Hypothesizing that histone deacetylase inhibitors can be used to reverse multiple drug resistance

Medical Hypotheses 74 (2010) 92–94

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Medical Hypotheses journal homepage: www.elsevier.com/locate/mehy

Hypothesizing that histone deacetylase inhibitors can be used to reverse multiple drug resistance Zhi-Ping Jiang a, Ping Xu b, Guan-Ping Wang a, Xie-Lan Zhao a, Fang-Ping Chen a,* a

Laboratory of Clinical Pharmacology, Department of Hematology, Xiang-Ya Hospital, Central-South University, 87, Xiang-Ya Road, Changsha 410008, People’s Republic of China Clinical Pharmacy and Pharmacology Research Institute, the Second Xiangya Hospital, Central-South University, Renmin Middle Road 139, Changsha, Hunan Province 410011, People’s Republic of China b

a r t i c l e

i n f o

Article history: Received 20 July 2009 Accepted 22 July 2009

s u m m a r y It is well known that the mechanism of action of chemotherapeutic drugs and their ability to induce multidrug resistance (MDR) are of relevance to cancer treatment. Although MDR is a multifactorial process, the main obstacle is the expression of multidrug-efflux pumps that lowers the intracellular drug levels. P-glycoprotein (P-gp) is the longest identified efflux pump. Thus, P-gp has been looked as a well established mediator of MDR and it became a therapeutic target for circumventing multidrug resistance. However, the mechanism of adjusting the expression of P-gp is not clear yet. The results of the effect of genetic polymorphism on P-gp expression and function remain conflicting. More recently, studies on the regulation of MDR1 has widened to examine the role of epigenetics and some new results were found to support the effect of epigenetic variance in vitro. It is hence hypothesized that epigenetic variants play more important roles than genetic polymorphism, thus adjusting the epigenetic factors could alter the expression of MDR, leading to the reverse of MDR. And it is further hypothesized that histone deacetylase inhibitors could be another strategy to overcome MDR. The mechanism may include a bidirectional modulation of P-gp by histone deacetylase inhibitors. Ó 2009 Elsevier Ltd. All rights reserved.

Background It is well known that the mechanism of action of chemotherapeutic drugs and their ability to induce multidrug resistance (MDR) are of relevance to cancer treatment. The most striking phenotypic marker of MDR is the overexpression of P-glycoprotein (P-gp), a plasma membrane pump associated with multidrug resistance. P-gp can severely limit the efficacy of anti-cancer agents by acting as an efflux pump [1]. Thus, P-gp has been looked as a well established mediator of MDR and it became a therapeutic target for circumventing multidrug resistance [2]. The mechanism of adjusting the expression of P-gp also became a hot field of research. P-gp is encoded by the MDR1 gene. Although a number of nucleotide variants have been described in the MDR1 gene, most studies in this area have focused on the association of single nucleotide polymorphisms (SNPs) in the coding region with the altered expression of P-glycoprotein or pharmacokinetics of clinically used drugs [3]. Since Hoffmeyer et al. performed the first systematic screening of SNPs in MDR1 gene and reported that only C3435T in exon 26 was shown to correlate with decreased enterocyte P-gp expression, lower activity of P-gp and maximum concentra* Corresponding author. Tel.: +86 731 84327214; fax: +86 731 84327332. E-mail address: [email protected] (F.-P. Chen). 0306-9877/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved. doi:10.1016/j.mehy.2009.07.036

tion of digoxin in Caucasians [4], many experiments have been performed and up to 50 SNPs of MDR1 gene have been found [5]. However, the studies on the impact of SNPs on drug metabolism showed conflicting results [6]. The reason is unclear yet. And it is interesting that the synonymous SNP at exon 26 (C3435T) has been considered the most possible variant to be associated with altered protein expression and drug metabolism, although the SNP does not change the encoded amino acid. The mechanism behind this interesting phenomenon is also unknown. A possible explanation relied on linkage disequilibrium between C3435T polymorphism and other SNPs. It was demonstrated that there was linkage disequilibrium between C3435T polymorphism in exon 26 and G2677T/A in exon 21, which seemed in part support the ‘‘linkage disequilibrium” hypothesis [7]. However, in many cases, the missense polymorphisms alone did not affect Pglycoprotein expression or function. Thus, this hypothesis does not explain the observed alterations in P-glycoprotein. More recently, studies on the regulation of MDR1 has widened to examine the role of epigenetics; that is the role of DNA methylation, histone deacetylation and chromatin structure on the transcriptional regulation of MDR1. The results from in vitro studies demonstrated that methylation of MDR1 promotor region was linked with a lack of P-gp expression in several cell lines, as well as patient samples [8–13]. The research by Baker et al.

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demonstrated that chemotherapeutic drugs can actively induce epigenetic changes within the MDR1 promoter, and enhance the MDR phenotype [14]. Moreover, our recent investigation demonstrated a correlation between MDR1 genetic polymorphism C3435T and G2677T/A with methylation status of MDR1 promoter region [15]. Thus mechanism other than amino acid change could play a role in correlation between MDR1 genotype and phenotype. However, some research results support the important role of histone acetylation, instead of methylation status in P-gp expression and function. Takane et al. found no clear association between methylation status at CpG Sites in the MDR1 promoter region with MDR1 mRNA expression among human placental samples [16]. The experiments by Xiao et al. revealed a significant role of histone acetylation, instead of methylation status, in MDR1 transcription, which seems to mediate FK228 resistance [17]. Chen’s paper provided evidence for a major genomic alteration that changes the chromatin structure of the ABCB1 upstream promoter via acetylation of histone H3 initiating ABCB1 activation, further elucidating the genetic and epigenetic bases that determine chemotherapeutic response in drug-resistant derivatives of MES-SA cells [18]. The hypothesis We propose our hypothesis that epigenetic variants play more important roles than genetic polymorphism, thus adjusting the epigenetic factors could alter the expression of MDR, leading to the reverse of MDR. Since it was found that methylation status is inversely correlated with P-gp expression, while promotor methylation may lead to tumor suppressor silencing, adjusting methylation status could not be practical. Thus histone deacetylase inhibitors could be another strategy to overcome MDR. Evaluation of the hypothesis Our hypothesis is supported, at least partly, by the results of in vitro studies as follows. Ruefli et al. found SAHA, a histone deacetylase inhibitor, induced equivalent death in P-gp-positive cells compared with P-gp-negative cells. This data demonstrated that SAHA may be of value for the treatment of P-gp-expressing MDR cancers [19]. Furthermore, El-Khoury examined the effects of the histone deacetylase inhibitor trichostatin A (TSA) on MDR1 gene expression in small cell lung carcinoma (SCLC) drug-sensitive (H69WT) or etoposide-resistant (H69VP) cells. It was also found in this study that TSA induced an increase in P-gp expression in drug-sensitive cells, but strongly decreased it in drug-resistant cells. The up- and down regulations occurred at the transcriptional level. And the P-gp regulations were not effected by the methylation status of the promoter 50GC, 110GC, and Inr sites, and did not result in further changes to these methylation profiles. The results suggested that in H69 drug-resistant SCLC cell line TSA induces downregulation of P-gp expression through a transcriptional mechanism, independently of promoter methylation [20]. If genetic polymorphism does not play an important role in MDR1 expression, we have to face this interesting question that there is no rational explanation for why silent SNPs might have such effects, especially when no change in P-glycoprotein mRNA and protein expression levels has been observed. It was demonstrated by Kimchi-Sarfaty et al. that silent polymorphisms (in particular, C3435T) in MDR1 can alter P-glycoprotein conformation and protein substrate specificity by affecting the timing of cotranslational folding and insertion of P-gp into the membrane, thereby altering the structure of substrate and inhibitor interaction sites. This study is of immense importance since it demonstrates for the first time that naturally occurring silent SNPs can lead to the

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synthesis of protein product with the same amino acid sequence but different structural and functional properties. This results also support that factors other than amino acid sequence change can visibly affect the phenotype of certain gene [21]. And it should be noticed that some histone deacetylase inhibitors are also a substrate of P-gp. The study by Robey et al. suggested that depsipeptide, a kind of histone deacetylase inhibitor also substrate of P-gp, induced its own mechanism of resistance by up-regulation of the MDR1 gene. This result would mean that depsipeptide induces its own mechanism of resistance and thus provided a basis for clinical trials evaluating the action of depsipeptide, in combination with a P-gp inhibitor in vivo [22]. The research of Yu et al. indicated that TSA was a substrate and reversing agent for P-gp; and P-gp-mediated efflux of TSA into the gut lumen and the first-pass metabolism contributed to the low oral bioavailability [23]. Further studies are needed to explore the effect of induction of its own resistance on the pharmacokinetics and pharmacodynamics of histone deacetylase inhibitors in vivo. Discussion In the last decades, multidrug resistance (MDR) was looked as one of the main problems in the anti-cancer therapy with cytostatically active agents. Although MDR is a multifactorial process, the main obstacle is the expression of multidrug-efflux pumps that lowers the intracellular drug levels. P-glycoprotein (P-gp) is the longest identified efflux pump. Although several other mechanisms for MDR are elucidated in recent years, considerable efforts attempting to inverse MDR are involved in exploring P-glycoprotein modulators and suppressing P-glycoprotein expression. A potential strategy is to co-administer efflux pump inhibitors, although such reversal agents might actually increase the side effects of chemotherapy by blocking physiological anti-cancer drug efflux from normal cells [24]. Although many efforts to overcome MDR have been made by using first and second generation reversal agents comprising drugs already in current clinical use for other indications (e.g. verapamil, cyclosporine A, quinidine) or analogues of the first-generation drugs (e.g. dexverapamil, valspodar, cinchonine), few significant advances have been made. Clinical trials with third generation modulators (e.g. biricodar, zosuquidar, and laniquidar) specifically developed for MDR reversal are ongoing. The results however are not encouraging and it may be that the perfect reverser does not exist [2]. Our hypothesis provides a new consideration of strategy to overcome MDR. It should be noticed that P-gp is also involved in normal physiologic functions thus inhibiting it could lead to varieties of side effects and toxicity [25]. However, TSA expressed bidirectional modulation of P-gp. That is, TSA induced an increase in P-gp expression in drug-sensitive cells, but strongly decreased it in drug-resistant cells [20]. Thus, if histone deacetylase inhibitor like TSA or its analogues would be used as efflux pump modulators, the dilemma that MDR reverse agents enhance the effect of chemotherapy while increasing the toxicity of chemotherapy could be at least partly overcomed. However, there were only the results of adjust P-gp expression by using histone deacetylase inhibitors in vitro studies, further studies are needed to investigate the effect of modulation of P-gp by histone deacetylase inhibitors in vivo. Furthermore, the mechanism of bidirectional effects of histone deacetylase inhibitor should also be clarified. Conflicts of interest statement We declare that we have no financial and personal relationships with other people or organizations that can inappropriately influence our work, there is no professional or other personal inter-

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est of any nature or kind in any product, service and/or company that could be construed as influencing the position presented in, or the review of, the manuscript entitled.

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