Modification of daunorubicin‐GnRH‐III bioconjugates with oligoethylene glycol derivatives to improve solubility and bioavailability for targeted cancer chemotherapy

Abstract

Daunorubicin-GnRH-III bioconjugates have recently been developed as drug delivery systems with potential applications in targeted cancer chemotherapy. In order to improve their biochemical properties, several strategies have been pursued: (1) incorporation of an enzymatic cleavable spacer between the anticancer drug and the peptide-based targeting moiety, (2) peptide modification by short chain fatty acids or (3) attachment of two anticancer drugs to the same GnRH-III derivative. Although these modifications led to more potent bioconjugates, a decrease in their solubility was observed. Here we report on the design, synthesis and biochemical characterization of daunorubicin-GnRH-III bioconjugates with increased solubility, which could be achieved by incorporating oligoethylene glycol-based spacers in their structure. First, we have evaluated the effect of an oligoethylene glycol-based spacer on the solubility, enzymatic stability/degradation, cellular uptake and in vitro cytostatic effect of a bioconjugate containing only one daunorubicin attached through a GFLG tetrapeptide spacer to the GnRH-III targeting moiety. Thereafter, more complex compounds containing two copies of daunorubicin, GFLG spacers as well as Lys(nBu) in position 4 of GnRH-III were synthesized and biochemically characterized. Our results indicated that all synthesized oligoethylene glycol-containing bioconjugates had higher solubility in cell culture medium than the unmodified analogs. They were degraded in the presence of rat liver lysosomal homogenate leading to the formation of small drug containing metabolites. In the case of bioconjugates containing two copies of daunorubicin, the incorporation of oligoethylene glycol-based spacers led to increased in vitro cytostatic effect on MCF-7 human breast cancer cells.

On the design principles of peptide–drug conjugates for targeted drug delivery to the malignant tumor site

“…The resulted PDC presented a reduced aqueous solubility, thus an oligoethylene glycol linker was inserted between the spacer and the tumor-homing peptide [ 154 ]. This PDC showed the best in vitro cytostatic effects among the oxime-linked Dau-containing conjugates, but the improvement of the synthetic process to lead to higher amounts of this PDC is required to proceed for in vivo studies.…”

Sequence modification of heptapeptide selected by phage display as homing device for HT-29 colon cancer cells to improve the anti-tumour activity of drug delivery systems

“…The description of the in vitro cytostatic effect study can be seen in [21], except for the incubation time with the conjugates that was in this case 24 h (followed by a washing step and 48 h additional incubation).…”

“…For determining cellular uptake of the bioconjugates by HT-29 cells, flow cytometry was used (details can be seen in [21]), except for the incubation time with the conjugates that was 3 h.…”

Enhanced In Vitro Antitumor Activity of GnRH-III-Daunorubicin Bioconjugates Influenced by Sequence Modification

“…In the past decade, a variety of GnRH-III-Dau conjugates containing an oxime bond have been developed in our laboratories, whereby Dau was attached to an incorporated aminooxyacetyl moiety (Aoa) and 8 Lys was mainly used as the conjugation site. To improve the antitumor activity and other biochemical properties of the compounds, many different strategies have been followed including the insertion of cathepsin B labile peptide linkers or oligoethylene glycol-based spacer as well as the replacement of 4 Ser by an acylated lysine or dimeric GnRH-III conjugates [29,33,34,35,36]. One of our most potent and efficient bioconjugates is GnRH-III-[ 4 Lys(Bu), 8 Lys(Dau=Aoa)] ( K2 ), in which the Ser in position four was replaced by a butyrylated lysine [34].…”