In Silico and In Vitro Identification of P-Glycoprotein Inhibitors from a Library of 375 Phytochemicals

Abstract

Cancer therapy with clinically established anticancer drugs is frequently hampered by the development of drug resistance of tumors and severe side effects in normal organs and tissues. The demand for powerful, but less toxic, drugs is high. Phytochemicals represent an important reservoir for drug development and frequently exert less toxicity than synthetic drugs. Bioinformatics can accelerate and simplify the highly complex, time-consuming, and expensive drug development process. Here, we analyzed 375 phytochemicals using virtual screenings, molecular docking, and in silico toxicity predictions. Based on these in silico studies, six candidate compounds were further investigated in vitro. Resazurin assays were performed to determine the growth-inhibitory effects towards wild-type CCRF-CEM leukemia cells and their multidrug-resistant, P-glycoprotein (P-gp)-overexpressing subline, CEM/ADR5000. Flow cytometry was used to measure the potential to measure P-gp-mediated doxorubicin transport. Bidwillon A, neobavaisoflavone, coptisine, and z-guggulsterone all showed growth-inhibitory effects and moderate P-gp inhibition, whereas miltirone and chamazulene strongly inhibited tumor cell growth and strongly increased intracellular doxorubicin uptake. Bidwillon A and miltirone were selected for molecular docking to wildtype and mutated P-gp forms in closed and open conformations. The P-gp homology models harbored clinically relevant mutations, i.e., six single missense mutations (F336Y, A718C, Q725A, F728A, M949C, Y953C), three double mutations (Y310A-F728A; F343C-V982C; Y953A-F978A), or one quadruple mutation (Y307C-F728A-Y953A-F978A). The mutants did not show major differences in binding energies compared to wildtypes. Closed P-gp forms generally showed higher binding affinities than open ones. Closed conformations might stabilize the binding, thereby leading to higher binding affinities, while open conformations may favor the release of compounds into the extracellular space. In conclusion, this study described the capability of selected phytochemicals to overcome multidrug resistance.

Autoren

Julia Schäfer
Vincent Julius Klösgen
Ejlal A Omer
Onat Kadioglu
Armelle T Mbaveng
Victor Kuete
Andreas Hildebrandt
Thomas Efferth

DOI

10.3390/ijms241210240

eISSN

1422-0067

Ausgabe der Veröffentlichung

12

Zeitschrift

International Journal of Molecular Sciences

Sprache

en

Online publication date

2023

Paginierung

10240 - 10240

Status

Published online

Herausgeber

MDPI AG

Herausgeber URL

http://dx.doi.org/10.3390/ijms241210240

Datum der Datenerfassung

2023

Titel

In Silico and In Vitro Identification of P-Glycoprotein Inhibitors from a Library of 375 Phytochemicals

Ausgabe der Zeitschrift

24

Data source: Crossref

Abstract

Cancer therapy with clinically established anticancer drugs is frequently hampered by the development of drug resistance of tumors and severe side effects in normal organs and tissues. The demand for powerful, but less toxic, drugs is high. Phytochemicals represent an important reservoir for drug development and frequently exert less toxicity than synthetic drugs. Bioinformatics can accelerate and simplify the highly complex, time-consuming, and expensive drug development process. Here, we analyzed 375 phytochemicals using virtual screenings, molecular docking, and in silico toxicity predictions. Based on these in silico studies, six candidate compounds were further investigated in vitro. Resazurin assays were performed to determine the growth-inhibitory effects towards wild-type CCRF-CEM leukemia cells and their multidrug-resistant, P-glycoprotein (P-gp)-overexpressing subline, CEM/ADR5000. Flow cytometry was used to measure the potential to measure P-gp-mediated doxorubicin transport. Bidwillon A, neobavaisoflavone, coptisine, and z-guggulsterone all showed growth-inhibitory effects and moderate P-gp inhibition, whereas miltirone and chamazulene strongly inhibited tumor cell growth and strongly increased intracellular doxorubicin uptake. Bidwillon A and miltirone were selected for molecular docking to wildtype and mutated P-gp forms in closed and open conformations. The P-gp homology models harbored clinically relevant mutations, i.e., six single missense mutations (F336Y, A718C, Q725A, F728A, M949C, Y953C), three double mutations (Y310A-F728A; F343C-V982C; Y953A-F978A), or one quadruple mutation (Y307C-F728A-Y953A-F978A). The mutants did not show major differences in binding energies compared to wildtypes. Closed P-gp forms generally showed higher binding affinities than open ones. Closed conformations might stabilize the binding, thereby leading to higher binding affinities, while open conformations may favor the release of compounds into the extracellular space. In conclusion, this study described the capability of selected phytochemicals to overcome multidrug resistance.

Addresses

Department of Pharmaceutical Biology, Institute of Pharmaceutical and Biomedical Sciences, Johannes Gutenberg University, Staudinger Weg 5, 55128 Mainz, Germany.

Autoren

Julia Schäfer
Vincent Julius Klösgen
Ejlal A Omer
Onat Kadioglu
Armelle T Mbaveng
Victor Kuete
Andreas Hildebrandt
Thomas Efferth

DOI

10.3390/ijms241210240

eISSN

1422-0067

Externe Identifier

PubMed Identifier: 37373385
PubMed Central ID: PMC10299530

Funding acknowledgements

Alexander von Humboldt Foundation:
German Academic Exchange:
Marc Strobel, Frankfurt, Germany: private donation

Open access

true

ISSN

1422-0067

Ausgabe der Veröffentlichung

12

Zeitschrift

International journal of molecular sciences

Schlüsselwörter

Cell Line, Tumor
Humans
Neoplasms
Doxorubicin
Drug Resistance, Neoplasm
Molecular Docking Simulation
Phytochemicals
ATP Binding Cassette Transporter, Subfamily B

Sprache

eng

Medium

Electronic

Online publication date

2023

Open access status

Open Access

Paginierung

10240

Datum der Veröffentlichung

2023

Status

Published

Publisher licence

CC BY

Datum der Datenerfassung

2023

Titel

In Silico and In Vitro Identification of P-Glycoprotein Inhibitors from a Library of 375 Phytochemicals.

Sub types

research-article
Journal Article

Ausgabe der Zeitschrift

24

Files

https://www.mdpi.com/1422-0067/24/12/10240/pdf?version=1686914366 https://europepmc.org/articles/PMC10299530?pdf=render

Data source: Europe PubMed Central

Abstract

Cancer therapy with clinically established anticancer drugs is frequently hampered by the development of drug resistance of tumors and severe side effects in normal organs and tissues. The demand for powerful, but less toxic, drugs is high. Phytochemicals represent an important reservoir for drug development and frequently exert less toxicity than synthetic drugs. Bioinformatics can accelerate and simplify the highly complex, time-consuming, and expensive drug development process. Here, we analyzed 375 phytochemicals using virtual screenings, molecular docking, and in silico toxicity predictions. Based on these in silico studies, six candidate compounds were further investigated in vitro. Resazurin assays were performed to determine the growth-inhibitory effects towards wild-type CCRF-CEM leukemia cells and their multidrug-resistant, P-glycoprotein (P-gp)-overexpressing subline, CEM/ADR5000. Flow cytometry was used to measure the potential to measure P-gp-mediated doxorubicin transport. Bidwillon A, neobavaisoflavone, coptisine, and z-guggulsterone all showed growth-inhibitory effects and moderate P-gp inhibition, whereas miltirone and chamazulene strongly inhibited tumor cell growth and strongly increased intracellular doxorubicin uptake. Bidwillon A and miltirone were selected for molecular docking to wildtype and mutated P-gp forms in closed and open conformations. The P-gp homology models harbored clinically relevant mutations, i.e., six single missense mutations (F336Y, A718C, Q725A, F728A, M949C, Y953C), three double mutations (Y310A-F728A; F343C-V982C; Y953A-F978A), or one quadruple mutation (Y307C-F728A-Y953A-F978A). The mutants did not show major differences in binding energies compared to wildtypes. Closed P-gp forms generally showed higher binding affinities than open ones. Closed conformations might stabilize the binding, thereby leading to higher binding affinities, while open conformations may favor the release of compounds into the extracellular space. In conclusion, this study described the capability of selected phytochemicals to overcome multidrug resistance.

Date of acceptance

2023

Autoren

Julia Schäfer
Vincent Julius Klösgen
Ejlal A Omer
Onat Kadioglu
Armelle T Mbaveng
Victor Kuete
Andreas Hildebrandt
Thomas Efferth

Autoren-URL

https://www.ncbi.nlm.nih.gov/pubmed/37373385

DOI

10.3390/ijms241210240

eISSN

1422-0067

Externe Identifier

PubMed Central ID: PMC10299530

Funding acknowledgements

Marc Strobel, Frankfurt, Germany: private donation

Ausgabe der Veröffentlichung

12

Zeitschrift

Int J Mol Sci

Schlüsselwörter

cancer
chemotherapy
molecular docking
multidrug resistance
natural products
virtual drug screening
Humans
Molecular Docking Simulation
Drug Resistance, Neoplasm
Doxorubicin
Neoplasms
Phytochemicals
ATP Binding Cassette Transporter, Subfamily B
Cell Line, Tumor

Sprache

eng

Country

Switzerland

PII

ijms241210240

Datum der Veröffentlichung

2023

Status

Published online

Datum, an dem der Datensatz öffentlich gemacht wurde

2023

Titel

In Silico and In Vitro Identification of P-Glycoprotein Inhibitors from a Library of 375 Phytochemicals.

Sub types

Journal Article

Ausgabe der Zeitschrift

24

Data source: PubMed

In Silico and In Vitro Identification of P-Glycoprotein Inhibitors from a Library of 375 Phytochemicals

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