Identifying a Cardiac PTD Figure 7. CTP targets fluospheres to the heart. Whole mouse in vivo imaging was performed following intra-cardiac injection of fluospheres alone, CTP+fluospheres and RAN+fluospheres at 30, 60, 120 and 180 minutes, at a dose of 10 mg/Kg of body weight (for peptides). (N = 3 in each group). doi:10.1371/journal.pone.0012252.g007 gene delivery using viral-based vectors. In addition, there are issues regarding the presence of pre-existing neutralizing antibod- ies or immune responses to certain viral vectors. The well- characterized cell penetrating peptides, like TAT from HIV coat protein, homopolymers of arginine or lysine, are not cell specific and transduce hepatocytes and multiple other organs in addition to the heart. Therefore, identifying a peptide with transduction capabilities specific for the heart would allow for new approaches for effective cardiac delivery of therapeutics. We previously have reported the ability to identify a synovial specific transduction peptide by screening an M13 phage peptide display library for internalized phage [8]. Thus we screened a large phage peptide display library in order to identify novel peptides potentially able to transduce cardiomyocytes in vivo. Indeed, we report here the identification of a specific peptide, termed CTP, which 15 or 30 minutes post-peripheral intravenous injection can efficiently and specifically transduce cardiac tissue (Fig. 4 and 5 respectively). Transduction of cardiomyocytes on confocal microscopy of cross-sections of the mouse heart occurred in a diffuse manner, though there appeared to be some preference for the subendocardial and subepicardial regions at earlier time point of 15 minutes. No other organ showed uptake except kidney glomeruli, limited to the cortex, and rare lung capillaries, to a much lesser extent than heart tissue. Furthermore, CTP was able to transduce heart tissue in vivo far more efficiently and in a tissue- specific manner as compared to 8-Lysine, a known PTD (Fig. 7). Since the initial description of in vivo screening of phage display libraries by Pasqualini and Ruoslahti [13], this approach has been utilized to identify peptides that target tumor vasculature [5], adipose tissue [6], pancreatic islet cells [7], synoviocytes [8], PLoS ONE | www.plosone.org 9 August 2010 | Volume 5 | Issue 8 | e12252