherapy alone, 59% versus 11%.68 The majority of hemorrhagic complications were grade 1 or 2 episodes of epistaxis. In the phase II study by Kabbinavar, 10% of individuals experienced episodes of gastrointestinal hemorrhage, 43% of which were grade 3 or 4; 68 in the larger phase III trial, the incidence of grade 3 or 4 bleeding was not higher than that of the control group.74 Although rare, the bleeding can be severe. It is important to note that patients with central nervous system metastases were excluded from the clinical trials, and the safety of this therapy in that group has not been established.
The consequences of inhibition of VEGF signaling can be complex, since both hemorrhagic and thrombotic complications have been recognized with bevacizumab. A possible mechanism for these adverse events relates to the decreased ability of endothelial cells to respond to an injury when VEGF is inhibited, thereby leading to a hemorrhagic tendency. On the other hand, if vascular injury occurs, the coagulation system may be activated by exposure of tissue factor, and a thrombotic event may result.75 In clinical trials to date, thrombotic events have included deep venous thrombosis, pulmonary embolism, catheter-related thrombosis, stroke, and transient ischemic events. On August 4, 2004, Genentech issued a warning notifying physicians of an increased incidence of arterial thromboembolic events, including strokes, transient ischemic attacks, myocardial infarctions, and angina. In addition to bevacizumab exposure, risk factors for these arterial events were history of prior arterial thromboembolism or age greater than 65. If patients develop a thrombosis requiring anticoagulation while on bevacizumab, a recent report suggests that there is no increased risk of bleeding. Patients from the Hurwitz study74 who developed a thrombosis were placed on full-dose anticoagulation; 55% of patients receiving IFL and 83% of patients treated with IFL plus bevacizumab continued in the study. While on anticoagulation, grade 3 or 4 bleeding developed in 2 of 30 patients (6.7%) on IFL and in 2 of 58 patients (3.8%) receiving bevacizumab.76 Despite this report, a better understanding of the effects of anti-VEGF therapy on the hemostatic system is necessary before recommendations can be made for safe administration of this therapy with anticoagulation.
Bevacizumab has also shown promise in non–small cell lung cancer. Patients with stage IIIB or IV non–small cell lung cancer were randomized to one of three groups: carboplatin/paclitaxel every 3 weeks or carboplatin/paclitaxel plus bevacizumab 7.5 or 15 mg/kg.70 The bevacizumab was given after each cycle of carboplatin/paclitaxel. The difference in the median time to progression between the higher dose bevacizumab group and the control arm was statistically significant (7.4 months vs. 4.2 months, P = .023).70 The differences in overall response and survival were not statistically significant. The toxicities were similar to those reported in previous clinical trials with bevacizumab. It is important to note that major life-threatening bleeding occurred in 9% of patients receiving study drug, and fatal hemorrhage, primarily from hemoptysis, occurred in 6% of bevacizumab-treated patients. The bevacizumab doses in both arms of this study were higher than the 5-mg/kg dose that has been approved for the treatment of colorectal cancer.
A randomized phase III study in patients with metastatic breast cancer who had progressed after anthracycline and taxane therapy was conducted, and capecitabine was administered alone or with bevacizumab 15 mg/kg every 3 weeks.77 The overall response rate was greater in the combination arm (20% vs. 9%), but there was no improvement in progression-free survival. Additional phase III studies are currently being conducted in breast, colorectal, non–small cell lung, and renal cell cancers. In addition, numerous phase II studies are ongoing in patients with small cell lung, head and neck, pancreatic, hepatocellular, and ovarian cancers, melanoma, carcinoid tumors, soft-tissue sarcoma, Kaposi's sarcoma, mesothelioma, non-Hodgkin's lymphoma, chronic myelogenous leukemia, multiple myeloma, and myelodysplastic syndrome (MDS). We await the results of these numerous studies to further define the role of this promising therapy. In addition, an effort should be made to better define a subset of patients who would be more likely to benefit from therapy. In the phase I trials, there was no apparent correlation between baseline levels of serum VEGF and response.67 Some of the ongoing studies measure biologic surrogates such as serum or urine VEGF levels. Future studies should focus on identifying the utility of biologic markers in predicting response to therapy.
VEGF-Trap is an engineered protein that binds VEGF. It was created by fusing a portion of the extracellular domain of VEGFR1 and VEGFR2 to a human IgG1.78It has a very high affinity for all variants of VEGF-A, also binds placental growth factor, and is more efficacious than the VEGFR-2 monoclonal antibodies in binding VEGF.78 In a neuroblastoma murine model, VEGF-Trap was more effective than an anti-VEGF antibody in inhibiting tumor growth.79 Phase I trials with this agent are ongoing in patients with advanced malignancies. The dose-escalation study recruited 38 patients who were treated at seven dose levels, from 25 to 800 µg/kg subcutaneously once per week and 800 µg/kg biweekly.80 A variety of malignancies were represented, including renal cell cancer in 9 patients and colon cancer in 5. The half-life of the drug at the 800 µg/kg per week dose is approximately 25 ± 3 days. The maximum tolerated dose has not yet been achieved. Adverse events of note include proteinuria (14/33), hypertension (5/33, 2 of the 5 with grade 3 or 4 hypertension), grade 3 or 4 thrombosis (2/33), and grade 1 or 2 hemorrhage (1/33). Other grade 1 or 2 toxicities included fatigue, constipation, nausea, vomiting, anorexia, arthralgias, and diarrhea.81 No patients have developed antibodies to the VEGF-Trap. No responses have been seen, but at the higher dose of 800 µg/kg once or twice per week, 8 of 10 patients demonstrated stable disease after 10 weeks on therapy.
PTK787/ZK 222584 (Vatalanib)
PTK787/ZK 222584 (PTK/ZK) is an oral inhibitor of both VEGF-R1 (Flt-1) and VEGFR-2 (KDR). In animal models, this agent inhibits tumor growth, tumor vessel density, and development of metastases.82, 83, 84 In addition, there is evidence from the preclinical study that treatment resulted in decreased tumor blood flow as detected by color Doppler ultrasound and that this correlated with a decrease in vessel density.83 In phase I trials of PTK/ZK, this oral agent was well tolerated by patients with a variety of advanced malignancies, including glioblastoma and colorectal cancer.85, 86 Doses have been escalated from 50 to 2,000 mg daily. Toxicities were rare; the only grade 3 toxicities reported to date include deep venous thrombosis, pedal edema, and elevation of liver enzymes.85The drug is absorbed within 2 hours, and the average half-life is approximately 6 hours.86 Dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) was performed during administration of PTK/ZK in phase I studies in an attempt to identify early biologic response. The reduction in DCE-MRI contrast enhancement was dose-dependent, and there was a correlation between reduction of enhancement and tumor response.87 Further studies of this technique are required to determine if imaging results will correspond to disease improvement. The potential use for this technique in optimizing dose of antiangiogenic therapy needs to be investigated.
PTK/ZK has also been safely administered with oxaliplatin, 5-FU, and leucovorin (FOLFOX-4) in patients with advanced colorectal cancer.88 A total of 35 patients who were previously untreated received oral PTK/ZK at doses from 500 to 2,000 mg daily. Dose-limiting toxicities were observed at doses greater than 1,250 mg/day; these included ataxia, dizziness, and expressive dysphasia. For 28 evaluable patients, 1 (4%) had a complete response and 14 (50%) partial responses. As a result of these data, phase III randomized trials are planned in patients with advanced colorectal cancer. Other phase II studies continue in advanced malignancies such as mesothelioma, AML, and MDS.
ZD6474 is an orally bioavailable small-molecule tyrosine kinase inhibitor of VEGFR-2, VEGFR-3, and the epidermal growth factor receptor (EGFR). Early animal studies indicated that this agent was capable of inhibiting VEGF and angiogenesis in bone growth, and a number of tumor xenografts, including lung, colon, breast, and prostate, were inhibited by the ingestion of ZD6474.89 Due to promising preclinical data, phase I clinical trials were conducted.90 Doses were escalated from 50 to 600 mg daily in six cohorts. Patients with a variety of advanced malignancies were enrolled (31% of patients had colorectal cancer). The half-life of ZD6474 was estimated to be 120 hours. In normal healthy volunteers, absorption was not found to be affected by concurrent food intake.91Toxicities associated with this agent include asymptomatic QTc prolongation, seen in 7 of 49 patients. Grade 3 toxicities included rash (2 patients), diarrhea (2), and thrombocytopenia (1). The maximum tolerated dose was 500 mg per day.90 A smaller study in Japan displayed similar toxicities, the most common being rash (78%), asymptomatic QTc prolongation (61%), diarrhea (56%), proteinuria (56%), and hypertension (39%).92 Since ZD6474 is well tolerated, phase II clinical trials have proceeded. A pilot study found no increased toxicity or interactions between docetaxel and ZD6474 in patients with lung cancer.93Therefore, a trial is now being conducted that randomly assigns patients with advanced non–small cell lung cancer to either docetaxel with placebo or ZD6474. Other phase II trials are also being conducted that combine ZD6474 with chemotherapy or radiation in patients with lung cancer.
IMC-1C11 is a chimeric monoclonal antibody that targets the vascular endothelial growth factor receptor-2 (VEGFR-2), also known as Flk-1 in mice and KDR in humans. VEGFR-2 is present on endothelial cells and some tumor cells and is thought to be the main mediator of VEGF-stimulated angiogenesis. Animal studies were performed with DC101, a murine monoclonal antibody to VEGFR-2 (flk-1). A variety of tumors, including lung and pancreatic cancers, melanoma, glioblastoma, and lymphoma, were inhibited in mice treated with DC101.66, 94 As a result, chimeric and human anti-VEGFR-2 antibodies were developed.95Preclinical studies demonstrated that inhibition of VEGFR-2 by these antibodies decreased leukemia cell migration in vitro and significantly improved survival in a mouse model of leukemia, although leukemia was not eradicated in any of the mice.95
IMC-1C11, the chimeric monoclonal antibody, has been tested in a phase I dose-escalation clinical trial in patients with colorectal cancer metastatic to the liver.96 Fourteen patients were treated with IMC-1C11 at doses from 0.2 to 4 mg/kg weekly, administered intravenously over 1 hour. Pharmacokinetic studies revealed a very short half-life at the lower doses, while at the highest dose (4 mg/kg), the half-life was 3 days.96 The infusion was well tolerated, with no acute reactions. There were no dose-limiting toxicities in this trial. Minor bleeding, including epistaxis, blood-streaked sputum, and hematuria, was seen in 4 of 14 patients. A human antibody to chimera antibody (HACA) developed in 50% of patients, primarily those who received the lower doses. No patients had a response to therapy, although stable disease was noted in 4. In an attempt to identify surrogate markers for response, this study measured vascular perfusion of the liver metastases by DCE-MRI. Perfusion appeared to decrease with therapy, yet there was no significant radiographic tumor response. This method has been shown to correlate with microvessel density and tumor grade in colorectal cancer.97 It warrants investigation in larger studies and with other antiangiogenic therapies to determine its role in assessing treatment efficacy. In summary, IMC-1C11 was well tolerated, and further studies with it and other human monoclonal antibodies to VEGFR-2 are planned.
Thalidomide has shown promise as an oral antiangiogenic agent. In the early 1990s, in vivo studies reported that thalidomide inhibited angiogenesis induced by basic fibroblast growth factor (bFGF) in a rabbit corneal micropocket assay.98 Numerous subsequent preclinical studies demonstrated the antiangiogenic properties of thalidomide, although all the mechanisms of its action are not known. Thalidomide has been shown to down-regulate expression of VEGF and bFGF99 and inhibit endothelial cell proliferation, which is associated with inhibition of NF-κB activation.100 The teratogenic effects of thalidomide have been linked to its generation of reactive oxygen species.101, 102 This free-radical oxidative stress may have a role in thalidomide's tumor antiangiogenic activity. In addition to its antiangiogenic properties, thalidomide has an immunomodulatory effect, which may account for some of its antitumor efficacy. Thalidomide decreases the expression of cell surface adhesion molecules.103 Also, thalidomide inhibits TNF-α, which may account for its anti-inflammatory effect but may not correlate with antitumor activity.104 Finally, thalidomide's effect on immune regulation may be due to its action as a costimulator of T cells, primarily of the CD8+ subset.105
Thalidomide is a synthetic derivative of glutamic acid and is orally bioavailable. It has low solubility in the gastrointestinal tract, which limits its absorption, but higher doses do result in higher plasma concentrations.106 The drug undergoes hydrolysis into a number of metabolites, which are renally excreted. At a dose of 200 mg, the half-life is approximately 6 to 7 hours.106, 107
The first successes with thalidomide as an antitumor agent were described in multiple myeloma. In 1999, Singhal et al. reported activity of thalidomide in previously treated patients with multiple myeloma; 90% of these patients had progressed after high-dose chemotherapy followed by autologous stem cell transplantation.108 A total of 84 patients were treated with thalidomide starting at a dose of 200 mg per day, and it was escalated as tolerated to a maximum dose of 800 mg, which was achieved in 55% of patients. Ten percent of patients had a complete or near complete response, while 32% had at least a 25% drop in paraprotein levels. Therapy was relatively well tolerated, and grade 3 or 4 toxicities were very rare. This response was encouraging given the advanced nature of disease in this cohort of patients. Further studies have demonstrated similar activity with single-agent thalidomide in both previously treated and untreated multiple myeloma. Overall response rates in these studies are reported to be 25 to 35%.109, 110, 111 Adverse effects are dose-related and have included constipation, fatigue, somnolence, depression, tremor, and sensory neuropathy.108, 109, 111 With thalidomide alone, less than 5% of patients developed thrombotic events. Response to thalidomide has not correlated with reduction in angiogenic cytokines such as bFGF, VEGF, and TNF-α.112 On the other hand, there has been some evidence of reduction in bone marrow microvessel density in patients who respond to thalidomide.108, 113 Therefore, the exact mechanism of thalidomide's action is not well understood.
Other agents such as dexamethasone or combination chemotherapy have been studied with thalidomide in both untreated and previously treated patients with multiple myeloma. The response rates with combination therapy are higher than with thalidomide alone; overall response rates from 55 to 72% have been observed.111, 114, 115, 116 It is not clear that overall survival is better with combination therapy, but phase III trials are ongoing. The neurologic toxicities appear to decrease with combination therapy, particularly dexamethasone plus thalidomide, but the incidence of thrombotic events increases significantly. Studies have consistently reported that up to 16% of patients develop deep venous thrombosis with dexamethasone plus thalidomide111, 116, 117 and up to 28% with multiagent chemotherapy plus thalidomide.118 Therefore, prophylactic anticoagulation is warranted with these regimens.
Thalidomide has been evaluated in a number of other hematologic disorders, including AML,119 myelofibrosis with myeloid metaplasia,120 and MDS.121, 122, 123In myelofibrosis with myeloid metaplasia (MMM), thalidomide has shown efficacy in a number of phase II studies. The largest study treated 63 patients with thalidomide, at a dose of 50 mg/day escalated to a maximum of 400 mg/day as tolerated.120 The median maximum tolerated dose was 100 mg/day. Responses were characterized by an improvement in anemia (26%), platelet increase by more than 50 × 109/L (41%), and 50% reduction in splenomegaly (19%).120These results warrant further investigation of thalidomide in MMM.
Responses to thalidomide have also been seen in patients with AML or MDS. In 20 AML patients who were refractory to cytotoxic therapy or who were not candidates for cytotoxic therapy, 25% demonstrated a partial response to single-agent thalidomide.119 In patients who responded, there was a decrease in bone marrow microvessel density. Studies of single-agent thalidomide have also been promising in patients with MDS; 19 to 55% of patients achieved a hematologic improvement and a subset of patients even became red blood cell transfusion–independent.121, 122, 123 In both the AML and MDS studies, patients experienced significant toxicity, which limited the dose that could be administered. The majority of patients could not tolerate doses above 200 mg/day. The adverse effects were similar to those described in the previous studies, including fatigue, constipation, fluid retention, dizziness, and neuropathy. A study of thalidomide and darbepoetin-α in MDS patients was stopped early due to thromboembolic events in three of the first seven patients, including one fatal pulmonary embolism.124
Thalidomide's activity in a number of solid tumors has also been investigated. Single-agent thalidomide or combination therapy has demonstrated antitumor activity in glioblastoma multiforme, melanoma, renal cell cancer, and prostate cancer. Patients with glioblastoma multiforme were treated with either thalidomide alone (19 patients) or thalidomide with temozolomide (25 patients) after resection and irradiation.125 Although this was not a randomized study, overall survival was significantly better in the combination arm, 103 weeks versus 63 weeks. Temozolomide is an oral alkylating agent that is well tolerated. Only 1 patient of 25 had grade 3 myelosuppression. Other toxicities were those commonly reported with thalidomide in particular, sedation and thrombotic events.125 The combination of thalidomide and temozolomide has also been effective in metastatic melanoma, as demonstrated in two separate phase II studies.126, 127 Danson and colleagues randomized patients with metastatic melanoma to either temozolomide alone (59), temozolomide and interferon α-2b (62), or temozolomide and thalidomide (60); objective responses were observed in 9%, 18%, and 15% of patients, and median survival was 5.3, 7.7, and 7.3 months, respectively. The temozolomide-thalidomide arm was better tolerated due to the low frequency of hematologic adverse events.127 The treatment doses in this study were 100 mg of thalidomide daily with temozolomide 150 mg/m2 daily for 5 days during the first cycle, increased to 200 mg/m2 daily for 5 days in each subsequent cycle. Higher doses of thalidomide are also tolerated in this patient population; 200 mg daily of thalidomide was escalated to 400 mg daily, and the temozolomide dose was 75 mg/m2 daily for 6 weeks followed by a 2-week rest.126 The response rate in this single-arm phase II trial was 32%, including a complete response lasting over 25 months. Further studies to determine the optimal dosing and treatment schedule are necessary in this group of patients.
Renal cell carcinoma is another vascular tumor in which thalidomide has been tested. When thalidomide was used as a single agent, responses were modest, with rare partial responses and a small subset of patients with stable disease.128, 129 In these studies, thalidomide was dose escalated from 400 to 1,200 mg daily. Significant toxicity, particularly neurotoxicity, was observed. In addition, venous thromboembolic events, which have been rare in other studies with single-agent thalidomide, occurred in 9 of 40 patients (23%) in the study by Escudier and colleagues.128 Better results have been seen when thalidomide was combined with interferon-α, but the studies have been small. Among 30 patients treated with interferon-α 1.2 million units three times daily and thalidomide 300 mg once daily, 20% had a partial response, and 63% had stable disease for at least 3 months.130 The added benefit of thalidomide in this group of patients is not clear, and results from randomized phase III trials are anticipated.
In hormone-refractory prostate cancer, thalidomide at low doses (100 to 200 mg daily) has shown some activity as a single agent, resulting in PSA decline.131,132 Thalidomide has been successfully combined with docetaxel in this same population. Patients were randomly assigned to receive either docetaxel alone (30 mg/m2 weekly for 3 weeks of a 4-week cycle) or docetaxel plus thalidomide (200 mg daily).133 Overall survival at 18 months was 43% in the docetaxel arm versus 68% in the docetaxel-thalidomide arm (P = .11). The therapy was relatively well tolerated; only 8% of patients had grade 3 hematologic toxicity in the combination group. Thromboembolic events were significantly increased in the combination group. Twelve of the first 43 patients in this group developed a venous thrombosis (9) or transient ischemic event/ stroke (3). The subsequent 6 patients treated on the combination arm received prophylactic low molecular weight heparin during therapy.
The role of thalidomide in these diseases is promising but needs to be better defined. Ongoing studies continue with thalidomide in a variety of other solid tumors, including small and non–small cell lung cancer, ovarian cancer, sarcoma, and hepatocellular cancer.
IMiDs: CC-5013 (Revlamid) and CC-4047 (Actimid)
A number of thalidomide derivatives have been developed, and these make up at least two new classes of drugs: immunomodulatory drugs (IMiDs) and selective cytokine-inhibitory drugs (SelCIDs). They exhibit effects on immune regulation, cytokine production, angiogenesis, and tumor growth in the preclinical setting that are similar to or more potent than those of thalidomide.134, 135, 136, 137, 138 T-cell stimulation is observed only in the IMiD class. Antiangiogenic activity, which has been seen in both classes in vivo and in vitro, appears to be distinct from the immunomodulatory effects.138 Phase I studies of the IMiDs that show better tolerability and potential efficacy (CC-5013 and CC-4047) have been conducted.139, 140, 141
To date, most clinical experience has been with CC-5013 (Revlamid). Pharmacokinetic studies in patients with multiple myeloma have demonstrated that the drug is absorbed within 2 hours and that the elimination half-life is between 3 and 6 hours.139, 142 The maximum tolerated dose in patients with multiple myeloma is 25 mg daily. Myelosuppression, predominantly neutropenia and thrombocytopenia, is the dose-limiting toxicity. Lethargy, constipation, and neuropathy have not been observed. In 17 of 24 patients (71%), the paraprotein concentration decreased by at least 25%, and the decrease was seen in 11 patients who had previously received thalidomide.139 Preliminary results in patients with MDS are very promising. A total of 45 patients who were transfusion-dependent or had symptomatic anemia were treated with one of three doses of CC-5013: 25 mg daily, 10 mg daily, or 10 mg daily for 3 weeks followed by a 1-week break.143 In this study, 88% of patients were in the Low/Intermediate-1 International Prognostic Scoring System (IPSS) group. A major hematologic response was seen in 19 patients. In addition, cytogenetic responses were observed in 11 patients, with 10 patients converting to a normal karyotype. Myelosuppression was the most common and significant adverse event. Phase I studies have also been conducted in patients with malignant melanoma, high-grade gliomas, and other advanced solid tumors.140, 144 CC-5013 is safe and tolerable in this cohort. We await the results of ongoing studies to define the efficacy of CC-5013 and its role in treatment algorithms for multiple myeloma, MDS, and other malignancies.
The experience with CC-4047 (Actimid) is less extensive. In a phase I clinical trial in 24 patients with multiple myeloma, CC-4047 was well tolerated.141 It is orally bioavailable, with an elimination half-life of about 7 hours. The maximum tolerated dose was determined to be 2 mg per day. Myelosuppression, primarily neutropenia, was the dose-limiting toxicity. In addition, 4 patients developed a deep venous thrombosis while receiving doses of 1, 2, or 5 mg. Four patients achieved a complete response, while 9 had a partial response. The results from this phase I study are encouraging, and further studies are being conducted in multiple myeloma, MDS, prostate cancer, and other solid tumors.
AE-941 is an antiangiogenic agent that has been purified from shark cartilage. In animal studies, this agent decreased both angiogenesis and metastases.145It exerts its effects through a number of mechanisms. In vitro and in vivo animal studies have demonstrated that AE-941 competitively inhibits binding of VEGF to the VEGF receptor-2.146 In addition, AE-941 inhibits MMP-2, -9, and -12147 and induces endothelial cell apoptosis.148 It is an orally bioavailable agent and has been well tolerated in a number of trials that have included over 800 patients total.149
A phase I/II dose escalation study was conducted in patients who had advanced non–small cell lung cancer and were refractory to previous therapy. No dose-limiting toxicity was identified in the 80 patients.150 The most common side effects included nausea (9%), pruritus (5%), anorexia (4%), and emesis (4%). Although there were no tumor responses with therapy, 26% of patients in the highest dose group (240 mL/day) had stable disease, as compared with 14% in the lower dose groups. A phase III trial of induction chemoradiotherapy with or without AE-941 in unresectable non–small cell lung cancer continues to accrue patients.
Results of using this agent in patients with renal cell cancer have shown promise. A phase II trial was reported in patients with advanced renal cell cancer.151Initially patients were treated with AE-941 60 mL per day; due to results from other trials, 14 months into the study the dose was increased to 240 mL per day, given in a divided dose. A total of 22 patients were enrolled, and 14 received the higher dose. Median survival was improved in the cohort that received 240 mL per day (16.3 vs. 7.1 months, P = .01)151 The regimen was well tolerated, and the most common adverse event was altered taste (14% of patients). The only grade 3 event was peripheral edema in one patient. A phase III trial in over 300 renal cell cancer patients has been completed and results are pending.152
A series of experiments performed by O'Reilly and colleagues identified an endogenous inhibitor of tumor growth and angiogenesis, angiostatin.153 It was found to be a 38-kd fragment composed of the first four kringle domains of plasminogen. The antiangiogenic effect was unique to angiostatin and was not demonstrated by treatment with intact plasminogen.153 The mechanism of action of angiostatin is not well understood, but three endothelial cell surface receptors have been identified. These include ATP synthase, angiomotin, and integrin αvβ3.154 Angiostatin binding likely results in apoptosis. In preclinical investigations, recombinant angiostatin administered to mice inhibited the growth of both primary tumors and metastases.155, 156
Recombinant human angiostatin (rhAngiostatin) has been developed and is in early clinical trials. The first reported trial included 15 patients with a variety of advanced malignancies.157 rhAngiostatin was administered daily as an intravenous infusion, at doses from 15 to 120 mg/m2, with no dose-limiting toxicities observed. VEGF and bFGF levels decreased in some of the treated patients.157 A subsequent study treated 24 patients with twice daily subcutaneous injections of rhAngiostatin, at doses of either 7.5, 15, or 30 mg/m2 per day.158 The therapy was well tolerated, and the most commonly observed toxicity was grade 1 or 2 erythema at the injection site (54% of patients). Other significant toxicities included 2 patients with hemorrhage into brain metastases and 2 patients who developed a deep venous thromboembolism. Twenty-five percent of patients had stable disease for over 6 months. In this study, treatment did not decrease serum or urine bFGF or VEGF levels. A phase II trial of combination chemotherapy plus rhAngiostatin is currently underway in patients with advanced non–small cell lung cancer.159 Patients with stage IIIB or IV disease who had never previously received chemotherapy participated. Treatment included paclitaxel 175 mg/m2 and carbolplatin AUC 5 on day 1, with administration of rhAngiostatin at either 15 or 60 mg subcutaneously twice daily for a maximum of six cycles. If patients responded or had stable disease on this therapy, they continued to receive maintenance rhAngiostatin until progression. Patients received rhAngiostatin for a median of 125 days. The most commonly observed adverse effect was a skin rash (grade 1 to 3) in 92% of patients. Grade 3 or 4 toxicities included: neutropenia (42%), fatigue (42%), and dyspnea (25%). There were no complete responses, but 39% of the patients (9) had a partial response, and stable disease occurred in 39%.159 This study demonstrated the feasibility of rhAngiostatin administration with combination chemotherapy. Further studies will be necessary to determine whether these results are better than those attainable with chemotherapy alone.
Endostatin is an endogenous inhibitor of angiogenesis and tumor growth first discovered by O'Reilly and colleagues.160 It is a 20-kd fragment of collagen XVIII. In mouse models, endostatin administered subcutaneously resulted in tumor regression.161 Recombinant human endostatin (rh-Endostatin) has been produced in yeast and has been tested in phase I clinical trials. In three clinical trials, rh-Endostatin was administered to 61 patients as a daily intravenous infusion over either 20 or 60 minutes.162, 163, 164 Doses were escalated from 15 to 600 mg/m2 daily and were all well tolerated. Doses over 300 mg/m2 daily resulted in an AUC that was therapeutic in preclinical studies.162, 164 Grade 3 toxicities were rare; they included anemia, deep venous thrombosis, and dyspnea each in only one patient.162 No significant clinical tumor responses were observed in these studies. There was no consistent reduction of VEGF or bFGF levels in patients treated with endostatin.162, 164 In addition, tumor blood flow analysis by PET scan and biopsies to assess tumor and endothelial cell apoptosis were performed in patients treated with endostatin.165 Further studies will be required to demonstrate whether these measures correlate to tumor response. Based on preclinical mouse tumor models suggesting that efficacy is improved with continuous infusion,166 new clinical trials were designed to administer endostatin continuously. Preliminary results are also available from a phase II study of 41 patients with advanced neuroendocrine tumors. Rh-Endostatin, was administered as a subcutaneous injection at a dose of 30 mg/m2 twice a day.167 Only 5% of patients had a minor radiographic response, and 62% had stable disease. Results in the clinical trials have been disappointing, as they did not reflect the promising results observed in preclinical models.
A number of agents with antiangiogenic activity show promise in the treatment of a wide variety of malignancies. One of the challenges facing clinical researchers interested in an agent that may inhibit angiogenesis is determining the optimal way to assess the clinical efficacy of the agent.168 The traditional approach for evaluating a new cytotoxic agent is to perform a phase I dose-escalation trial that identifies the maximal tolerated dose (MTD) of drug. With conventional cytotoxic agents, the MTD frequently correlates with maximal antitumor activity. With agents that inhibit angiogenesis, however, tumor regression may or may not be observed. As a result, the MTD may not accurately reflect the optimal biologic effect of the drug. Although identifying both acute and long-term toxicities associated with any new agent is important, an angiogenesis inhibitor may have a defined optimal biologic dose that is very different from the MTD.
Identifying plasma concentrations of the angiogenesis inhibitor in animals that are optimal for the desired biologic effect is important. Because angiogenesis inhibitors may not cause tumor shrinkage when administered alone, the clinical development of an agent may be abandoned if surrogate endpoints that reflect the activity of the agent are not incorporated into the trial design. Potential biologic markers for antiangiogenic activity have included measurement of serum markers of angiogenesis (VEGF, bFGF, VCAM-1 [vascular cell adhesion molecule], E-selectin) or circulating endothelial cells, assessment of endothelial cell and tumor cell apoptosis on biopsy, PET scan assessment of tumor blood flow, and DCE-MRI contrast enhancement as a reflection of microvessel density. Whether these effects of a new agent legitimately reflect the antiangiogenic or antitumor activity remains unclear. Yet these or other surrogate markers will likely be important in assessing the activity of angiogenesis inhibitors in early clinical trials.
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