Targeting G-quadruplex DNA with synthetic dendritic peptide: modulation of the proliferation of human cancer cells.

Soumi Biswas,Apurba K Das,Jishu Naskar,Satyabrata Samui,K Muniyappa,Sanjeev Pasadi

doi:10.1039/d0ra04780e

Soumi Biswas, Apurba K Das + Show 4 more

Open Access

https://doi.org/10.1039/d0ra04780e

Copy DOI

Abstract

Telomerase, a reverse transcriptase enzyme, is found to over express in most cancer cells. It elongates the telomere region by repeated adding of TTAGGG in the 3′-end and leads to excess cell proliferation which causes cancer. G-quadruplex (G4) formation can inhibit such telomere lengthening. So, stabilization of G4 structure as well as inhibition of telomerase activity is very promising approach in targeted cancer therapy. Herein, the aptitude of a synthetic dendritic peptide, Cδ2–(YEE)–E (peptide 1), to target specifically the human telomeric G4 DNA, dAGGG(TTAGGG)3, has been evaluated. Both biochemical and biophysical techniques including gel mobility shift assay, isothermal titration calorimetry and fluorescence spectroscopy have been employed for the purpose. Circular dichroism study reveals that the targeting results an increase in thermal stability of G4 DNA. Interestingly, replacement of N-terminal tyrosine residue of peptide 1 by valine, Cδ2–(VEE)–E, (peptide 2) consequences in loss of its G4 DNA targeting ability, although both the peptides exhibit comparable affinity toward double-stranded DNA. Of note, peptide 1 causes cessation of growth of human cancer cells (HeLa and U2OS) and induces apoptosis in vitro. But it has no significant inhibitory effect on the growth of normal human embryonic kidney 293 cells. Mechanistically, Telomeric Repeat Amplification Protocol (TRAP) assay indicates that peptide 1 effectively inhibits the telomerase activity in human cell extracts. Overall, this study demonstrates the usefulness of a synthetic dendritic peptide as an inhibitor of tumor cell growth by inducing apoptosis upon targeting the telomeric G4 DNA.

Highlights

The G-quadruplexes (G4) are non-canonical DNA/RNA structures that play important roles in DNA replication,1a recombination,1b transcriptional regulation,1c maintenance of genomic stability1d and aging.1e The G4 forming sequences occur throughout the human genome, but they are most prevalent in the telomeres,2a immunoglobulin switch regions2b and in promoters of proto-oncogenes.2c The telomerase activity is suppressed in most human somatic cells, except in stem cells and lymphocytes,3a but it becomes up-regulated in most tumor cells.3b The unfolded single-stranded DNA is required for optimal telomerase activity; whereas, G-quartet formation inhibits the telomerase activity.[4]
Peptide 2, Cd2–(VEE)–E, has been designed to address whether only hydrogen bonding interaction is enough to stabilize the G4 structure. Both the peptides have been synthesized by solution phase racemization free fragment condensation strategy and nal compounds have
We describe the interaction of two synthetic dendritic peptides with human telomeric G4 DNA

Summary

Introduction

Scheme 1 The schematic presentation of the synthesis of peptides. (a) DCC, HOBt, H-Glu(OMe)[2], (b) MeOH, NaOH (c) DCC, HOBt, H-Glu(OMe)[2] and (d) MeOH, NaOH followed by treatment with trifluoroacetic acid (TFA). We explore the ability of a synthetic dendritic peptide as ligand to stabilize the G4 DNA structure. The assay has been carried out by incubating 30 mM of G4 DNA with increasing concentrations (0, 15, 30, 45, 60 and 120 mM) of peptides for 30 min at room temperature and the DNA binding ability of the peptides is assessed by shi (retardation) in the electrophoretic migration of G4 DNA.

Results and discussions

Â 10À11

Conclusion