Background
Ovarian carcinoma remains the most mortality in gynecologic tumors [
1]. There are 225,000 new cases diagnosed and 140,000 deaths of ovarian carcinoma annually worldwide [
1]. The standard treatment remains surgery followed by platinum-based chemotherapy [
2]. Acquired drug resistance and cancer recurrence become the main hurdles for ovarian carcinoma treatment currently [
3]. As a result, new reagents targeting the chemo-resistant cells are needed.
Aldehyde dehydrogenases (ALDH) are a group of enzymes that catalyse dehydrogenation of aldehydes to their corresponding carboxylic acids. To date, nineteen ALDH genes which encode several isozymes have been identified in human genome. Aldehyde dehydrogenase 1 (ALDH1) gene encodes a cytosolic isoform localized in the cytoplasm and ALDH2 gene encodes a mitochondrial isoform located in mitochondrial matrix. Nevertheless, ALDH1 in human is not limited to the metabolic enzyme. It should be noted that ALDH1 is involved in regulating cell differentiation [
4,
6], proliferation and motility [
6,
7]. Its regulation role in stem cells is particularly through the retinoid signaling pathway [
8,
9]. It is also reported that inhibition of ALDH1-mediated retinoid signaling impairs human fetal islet cell differentiation and survival [
5]. ALDH1 may contribute to tumor initiation and chemoresistance [
10]. In addition, it is regarded as a cancer stem cell (CSC) marker in a variety of cancers [
11], including ovarian carcinoma [
12,
13], lung cancer [
14,
15], rectal cancer [
16] and others [
17,
18]. CSCs are a subpopulation of cancer cells which have the properties of self-renewal and tumorigenicity, and thus may play a key role in cancer metastasis, chemoresistance and relapse. Therapeutic modalities targeting CSCs are becoming a hot topic in recent years to prevent cancer relapse and vastly improve cancer survival probability [
19]. Targeting CSC specific markers is one of the most important and easily achievable ways to identify putative CSCs. ALDH1, as a largely used stem cell marker in recent CSC studies, is mostly regarded as a poor prognostic factor in a variety of cancers [
17,
18,
20,
21].
However, it remains debatable whether ALDH1 as a single marker can be sufficient to identify CSCs [
22]. Futhermore, different isoforms of ALDH1 may serve variable roles in CSCs [
23]. To date, the predictive role of ALDH1 in ovarian carcinoma cells and stromal cells are still obscure. While Chang et, al. found it was significantly associated with favorable clinical outcomes and better survivals in 442 cases of primary ovarian carcinoma patients [
24], others insisted that it is an unfavorable prognostic factor in ovarian carcinomas [
20,
21].
To further understand the prognositic role of ALDH1 in ovarian carcinoma cells and the stromal cells, we randomly enrolled 248 cases of primary ovarian carcinoma and investigated the expression of ALDH1 in these tissues by immunohistochemistry (IHC). The staining of ALDH1 in both carcinoma cells and stromal cells were evaluated and their associations with clinicalpathological parameters were analyzed by SPSS software.
Discussion
Ovarian carcinoma is actually one of the most chemosensitive solid malignancies [
31], but it is still the most lethal gynecologic malignancy worldwide. Although most ovarian carcinoma cells are initially sensitive to chemotherapy, there is always a small population cells that always survives and initiates new tumors which causes recurrence [
31]. The verification of tumor heterogeneity further enhances the hypothesis of CSCs. Compelling evidence has shown that ovarian carcinomas with enriched CSCs exhibit aggressive features
in vitro and predict poor outcomes in patients [
21,
32,
33].
Theoretically a high proportion of CSCs in tumor should be correlated with poor prognosis. However, the CSC markers are not always universal in a given tumor. The candidates raised up to characterize ovarian CSCs include CD44, epithelial cell adhesion molecule (EpCAM), CD133, CD117, CD90 (Thy-1), CD24, ABCG2, LY6A, AGR5 and ALDH1, etc. [
34,
35]. However, it still remains challenging to identify one single marker or several combined markers to specifically identify all the CSCs in ovarian tumor, and the exact roles of these ‘stemness’ related markers, are still poorly understood due to either a current lack of understanding of the biological functions of the markers, or frequently the lack of information correlating the varied isoforms, splicing variants or substrates to stem cell function [
34,
35].
ALDH1 is initially found as a cytosolic isozyme located in the cytoplasm, mainly functioning as the second enzyme after alcohol dehydrogenase in the major pathway of alcohol metabolism in liver. In recent years, soon after the rise of CSC theory, some specific CSC markers were being discovered to identify putative CSCs, and ALDH1 is becoming to act as a CSC marker in the CSC studies of variable tumor types
in vitro and
in vivo. Epithelial-to-mesenchymal transition (EMT) is an important driver of tumor invasion and metastasis, which may be a feature of CSC. Compared to ALDH1(−) EMT cells, only ALDH1(+) EMT cells had the ability to initiate a new epithelial tumor [
36]. Both EMT and other CSC properties of ALDH1(+) lung CSCs can be repressed through Fibulin3 treatment [
14]. Although ALDH1 was vastly studied as a CSC marker in other solid tumors, it has been identified as a CSC marker in ovarian carcinoma in recent years [
32]. In their study, dual positive cells of ALDH1 and another traditional ovarian CSC marker CD133 were isolated directly from human tumor to initiate tumor in mice, and these cells displayed enhanced angiogenesis and tumorigenicity like other CSCs [
32]. Moreover, the patients with CD133(+)/ALDH1(+) tumor cells displayed reduced PFS and OS [
32].
Distinct expression levels and patterns of ALDH1 in various human epithelial cancers and the corresponding normal tissues were explored [
16,
37]. Unlike breast, lung or colon cancers, ovarian cancer displayed a significantly reduced ALDH1 expression compared to benign tumors and normal ovary [
38], indicating a possibly different role of ALDH1 in ovarian cancer.
Our present immunohistochemical study of 248 well-characterized patients showed high levels of ALDH1 expression in ovarian carcinoma cells, which were observed in 15.7 % of the total cases, was associated with early-stage tumor, well-differentiated tumor and better survivals, although ALDH1 was not an independent risk factor in multivariate analysis. The patients involved in the current study were followed up for more than 12 years. To our best knowledge, it is a long follow up study, which has more predictive meanings in retrospective studies.
Our results were similar with a previous immunohistochemical study from The University of Texas MD Anderson Cancer Center. In their long-term follow-up study, The same mouse monoclonal anti-human ALDH Clone 44 from BD Transduction Laboratories™ was used, and it turned out that high level of ALDH1 was detected in 19 % out of the total 442 ovarian carcinoma samples, and it was significantly associated with endometrioid adenocarcinoma, early-stage tumor, complete response to chemotherapy, low serum CA125 level and favorable survivals [
24]. Similar results were obtained by immunofluorenscence-based and quantitative approach in Rimm’s group from Yale University School of Medicine, a better prognosis in ALDH1(+) patients with non-small cell lung cancer, using the same primary antibody with us [
39]. Thus, the choice of antibody may potentially explain the variably predictive and prognostic role of ALDH1 in human epithelial cancers. The current antibody we used has been proved specific and cytoplasmic with homogeneous staining pattern in different areas from the same case [
39]. In this study, the homogeneously strongly positive cases (with total score 7 and 8) showed a significantly better survival probability.
However, there were other studies indicating a better clinical outcome in ALDH1(+) cancer patients, using different antibodies. Our previous study, using rabbit polyclonal antibody against human ALDH1A1 (ab63026, Abcam, Cambridge, UK) has proved that ALDH1 expression in vulvar squamous cell carcinomas predicted a significantly better survival than the ALDH1 negative cases [
28]. Similarly, Hessman and coworkers, using antibody from Abcam, Cambridge, found that ALDH1 was highly expressed in early stage colorectal cancer in contrast with advanced stages [
16].
It should be noted that ALDH1 in normal stem cells has a function of activating cell differentiation through retinoid acid signaling pathway, and the inhibition of ALDH1-mediated retinoid signaling impairs human fetal islet cell differentiation and survival [
5]. It is also known that cancer stem cells share features of normal stem cells. Therefore, it can’t be excluded in the ovarian cancer cells that ALDH1 exerts its role through the same molecular mechanism, by such contributing to the better survival in ovarian cancers, although other unknown molecular mechanisms should be explored.
ALDH1 expression in stromal cells was previously reported to be frequently and strongly expressed in both non-malignant and tumor-associated stromal cells [
28,
40‐
42], which was confirmed in our present study. However, the potential role of ALDH1 expression in the tumor microenvironment is rather different in the above findings. Resetkova et al. hold the opinion that high expression of ALDH1 in breast cancer stromal cells had a best disease free survival and a trend of better overall survival [
42], De Brot et al. confirmed that ALDH1 frequent expression in tumor-associated stromal cells of triple negative breast cancer predicted a better outcome [
41]. On the other hand, Woodward and Ohi together with their colleages insisted that ALDH1 expression in stromal cells of breast cancer was not associated to any clinical outcomes [
43,
44]. In the present study, high expression level of ALDH1 in stromal cells was frequently observed in ovarian carcinoma, but the expression levels had no associations with the clinical parameters, and it is not associated with survival probabilities, which was in accordance with the findings from Woodward et al. [
43] and Ohi et al. [
44].
The current study has several limitations. First of all, although Allred scoring system combines the percentage and intensity of positive cells, as a manual scoring system, it may induce a level of subjectivity, especially the cut-off points were always a matter of discussion. Second, histological heterogeneity of ovarian cancers was not able to be addressed in the present study.
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Competing interests
The authors declare that they have no competing interests.
Authors’ contributions
Conceived and designed the study: RHuang, XL, RHolm, JMN, CGT, ZS. Performed the experiments: RHuang, XL. Evaluated the slides: RHuang, JMN, ZS. Analyzed the data: CGT. Wrote the manuscript: RHuang, XL, RHolm, JMN, CGT, ZS. All authors read and approved the final manuscript. All authors read and approved the final manuscript.