THESIS
2012
xiv, 102 p. : ill. (some col.) ; 30 cm
Abstract
Membrane lytic peptides are a novel class of anticancer agents which have the potential to overcome drug resistance. We aim to improve the selectivity, potency and pharmacokinetic profile of membrane lytic peptides in anticancer treatment and explore the underlying mechanism in this work....[
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Membrane lytic peptides are a novel class of anticancer agents which have the potential to overcome drug resistance. We aim to improve the selectivity, potency and pharmacokinetic profile of membrane lytic peptides in anticancer treatment and explore the underlying mechanism in this work.
The first part is focused on selectivity mediated by tumor-associated protease. An 18 residue membrane lytic peptide was cyclized with a linker cleavable by tumor overexpressed MT1-MMP. The activity of peptide was suppressed upon cyclization and could be recovered upon enzymatic digestion. And MT1-MMP-positive cells were preferentially killed by the peptide.
In the second part, we tried to improve the pharmacokinetic performance of lytic peptide with the strategy of polyvalency and polymeric therapeutics. A short amphipathic sequence (KW)
3 was conjugated to dextran in multiple copies to afford a polyvalent conjugate Dex-(KW)
3. The conjugate exhibited 500-fold potency enhancement compared to monomeric (KW)
3 in MTT assay. No hemolytic activity was detected. Thermodynamic study revealed stronger membrane binding and deeper membrane penetration of Dex-(KW)
3, which were attributed to the high local concentration of peptides on a nano-sized carrier.
We further investigated the mechanism of how the conjugate affects cancer cells in the third part of this work. The membrane integrity and metabolic activity of cells were examined at different concentrations of conjugate. Membrane damage occurred at high concentration range. The decrease of metabolic activity began at low concentration when the plasma membrane was still intact. A dual-function mechanism was proposed and proven with a polyelectrolyte complex between Dex-(KW)
3 and poly-γ-glutamic acid (PGA). The complexation neutralized the positive charge of Dex-(KW)
3 and efficiently suppressed the plasma membrane lysing activity. But it did not change the intracellular activity of conjugate, namely decreasing the cell metabolic activity and mitochondrial membrane potential. We hypothesized the complex Dex-(KW)3/PGA was internalized and dissociate in the acidic environment of endosome. And it was supported by thermodynamic study, in which weaker binding between conjugate and PGA at pH 5.5 than at pH 7.4 was found. The complexation inhibited the effect on plasma membrane while maintained the intracellular activity. The strategy separates one effect from the other thus allows better understanding and control of the novel conjugate.
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