Mechanisms of chemoresistance in epithelial ovarian cancer: recent discoveries

Keywords::

• chemoresistance
• cisplatin
• miRNA
• ovarian cancer
• paclitaxel

Epithelial ovarian cancer (EOC), the most common form of ovarian cancer, has the highest mortality rate among all gynecological malignancies. One of the reasons that women with EOC have a low survival rate is due to resistance to chemotherapy. Currently, the most common chemotherapy used for patients with advanced EOC is a combination of a platinum agent, cisplatin or carboplatin, with a taxane, such as paclitaxel [1]. Cisplatin and carboplatin exert cytotoxic effects by inducing DNA damage, resulting in growth inhibition and ultimately cell death [2]. On the other hand, paclitaxel induces cell cycle arrest and apoptosis by stabilizing the microtubules through binding to β-tubulin [3]. Although the initial response rate to the chemotherapy is high, the majority of patients develop recurrent disease, which is often characterized by chemoresistance [4]. Therefore, understanding the mechanisms underlying chemoresistance is imperative for designing effective chemotherapies to treat EOC patients.

Several mechanisms have been suggested to be involved in ovarian cancer chemoresistance, including an increase in drug efflux and drug inactivation, defective growth factor signaling, altered DNA repair activity, and abnormal cell cycle and apoptosis control [5]. Emerging evidence suggests that miRNAs, a class of small noncoding RNAs that regulate gene expression, primarily at the post-transcriptional level [6], are implicated in drug resistance in EOC. It was recently reported that miRNAs are differentially expressed between chemosensitive and chemoresistant ovarian tumors and that several miRNAs can alter the sensitivity of EOC cells to chemotherapeutic agents. These discoveries raise the exciting possibility of using miRNAs to predict and/or to enhance patients’ response to chemotherapies.

miRNAs as potential markers of chemosensitivity

miRNAs have been suggested to serve as biomarkers for diseases, including ovarian cancer [7,8]. Recent miRNA-profiling studies suggest that miRNAs may also be used as biomarkers to predict the response of EOC patients to chemotherapies. Microarray analysis of miRNA expression profile in stage III serous papillary ovarian adenocarcinomas identified 52 miRNAs that were overexpressed and eight miRNAs that were underexpressed in recurrent tumors compared with primary tumors [9]. Interestingly, unsupervised hierarchical clustering of these samples revealed that the recurrent tumors are clustered with primary chemoresistant tumors [9]. The most strongly upregulated miRNA in recurrent tumors is miR-223. In another study involving a larger sample set, a number of miRNAs were also found to be differentially expressed between chemosensitive and chemoresistant tumors [10]. Some of the miRNAs identified to be upregulated in chemoresistant tumors, including miR-223, are consistent with the findings by Laios et al.[9]. However, there are still considerable differences in the miRNAs identified to be related to chemosensitivity in these two studies. For example, let-7i, which was found to be strongly upregulated in recurrent tumors [9], had a significantly lower expression level in chemoresistant patients [10]. Thus, more studies are needed to determine the ‘miRNA signature for chemosensitivity’. Specifically, larger sample sizes are required for profiling studies. Histological types, clinical stages and pathological grades of the tumors, as well as the age of the patients, should be taken into consideration when comparing chemosensitive and chemoresistant samples.

miRNAs are detectable in serum and therefore may be used as biomarkers for diseases [11], including ovarian cancer [12]. It would be interesting to determine whether serum miRNAs could also be used as biomarkers for chemosensitivity. Once miRNAs identified from serum and/or tumor samples of patients are confirmed to be prognostic indicators of patients’ response to chemotherapy, more effective personalized therapy for EOC patients can be designed.

miRNAs modulate the sensitivity of EOC cells to chemotherapeutic agents

Several recent studies have demonstrated that altered expression levels of certain miRNAs within EOC cells modulate the response of these cells to chemotherapeutic drugs. Let-7i enhanced [10], while miR-214 [13] and miR-376c (previously known as miR-368) reduced [14], the effect of cisplatin on inducing EOC cell death. Responses to microtubule-targeting agents, such as paclitaxel, vincristine and epothilone B, were strongly increased by overexpression of miR-200c [15]. On the other hand, miR-27a blocked the effect of paclitaxel [16]. These findings suggest that manipulation of miRNA levels within tumor cells may enhance the efficiency of chemotherapeutic agents. However, all studies reported to date have been carried out using established cell lines. It will be important to determine whether similar results can be obtained from primary cultures of EOC cells from patients who are resistant to cisplatin and/or paclitaxel treatments. The effect of chemotherapeutic agents on mice grafted with EOC cells stably transfected with a miRNA or an anti-miRNA would further validate the modulatory effect of miRNAs in drug sensitivity. In the future, it is possible to incorporate miRNAs into chemotherapies. Either miRNA mimics (for miRNAs that enhance chemosensitivity) or miRNA inhibitors (for miRNAs that inhibit responses to chemotherapeutic agents) can be used to enhance the sensitivity of tumor cells to the chemotherapeutic drugs and to reduce the toxicity of these drugs to normal cells. Many obstacles remain to be overcome, such as the development of methods to efficiently deliver miRNA mimics and inhibitors into tumor cells in vivo.

Mechanisms underlying the effects of miRNAs

miRNAs exert their effects on cellular processes via regulation of gene expression. Depending on the degree of complementarity between miRNAs and their target mRNAs, miRNAs can induce translational repression and/or mRNA cleavage to inhibit gene expression [17]. Differential expression of miRNAs between chemosensitive and chemoresistant tumors, therefore, is expected to cause changes in the expression levels of their target genes within the cells. Indeed, several studies have linked miRNAs with genes that are known to contribute to chemoresistance in EOC.

The multidrug-resistance gene, MDR1 (also known as ABCB1 and P-glycoprotein), is an ATP-dependent transporter that pumps cytotoxic drugs out of cells. Expression of MDR1 is associated with chemoresistance in EOC cells [18]. miR-27a induced drug resistance by inhibiting the expression of homeodomain-interacting protein kinase-2 and increasing the expression of MDR1 [16]. Based on bioinformatics prediction and published literature, the authors suggested that miR-27a may target homeodomain-interacting protein kinase-2, which in turn, increases MDR1.

Abnormal growth factor signaling has been linked to chemoresistance [19]. Two miRNAs, miR-376c and miR-214, have been reported to promote chemoresistance in EOC cells by targeting growth factor signaling pathways. In EOC cells, Nodal induces apoptosis by activating activin receptor-like kinase 7 (ALK7) [20,21]. MiR-376c inhibited ALK7 expression by binding to the 3´ untranslated region of ALK7 mRNA, resulting in the suppression of Nodal-induced apoptosis [14]. Forced overexpression of miR-376c reduced, whereas inhibition of endogeneous miR-376c increased, the effect of cisplatin on cell death. Furthermore, silencing of ALK7 mimicked, while overexpression of ALK7 reversed, the effect of miR-376c on chemosensitivity of EOC cells. These findings demonstrated that the Nodal–ALK7 signaling pathway is involved in cisplatin-induced cell death and that miR-376c promotes chemoresistance by suppressing the expression of ALK7 [14]. AKT, a downstream mediator of many growth factors, is known to exert prosurvival functions [22] and promote chemoresistance in EOC cells [23]. A mechanism by which miR-214 induces cisplatin resistance is via inhibition of PTEN expression and subsequent activation of AKT [13].

Paclitaxel exerts its cytotoxic effects by binding to β-tubulin [24]. Abnormal expression of class III β-tubulin (TUBB3) has been linked to primary chemoresistance and poor survival in EOC patients [25]. Interestingly, miR-200c directly targets TUBB3 and dramatically sensitizes EOC cells to paclitaxel and related compounds [26].

A miRNA is known to target many genes; therefore, manipulating a miRNA level within tumor cells will result in changes in the level of many proteins. Studies conducted to date on ovarian cancer cells have been mainly focused on the interaction between one miRNA and one target. Global protein-profiling studies using mass spectrometry-based methods will provide further insight into the network of proteins regulated by a particular miRNA.

Conclusion

In summary, although chemoresistance remains a major challenge for the management of EOC, recent discoveries regarding the mechanisms of chemoresistance, particularly the involvement of miRNAs, may provide new tools to design better chemotherapeutic strategies to fight this deadly disease. In the future, it may be possible to test a patient’s miRNA signature to design personalized therapy that includes both traditional chemotherapeutic agents and miRNA mimics and/or inhibitors.

Financial & competing interests disclosure

The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.

No writing assistance was utilized in the production of this manuscript.