Basic Res Cardiol 97: 348 – 358 (2002) DOI 10.1007/s00395-002-0360-0 ORIGINAL CONTRIBUTION Jing Zhao Gavin J. Pettigrew Joan Thomas Jamie I. Vandenberg Luc Delriviere Eleanor M. Bolton Andrew Carmichael Jody L. Martin Michael S. Marber Andrew M. L. Lever Lentiviral vectors for delivery of genes into neonatal and adult ventricular cardiac myocytes in vitro and in vivo  Abstract Vectors based on lentiviruses such as human immunodeficiency virus (HIV) type-1 have many advantages for gene therapy, including the abil- ity to infect non-dividing cells, long-term transgene expression and the absence of induction of an inflammatory/immune response. This study was initiated to determine whether lentiviruses would efficiently transfer genes to both neonatal and adult cardiac cells in culture and, by direct injection, to the heart in vivo. A three-plasmid expression system, including a packaging defective helper construct, a plasmid coding for a heterologous (VSV-G) envelope protein and a vector construct harboring reporter genes – E-GFP (enhanced green fluorescent protein) and puro (puromycin-resistance pro- tein) was used to generate pseudotyped HIV-1 particles by transient trans- fection of human embryonic kidney 293T cells. We demonstrated efficient gene transfer into neonatal and adult cardiac myocytes in vitro and identified conditions in which virtually 100 % of cultured neonatal and 70 % of adult car- diac myocytes express the reporter gene. Transduction of adult cardiac myo- cytes with high titre lentiviral vectors did not affect the cell number, mor- phology or viability compared to untransduced cells. We delivered HIV-1- based vectors to the intact heart by direct injection. Hearts transduced with pseudotyped HIV-1 vectors showed levels of transgene expression compara- ble to that achieved by adenovirus vectors. This study demonstrates for the first time that lentivirus-based vectors can successfully transduce adult cardiomyocytes both in vitro and in vivo, and opens up the prospect of lentivirus-based vectors becoming an important gene delivery system in the cardiovascular field.  Key words HIV-1 – lentiviral vector – cardiac myocytes – gene transfer – heart BRC 360 Prof. A. M. L. Lever () · J. Zhao · J. Thomas A. Carmichael Department of Medicine Addenbrooke’s Hospital Hills Road Cambridge, CB2 2QQ, UK Tel.: 01223/336747 Fax: 01223/336846 E-Mail: amll1@mole.bio.cam.ac.uk G. J. Pettigrew · L. Delriviere · E. M. Bolton University of Cambridge Department of Surgery Addenbrooke’s Hospital Hills Road Cambridge, CB2 2QQ, UK J. I. Vandenberg University of Cambridge Department of Biochemistry Building O, Downing Site Cambridge, CB2 1QW, UK J. L. Martin · M. S. Marber Department of Cardiology King’s College London Rayne Institute St Thomas Hospital London, SE1 7EH, UK Received: 1 October 2001 Returned for 1. revision: 18 October 2001 1. Revision received: 19 November 2001 Returned for 2. revision: 6 December 2001 2. Revision received: 13 February 2002 Accepted: 6 March 2002 Introduction Gene delivery to cardiac myocytes has the potential both to therapeutically correct genetic defects and, investi- gationally, to study cardiac muscle physiology. It has proven difficult to deliver genes to cardiac myocytes as they are terminally differentiated, they do not divide, and, in vitro, have a relatively short life span (19). Gene delivery to cardiac myocytes is further hampered by tox- icity associated with delivery vectors; adenoviruses, for example, may disrupt the physiology of the cell (54).