Targeting pericytes with a PDGF-B aptamer in human ovarian carcinoma models

Abstract

Purpose: On the basis of the known role of platelet-derived growth factor (PDGF)-BB/PDGF receptor (PDGFR) beta in pericyte regulation, highly specific inhibitors of this target are needed. We tested the efficacy of a highly selective aptamer against PDGF-B with or without anti-VEGF therapy in ovarian cancer models.

Results: Bevacizumab inhibited tumor growth by 45% and 48% in the HeyA8 and SKOV3ip1 models, respectively. AX102 had minimal effect on the HeyA8 model, but increased tumor growth in the SKOV3ip1 model. However, bevacizumab plus AX102 was more effective than bevacizumab alone, and resulted in 76-88% inhibition of tumor growth in both models. A longitudinal study in the HeyA8 model using bioluminescence imaging showed that combination of bevacizumab, AX102 and paclitaxel caused tumor reduction by 65% (based on bioluminescence imaging). In the HeyA8 model, MVD and PCNA counts were significantly reduced in the bevacizumab treatment groups, and pericyte coverage was significantly decreased in the AX102 treatment groups. In the SKOV3ip1 model, MVD and PCNA was significantly reduced in the bevacizumab treatment group, and even lower in the bevacizumab and AX102 combination treatment group.

Experimental design: The therapeutic efficacy of targeting endothelial cells (bevacizumab) and/or pericytes (PDGF-aptamer, AX102) was examined using HeyA8 and SKOV3ip1 orthotopic models of ovarian cancer metastasis. Following therapy, tumors were examined for microvessel density (MVD), proliferating cell nuclear antigen (PCNA) and vascular maturation (pericyte coverage).

Conclusions: Dual targeting of endothelial cells and pericytes holds potential as an anti-vascular therapeutic approach in ovarian carcinoma.

Figure 1

Effect of dual targeting of endothelial cells and pericyte on human ovarian cancer growth. (A) Mean tumor weights with standard errors are shown. (B) Mean number of tumor nodules with standard errors is shown. Mice were injected with HeyA8, or SKOV3ip1 human ovarian cancer cells. Seven days later, mice were randomized to receive treatment: (1) vehicle control; (2) bevacizumab alone (6.25 mg/kg, i.p. twice per week); (3) AX102 (50 mg/kg, i.p., daily) alone; (4) bevacizumab (6.25 mg/kg, i.p. twice per week) plus AX102 (50 mg/kg, i.p., daily). *p < 0.05 compared to vehicle controls; ^†p < 0.05 compared to bevacizumab alone. (C) Longitudinal assessment of tumor growth. Mice were injected i.p. with HeyA8-Luc cells, and seven days later were randomized to receive treatment: (1) Vehicle control; (2) bevacizumab (6.25 mg/kg, i.p. twice per week) alone; (3) bevacizumab (6.25 mg/kg, i.p. twice per week) plus docetaxel (2 mg/kg, i.p. weekly); (4) bevacizumab (6.25 mg/kg, i.p. twice per week) plus AX102 (50 mg/kg, i.p., daily) plus docetaxel (2 mg/kg, i.p. weekly). The in vivo bioluminescence imaging was conducted longitudinally at different time points. *p < 0.05 compared to other groups.

Figure 2

Effect of dual targeting of endothelial and pericyte on tumor vasculature. (A) Frozen tumor sections were stained for CD31 to reveal microvessel density using immunohistochemical staining in HeyA8 model. (B) In SKOV3 ip1 model, paraffin embedded sections were used for CD31 staining. Microvessels were counted and graphed. (C) Dual immunofluorescence staining for CD31 (endothelial marker, red) and desmin (pericyte marker, green) was performed to determine the extent of pericyte coverage in HeyA8 model. The percentage of vessels with at least 50% coverage of associated desmin-positive cells were recorded. *p < 0.05 compared to vehicle controls. ^†p < 0.05 compared to bevacizumab alone. Images were taken at original magnification x200.

Figure 3

Effect of dual targeting of endothelial cells and pericytes on tumor proliferation. Tumor sections were stained for PCNA to reveal tumor cell proliferation in (A) HeyA8 model and (B) SKOV3 ip1 model. PCNA positive cells were counted and graphed. *p < 0.05 compared to vehicle controls. ^†p < 0.05 compared to bevacizumab alone. Images were taken at original magnification x400 for (A) and x200 for (B).