Synthesis and in vitro biochemical evaluation of oxime bond-linked daunorubicin-GnRH-III conjugates developed for targeted drug delivery

TY - JOUR

T1 - Synthesis and in vitro biochemical evaluation of oxime bond-linked daunorubicin-GnRH-III conjugates developed for targeted drug delivery

AU - Schuster, Sabine

AU - Biri-Kovács, Beáta

AU - Szeder, Bálint

AU - Farkas, Viktor

AU - Buday, László

AU - Szabó, Zsuzsanna

AU - Halmos, Gábor

AU - Mező, Gábor

N1 - Funding Information: This project has received funding from the European Union's Horizon 2020 research and innovation program under the Marie Sklodowska-Curie grant agreement No 642004, from the Hungarian National Science Fund (OTKA K104045) and National Research, Development and Innovation Office (NKFIH K119552). The work is also supported by the GINOP-2.3.2-15-2016-00043 (G.H.) project. The project is co-financed by the European Union and the European Regional Development Fund. The authors would like to acknowledge the help of Szilvia Bősze and László Kőhidai in cell culture studies, János Gardi for his help in preparation of radioligand as well as Ildikó Szabó for providing triptorelin reagent. Publisher Copyright: © 2018 Schuster et al.; licensee Beilstein-Institut. Copyright: Copyright 2018 Elsevier B.V., All rights reserved.

PY - 2018/4/4

Y1 - 2018/4/4

N2 - Gonadotropin releasing hormone-III (GnRH-III), a native isoform of the human GnRH isolated from sea lamprey, specifically binds to GnRH receptors on cancer cells enabling its application as targeting moieties for anticancer drugs. Recently, we reported on the identification of a novel daunorubicin-GnRH-III conjugate (GnRH-III-[4Lys(Bu), 8Lys(Dau=Aoa)] with efficient in vitro and in vivo antitumor activity. To get a deeper insight into the mechanism of action of our lead compound, the cellular uptake was followed by confocal laser scanning microscopy. Hereby, the drug daunorubicin could be visualized in different subcellular compartments by following the localization of the drug in a time-dependent manner. Colocalization studies were carried out to prove the presence of the drug in lysosomes (early stage) and on its site of action (nuclei after 10 min). Additional flow cytometry studies demonstrated that the cellular uptake of the bioconjugate was inhibited in the presence of the competitive ligand triptorelin indicating a receptor-mediated pathway. For comparative purpose, six novel daunorubicin-GnRH-III bioconjugates have been synthesized and biochemically characterized in which 6Asp was replaced by D-Asp, D-Glu and D-Trp. In addition to the analysis of the in vitro cytostatic effect and cellular uptake, receptor binding studies with 125I-triptorelin as radiotracer and degradation of the GnRH-III conjugates in the presence of rat liver lysosomal homogenate have been performed. All derivatives showed high binding affinities to GnRH receptors and displayed in vitro cytostatic effects on HT-29 and MCF-7 cancer cells with IC50 values in a low micro-molar range. Moreover, we found that the release of the active drug metabolite and the cellular uptake of the bioconjugates were strongly affected by the amino acid exchange which in turn had an impact on the antitumor activity of the bioconjugates.

AB - Gonadotropin releasing hormone-III (GnRH-III), a native isoform of the human GnRH isolated from sea lamprey, specifically binds to GnRH receptors on cancer cells enabling its application as targeting moieties for anticancer drugs. Recently, we reported on the identification of a novel daunorubicin-GnRH-III conjugate (GnRH-III-[4Lys(Bu), 8Lys(Dau=Aoa)] with efficient in vitro and in vivo antitumor activity. To get a deeper insight into the mechanism of action of our lead compound, the cellular uptake was followed by confocal laser scanning microscopy. Hereby, the drug daunorubicin could be visualized in different subcellular compartments by following the localization of the drug in a time-dependent manner. Colocalization studies were carried out to prove the presence of the drug in lysosomes (early stage) and on its site of action (nuclei after 10 min). Additional flow cytometry studies demonstrated that the cellular uptake of the bioconjugate was inhibited in the presence of the competitive ligand triptorelin indicating a receptor-mediated pathway. For comparative purpose, six novel daunorubicin-GnRH-III bioconjugates have been synthesized and biochemically characterized in which 6Asp was replaced by D-Asp, D-Glu and D-Trp. In addition to the analysis of the in vitro cytostatic effect and cellular uptake, receptor binding studies with 125I-triptorelin as radiotracer and degradation of the GnRH-III conjugates in the presence of rat liver lysosomal homogenate have been performed. All derivatives showed high binding affinities to GnRH receptors and displayed in vitro cytostatic effects on HT-29 and MCF-7 cancer cells with IC50 values in a low micro-molar range. Moreover, we found that the release of the active drug metabolite and the cellular uptake of the bioconjugates were strongly affected by the amino acid exchange which in turn had an impact on the antitumor activity of the bioconjugates.

KW - Cytostatic effect

KW - Daunorubicin

KW - Drug-targeting

KW - GnRH derivatives

KW - Oxime linkage

UR - https://www.scopus.com/pages/publications/85045194815

U2 - 10.3762/bjoc.14.64

DO - 10.3762/bjoc.14.64

M3 - Article

AN - SCOPUS:85045194815

SN - 1860-5397

VL - 14

SP - 756

EP - 771

JO - Beilstein Journal of Organic Chemistry

JF - Beilstein Journal of Organic Chemistry

ER -