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  Cylop-1 Peptide: a Tool for the Development of Targeted Intracellular Molecular Imaging Probes

Dhingra, K., Jha, D., Ugurbil, K., & Engelmann, J. (2009). Cylop-1 Peptide: a Tool for the Development of Targeted Intracellular Molecular Imaging Probes. Poster presented at 4th Europen Molecular Imaging Meeting (EMIM 2009), Barcelona, Spain.

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Item Permalink: http://hdl.handle.net/21.11116/0000-0003-14D2-9 Version Permalink: http://hdl.handle.net/21.11116/0000-0003-14D3-8
Genre: Poster

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Dhingra, K1, 2, Author              
Jha, D2, 3, Author              
Ugurbil, K, Author
Engelmann, J2, 3, Author              
Affiliations:
1Department Physiology of Cognitive Processes, Max Planck Institute for Biological Cybernetics, Max Planck Society, ou_1497798              
2Max Planck Institute for Biological Cybernetics, Max Planck Society, Spemannstrasse 38, 72076 Tübingen, DE, ou_1497794              
3Former Department MRZ, Max Planck Institute for Biological Cybernetics, Max Planck Society, ou_2528700              

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 Abstract: introduction: Molecular imaging of cellular processes requires the interaction of imaging probes with intracellular targets like proteins, mrna or dna. The plasma membrane of cells poses an impervious barrier to the intracellular delivery of such probes. Cell penetrating peptides (CPPs) gained importance as vectors for transport of cargos attached to them into cells primarily by endocytosis. however, confi nement of biomolecules into endosomes limited their use for intracellular targeting. We present here the development of a novel cysteine-rich peptide, CyloP-1, that shows the ability to deliver into the cytosolic compartment of the cell. The transport of gd-doTa, a contrast agent for magnetic resonance imaging (Mri), after conjugation with CyloP-1 will be discussed as well. Methods: The conjugates were synthesized by continuous solid phase synthesis using fmoc chemistry and n-terminally labeled with fl uorescein isothiocyanate. Cellular uptake of compounds was confi rmed by fl uorescence microscopy and spectroscopy in nih-3T3 mouse fi broblast cells plated in 96well plates. internalized fl uorescence was measured in a multiplate reader, and microscopic images were made to determine the cellular localization of conjugates. Mr analysis of gd-doTa conjugates was also performed on labeled cells in eppendorf tubes. Mri of cell pellets was conducted at 3T in a Siemens human Mr scanner using T1- and T2-weighted spin-echo sequences. relaxation rates were obtained from axial slices as well as images of sagittal slices were made. results: The novel cysteine rich CPP was derived by Structure activity relationship (Sar) studies of a sequence from the polypeptide Crotamine [1]. CyloP-1 is markedly distinct showing an effi cient uptake at low concentrations (2.5 μM) and a cytosolic distribution along with vesicular uptake (fig. 1a) unlike other common CPPs (e.g. Tat or antennapedia) at these concentrations. The localization of CyloP-1 into the cytosol was maintained even at 4°C while the vesicular uptake was much reduced. additional coupling of gd-doTa to this peptide showed profi cient uptake maintaining the cytosolic localization of the conjugate (fig. 1b). furthermore, fl uorescence spectroscopic data showed that the internalization effi cacy at 2.5 μM increased signifi cantly by about 50% in comparison to the conjugate containing the d-form of Tat peptide, known to be an effi cient CPP. Conclusions: These results demonstrate that our novel peptide might prove useful for efficient trans-membrane delivery of molecular imaging probes directed to cytosolic targets.

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 Dates: 2009-05
 Publication Status: Published online
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Title: 4th Europen Molecular Imaging Meeting (EMIM 2009)
Place of Event: Barcelona, Spain
Start-/End Date: 2009-05-27 - 2009-05-30

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Title: 4th Europen Molecular Imaging Meeting (EMIM 2009)
Source Genre: Proceedings
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Pages: - Volume / Issue: - Sequence Number: 38 Start / End Page: 198 Identifier: -