Abstract

Background:

renal cell carcinoma (RCC) is a potential target for anti-angiogenic drugs because of its high vascularization. Neovastat (Æ-941) is an inhibitor of angiogenesis with a mechanism of action that could prove beneficial in the treatment of RCC.

Patients and design

A phase II trial was conducted to identify the long-term safety profile of Neovastat in advanced cancer patients and to obtain preliminary information on its efficacy in solid tumors refractory to standard treatments. Neovastat (60 or 240 ml/day) was administered orally (b.i.d.) to 144 patients with solid tumors refractory to standard therapies or for whom no standard treatments were available.

Results:

A survival analysis was conducted on 22 patients with a primary diagnosis of refractory RCC to determine whether the dose of Neovastat had any effect. A significant relationship between dose and survival was observed; the median survival time was significantly longer (16.3 versus 7.1 months; P = 0.01) in patients treated with Neovastat 240 ml/day (n = 14) compared with patients receiving 60 ml/day (n = 8). No dose-limiting toxicity was reported. The most frequent adverse event was taste alteration (13.6%).

Conclusions:

Neovastat is well tolerated by advanced cancer patients at doses of 60 and 240 ml/day. The higher dose of Neovastat administered in this trial is associated with a survival benefit in RCC, which is not explained by differences in major prognostic factors.

Received 3 October 2001; revised 19 December 2001; accepted 9 January 2002

Introduction

Renal cell carcinoma (RCC) is the seventh most common cancer and accounts for 3% of all malignancies and 80–85% of kidney tumors in adults [1]. In 2001, an estimated 35 000 new cases of RCC are expected to be diagnosed in Canada and the USA, and 13 500 people will die from this disease [2, 3]. Despite the introduction of biological therapies, of which interleukin-2 (IL-2) has been the principal model with a 7–9% long-term cure rate in a highly selected population of metastatic RCC, the effectiveness of all combined systemic modalities (e.g. IL-2 alone, IL-2 and/or interferon, biochemotherapies) continues to be unsatisfactory.

Angiogenesis, the outgrowth of new capillaries from pre-existing blood vessels, is a complex, multistep process that is tightly controlled by a balance between angiogenesis stimulators and inhibitors. It is characterized by a number of independent but interrelated steps, such as the dissolution of the basement membrane, the proliferation and migration of endothelial cells and the formation of microvessels. It plays an important role in vessel development and homeostasis during wound healing, female reproduction and tissue regeneration. However, the up-regulation of angiogenesis plays a central role in the development of several pathological conditions, such as age-related macular degeneration, psoriasis and rheumatoid arthritis [4, 5].

Experimental evidence suggests that angiogenesis plays an important role in the growth, progression and metastasis of highly vascularized tumors, including RCC. Increased serum levels of basic fibroblast growth factor (bFGF), vascular endothelial growth factor (VEGF) and other angiogenic factors have been detected in the serum of patients with RCC. In a trial including 76 patients with RCC and 41 healthy controls, patients with RCC were shown to have significantly higher serum levels of bFGF and VEGF than healthy subjects [6]. Moreover, patients with disseminated cancer exhibited significantly higher serum levels of bFGF and VEGF than did those with undisseminated cancer. Another trial showed that serum bFGF levels were significantly higher in advanced RCC patients with pulmonary metastasis than in those lacking pulmonary lesions [7]. Fujimoto et al. [8] confirmed that advanced RCC is characterized by increased serum levels of bFGF, compared with less advanced cases. Interestingly, in five of seven subjects who underwent nephrectomy and whose serum bFGF levels were elevated prior to the surgery, bFGF levels returned to normal within 2 weeks after resection of the tumor. Of the two patients whose bFGF levels did not decrease post-nephrectomy, one had a large residual tumor and the other had postoperative complications, which suggests that bFGF is also actively involved in wound healing.

Increased understanding of the complex factors involved in angiogenesis has led to the development of agents that can interfere with specific components of the angiogenic process [4, 9]. Among the few angiogenesis inhibitors in late stage clinical development, Neovastat (Æ-941) is composed of water-soluble molecules extracted from cartilage (Dupont É et al., US Patent No. 5 618 925), and has the advantages of being administered orally and of having multiple anti-angiogenic mechanisms of action. It selectively inhibits matrix metalloproteinases (MMPs)-2, -9 and -12, and stimulates tissue plasminogen activator enzymatic activities [10, 11]. It also selectively competes for the binding of VEGF to its receptor (VEGFR) [12] causing disruption of this signaling pathway. This has been demonstrated by the inhibition of VEGFR-2 (Flk-1/KDR) phosphorylation and inhibition of VEGF-induced plasma extravasation in mice [12]. Finally, Neovastat induces apoptotic activities in endothelial cells [13]. The antitumor effect of Neovastat has been confirmed in several different in vivo animal models. Oral treatment with Neovastat induced a dose-dependent decrease in tumor growth in mice grafted with mouse mammary adenocarcinoma cells DA3 and metastatic invasion in mice grafted with M27 Lewis lung carcinoma cells [14, 15]. An important effect on tumors has also been observed following oral administration of Neovastat in human glioblastoma models in mice (HGD model) that was correlated with a decrease in angiogenesis [11].

Previously published clinical studies with dry powder preparations of crude cartilage were inconclusive [16] or demonstrated that such products were inactive in patients with advanced stage cancers [17]. This is particularly troublesome in view of the recent findings that cartilage is one of the most often used unconventional therapies among cancer patients [18]. These studies were performed using powdered and lyophilized cartilage. This paper describes a study using a liquid extract obtained under controlled and reproducible conditions, which has demonstrable biological activity in the pre-clinical setting. A dose-dependent increase in median survival time has been reported in a phase I/II trial involving unresectable stage III and IV non-small-cell lung cancer patients treated with Neovastat [19]. In this report, we provide data on the long-term side-effect profile of oral Neovastat in patients with RCC. We also describe an exploratory analysis aimed at determining whether the dose level had any potential impact on survival in these patients.

Patients and methods

This study (protocol number CT/Æ-941/002) was designed as a phase II multicenter, open-label, open-ended treatment study. The objective was to establish the long-term safety profile of Neovastat and to obtain preliminary survival data from a wide variety of advanced cancer patients. Patient recruitment was opened in four investigative centers in Canada from September 1997 to May 2000. The protocol was approved by Health Canada’s Therapeutic Products Program and by the Institutional Review Boards of the participating sites. Informed consent was obtained from all patients enrolled onto the trial.

Eligibility criteria

Patients were eligible for the trial if they had a histologically confirmed solid tumor refractory to standard therapies or if they refused conventional anticancer therapy after being fully informed of the available conventional therapies and their greater likelihood of efficacy. Patients with known hypersensitivity to fish products were excluded. Pregnant women and lactating mothers were also excluded.

Study design

Baseline evaluation included a complete clinical history and physical examination. Specific details of all prior anticancer therapies were obtained. Medical imaging studies appropriate for the specific tumor site were obtained to document the extent of disease. Cross-dimensional diameters of all measurable lesions were determined whenever possible. Assessable but non-measurable lesions (e.g. positive bone scans) were monitored. Laboratory tests were mandatory while chemotherapy and/or radiotherapy was administered in combination with Neovastat.

Neovastat was manufactured by Les Laboratoires Æterna (Québec, Canada). Patients who were enrolled were initially treated with Neovastat 60 ml/day. In November 1998, on the basis of the results of two separate phase I tolerability studies [20, 21],the dose in patients still being treated with Neovastat at that time in the study was increased from 60 to 240 ml/day. Patients enrolled after November 1998 were administered Neovastat at a dose of 240 ml/day.

Patients were instructed to take Neovastat orally, in two divided doses (b.i.d.) in a fasting state. They were treated as outpatients and were reviewed monthly for the first 6 months, then every 3 months for the remainder of the trial by clinical history, physical examination and imaging studies in accordance with standard practice according to the type of cancer. Use of concomitant drugs and occurrence of adverse events were also reviewed. Compliance was determined by direct questioning of patients and confirmation from pharmacy records. Following the end of Neovastat treatment, patients were followed for vital status at 3-month intervals.

Criteria for withdrawing from the study were intolerable drug-related toxicity, progressive disease, the patient’s desire to withdraw or non-compliance. Adverse events were defined as those that were possibly, probably or definitely related to Neovastat, as judged by the investigators.

Statistical analysis

A survival analysis was performed on the group of patients (n = 22) with a primary diagnosis of RCC who were enrolled on the trial, using a Cox survival model. Survival time was defined as the time between the start of Neovastat treatment and death (or date of last follow-up). The Cox survival model was adjusted for age, gender, number of organs with metastasis, use of immunotherapy and time since initial diagnosis.

Seven patients enrolled on to the trial received only Neovastat 60 ml/day and nine patients received only Neovastat 240 ml/day. As explained above, the dose was increased from 60 to 240 ml/day for six patients. Among these six patients, five were assigned to the 240 ml/day group for the purpose of the survival analysis, since they received that dose for most of the duration of their treatment (median percentage of time on the dose of 240 ml/day was 76%). The other patient was assigned to the 60 ml/day group as he received 240 ml/day for only five of the 117 days of treatment. Thus this assignment resulted in eight patients in the 60 ml/day group and 14 patients in the 240 ml/day group. For patients who switched dose, survival analysis was performed with both starting points: time from start of Neovastat treatment and time from the start of Neovastat treatment at the dose of 240 ml/day.

Analyses were performed using the following software: SAS (version 6.12; SAS Institute, Cary, NC, USA) and S-PLUS 2000 (Mathsoft 1999, Seattle, WA, USA).

Results

The demographics of patients with a primary diagnosis of RCC are presented in Table 1. Half of the patients were male; the mean time since diagnosis was ∼4.5 years; over 80% had undergone nephrectomy and 68% had an Eastern Cooperative Oncology Group (ECOG) performance status of <2. Non-parametric comparative testing (Fisher’s exact test) on demographic parameters did not identify any differences between groups. Apart from Neovastat, no other type of anticancer treatment was taken by the RCC patients during the trial. The incidence of lung and nodal metastases is comparable in the two groups of patients. With regard to extrapulmonary disease, there are differences, although the small numbers make it difficult to determine their significance. While there are more patients with liver metastases in the low dose treatment group (37.5% versus 21%), the frequency of multiple bone metastases is greater in the higher dose group (50% versus 12.5%).

Efficacy

In patients with RCC, there were two objective responses amongst the 14 patients who received Neovastat 240 ml/day and none in the 60 ml/day group. One was a large soft-tissue facial metastasis which did not change 3 months after radiation therapy, nor during a period of 2 months on the low dose. During 8 months on the 240 ml/day dose of Neovastat, however, this large facial metastasis completely resolved, while concurrent pulmonary metastases remained stable. The other clinical response was in an enlarging, though unbiopsied, supraclavicular node. After a brief period at the low dose, during which the lymph node continued to grow, the patient was increased to the 240 ml/day dose and the node progressively shrank over a period of 7 months, and remained thus for >24 months. While these are only anecdotal reports, they demonstrate biological activity and suggest of a dose effect.

The Cox survival model was used to compare the survival time of RCC patients receiving Neovastat 240 ml/day (n = 14) with 60 ml/day (n = 8). A statistically significant longer survival time (P = 0.01) was observed in patients treated with Neovastat 240 ml/day (16.3 months) compared with patients receiving 60 ml/day (7.1 months). When survival time in patients who were dose escalated from 60 to 240 ml/day is calculated only from the actual start of the 240 ml/day regimen, the difference in median survival time between the two groups is also statistically highly significant (14.4 versus 7.1 months; P = 0.024).

Survival rate at 2 years was 0% in the 60 ml/day group and 36% in the 240 ml/day group. The unadjusted differences in survival time are summarized in the Kaplan–Meier curves presented in Figure 1.

We recognize that these groups were not randomized, and thus we have examined important prognostic factors. Most are roughly evenly balanced, given the small numbers. As expected, the presence of liver metastases has a highly significant impact on survival (P = 0.0068). Amongst the three patients in each dose group with liver metastases, the median survival was 6.8 months (2.3, 6.8 and 7.2 months, respectively) in the lower dose group and 8.6 months in patients who received Neovastat 240 ml/day (3.5, 8.6 and 11.6 months, respectively). These are small numbers, but it is noteworthy that the trend favoring the higher dose is still present.

When we examined the survival amongst patients without liver metastases, median survival at Neovastat 60 ml/day amongst five patients was 8.9 months, and median survival at Neovastat 240 ml/day amongst 11 patients was 21.9 months, which is highly statistically significant (P = 0.024). Furthermore, when we incorporate liver metastasis as a covariate in the Cox model, the difference between median survival times (7.1 and 16.3 months in the Neovastat 60 and 240 ml/day groups, respectively) remains highly significant: P = 0.016 if survival is calculated from the start of Neovastat administration or P = 0.022 if survival is calculated from the start of the 240 ml/day regimen in those that were switched to the higher dose.

Adverse events

Adverse events were reported in six of 22 RCC patients (Table 2). No deaths or grade 3 or 4 laboratory test abnormalities related to Neovastat were reported. One serious adverse event was reported, namely a hypoglycemic episode in a known type II diabetic patient. Only one patient experienced a grade 3 adverse event, which consisted of peripheral edema and led to study discontinuation. No patient had a grade 4 adverse event. There was no relationship between the incidence or severity of adverse events and the dose of Neovastat.

Discussion

In this trial, no dose-limiting toxicity was observed following the oral administration of Neovastat at doses ≤240 ml/day. These findings are consistent with the favorable safety profile which has been reported with Neovastat in other trials involving >800 patients, some receiving the investigative agent for treatment periods >4 years. This would also be necessary for any agent for which long-term use is envisioned, as in the case with anti-angiogenic therapies.

An analysis of survival time for the 22 patients with RCC was carried out. A dose of 240 ml/day was associated with a significantly longer median survival than was observed in those receiving 60 ml/day (16.3 versus 7.1 months; P = 0.01). When survival time in patients who were dose escalated from 60 to 240 ml/day was calculated only from the start of the 240 ml/day regimen, the difference in median survival time was still highly significant (14.4 versus 7.1 months; P = 0.024).

Since these treatment dose groups were not randomized, we examined important prognostic factors, many of which were evenly balanced, given the small numbers. There were three patients with the particularly grave prognostic factor of liver metastases, which represents a higher incidence in the low-dose group. Interestingly, the median survival is almost 2 months higher for the high-dose group, even amongst the patients with liver metastases. When we incorporate the effect of liver metastasis on the analysis of Neovastat dose versus survival, the dose–effect remains highly statistically significant, even with these small numbers of patients. Thus while we recognize the limitations of these analyses, it is evident that in all cases the pattern appeared to be the same, and it always suggested a positive dose effect.

Amongst the many clinical trials underway targeting tumor angiogenesis, there have been very few (in fact rare) demonstrable tumor responses. Rather, most phase I/II trials are providing safety data as well as information on tumor status and survival that can only be compared with historical results. This manuscript describes some provocative data that take advantage of the fact that similar patients were treated at two different dose levels. There appears to be a dose–survival relationship. Although this was not a randomized trial, we feel these data are nonetheless of interest.

Although the design of our study does not allow firm conclusions to be drawn, the observation is provocative and justifies further clinical research to determine if Neovastat favorably influences the survival of patients with refractory RCC. A phase III trial is currently ongoing in centers in Canada, USA and several European countries. Patients refractory to immunotherapy are being treated with either Neovastat or placebo.

Acknowledgements

This study was sponsored by Les Laboratoires Æterna (Québec, Canada).

+

Correspondence to: Dr G. Batist, McGill Center for Translational Research in Cancer, Jewish General Hospital, 3755 Côte Ste-Catherine, Montréal, Quebec, Canada, H3T 1E2. Tel: +1-514-340-7915; Fax: +1-514-340-7916; E-mail: gbatist@onc.jgh.mcgill.ca

Table 1.

 Demographics of study patients

Parameter Neovastat (ml/day) 
 60 240 
Total number of patients recruited  8  14 
Male (%)  4 (50)   7 (50) 
Age (years)   
 Median 57.7  61.5 
 Range 45–74  38–76 
Mean time since diagnosis (years)  4.5   4.6 
Prior immunotherapy (%)  6 (75)   6 (43) 
Nephrectomy (%)  7 (88)  11 (79) 
Time since nephrectomy (years)   
 Median  5.1   2.9 
 Range  1.0–8.5   0.9–16.7 
Mean number of metastatic sites (%)  2.13   2.07 
 Bone  1 (12.5)   7 (50) 
 Liver  3 (37.5)   3 (21) 
 Lung  5 (62.5)   8 (57) 
 Lymph node  3 (37.5)   6 (43) 
 Renal mass  1 (12.5)   3 (21) 
 Other  3 (37.5)   3 (21) 
ECOG performance statusa (%)   
 0  1 (12.5)   3 (21) 
 1  3 (37.5)   8 (57) 
 2  3 (37.5)   2 (14) 
 3  0   1 (7) 
Parameter Neovastat (ml/day) 
 60 240 
Total number of patients recruited  8  14 
Male (%)  4 (50)   7 (50) 
Age (years)   
 Median 57.7  61.5 
 Range 45–74  38–76 
Mean time since diagnosis (years)  4.5   4.6 
Prior immunotherapy (%)  6 (75)   6 (43) 
Nephrectomy (%)  7 (88)  11 (79) 
Time since nephrectomy (years)   
 Median  5.1   2.9 
 Range  1.0–8.5   0.9–16.7 
Mean number of metastatic sites (%)  2.13   2.07 
 Bone  1 (12.5)   7 (50) 
 Liver  3 (37.5)   3 (21) 
 Lung  5 (62.5)   8 (57) 
 Lymph node  3 (37.5)   6 (43) 
 Renal mass  1 (12.5)   3 (21) 
 Other  3 (37.5)   3 (21) 
ECOG performance statusa (%)   
 0  1 (12.5)   3 (21) 
 1  3 (37.5)   8 (57) 
 2  3 (37.5)   2 (14) 
 3  0   1 (7) 

aNot available in one patient.

ECOG, Eastern Cooperative Oncology Group.

Table 2.

 Most frequent adverse event

Event Neovastat (ml/day) 
 60 (n = 13)a 240 (n = 15) Overall (n = 22)b 
Taste alteration (%) 1 (7.7) 2 (13.3) 3 (13.6) 
Anorexia (%) 1 (7.7) – 1 (4.5) 
Peripheral edema (%) 1 (7.7) – 1 (4.5) 
Erythema (%) 1 (7.7) – 1 (4.5) 
Hypoglycemia (%) 1 (7.7) – 1 (4.5) 
Nausea (%) 1 (7.7) – 1 (4.5) 
Event Neovastat (ml/day) 
 60 (n = 13)a 240 (n = 15) Overall (n = 22)b 
Taste alteration (%) 1 (7.7) 2 (13.3) 3 (13.6) 
Anorexia (%) 1 (7.7) – 1 (4.5) 
Peripheral edema (%) 1 (7.7) – 1 (4.5) 
Erythema (%) 1 (7.7) – 1 (4.5) 
Hypoglycemia (%) 1 (7.7) – 1 (4.5) 
Nausea (%) 1 (7.7) – 1 (4.5) 

aNumber of patients treated at that dose.

bThe total number of patients of each column does not add up to 22 patients as six patients received Neovastat at the dose of 60 and 240 ml/day.

Figure 1. Kaplan–Meier survival graph of patients with renal cell carcinoma receiving Neovastat.

Figure 1. Kaplan–Meier survival graph of patients with renal cell carcinoma receiving Neovastat.

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Author notes

1McGill Center for Translational Research in Cancer, McGill University, Montréal; 2Les Laboratoires Æterna, Québec; 33210308 Canada Inc., Montréal, Canada; 4Institut Gustave Roussy, Villejuif, France