Reverse translation of phase I biomarker findings links the activity of angiotensin-(1-7) to repression of hypoxia inducible factor-1α in vascular sarcomas

Abstract

Background: In a phase I study of angiotensin-(1-7) [Ang-(1-7)], clinical benefit was associated with reduction in plasma placental growth factor (PlGF) concentrations. The current study examines Ang-(1-7) induced changes in biomarkers according to cancer type and investigates mechanisms of action engaged in vitro.

Methods: Plasma biomarkers were measured prior to Ang-(1-7) administration as well as 1, 2, 3, 4, and 6 hours after treatment. Tests for interaction were performed to determine the impact of cancer type on angiogenic hormone levels. If a positive interaction was detected, treatment-induced biomarker changes for individual cancer types were assessed. To investigate mechanisms of action, in vitro growth assays were performed using a murine endothelioma cell line (EOMA). PCR arrays were performed to identify and statistically validate genes that were altered by Ang-(1-7) treatment in these cells.

Results: Tests for interaction controlled for dose cohort and clinical response indicated a significant impact of cancer type on post-treatment VEGF and PlGF levels. Following treatment, PlGF levels decreased over time in patients with sarcoma (P = .007). Treatment of EOMA cells with increasing doses of Ang-(1-7) led to significant growth suppression at doses as low as 100 nM. PCR arrays identified 18 genes that appeared to have altered expression after Ang-(1-7) treatment. Replicate analyses confirmed significant changes in 8 genes including reduction in PlGF (P = .04) and hypoxia inducible factor 1α (HIF-1α) expression (P < .001).

Conclusions: Ang-(1-7) has clinical and pre-clinical activity for vascular sarcomas that is linked to reduced HIF-1α and PlGF expression.

Figure 1

Post-treatment PlGF changes according to cancer type on day 1 of treatment. The lines shown represent trend lines for median normalized values. Red color indicates a significant trend (P < .05) based on the regression statistic. For cancer types with a significant trend, median values and standard deviation error bars are shown for each time point. * indicates a significant difference in biomarker value (P < .05) compared to the time 0 biomarker value based on a two-sample t-test.

Figure 2

(A) Radiographic changes in a patient with metastatic hemangiopericytoma after two cycles of treatment. (B) Decreases in PlGF concentrations are shown for the same patient on day 1 of Ang-(1–7) treatment. Error bars represent the standard deviations of repeated ELISA analyses from a single plasma sample collected at each time point from this patient.

Figure 3

Changes in the proliferation of murine endothelioma (EOMA) cells following treatment with either Ang-(1–7) (black bars) or Ang II (white bars) are shown. * indicates a significant difference in proliferation (P < .05) compared to the proliferation rate of untreated cells.

Figure 4

Changes in expression of target genes following treatment with Ang-(1–7). Error bars represent the 95% confidence interval. * indicates a significant difference in target gene expression (P < .05) in Ang-(1–7) treated cells as compared to untreated cells.