Platelets Take Up the Monoclonal Antibody Bevacizumab

^{1 1}The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins and ²Department of Radiology, Johns Hopkins Medical Institutions, Baltimore, Maryland; ³Department of Medical Oncology and ⁴Thrombosis and Hemostasis Laboratory, University Medical Center Utrecht, the Netherlands

^* To whom correspondence should be addressed. E-mail: h.m.w.verheul{at}umcutrecht.nl.

	Abstract

Purpose: One of the key factors that promotes angiogenesis is vascular endothelial growth factor (VEGF). Platelets are the main source of VEGF in blood and contribute to angiogenesis by release of growth factors, including VEGF, from their -granules on activation. The monoclonal antibody bevacizumab blocks VEGF in the blood of patients within hours after administration. Platelets are known to endocytose plasma proteins including immunoglobulins. We tested the hypothesis that platelets take up bevacizumab.

Experimental Design: Fluorescence-activated cell sorting analysis, immunofluorescence imaging, and Western blotting were used to study uptake and release of bevacizumab by platelets in vitro and in vivo. The angiogenic activity of platelets preincubated with bevacizumab was studied in endothelial proliferation assays. Finally, we determined whether treatment with bevacizumab neutralizes VEGF in platelets from cancer patients.

Results: We found that platelets are able to take up bevacizumab. Activation of platelets preincubated with bevacizumab resulted in release of the antibody and release of VEGF neutralized by bevacizumab. Immunofluorescence microscopy revealed that FITC-labeled bevacizumab and P-selectin colocalize, indicating -granule localization. In addition, bevacizumab uptake inhibited platelet-induced human endothelial cell proliferation. In in vivo rabbit experiments, FITC-labeled bevacizumab was present in platelets after 2 h and up to 2 weeks following i.v. administration. Finally, we found that platelets take up bevacizumab in patients receiving bevacizumab treatment. Within 8 h after bevacizumab administration, platelet VEGF was almost completely neutralized due to this uptake.

Conclusion: These studies show that bevacizumab is taken up by platelets and may explain its clinical effect on wound healing and tumor growth.

Key Words: therapeutic antibodies, bevacizumab, platelet, angiogenesis, VEGF

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