Vaccine 24 (2006) 4942–4950 Intramuscular immunization with a DNA vaccine encoding a 26-amino acid CETP epitope displayed by HBc protein and containing CpG DNA inhibits atherosclerosis in a rabbit model of atherosclerosis Dan Mao, Gu Kai, Qi Gaofu, Zhu Zheng, Zhong Li, Wu Jie, Liu Jingjing, Cao Rongyue ∗ Minigene Pharmacy Laboratory, School of Life Science and Technology, China Pharmaceutical University, Tong Jia Xiang 24, Nanjing 210009, Jiangsu, PR China Received 15 October 2005; received in revised form 3 March 2006; accepted 20 March 2006 Available online 18 April 2006 Abstract Rabbits were intramuscularly immunized with the plasmid pCR-X8-HBc-CETP encoding a B-cell epitope of cholesteryl ester transfer protein (CETP) C-terminal fragment (CETPC) displayed by Hepatitis B virus core (HBc) particle. This plasmid also contained immunostim- ulatory sequences (ISS) which included eight CpG motifs 5 ′ -GACGTT-3 ′ , functioning as immunomodulators. After anti-CETP antibodies were successfully produced, rabbits were fed with a high-cholesterol diet for 15 weeks, and then the antiatherogenic effects of this DNA immunization were evaluated. The results showed that the fraction of plasma cholesterol in HDL significantly increased and the fraction of plasma cholesterol in LDL decreased in the pCR-X8-HBc-CETP immunized rabbits compared with those in the saline control group and one group treated with the plasmid pCR-X8-HBc containing ISS but lacking CETP epitope. More importantly, DNA immunization with pCR-X8-HBc-CETP markedly reduced the average percentage of aortic lesions in the entire aorta area by 80.60% compared with the saline control (3.78% versus 19.48%) and the average thickness of hyperplastic coronary artery in this group was also significantly less than in the saline control group (146 ± 11 m versus 248 ± 18 m). Our data also showed that CpG DNA alone could be antiatherogenic in this model because the average percentage of aortic lesions in pCR-X8-HBc immunized rabbits was 16.53% lower than that of the saline control group and the average thickness of hyperplastic coronary artery was also substantially lower than saline control group (155 ± 13 m versus 248 ± 18 m). Thus, plasmid pCR-X8-HBc-CETP could significantly inhibit the progression of atherosclerosis and be potentially developed as a suitable DNA vaccine against atherosclerosis. © 2006 Elsevier Ltd. All rights reserved. Keywords: DNA immunization; Atherosclerosis; Vaccine; Cholesteryl ester transfer protein; Immunostimulatory sequence; CpG 1. Introduction Atherosclerosis is a complex histopathological process, which is considered to be the most common underlying one in cardiovascular morbidity and mortality [1]. During this multi- factorial process, lipid abnormality has been demonstrated as a predictor of increasing risk of atherosclerotic cardiovascular diseases. A decrease in high-density lipoprotein cholesterol (HDL-C) or an increase in low-density lipoprotein choles- ∗ Corresponding author. Tel.: +86 25 83271242; fax: +86 25 83204240. E-mail addresses: liujingj@publicl.ptt.js.cn (L. Jingjing), caorongyuenanjing@yahoo.com.cn (C. Rongyue). terol (LDL-C) remarkably promoted atherosclerosis [2]. Cholesteryl ester transfer protein (CETP) is a 74 kDa plasma glycoprotein that mediates transfer of neutral lipids and phos- pholipids between lipoproteins and functions in the plasma to lower the concentration of HDL-C by moving cholesteryl esters from HDL to very low-density lipoproteins (VLDL) and LDL [3,4]. So CETP promotes atherosclerosis formation when its enzyme activity is excessively high and partial inhi- bition of CETP activity to the normal level is important to pre- vent atherosclerosis [5,6]. Several studies have demonstrated that inhibition of CETP activity by antibodies could decrease the development of atherosclerosis by increasing HDL-C [7,8] and no data suggested that pharmacological inhibition of 0264-410X/$ – see front matter © 2006 Elsevier Ltd. All rights reserved. doi:10.1016/j.vaccine.2006.03.082