Beyond diagnostic applications, several antitumor therapies are known from the literature that act through the direct inhibition of the APN/CD13 molecule, and also various NGR-conjugated therapeutic brokers have been developed that deliver different cytotoxins (e.g., NGR-coated liposomes, doxorubicin-NGR conjugates, and NGR-TNF alpha conjugates) to APN/CD13-positive solid tumors . Two main types of APN/CD13 inhibitors are known, the synthetic and the naturally occurring form. metastases due to their APN/CD13-specific inhibitor properties. Our previous studies have already shown that 68Ga-labeled NGR peptides bind specifically to APN/CD13 expressing tumor cells. The APN/CD13 specificity of 68Ga-NGR radiopharmaceuticals enables the following of the efficacy of antiangiogenic therapy with APN/CD13-specific inhibitors using positron emission tomography (PET). The aim of this study was to assess the antitumor effect of bestatin and actinonin treatment in subcutaneous ELX-02 sulfate transplanted HT1080 and B16-F10 tumor-bearing animal models using 68Ga-NODAGA-c(NGR). Materials and Methods Three days ELX-02 sulfate after the inoculation of HT1080 and B16-F10 cells, mice were treated with intraperitoneal injection of bestatin (15?mg/kg) or actinonin (5?mg/kg) for 7 days. Around the 5th and 10th day, PET scans and biodistribution studies were performed 90?min after intravenous injection of 5.5 0.2?MBq68Ga-NODAGA-c(NGR). Results Control-untreated HT1080 and B16-F10 tumors were clearly visualized by the APN/CD13-specific 68Ga-NODAGA-c(NGR) radiopharmaceutical. The western blot analysis also confirmed the strong APN/CD13 positivity in the investigated tumors. We found significantly ( 0.05) lower radiopharmaceutical uptake after bestatin treatment and higher radiotracer accumulation in the actinonin-treated HT1080 tumors. In contrast, significantly lower ( 0.01) 68Ga-NODAGA-c(NGR) accumulation was observed in both bestatin- and actinonin-treated B16-F10 melanoma tumors compared to the untreated-control tumors. Bestatin inhibited tumor growth and 68Ga-NODAGA-c(NGR) uptake in both tumor models. Conclusion The bestatin treatment is suitable for suppressing the neoangiogenic process and APN/CD13 expression of experimental HT1080 and B16-F10 tumors; furthermore, 68Ga-NODAGA-c(NGR) is an applicable radiotracer for the monitoring of the efficacy of the APN/CD13 inhibition-based anticancer therapies. 1. Introduction Angiogenesisthe new blood vessel formation from a preexisting capillary systemplays an important role in different physiological processes such as wound healing  and the action of the female reproduction system , but it can emerge in malignant processes such as psoriasis [3, 4], rheumatoid arthritis , retinopathies , and cancers [7, 8]. During the angiogenic process, several receptors and molecules (e.g., VEGF, integrins, and APN/CD13) appear in the cell surface, which provide opportunities to detect and treat malignant tumors [9C12]. Among these angiogenic molecules, aminopeptidase N (APN/CD13) showed a strong correlation with tumor-associated neoangiogenesis [13C15]. APN/CD13 is usually a 160?kDa weighted and glycosilated, zinc-dependent transmembrane ectopeptidase. It has three main functions: enzyme, receptor, and signaling molecule . As an enzyme, it plays an important role in peptide cleavage, such as angiotensins, kinins, enkephalins, cytokines, and chemokines. Furthermore, APN/CD13 participates in extracellular matrix protein degradation, which facilitates tumor cell invasion and migration. As a receptor, APN/CD13 is involved in endocytosis during viral contamination; moreover, as a signaling molecule, it attends in adhesion, phagocytosis, and angiogenic processes . APN/CD13 is usually physiologically expressed in the epithelial cells of the liver, intestine, and kidney and in the synaptic membranes and pericytes of the central nervous system . Several studies reported that APN/CD13 is usually overexpressed in the endothelial cells of tumor vasculature and in several solid tumors, such as melanoma [18, ELX-02 sulfate 19], prostate carcinoma , lung cancer Rabbit Polyclonal to RANBP17 , pancreas adenocarcinoma , ovarian cancer , breast cancer , colon cancer , thyroid cancer , and fibrosarcomas . Due to its elevated expression, APN/CD13 was reviewed as an important clinical marker in several inflammatory diseases and malignant cancers [22, 27, 28], and it has been considered as a suitable target for anticancer and anti-inflammatory therapy [29C32]. In antiangiogenesis therapy, the most frequently administered natural variants of APN/CD13 inhibitors are actinonin, amastatin, bestatin, phebestin, probestin, and curcumin, most of which are originated from bacteria or plants . Bestatin is usually a well-known and potent APN/CD13 inhibitor which has already ELX-02 sulfate been investigated by several authors in and studies. Bestatin, due to its competitive, reversible protease inhibitor properties, has an antiangiogenic effect through the inhibition of APN/CD13’s activity in numerous tumors (e.g., murine colon adenocarcinoma and myeloid leukemia , human promyelocytic leukemia , human choriocarcinoma , murine melanoma ELX-02 sulfate [36, 37], murine lung carcinoma , human.