Dindyal Mandal

Design and Biological Evaluation of Cell-Penetrating Peptide–Doxorubicin Conjugates as Prodrugs

Doxorubicin (Dox) is a hydrophilic anticancer drug that has short retention time due to the efficient efflux in some cancer cells (e.g., ovarian adenocarcinoma SK-OV-3). Cyclic [W(RW)(4)] and the corresponding linear peptide (RW)(4) were conjugated with Dox through an appropriate linker to afford cyclic [W(RW)(4)]-Dox and linear (RW)(4)-Dox conjugates to enhance the cellular uptake and cellular retention of the parent drug for sustained anticancer activity. Comparative antiproliferative assays between covalent (cyclic [W(RW)(4)]-Dox and linear (RW)(4)-Dox) and the corresponding noncovalent physical mixtures of the peptides and Dox were performed. Cyclic [W(RW)(4)]-Dox inhibited the cell proliferation of human leukemia (CCRF-CEM) (62-73%), ovarian adenocarcinoma (SK-OV-3) (51-74%), colorectal carcinoma (HCT-116) (50-67%), and breast carcinoma (MDA-MB-468) (60-79%) cells at a concentration of 1 μM after 72-120 h of incubation. Cyclic [W(RW)(4)]-Dox exhibited higher antiproliferative activity than linear (RW)(4)-Dox in all cancer cells with the highest activity observed after 72 h. Flow cytometry analysis showed 3.6-fold higher cellular uptake of cyclic [W(RW)(4)]-Dox than Dox alone in SK-OV-3 cells after 24 h incubation. The cellular hydrolysis study showed that 99% of cyclic [W(RW)(4)]-Dox was hydrolyzed intracellularly within 72 h and released Dox. These data suggest that cyclic [W(RW)(4)]-Dox can be used as a potential prodrug for improving the cellular delivery and retention of Dox.

Cyclic peptide-capped gold nanoparticles as drug delivery systems.

A number of cyclic peptides were synthesized and evaluated as simultaneous reducing and capping agents for generation of cyclic peptide-capped gold nanoparticles (CP-AuNPs). Among them, direct dissolution of cyclic peptides containing alternate arginine and tryptophan [WR](n) (n = 3-5) into an aqueous solution of AuCl(4)(-) led to the formation of CP-AuNPs, through the reducing activity of tryptophan residues and attraction of positively charged arginine residues toward chloroaurate anions in the reaction environment. Differential interference contrast microscopy of fluorescence-labeled lamivudine in the presence of [WR](4)-capped AuNPs showed significantly higher cellular delivery of antiviral drug versus that of parent drug alone. Flow cytometry studies also showed that the cellular uptake of fluorescence-labeled lamivudine, emtricitabine, and stavudine was significantly enhanced in human ovarian adenocarcinoma (SK-OV-3) cells in the presence of [WR](4)-AuNPs. For example, fluorescence labeled lamivudine-loaded [WR](4)-AuNPs exhibited approximately 12- and 15-times higher cellular uptake than that of fluorescence labeled lamivudine alone in CCRF-CEM cells and SK-OV-3 cells, respectively. Confocal microscopy revealed that the presence of the [WR](4)-AuNPs enhanced the retention and nuclear localization of doxorubicin in SK-OV-3 cells after 24 h. These data suggest that these complexes can be used as potential noncovalent prodrugs for delivery of antiviral and anticancer agents.

Cyclic Peptide-Capped Gold Nanoparticles for Enhanced siRNA Delivery

Previously, we have reported the synthesis of a homochiral l-cyclic peptide [WR]5 and its use for delivery of anti-HIV drugs and biomolecules. A physical mixture of HAuCl4 and the peptide generated peptide-capped gold nanoparticles. Here, [WR]5 and [WR]5-AuNPs were tested for their efficiency to deliver a small interfering RNA molecule (siRNA) in human cervix adenocarcinoma (HeLa) cells. Flow cytometry investigation revealed that the intracellular uptake of a fluorescence-labeled non-targeting siRNA (200 nM) was enhanced in the presence of [WR]5 and [WR]5-AuNPs by 2- and 3.8-fold when compared with that of siRNA alone after 24 h incubation. Comparative toxicity results showed that [WR]5 and [WR]5-AuNPs were less toxic in cells compared to other available carrier systems, such as Lipofectamine.

Self-Assembled Surfactant Cyclic Peptide Nanostructures as Stabilizing Agents

A number of cyclic peptides including [FR]4, [FK]4, [WR]4, [CR]4, [AK]4, and [WK]n (n = 3-5) containing L-amino acids were produced using solid-phase peptide synthesis. We hypothesized that an optimal balance of hydrophobicity and charge could generate self-assembled nanostructures in aqueous solution by intramolecular and/or intermolecular interactions. Among all the designed peptides, [WR]n (n = 3-5) generated self-assembled vesicle-like nanostructures at room temperature as shown by transmission electron microscopy (TEM), scanning electron microscopy (SEM), and/or dynamic light scattering (DLS). This class of peptides represents the first report of surfactant-like cyclic peptides that self-assemble into nanostructures. A plausible mechanistic insight into the self-assembly of [WR]5 was obtained by molecular modeling studies. Modified [WR]5 analogues, such as [WMeR]5, [WR(Me)2]5, [WMeR(Me)2]5, and [WdR]5, exhibited different morphologies to [WR]5 as shown by TEM observations. [WR]5 exhibited a significant stabilizing effect for generated silver nanoparticles and glyceraldehyde-3-phosphate dehydrogenase activity. These studies established a new class of surfactant-like cyclic peptides that self-assembled into nanostructures and could have potential applications for the stabilization of silver nanoparticles and protein biomolecules.

Cyclic peptides containing tryptophan and arginine as Src kinase inhibitors

A number of cyclic and linear peptides containing various combinations of amino acids were evaluated for their Src kinase inhibitory potency. Among all the peptides, cyclic decapeptide C[RW]5 containing alternative arginine (R) and tryptophan (W) residues was found to be the most potent Src kinase inhibitor. C[RW]5 showed higher inhibitory activity (IC50=2.8 μM) than C[KW]5, L(KW)5, C[RW]4, and C[RW]3 with IC50 values of 46.9, 69.1, 21.5, and 25.0 μM, respectively, as determined in a fluorescence intensity-based assay. Thus, the cyclic nature, the presence of arginine, ring size, and the number of amino acids in the structure of the peptide were found to be critical in Src kinase inhibitory potency. The IC50 value of C[RW]5 was found to be 0.8 μM in a radioactive assay using [γ-(32)P]-ATP and polyE4Y as the substrate. C[RW]5 was a noncompetitive Src kinase inhibitor, showing approximately fourfold more selectivity towards Src than Abl.

Photo-bioreduction of Ag(+) ions towards the generation of multifunctional silver nanoparticles: Mechanistic perspective and therapeutic potential.

In this article, light induced plant extract mediated one pot synthesis of silver nanoparticles (AgNPs) has been demonstrated and potential mechanistic insight in the synthesis has been investigated. Bioactive molecules containing medicinal plant Cassytha filiformis has been explored for the synthesis of silver nanoparticles. The as-synthesized silver nanoparticles were characterized by various analytical techniques including Ultraviolet-visible spectroscopy (UV-Vis), High Resolution Transmission Electron Microscopy (HR-TEM), Dynamic Light Scattering (DLS) and Fourier Transform Infrared Spectroscopy (FT-IR). Among different light sources (sunlight, room light, UV) applied the sunlight was found to be efficient external stimuli to induce rapid synthesis of AgNPs at room temperature. Modified DPPH assay indicated that polyphenolic compounds were most likely involved in the synthesis of AgNPs. Possible molecule responsible for the synthesis of AgNPs was identified, purified and characterized. Potential biomedical applications such as antibacterial, antifungal and anticancer activities of AgNPs have been evaluated. Irrespective of nature of pathogenic strains nanoparticles exhibited significant antibacterial activities against Gram positive (Streptococcus aureus) and Gram negative (Escherichia coli) bacterial pathogens. It showed higher activity on E. coli than on S. aureus. Distinct antifungal activity (MIC=5.244μg/ml) and remarkable anticancer activity (IC50=10μg/ml) was found against Candida albicans and HCT116 (colorectal carcinoma) cells, respectively. Taken together, these findings suggested that light induced plant generated silver nanoparticles could be used for various biomedical purposes.

Green Synthesized Nanoparticles as Potential Nanosensors

The recognition of chemical and biological entities is an important step in biomedical, forensic, and environmental sciences. The generation of extremely sensitive and cost-effective sensors needs sophisticated equipment coupled with basic understanding of chemistry, biology, and material science. Nanoparticles especially noble metal nanoparticles (gold, silver) have drawn great attention in this area due to their distinctive physicochemical properties and can be used in constructing innovative identification and transduction process for chemical and biological sensors. The use of toxic chemicals in traditional synthesis restricts their use in the real samples. Thus, to overcome the limitation, green synthetic pathways of nanoparticles have gained tremendous attention, which provide some important features including easy fabrication and high biocompatibility. Herein, potential applications of green synthesized nanoparticles such as silver nanoparticles, gold nanoparticles, and carbon dots have been described.

Cyclic peptide-based nanostructures as efficient siRNA carriers

Abstract RNA interference shows a great strategy for biological studies; however, delivering of small interfering RNA (siRNA) remains challenging. Although several delivery vehicles, including cell-penetrating peptides, have been developed, their implementation is often restricted because of their endosomal entrapment. Herein, we report the formation of self-assembled nanostructures from rationally designed cyclic peptides and explore them for efficient delivery of functional biomacromolecules such as siRNA into mammalian cells. The newly obtained soft materials make stable complexes with siRNAs, thereby increasing their stability and deliver fluorescent labelled siRNA inside the cells as evident from confocal microscopy analysis. Flow cytometry analysis reveals that significant uptake of FAM-siRNA occurs in the presence of peptide nanostructures compared with siRNA alone. Peptide nanostructure-mediated delivery of very low concentration of siRNA causes significant knockdown of the target gene as observed at protein level by Western blot analysis, which is comparable to lipofectamine, commercially available transfection agent.