Date of Award

Spring 8-2021

Document Type


Degree Name

Master of Science (MS)


Pharmaceutical Sciences

First Advisor

Keykavous Parang, Pharm.D., Ph.D.

Second Advisor

Rakesh Tiwari, Ph.D.

Third Advisor

Hamidreza Montazeri Aliabadi, Ph.D.


The cell membrane properties create a significant obstacle in intracellular delivery of cell-impermeable and negatively charged molecules. Amphiphilic cyclic peptides containing alternative arginine and tryptophan such as [WR]5 have been shown to enhance the transport of cargo molecules across the cell membrane. Herein, we report the synthesis and biological evaluation of a novel series of hybrid cyclic-linear peptides containing alternative positive and hydrophobic amino acids on the ring and side-chain [(RW)5]K(RW)X (X=1-5) to compare their molecular transporter efficiency. The peptides were synthesized through Fmoc solid-phase peptide synthesis. Final compounds were purified by a reversed-phase HPLC (High Performance Liquid Chromatography). The chemical structures of the final products were confirmed by MALDI-TOF (Matrix Assisted Laser Desorption/Ionization).

In vitro cytotoxicity of the peptides was evaluated in human leukemia carcinoma cell line (CCRF-CEM), human ovarian adenocarcinoma cells (SK-OV-3), human epithelial embryonic kidney healthy (HEK-293), and human epithelial mammary gland adenocarcinoma cells (MDA-MB-231) using the MTS assay. The peptides did not exhibit any significant cytotoxicity at the concentration of 10 μM in CCRF-CEM, HEK-293, MDA-MB-231, and SK-OV-3 cells after 3 h incubation.

The cellular uptake of a fluorescence-labeled phosphopeptide (F′-GpYEEI) and anti-HIV drugs (lamivudine (F′-3TC), emtricitabine (F′-FTC), Stavudine (F′-d4T)), where F′ is carboxyfluorescein, were measured in the presence of the peptides in CCRF-CEM and SK-OV-3 cells. Among all peptides, [(RW)5K](RW)5 (10 μM) was the most efficient transporter and improved the cellular uptake of F′-GpYEEI (2 μM) by 18 and 11-fold in CCRF-CEM and SK-OV-3, respectively, when compared with F′-GpYEEI alone and in the presence of [WR]5.

The cellular uptake of F′-[(RW)5K](RW)5 was increased in a time- and concentration-dependent manner. The FACS (fluorescence-activated cell sorting) analysis and confocal microscopy results indicate that the cellular uptake of fluorescent-labeled peptide (F'-[(RW)5K](RW)5) was partially inhibited by chlorpromazine endocytosis inhibitors after 3 h incubation in MDA-MB-231 cells, which suggests the partial uptake through the clathrin-mediated endocytosis pathway. Intracellular localization (mostly in the cytosol) of F'-[(RW)5K](RW)5 after 3 h incubation in MDA-MB-231 cells was confirmed by confocal microscopy. These data suggest the potential of this series of hybrid cyclic-linear peptides compared to [WR]5 as molecular transporters of large molecules, such as a negatively charged phosphopeptide.

Creative Commons License

Creative Commons License
This work is licensed under a Creative Commons Attribution-Noncommercial-No Derivative Works 4.0 License.



To view the content in your browser, please download Adobe Reader or, alternately,
you may Download the file to your hard drive.

NOTE: The latest versions of Adobe Reader do not support viewing PDF files within Firefox on Mac OS and if you are using a modern (Intel) Mac, there is no official plugin for viewing PDF files within the browser window.