Molecular Imaging of Changes in the Prevalence of Vascular Endothelial Growth Factor Receptor in Sunitinib-Treated Murine Mammary Tumors

Abstract

Several drugs targeting vascular endothelial growth factor (VEGF) and its receptors (VEGFRs) are approved for cancer treatment. However, these drugs induce relatively modest and frequently unpredictable tumor responses. In this work, we explored whether noninvasive imaging of VEGFR, a direct target of antiangiogenic drugs, can provide real-time information on responses to the treatment with sunitinib, a small-molecule VEGFR inhibitor approved by the Food and Drug Administration. Methods: We imaged VEGFR in an orthotopic mammary tumor model during the course of treatment with sunitinib using a recently developed SPECT tracer, a 99m Tc-labeled single-chain VEGF (scVEGF), that binds to and is internalized by VEGFR. Tumors from imaged mice were harvested and cryosectioned, and alternating sections were analyzed by autoradiography and immunohistochemistry to determine the expression of endothelial cell markers VEGFR-2 and CD31. Results: In vitro assays with endothelial cells overexpressing VEGFR-2 established that sunitinib does not inhibit VEGFR-2-mediated uptake of scVEGFbased tracers. SPECT and autoradiography with 99m Tc-scVEGF of tumor cryosections revealed a 2.2-to 2.6-fold decrease in tracer uptake after 4 daily doses of sunitinib. However, once treatment was discontinued, tracer uptake rapidly (3 d) increased, particularly at the tumor edges. Immunohistochemical analysis of VEGFR-2 and CD31 supported SPECT and autoradiographic imaging findings, revealing the corresponding depletion of VEGFR-2-and CD31-positive endothelial cells from tumor vasculature during therapy and the rapid reemergence of VEGFR-2-and CD31-positive vasculature at the tumor edges after discontinuation of treatment. Conclusion: Our findings suggest that imaging with 99m Tc-scVEGF might be useful for monitoring VEGFR responses to antiangiogenic treatment regimens.

Targeted Systemic Radiotherapy with scVEGF/177Lu Leads to Sustained Disruption of the Tumor Vasculature and Intratumoral Apoptosis

“…Orthotopic 4T1luc breast tumors in 5-to 8-wk-old female BALB/c mice and MDA231luc breast tumors in 5-to 8-wk-old ICR (Institute of Cancer Research, Fox Chase)-severe combined immune-deficient (SCID) mice (ICR and C.B-17-SCID background; Taconic) were generated as described (6,8). Tumor volume was calculated as V 5 0.52 · A 2 · B, where A is the short axis and B is the long axis of a tumor.…”

“…Cryosections (20 mm) were stained as described previously (7,8). For quantitative analysis of avascular areas, color images were acquired with a microscope objective (·4) and exported into Photoshop (Adobe).…”

“…We have previously exploited VEGFR overexpression in tumor vasculature for selective SPECT and PET of these receptors (6,7) and their response to antiangiogenic therapy (8,9), using scVEGF, an engineered single-chain (sc) form of VEGF (6)(7)(8)(9)(10) for the intracellular delivery of 64 Cu, 68 Ga, and 99m Tc. In the current study, we chose 177 Lu as our radiotherapeutic isotope because of its relatively mild b-emission (;1.5-mm maximum penetration in soft tissue) capable of inducing bystander cytotoxic effects in tumors but not in surrounding host tissue (11), its logistically convenient intermediate halflife (6.7 d), and its commercial availability under good manufacturing practice conditions for possible future clinical development.…”

In vivo small animal imaging: Current status and future prospects

“…248 It is here where in vivo imaging may illuminate the key mechanisms and bridge the gap between success in animal and failure in human trials and several recent publications support this hypothesis. Although numerous studies are using molecular imaging as a technique to qualitatively assess and quantify the efficacy of angiogenic treatment, [249][250][251][252] we will focus on some examples, where imaging is combined with therapy.…”

Mind the Gap: Potential for Rebounds during Antiangiogenic Treatment Breaks

“…1). Previously, preclinical models have shown that halting antiangiogenic therapy could lead to rapid vascular regrowth (2), and some studies have shown tumors can quickly grow after therapy is stopped (3,4). Furthermore, clinical examples of tumor "flares" have been noted during drug holidays or upon treatment discontinuation (5), and isolated case reports show that occult metastatic disease, if missed during cytoreductive sunitinib therapy prior to nephrectomy, can rapidly regrow during prolonged drug washout periods prior to surgery (6).…”