Prognostic Significance of Plasma Vascular Endothelial Growth Factor Levels in Patients with Hormone-refractory Prostate Cancer Treated on Cancer and Leukemia Group

Purpose: Plasma vascular endothelial growth factor (VEGF) levels are significantly elevated in patients with hormone-refractory prostate cancer (HRPC) compared with patients with localized disease and have been associated with disease progression in other cancer patient populations. Therefore, we measured VEGF levels in plasma prospectively collected from patients enrolled in Cancer and Leukemia Group B 9480, an intergroup study of suramin in patients with HRPC, to determine whether these levels had prognostic significance. Experimental Design: Pretreatment plasma was collected from patients with HRPC enrolled in Cancer and Leukemia Group B 9480. In a subset of samples representative of the entire cohort, plasma VEGF levels were determined in duplicate using a Quantiglo chemiluminescent ELISA kit (R&D Systems, Minneapolis, MN). Statistical analyses were performed to determine the correlation between pretreatment plasma VEGF levels and time of overall survival. The proportional hazards model was used to assess the prognostic significance of various cut points in multivariate models. Results: Plasma VEGF levels in this population ranged from 4–885 pg/ml, with a median level of 83 pg/ml. As a continuous variable, plasma VEGF levels inversely correlated with survival time (P 5 0.002). Using various exploratory cut points, plasma VEGF levels appeared to correlate with survival. In multivariate models in which other prognostic factors (serum prostate-specific antigen, alkaline phosphatase, evidence of measurable disease, and hemoglobin) were included, plasma VEGF levels were significant at various cut points tested. Conclusion: Although these data are exploratory and need to be confirmed in an independent data set, they suggest that VEGF may have clinical significance in patients with HRPC. INTRODUCTION HRPC is a uniformly fatal disease accounting for an estimated 31,900 deaths annually in the United States (1), with a median survival slightly over 1 year (2–5). Because of both selective and adaptive processes, HRPC is a heterogeneous disease. Factors that predict the outcome of HRPC patients qualitatively reflect either the overall tumor burden (e.g., elevated PSA levels, alkaline phosphatase, or bone scan findings) or condition of the host (e.g., performance status, plasma hemoglobin level, or weight loss). However, biological behavior of individual prostate cancers is not assessed directly by such factors (6). Circulating biomarkers produced by tumors may correlate with disease progression and predict a specific biological phenotype. Such markers might better characterize this heterogeneous patient population and might represent new biological targets for therapy. One such marker is VEGF. VEGF is a homodimeric cytokine that was originally identified by its effects on endothelial cell proliferation and vascular permeability (7, 8). Since its discovery, VEGF has been demonstrated to bind to two tyrosine kinase receptors, FMS-like tyrosine receptor-1 and kinase domain receptor (or VEGFR-1 and 2, respectively), on the surface of endothelial cells to regulate physiological and pathophysiological angiogenesis (9). Microvessel density is a surrogate marker of tumor angiogenesis that correlates with disease progression and survival in patients with localized prostate cancer (10–13). However, in the metastatic setting there are no surrogate markers of tumor angiogenesis. Circulating levels of VEGF, which can be detected in patients with many solid tumor cell types including prostate cancer, may be a marker of the degree and activity of tumor angiogenesis. Received 2/1/01; revised 4/20/01; accepted 4/23/01. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 1 Supported in part by a CaPCURE Young Investigator Award [to D. J. G.] and the Gelb Center for Translational Research at the DanaFarber Cancer Institute. 2 To whom requests for reprints should be addressed, at 1230 Dana Building, Dana-Farber Cancer Institute, 44 Binney Street, Boston, MA 02115. Phone: (617) 632-3466; Fax: (617) 632-2165. 3 The abbreviations used are: HRPC, hormone-refractory prostate cancer; VEGF, vascular endothelial growth factor; CALGB, Cancer and Leukemia Group B; PSA, prostate-specific antigen; HR, hazard ratio; CI, confidence interval. 1932 Vol. 7, 1932–1936, July 2001 Clinical Cancer Research Research. on July 17, 2017. © 2001 American Association for Cancer clincancerres.aacrjournals.org Downloaded from Initial studies (14, 15) evaluating circulating VEGF levels in various cancer patient populations were performed in serum; however, results were inconclusive, because VEGF is present in platelets and is released during the clotting process (16). One study (17) suggests that quantitative measurements of VEGF levels by ELISA in plasma offer the most reliable measure of circulating VEGF levels. Studies (18, 19) in patients with gastric and colorectal cancer have demonstrated an association between elevated plasma VEGF levels and advanced disease. However, the functional significance of these elevated VEGF levels is unknown. Duque et al. (20) recently studied the relationship between plasma VEGF levels in patients with prostate cancer and demonstrated that median levels were significantly higher in patients with metastatic disease compared to patients with localized prostate cancer. To explore the use of circulating VEGF levels as a prognostic factor for survival, we measured plasma VEGF in patients with HRPC enrolled into a multicenter (intergroup) study and correlated these levels with outcomes, including duration of survival. PATIENTS AND METHODS Patient Selection. An intergroup Phase III trial (CALGB 9480) of three different fixed doses of suramin was conducted (21). Between February 1996 and July 1998, 390 patients with metastatic HRPC were randomized with equal probability to receive one of three fixed dose regimens of suramin. Randomization was stratified by site (bone only versus soft tissue), performance status (0–1 versus 2), and number of prior hormonal manipulations (1–2 versus 3). Suramin was given on days 1, 2, 8, and 9 of a 28-day cycle for three cycles, with total cumulative doses of 3.19 g/m (low-dose arm), 5.32 g/m (intermediate-dose arm), and 7.66 g/m (high-dose arm). All of the patients received 40 mg/day of hydrocortisone, whereas patients in the high-dose arm also received 0.1 mg/day of fludrocortisone. Patients were eligible if they had evidence of progressive metastatic adenocarcinoma of the prostate, a life expectancy of at least 3 months, a CALGB performance status of 0–2, and adequate hematological, renal, hepatic, and clotting function. Patients were allowed no more than three prior hormonal manipulations and no prior chemotherapy, immunotherapy, or nonhormonal therapy. If patients had been treated with strontium-89 or radiation therapy, it must have been completed at least 8 weeks and 4 weeks before enrollment, respectively. The end points of the study were objective and PSA responses, progression-free survival, and overall survival. Pretreatment Blood Collection. During the accrual period, an amendment was added to that allowed for a pretreatment blood sample to be drawn for correlative studies. Blood (7 ml) drawn into glass vacutainer tubes containing EDTA was collected at various affiliated institutions and transferred to Dana-Farber Cancer Institute for plasma preparation and biomarker assessment. Within 12 h of arrival, samples were spun at 2000 3 g for 15 min. Plasma was removed, aliquotted into 500-ml microtubes, stored at 220°C, and thawed just before testing. In total, samples from 197 patients were received for these studies. Assessment of Plasma VEGF Levels. Plasma VEGF levels were determined in duplicate using a Quantiglo chemiluminescent ELISA kit (R&D Systems, Minneapolis, MN) following manufacturer’s instructions. A MLX Luminometer (DYNEX Technologies, Chantilly, VA) was used to measure light intensity correlating with VEGF binding. Statistical Analysis. Survival time was defined as the time between study entry and death. Patients lost to follow-up were censored. Exploratory statistical methods were used to find different (than the median) cut points for VEGF. The KaplanMeier product limit estimator was used to estimate the survival distribution by the two groups of VEGF levels, and the log-rank statistic was used to test for differences in the distribution of the survival times between the two groups of low and high VEGF levels. In addition, the proportional hazards model was used to assess the prognostic importance of plasma VEGF for survival adjusting for important baseline predictors, such as baseline PSA, measurable disease, and alkaline phosphatase. The HR is the ratio of the failure probability among the high-risk group (above a certain cut point) compared with the low-risk group (less than or equal to a certain cut point); e.g., if for a given cut point, the HR is greater than one, then the failure rate for patients above the cut point is higher than the failure rate for patients below the cut point. All of the tests were performed using a two-sided a level 5 0.05.