Failure of annexin-based apoptosis imaging in the assessment of antiangiogenic therapy effects

Abstract

BackgroundMolecular apoptosis imaging is frequently discussed to be useful for monitoring cancer therapy. We demonstrate that the sole assessment of therapy effects by apoptosis imaging can be misleading, depending on the therapy effect on the tumor vasculature.MethodsApoptosis was investigated by determining the uptake of Annexin Vivo by optical imaging (study part I) and of 99 mTc-6-hydrazinonicotinic [HYNIC]-radiolabeled Annexin V by gamma counting (study part II) in subcutaneous epidermoid carcinoma xenografts (A431) in nude mice after antiangiogenic treatment (SU11248). Optical imaging was performed by optical tomography (3D) and 2D reflectance imaging (control, n = 7; therapy, n = 6). Accumulation of the radioactive tracer was determined ex vivo (control, n = 5; therapy, n = 6). Tumor vascularization was investigated with an optical blood pool marker (study part I) and contrast-enhanced ultrasound (both studies). Data were validated by immunohistology.ResultsA significantly higher apoptosis rate was detected in treated tumors by immunohistological terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling staining (area fraction: control, 0.023 ± 0.015%; therapy, 0.387 ± 0.105%; P < 0.001). However, both 2D reflectance imaging using Annexin Vivo (control, 13 ± 15 FI/cm2; therapy, 11 ± 7 FI/cm2) and gamma counting using 99 mTc-HYNIC-Annexin V (tumor-to-muscle ratio control, 5.66 ± 1.46; therapy, 6.09 ± 1.40) failed in showing higher accumulation in treated tumors. Optical tomography even indicated higher probe accumulation in controls (control, 81.3 ± 73.7 pmol/cm3; therapy, 27.5 ± 34.7 pmol/cm3). Vascularization was strongly reduced after therapy, demonstrated by contrast-enhanced ultrasound, optical imaging, and immunohistology.ConclusionsThe failure of annexin-based apoptosis assessment in vivo can be explained by the significant breakdown of the vasculature after therapy, resulting in reduced probe/tracer delivery. This favors annexin-based apoptosis imaging only in therapies that do not severely interfere with the vasculature.

Squamous Cell Carcinoma Xenografts: Use of VEGFR2-targeted Microbubbles for Combined Functional and Molecular US to Monitor Antiangiogenic Therapy Effects

“…Conversely, significant differences between tumor models and early antiangiogneic therapy are often difficult to determine by assessing tumor vascularization (eg, with nontargeted, contrastenhanced US). This can be explained by the fact that vascularization does not necessarily reflect the angiogenic state of tumors (30). In a tumor with low angiogenesis, slow growth, and high vessel maturation, many functional blood vessels may survive and thus this tumor appears better perfused than a rapidly growing tumor with high angiogenesis but low vessel maturation, where most angiogenic vessels rapidly undergo apoptosis and even do not carry Europe PMC Funders Author Manuscripts blood.…”

Early Prediction of Tumor Response to Treatment: Preclinical Validation of 99mTc-Duramycin

“…In the past, 99m Tc-annexin V proceeded to clinical trials, where it was used to predict and evaluate tumor response to chemotherapy (7,14) and radiotherapy (15,16) in cancer patients. Besides low and nonspecific tumor uptake of 99m Tc-annexin V, these clinical studies showed that the usefulness of this radiotracer for apoptosis imaging was limited to only specific tumor types, such as head and neck squamous cell carcinoma (15), and when using cancer therapies that have minor effects on the tumor vasculature, such as conventional chemotherapy and radiotherapy (17). Additional reasons for the failure of 99m Tc-annexin V to reach clinical practice include an inadequate biodistribution profile and low target-to-background ratio (18) resulting from the large protein structure of annexin V (36 kDa) and the fact that the list of chemical alternatives to improve the pharmacokinetics of this SPECT tracer is almost exhausted.…”

Radiopharmaceuticals as probes to characterize tumour tissue

“…Annexin V has been derivatized with a variety of SPECT radioisotopes and chelates since its characterization in the mid-1990s but from a clinical standpoint and despite reaching phase II/III clinical trials, has been plagued by poor biodistribution with high uptake in the liver and kidneys which precludes imaging in the abdominal region (Table 1) [64,125,126]. The probe is also unable to detect dying cells when accessibility is compromised because of disruption to vasculature in the context of antiangiogenic therapy [127]. Another similar but smaller PS binding protein based on the C2A domain of synaptotagmin I has been shown to be more specific than annexin V for dying cells in vitro [128].…”