In vitro and in vivo reversal of cancer cell multidrug resistance by the semi-synthetic antibiotic tiamulin

doi:10.1016/S0006-2952(98)00229-9

Biochemical Pharmacology

Volume 56, Issue 9, 1 November 1998, Pages 1219-1228

https://doi.org/10.1016/S0006-2952(98)00229-9 Get rights and content

Abstract

A large number of multidrug resistance (MDR) modulators, termed chemosensitizers, have been identified from a variety of chemicals, but most have been proven to be clinically toxic. Low concentrations of the pleuromutilin-derived semi-synthetic antibiotic tiamulin (0.1 to 10 μM) sensitized the three highly resistant P-glycoprotein (Pgp)-overexpressing tumor cell lines P388 (murine lymphoid leukemia), AS30-D (rat hepatoma), CEM (human lymphoblastic leukemia), and the barely resistant AS30-D/S cell lines to several MDR-related anticancer drugs. Flow cytometric analysis showed that tiamulin significantly increased the intracellular accumulation of daunomycin. When compared to reference modulating agents such as verapamil and cyclosporin A, tiamulin proved to be 1.1 to 8.3 times more efficient in sensitizing the resistant cell lines. Moreover, when given i.p. (1.6 μg/mg body weight), tiamulin increased the survival rate of adriamycin-treated mice bearing the P388/ADR25 tumor line by 29%. In the presence of an anticancer drug, tiamulin inhibited both ATPase and drug transport activities of Pgp in plasma membranes from tumor cells. Tiamulin is thus a potent chemosensitizer that antagonizes the Pgp-mediated chemoresistance in many tumor cell lines expressing the MDR phenotype at different levels and displays no toxic effects on contractile tissues at active doses, therefore providing the promise for potential clinical applications.

Section snippets

Chemicals

Cell culture media (Dulbecco’s modified Eagle’s medium and RPMI-1640) were from BioWhittaker and fetal bovine serum was from Sigma. ADR, COL, daunomycin, propidium iodide, antibiotic, and antimycotic solution (containing 10,000 units/mL of penicillin, 10 mg/mL of streptomycin, and 25 μg/mL of amphotericin B), were from Sigma. VBL was a gift from Roger Bellon Laboratories. Tiamulin (BM cyclin) was purchased from Boehringer Mannheim. All drugs were prepared in water under sterile conditions and

Tiamulin

Tiamulin (Fig. 1) or [3aS-(3aα,4β,5α,6α,8β,9α,9αβ,10S)]-[(2-(diethylamino)ethyl)thio]acetic acid 6-ethenyldecahydro-5hydroxy-4,6,9,10-tetramethyl-1-oxo-3a,9-propano-3aH-cyclopentacycloocten-8-yl ester] (C₂₈H₄₇NO₄S, molecular weight 493.76) is a semi-synthetic derivative of the natural antibiotic pleuromutilin isolated from the Basidiomycete Pleurotus mutilus[13]. It is frequently used in veterinary medicine to treat Gram-positive and mycoplasma infections of farm animals and is known to have no

Discussion

The results reported in the present study strongly support the notion that tiamulin, a semi-synthetic antibiotic used against Gram positive and mycoplasma infections in farm animals, promotes anticancer drug accumulation and modulates Pgp activity in Pgp-expressing or Pgp-overexpressing multidrug-resistant cancer cells in vitro and in vivo.

Although a common chemical basis for the modulating action of all chemosensitizers does not seem to exist, several “rules” have been defined for the design

Acknowledgements

The authors wish to thank Annick Berne, Jacques Vigouroux, Isabelle Guilhot, Blandine Lermite, and Aurélien Menuet for their help in cell culture, and Christophe Tenin for scientific discussion. Vinblastine was a gift from Roger Bellon Laboratories. We also thank Dr. Laurence J. Miller at the Mayo Clinic for stimulating discussion and improving the English of the manuscript. This work was supported by grants from the Association pour la Recherche contre le Cancer (Grant 6756 to L.G.B.), from

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Cited by (25)

Natural alkaloids as P-gp inhibitors for multidrug resistance reversal in cancer
2017, European Journal of Medicinal Chemistry
Citation Excerpt :
Moreover, up on i.p. administration (1.6 mg/mg body weight), tiamulin increased the survival rate of DOX-treated mice bearing the P388/DOX25 tumor line by 29%. In the presence of an anticancer drug, tiamulin inhibited both ATPase (IC50 = 0.54 μM) and [3H]-VBL transport (IC50 = 6.2 μM) in plasma membranes from tumor cells [145]. N-Arachidonoyl-ethanolamide (104) an endogenous canabinoid, modulates the P-gp inhibition activity at moderate levels in HEK-293 cells.
The biggest challenge associated with cancer chemotherapy is the development of cross multi-drug resistance to almost all anti-cancer agents upon chronic treatment. The major contributing factor for this resistance is efflux of the drugs by the p-glycoprotein pump. Over the years, inhibitors of this pump have been discovered to administer them in combination with chemotherapeutic agents. The clinical failure of first and second generation P-gp inhibitors (such as verapamil and cyclosporine analogs) has led to the discovery of third generation potent P-gp inhibitors (tariquidar, zosuquidar, laniquidar). Most of these inhibitors are nitrogenous compounds and recently a natural alkaloid CBT-01^® (tetrandrine) has advanced to the clinical phase. CBT-01 demonstrated positive results in Phase-I study in combination with paclitaxel, which warranted conducting it's Phase II/III trial. Apart from this, there exist a large number of natural alkaloids possessing potent inhibition of P-gp efflux pump and other related pumps responsible for the development of resistance. Despite the extensive contribution of alkaloids in this area, has never been reviewed. The present review provides a comprehensive account on natural alkaloids possessing P-gp inhibition activity and their potential for multidrug resistance reversal in cancer.
Establishment and characterization of an MDCK cell line stably-transfected with chicken Abcb1 encoding P-glycoprotein
2016, Research in Veterinary Science
Citation Excerpt :
It is well known that tiamulin, a semi-synthetic pleuromutilin commonly used in the control of pulmonary infections in chicken, produces a lethal interaction with ionophoric drugs. Later it has been found that tiamulin is an inhibitor of P-gp in human and rodent cell lines (Baggetto et al., 1998), and salinomycin is a substrate of P-gp (Lagas et al., 2008). This has clarified the issue of the drug-drug interactions described between ionophores and pleuromutilins.
Chicken P-glycoprotein (chP-gp), encoded by Abcb1, determines the bioavailability because of its effect on pharmacokinetics of various drugs. However, comprehensive studies on chP-gp are still limited. In this study, the chicken full-length cDNA was first successfully cloned and then stably expressed in MDCK cell line. The open reading frame of chicken Abcb1 consists of 3864 nucleotides, encoding for a 1287-amino acid protein. Sequence alignments analysis showed that chicken P-gp had high identities with the homologues of turkey (95%), human (72%), pig (72%), rat (71%) and cattle (68%). The efflux ratio of rhodamine123 (Rho123, a human P-gp substrate) in chAbcb1 transfected MDCK cells was significantly higher than that in the wild type MDCK cell (6.24 vs 1.64, P < 0.05), suggesting a good transporting function of chicken P-gp overexpressed in the transfected cell. Importantly, MDCK-chAbcb1 cells, unlike Caco-2 cells, exhibited biphasic saturation kinetics in transporting Rho123. In conclusion, an MDCK cell line stably expressing chAbcb1 was successfully established, which could provide a new cell model to screen its substrates and inhibitors and study the drug-drug interaction medicated via chicken P-gp.
Modulating cancer multidrug resistance by sertraline in combination with a nanomedicine
2014, Cancer Letters
Citation Excerpt :
A combination treatment between DOX and sertraline slows down tumor growth in NAR xenograft mouse model (Fig. 3A) and increased the survival (Fig. 3B) by 1.5-fold. Bearing in mind the dose-related problems with previous-generation chemosensitizers, we deliberately set out to test, in vivo, a low sertraline dose: 2 mg/kg body/day compared (for example) to the ranges of 5 to 30, 2 to 8, and 200 to 300 mg/kg body/dose for OC144–093, XR9756 and MS-209 respectively [22,34–36]. For treatment of depression, sertraline is prescribed at the dose range of 50–100 mg/person/day [37].
Inherent and acquired multiple drug resistance (MDR) to chemotherapeutic drugs is a major obstacle in cancer treatment. The ATP Binding Cassettes (ABC) transporter super family that act as extrusion pumps such as P-glycoprotein and multidrug-resistance-associated-proteins have prominent roles in cancer MDR. One of the most efficient strategies to modulate this active drug efflux from the cells is to physically block the pump proteins and thus change the balance between drug influx and efflux toward an accumulation of drug inside the cell, which eventually cumulates into cell death. MDR modulators (also known as chemosensitizers) were found among drugs approved for non-cancer indications. Yet, toxicity, adverse effects, and poor solubility at doses required for MDR reversal prevent their clinical application. Previous reports have shown that drugs belonging to the selective serotonin reuptake inhibitors (SSRI) family, which are clinically used as antidepressants, can act as effective chemosensitizers both in vitro and in vivo in tumor bearing mouse models. Here, we set out to explore whether sertraline (Zoloft®), a molecule belonging to the SSRI family, can be used as an MDR modulator. Combining sertraline with another FDA approved drug, Doxil® (pegylated liposomal doxorubicin), is expected to enhance the effect of chemotherapy while potentially reducing adverse effects. Our findings reveal that sertraline acts as a pump modulator in cellular models of MDR. In addition, in an aggressive and highly resistant human ovarian xenograft mouse model the use of sertraline in combination with Doxil® generated substantial reduction in tumor progression, with extension of the median survival of tumor-bearing mice. Taken together, our results show that sertraline could act as a clinically relevant cancer MDR inhibitor. Moreover, combining two FDA approved drugs, DOXIL®, which favor the influx of chemotherapy inside the malignant cell with sertraline, which blocks the extrusion pumps, could readily be available for clinical translation in the battle against resistant tumors.
Overcoming of P-glycoprotein-mediated multidrug resistance of tumors in vivo by drug combinations
2014, Synergy
Inhibition of P-glycoprotein represents an attractive possibility to modulate resistance of cancer cells to anticancer drugs. One major strategy to overcome P-glycoprotein-mediated multidrug resistance (MDR) of tumors is to increase intracellular concentrations of anticancer drugs. This can be achieved by blocking of P-glycoprotein-mediated drug efflux using synthetic or natural small molecules or monoclonal antibodies, which bind to various parts of the efflux channel. Another possibility to increase intracellular drug concentrations can be reached by nanoparticles. A further major strategy to overcome MDR involves the downregulation of P-glycoprotein expression either by therapeutic nucleic acids (antisense oligodeoxynucleotides, RNA interference) or by small molecule inhibitors of signal transduction kinases that govern P-glycoprotein expression (e.g. PI3K/Akt inhibitors). In the present review, we give a critical overview on in vivo experiments using murine syngeneic and human xenograft tumors. Strengths and weaknesses of in vivo experiments are critically discussed to further explore and utilize the full potential of animal experiments for better determination of requirements for successful clinical trials on MDR reversal in the future.
Implications of ABC transporters on the disposition of typical veterinary medicinal products
2008, European Journal of Pharmacology
Citation Excerpt :
It is well known to produce clinically important and often lethal interactions with ionophoric drugs, particularly the coccidiostats monensin and salinomycin, resulting in an increased cardiomyotoxicity as a common sign of ionophore toxicity (Ratz et al., 1997; Szucs et al., 2004). Tiamulin has later been identified as an inhibitor of P-gp in human and rodent cell lines (Baggetto et al., 1998) and is thus a potential inhibitor of P-gp in farm animal species. The recent finding that salinomycin is a P-gp substrate, suggests that the drug–drug interactions described between ionophores and pleuromutilins are not only related to inhibition of biotransformation, but also to inhibition of P-gp.
The ATP-Binding Cassette (ABC) transporters ABCB1, ABCC2 and ABCG2 are efflux transporters that facilitate the excretion of drugs, contribute to the function of biological barriers and maintain low cytoplasmic substrate concentrations in cells. ABC transporters modulate drug absorption, distribution and elimination according to the level of expression in the intestine, liver, kidney, and at biological barriers such as the blood-brain barrier. Moreover individual transporters are known to convey multi-drug resistance to tumour cells. While these diverse functions have been described in laboratory animal studies and in humans, the available information is very limited in animal species that are typical veterinary patients. This brief review summarizes the available data on organ distribution and expression levels in animals, genetic defects in dogs resulting in a non-functional P-gp expression, and describes examples of kinetic investigations directed to assess the clinical relevance of species differences in ABC-transporter expression.
The medicinal chemistry of multidrug resistance (MDR) reversing drugs
2002, Farmaco
Multidrug resistance (MDR) is a kind of resistance of cancer cells to multiple classes of chemotherapic drugs that can be structurally and mechanistically unrelated. Classical MDR regards altered membrane transport that results in lower cell concentrations of cytotoxic drug and is related to the over expression of a variety of proteins that act as ATP-dependent extrusion pumps. P-glycoprotein (Pgp) and multidrug resistance protein (MRP1) are the most important and widely studied members of the family that belongs to the ABC superfamily of transporters. It is apparent that, besides their role in cancer cell resistance, these proteins have multiple physiological functions as well, since they are expressed also in many important non-tumoural tissues and are largely present in prokaryotic organisms. A number of drugs have been identified which are able to reverse the effects of Pgp, MRP1 and sister proteins, on multidrug resistance. The first MDR modulators discovered and studied in clinical trials were endowed with definite pharmacological actions so that the doses required to overcome MDR were associated with unacceptably high side effects. As a consequence, much attention has been focused on developing more potent and selective modulators with proper potency, selectivity and pharmacokinetics that can be used at lower doses. Several novel MDR reversing agents (also known as chemosensitisers) are currently undergoing clinical evaluation for the treatment of resistant tumours. This review is concerned with the medicinal chemistry of MDR reversers, with particular attention to the drugs that are presently in development.

View all citing articles on Scopus

^‡: Present address: Center for Basic Research in Digestive Diseases, Mayo Clinic, Rochester, MN 55905, U.S.A.

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Biochemical Pharmacology

Abstract

Section snippets

Chemicals

Tiamulin

Discussion

Acknowledgements

References (22)

A surface glycoprotein modulating drug permeability in Chinese hamster ovary cell mutants

Biochem Biophys Acta

Multidrug resistance and chemosensitizationTherapeutic implications for cancer chemotherapy

Adv Pharmacol

Restoration of daunomycin retention in multidrug-resistant P388 cells by submicromolar concentrations of SDZPSC 833, a nonimmunosuppressive cyclosporin derivative

Exp Cell Res

Formation and utilization of acetoin, an unusual product of pyruvate metabolism by Ehrlich and AS30-D tumor mitochondria

J Biol Chem

Efficient purification and reconstitution of P-glycoprotein for functional and structural studies

J Biol Chem

Reduction of I_CaL and I_to1 density in hypertrophied right ventricular cells by simulated high altitude in adult rats

J Mol Cell Cardiol

Chemical studies with pleuromutilin

J Biol Chem

Transepithelial transport of vinblastine by kidney-derived cells. Application of new kinetic model to estimate in situ K_m of the pump

Biochim Biophys Acta

Competitive interaction of cyclosporins with the Vinca alkaloid-binding site of P-glycoprotein in multidrug-resistant cells

J Biol Chem

Isolation and expression of a complementary DNA that confers multidrug resistance

Nature

Pharmacologic circumvention of multidrug resistance

Cytotechnology

Cited by (25)

Natural alkaloids as P-gp inhibitors for multidrug resistance reversal in cancer

Establishment and characterization of an MDCK cell line stably-transfected with chicken Abcb1 encoding P-glycoprotein

Modulating cancer multidrug resistance by sertraline in combination with a nanomedicine

Overcoming of P-glycoprotein-mediated multidrug resistance of tumors in vivo by drug combinations

Implications of ABC transporters on the disposition of typical veterinary medicinal products

The medicinal chemistry of multidrug resistance (MDR) reversing drugs

Chemotherapy and Metabolic InhibitorsIn vitro and in vivo reversal of cancer cell multidrug resistance by the semi-synthetic antibiotic tiamulin

Abstract

Section snippets

Chemicals

Tiamulin

Discussion

Acknowledgements

Biochem Biophys Acta

Adv Pharmacol

Exp Cell Res

J Biol Chem

J Biol Chem

J Mol Cell Cardiol

J Biol Chem

Biochim Biophys Acta

J Biol Chem

Isolation and expression of a complementary DNA that confers multidrug resistance

Nature

Pharmacologic circumvention of multidrug resistance

Cytotechnology

Chemotherapy and Metabolic Inhibitors
In vitro and in vivo reversal of cancer cell multidrug resistance by the semi-synthetic antibiotic tiamulin