Cardiac Targeting Peptide, a Novel Cardiac Vector: Studies in Bio-Distribution, Imaging Application, and Mechanism of Transduction
November 14, 2018
Maliha Zahid, Kyle S. Feldman, Gabriel Garcia-Borrero, Timothy N. Feinstein, Nicholas Pogodzinski, Xinxiu Xu, Raymond Yurko, Michael Czachowski , Yijen L. Wu, Neale S. Mason and CeciliaW. Lo Biomolecules 2018, 8, 147; doi:10.3390/biom8040147 Received: 24 September 2018 / Accepted: 8 November 2018 / Published: 14 November 2018
Our previous work identified a 12-amino acid peptide that targets the heart, termed cardiac targeting peptide (CTP).We now quantitatively assess the bio-distribution of CTP, show a clinical application with the imaging of the murine heart, and study its mechanisms of transduction. Bio-distribution studies of cyanine5.5-N-Hydroxysuccinimide (Cy5.5) labeled CTP were undertaken in wild-type mice. Cardiac targeting peptide was labeled with Tc. 99m (99mTc) using the chelator hydrazino-nicotinamide (HYNIC), and imaging performed using micro-single photon emission computerized tomography/computerized tomography (SPECT/CT). Human-induced pluripotent stem cell (iPSC)-derived cardiomyocytes (CMCs) were incubated with dual-labeled CTP, and imaged using confocal microscopy. TriCEPs technology was utilized to study the mechanism of transduction. Bio-distribution studies showed peak uptake of CTP at 15 min. 99mTc-HYNIC-CTP showed heart-specific uptake. Robust transduction of beating human iPSC-derived CMCs was seen. TriCEPs experiments revealed five candidate binding partners for CTP, with Kcnh5 being felt to be the most likely candidate as it showed a trend towards being competed out by siRNA knockdown. Transduction efficiency was enhanced by increasing extracellular potassium concentration, and with Quin., a Kcnh5 inhibitor, that blocks the channel in an open position. We demonstrate that CTP transduces the normal heart as early as 15 min. 99mTc-HYNIC-CTP targets the normal murine heart with substantially improved targeting compared with 99mTc Sestamibi. Cardiac targeting peptide’s transduction ability is not species limited and has human applicability. Cardiac targeting peptide appears to utilize Kcnh5 to gain cell entry, a phenomenon that is affected by pre-treatment with Quin. and changes in potassium levels.
Maliha Zahid 1,*, Kyle S. Feldman 1, Gabriel Garcia-Borrero 1, Timothy N. Feinstein 1, Nicholas Pogodzinski 1, Xinxiu Xu 1, Raymond Yurko 2, Michael Czachowski 3, Yijen L. Wu 1, Neale S. Mason 3 and CeciliaW. Lo 1 1 Department of Developmental Biology, University of Pittsburgh, Pittsburgh, PA 15201, USA; firstname.lastname@example.org (K.S.F.); email@example.com (G.G.-B.); firstname.lastname@example.org (T.N.F.); email@example.com (N.P.);firstname.lastname@example.org (X.X.); email@example.com (Y.L.W.); firstname.lastname@example.org (C.W.L.) 2 Peptide Synthesis Facility, University of Pittsburgh, Pittsburgh, PA 15201, USA; email@example.com 3 Department of Radiology, University of Pittsburgh, Pittsburgh, PA 15201, USA; firstname.lastname@example.org (M.C.); email@example.com (N.S.M.)