Abstract 2276: Induction of apoptosis and inhibition of angiogenesis by novel fusion protein - AD-O54.9 as a new preclinical strategy in cancer treatment

• Published in: Cancer research (Chicago, Ill.) Ill.), 2014-10, Vol.74 (19_Supplement), p.2276-2276
• Main Authors: Rózga, Piotr, Pieczykolan, Jerzy, Zerek, Bartłomiej, Pieczykolan, Anna, Gałązka, Marlena, Bukato, Katarzyna, Szymanik, Michał, Jaworski, Albert, Pawlak, Sebastian, Teska-Kamińska, Małgorzata, Grochot-Przęczek, Anna
• Format: Article
• Language: English
• ISSN: 0008-5472; 1538-7445
• DOI: 10.1158/1538-7445.AM2014-2276

Background For almost two decades tumor necrosis factor-related apoptosis-inducing ligand (TRAIL/Apo2L) has been under extensive development as a potential therapeutic agent. TRAIL/Apo2L is a member of the TNF superfamily with unique ability to induce apoptosis in cancer cells while remaining neutral to normal cells. Tumor growth is tightly related to new blood vessel formation and tissue remodeling. Platelet derived growth factor (PDGF) is important in vascular physiological and pathological development. Inhibition of PDGF pathway blocks angiogenesis what results in reduction of tumor growth. We proposed a novel fusion protein (AD-O54.9) with dualistic proapoptotic and antiangiogenic activity. Our molecule consists of the soluble recombinant TRAIL/Apo2L variant linked with an effector peptide sequence and an activation motif recognized by tumor-specific proteases (MMPs, uPa) between. The effector peptide is composed of 19-amino acid fragment derived from human PDGF, which binds the PDGF receptors competitively to the natural ligand while being itself devoid of activity. As a consequence, angiogenic activity of PDGF is blocked, stimulation of new blood vessels formation does not occur and finally tumor growth is inhibited. Materials and methods AD-O54.9 protein was expressed in E. coli, using pET expression system, and purified by IEC chromatography. Cytotoxic activity of AD-O54.9 was examined using a propidium iodide assay and its antiangiogenic activity was evaluated by HUVEC tube formation and ring aortic assays. Safety was tested on human primary hepatocytes. The proapoptotic and antiproliferative activity was tested using molecular biology and flow cytometry methods. In vivo potential was examined on mice xenograft models of human colorectal adenocarcinoma (Colo205), human lung carcinoma (NCI-H460-luc2), human pancreatic carcinoma (MIA PaCa-2) and human oesophageal adenocarcinoma (OE19) cell lines where molecules showed anticancer activity. Results The molecule showed in vitro specific cytotoxic effect on various primary cancer cell lines at IC50 below 0.01 ng/ml. We demonstrated that AD-O54.9 is a very potent apoptosis inducer and inhibitor of angiogenesis. This fusion protein showed superior efficacy displaying significant tumor volume regression compared with TRAIL/Apo2L and standard chemotherapeutic agents. Conclusions The obtained results confirm that we developed a very promising fusion molecule with a high potential of anticancer activity that