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The protective effect of cardiac gene transfer of CuZn–sod in comparison with the cardioprotector monohydroxyethylrutoside against doxorubicin-induced cardiotoxicity in cultured cells

Cancer Gene Therapy volume 10, pages 270–277 (2003)Cite this article

Abstract

Doxorubicin-induced cardiotoxicity is related to its production of free radicals that specifically affect heart tissue because of its low antioxidant status. Monohydroxyethylrutoside (monoHER), a potent antioxidant flavonoid, is under development as a protector against doxorubicin-induced cardiotoxicity. The overexpression of high levels of superoxide dismutase (sod) protects against free radical damage in transgenic mice. Seeking alternatives besides the few cardioprotectors that are presently under investigation, the aim of the present study was to investigate the protective effect of cardiac gene transfer of CuZn–sod compared with that of the presently most promising cardioprotector monoHER against doxorubicin-induced cardiotoxic effects on neonatal rat cardiac myocytes (NeRCaMs) in vitro. NeRCaMs were infected with different multiplicity of infections (MOIs) of adenovirus encoding CuZn–sod (AdCuZn–sod). A control infection with an adenovirus vector encoding a nonrelated protein was included. The overexpression of CuZn–sod was characterized within 3 days postinfection.

For doxorubicin treatment, NeRCaMs were divided into three groups. The first group was infected with AdCuZn–sod before treatment with doxorubicin (0–50 μM). The second and third groups were treated with doxorubicin (0–50 μM) alone and with 1 mM monoHER, respectively. The LDH release and survival of treated cells were measured 24 and 48 hours after doxorubicin treatment. The beating rate was followed during the 3 days after doxorubicin (0–100 μM) treatment.

At the third day after infection with an MOI of 25 plaque-forming unit (PFU) of AdCuZn–sod/cell, the activity of CuZn–sod significantly increased (five-fold, P=.029). Higher MOI produced cytopathic effects (CPEs).

Doxorubicin alone produced significant concentration- and time-dependent reduction in NeRCaMs beating rate and survival (P<.0005). Doxorubicin (≥50 μM)-treated cells ceased to beat after 24 hours. This cytotoxicity was associated with an increase in the LDH release from the treated cells (P<.0005).

The five-fold increase in the activity of CuZn–sod did not protect against any of the cytotoxic effects of doxorubicin on NeRCaMs. In contrast, monoHER (1 mM) protected against the lethal effects of doxorubicin on the survival, LDH release and the beating rate of NeRCaMs (P<.004) during 48 hours after doxorubicin treatment. Doxorubicin-treated (≤100 μM) cells continued beating for >72 hours in the presence of monoHER.

The present study showed the lack of adenoviral CuZn–sod gene-transfer to protect myocardiocytes against doxorubicin-induced toxicity and confirms the efficacy of monoHER cardioprotection. Thus, a gene-therapy strategy involving overexpression of CuZn–sod to protect against doxorubicin-induced cardiotoxicity is not feasible with the currently available adenovirus vectors.

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Abbreviations

MonoHER :

7-monohydroxyethylrutoside

sod :

superoxide dismutase

NeRCaMs :

neonatal rat cardiac myocytes

AdCuZn–sod :

adenovirus encoding human CuZn–sod

pfu :

plaque-forming unit

MOI :

multiplicity of infection

LDH :

lactate dehydrogenase

μM :

micromol

CPE :

cytopathic effects

MTT :

3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide

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Authors and Affiliations

  1. Department of Medical Oncology, Free University Medical Center, PO Box 7057, Amsterdam, 1007 MB, The Netherlands

    M A I Abou El Hassan, M Heijn & W J F van der Vijgh

  2. Department of Pharmacology and Toxicology, University of Maastricht, PO Box 616, Maastricht, 6200 MD, The Netherlands

    A Bast

  3. Department of Molecular Cell Biology, Leiden University Medical Center, PO Box 9503, Leiden, 2300 RA, The Netherlands

    M A I Abou El Hassan, M J W E Rabelink & R C Hoeben

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Correspondence to M A I Abou El Hassan.

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Abou El Hassan, M., Heijn, M., Rabelink, M. et al. The protective effect of cardiac gene transfer of CuZn–sod in comparison with the cardioprotector monohydroxyethylrutoside against doxorubicin-induced cardiotoxicity in cultured cells. Cancer Gene Ther 10, 270–277 (2003). https://doi.org/10.1038/sj.cgt.7700564

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