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Corresponding author. Department of Traditional Chinese Medicine, Chengdu Military General Hospital. No. 270, Rongdu Avenue, Jinniu District, Chengdu, Sichuan, 610083, Sichuan Province, China
Aldehyde dehydrogenase 1 (ALDH1) has been identified as a putative cancer stem cell (CSC) marker in lung cancer. However, the clinicopathological and prognostic value of this protein in lung cancer patients remains controversial. Thus, we performed a systematic review and meta-analysis of studies assessing the clinical and prognostic significance of ALDH1 expression in lung cancer.
Methods
An identification and review of publications assessing clinical or prognostic significance of ALDH1 expression in lung cancer until September 1, 2014 was undertaken. A meta-analysis was performed to clarify the association between ALDH1 expression and clinical outcomes.
Results
A total of 14 publications met the criteria and comprised 1926 cases. Analysis of these data showed that ALDH1 expression was not significantly associated with the patient age (OR = 0.82, 95% confidence interval [CI]: 0.45–1.50, P = 0.52), tumour size (OR = 0.68, 95% CI: 0.22–2.06, P = 0.49), smoking status (OR = 1.37, 95% CI: 0.85–2.22, P = 0.19), or tumour grade (OR = 1.65, 95% CI: 0.83–3.26, P = 0.15). However, in the identified studies, ALDH1 expression was highly correlated with lymph node metastasis (OR = 1.97, 95% CI: 1.16–3.34, P = 0.01), tumour TNM staging (OR = 1.68, 95% CI 1.28–2.22, P = 0.0002), decreased overall survival (relative risk [RR]: 1.97,95% CI: 1.16–3.34, P =0.01) and decreased disease free survival (RR: 1.63, 95% CI: 1.01–2.64, P=0.05).
Conclusions
This meta-analysis shows ALDH1 expression in lung cancer is connected with decreased overall and disease free survival and thus marks a worse prognosis.
Although lung cancer treatments have rapidly developed in recent years, the overall prognosis of patients with lung cancer remains poor. Considerable evidence has supported the proposal for a model in which tumourigenesis is driven by cancer stem cells (CSC) that are derived from mutated adult stem cells. CSCs undergo self-renewal, recapitulate the phenotype of the tumour from which they were derived, develop into phenotypically diverse cancer cell populations, proliferate extensively, and drive both the continued expansion of malignant cells and chemotherapy resistance [
High expression of octamer-binding transcription factor 4A, prominin-1 and aldehyde dehydrogenase strongly indicates involvement in the initiation of lung adenocarcinoma resulting in shorter disease-free intervals.
European Journal of Cardio-Thoracic Surgery.2012; : ezs170
]. Therefore, the identification of CSCs has become an important issue, particularly in the context of potential therapeutic targeting.
The aldehyde dehydrogenase (ALDH) superfamily represents a diverse group of enzymes that metabolise and detoxify a wide variety of endogenous and exogenous aldehydes [
Genetic evidence that retinaldehyde dehydrogenase Raldh1 (Aldh1a1) functions downstream of alcohol dehydrogenase Adh1 in metabolism of retinol to retinoic acid.
Journal of Biological Chemistry.2003; 278: 36085-36090
]. RA signalling is linked to cellular differentiation during development and has an important function in the self-protection of stem cells throughout the lifespan of an organism [
Genetic evidence that retinaldehyde dehydrogenase Raldh1 (Aldh1a1) functions downstream of alcohol dehydrogenase Adh1 in metabolism of retinol to retinoic acid.
Journal of Biological Chemistry.2003; 278: 36085-36090
ALDH1 is important for normal development and homoeostasis in several organs and crucial during embryogenesis. It is an important detoxifying enzyme in the liver, also expressed in kidney, as well as haematopoeitic progenitor cells. ALDH1 is described to play a crucial role within normal differentiation of stem cells [
Identification of human aldehyde dehydrogenase 1 family member A1 as a novel CD8+ T-cell–defined tumor antigen in squamous cell carcinoma of the head and neck.
]. It has been one of the most frequently used biomarkers in CSC-related research; the use of ALDH originated from the isolation of ALDH+ CSCs from breast cancer [
]. In vitro experiments suggest that isolated lung cancer cells with high ALDH1 activity are associated with cancer stem cell characteristics, including capacities of proliferation, self-renewal and resistance to chemotherapy [
]. However, the correlations between ALDH1 and the clinicopothological features of lung cancer with their corresponding prognostic values remain controversial. Some researchers have concluded that ALDH1 expression is associated with favourable outcomes [
]. In the present study, we performed a systematic review and meta-analysis of the published literature to clarify the association of ALDH1 expression with clinicopathological features and the prognosis of lung cancer patients. The results may enable the prognostic stratification of lung cancer patients with adjuvant therapy while providing new insights into the potential cellular origin of lung cancer and its activated molecular pathways.
Materials and Methods
Search Strategy
The electronic databases of Pubmed, Embase, and Wanfang were searched for studies that investigated the association of clinicopathological parameters and prognosis with ALDH1 expression in lung cancer to be included in the present meta-analysis. Studies were examined, and an updated search was conducted on September 2014. The following search terms and combinations were used: “ALDH1” or “Aldehyde dehydrogenase 1,” as well as “lung neoplasms” or “lung cancer”. The citation lists from all the retrieved studies were used to identify other relevant publications. Review articles were also scanned to identify additional eligible studies. The title and abstract of each identified study were scanned to exclude any irrelevant publications. The remaining articles were reviewed to determine whether they contained information on the topic of interest.
Selection Criteria
Two of the authors carefully extracted information from all eligible publications independently and according to the inclusion criteria. Disagreements were resolved through consensus. The inclusion criteria were as follows: (1) articles dealing with the expression of ALDH1 and prognostic factors, overall survival (OS), or disease-free survival (DFS) relative to lung cancer; (2) articles containing sufficient data to enable the estimation of an odds ratio (OR) or a relative risk (RR) ratio of OS or DFS; (3) articles in English or Chinese; and (4) articles published as original research. Reviews, comments, duplicated studies, and articles that were not relevant to the present analysis were excluded. Studies with less than 50 patients and those with less than two years of follow-up were also excluded.
Data Extraction
The following information was extracted from the retrieved papers: author, publication year, country of the patient, time of collection, histological type, tumour pathological stage, number of patients, research technique used, the ages of the patients, and the choice of cutoff scores for the definition of positive staining or staining intensity. Two major groups were established on the basis of the objective. One group clarified the association between the expression of ALDH1 and clinicopathological parameters, including the ages of the patients, smoking status, tumour size, differentiation degree, tumour TNM stage, and lymph node status. Meanwhile, the other group investigated the association between the expression of ALDH1 and OS or DFS.
Statistical Analysis
The meta-analysis was performed as previously described. ORs with 95% CI were used to evaluate the association among stem cell markers, ALDH1, and the clinicopathological features for lung cancer, which included the ages of patients, smoking status, tumour size, differentiation degree, tumour TNM stage, and lymph node status. RR combined with the retrieved studies was used to assess the association of ALDH1 and OS or DFS. For the RRs that did not come directly from the published articles, the published data and figures from original papers were used to assess the RR according to the methods described by Parmar et al. [
]. The heterogeneity across the studies was evaluated using Q test and P values. ORs and RRs were calculated with the use of a random-effect model when the P value was less than 0.05. Otherwise, a fixed-effect model was used. The influence of individual studies on the summary effect estimate was displayed using the sensitivity analysis. In addition, funnel plots and the Egger's test were used to estimate possible publication bias [
]. Cochrane Review Manager version 5.2 (Cochrane Library, Oxford, UK) was used to calculate the ORs and RRs and their variations from each investigation.
Results
Description of Studies
A total of 363 articles were selected for the meta-analysis by browsing the databases PubMed, Embase, and Wanfang. Out of this total, 342 were excluded after the title and abstract were reviewed, and seven articles were excluded after the full publications were reviewed (Figure 1). The reasons for exclusion were: (a) studies were not associated with the topic of interest; (b) researchers of the article used neither histopathologic analysis nor close clinical and imaging follow-up for at least six months; (c) studies associated with other diseases (d); non-original articles; (e) data could not be extracted; and (f) repeated data from the same or similar population. Eventually, 14 publications met the criteria for the present analysis [
High expression of octamer-binding transcription factor 4A, prominin-1 and aldehyde dehydrogenase strongly indicates involvement in the initiation of lung adenocarcinoma resulting in shorter disease-free intervals.
European Journal of Cardio-Thoracic Surgery.2012; : ezs170
Expression of the potential cancer stem cell markers, CD133, CD44, ALDH1, and β-catenin, in primary lung adenocarcinoma – their prognostic significance.
]. The total number of patients was 1926, and each study had 50 to 282 patients. The main characteristics of the eligible studies are summarised in Table 1. A total of 13 articles dealt with clinicopathological factors. Moreover, the assessment of OS or DFS using Kaplan–Meier method was reported in 11 of these articles.
Figure 1Literature search strategy and selection of articles.
Correlation of ALDH1 Expression with Clinicopathological Parameters
The association between ALDH1 and several clinicopathological parameters is illustrated in Figure 2. ALDH1 expression had a high correlation with high tumour TNM stage (pooled OR = 1.68, 95% CI 1.28–2.22, P = 0.0002 fixed-effect) and lymph node metastasis (pooled OR = 1.97, 95% CI 1.16–3.34, P = 0.01 random-effect) (Figures 2A and 2B). However, ALDH1 expression was not associated with the patient age (pooled OR = 0.82, 95% CI 0.45–1.50, P = 0.52 fixed-effect) (Figure 2C), tumour size (pooled OR = 0.68, 95% CI 0.22–2.06, P = 0.49 random-effect) (Figure 2D), smoking status (pooled OR = 1.37, 95% CI 0.85–2.22, P = 0.19 random-effect) (Figure 2E), or tumour grade (pooled OR = 1.65, 95% CI 0.83–3.26, P = 0.15 random-effect) (Figure 2F).
Figure 2Forest plot depiction of ALDH1 expression and OR for clinical pathologic features. Clinicopathological parameters investigated are TMN classification (A), lymph node status (B), patient age (C), size of tumour (D), smoking status (E), tumour grade (F). OR with corresponding confidence intervals are shown.
With the use of the methods described above, the OS and/or DFS of 1620 patients in the 11 studies were analysed. The main results of this meta-analysis are shown in Figure 3. A five-year OS rate was extracted from six studies. The meta-analysis of the six studies for the prognostic value of ALDH1 expression showed that ALDH1 expression is associated with a poor OS. This result was obtained from the DerSimonian–Laird random-effect model with a value of 1.97 (95% CI: 1.16–3.34, P =0.01) (Figure. 3A). However, heterogeneity was found among the studies (I2 = 86%, Ph <0.0001).
Figure 3Analysis of ALDH1 expression and survival of lung cancer patients.
The meta-analysis of eight applicable studies showed that ALDH1 expression is associated with poor DFS (RR: 1.63, 95% CI: 1.01–2.64, P=0.05; Figure. 3B), even though the studies displayed heterogeneity (I2 = 80%, Ph <0.0001).
Table 2 shows the results of the subgroup meta-analyses. When grouped according to the ethnicity, the combined RRs of Asian studies in OS and DFS were 1.48 (1.16−1.90) and 2.08 (1.44–3.01), respectively, which shows that ALDH1 is an indicator of poor OS and DFS prognoses in Asian patients. The subgroup meta-analysis of studies with a cutoff > 10% staining showed that a high ALDH1 expression is associated with poor OS (RR, 4.06; 95% CI, 1.37-12.04) and DFS (RR, 1.73; 95% CI, 1.31–2.29) in lung cancer patients. When grouped according to the subtypes of the ALDH family, high ALDH1A1 expression is significantly associated with poor OS (RR, 2.33; 95% CI, 1.76–7.12) and DFS (RR, 2.20; 95% CI, 1.43-3.40).
Table 2Associations between ALDH1 expression and lung cancer prognosis grouped by selected factors.
A sensitivity analysis, in which one study was deleted at a time, was performed to gauge result stability. The results are shown in Figure 4. Both of the corresponding pooled RRs of OS and DFS did not significantly change, which suggests the robustness of the results.
Figure 4Sensitivity analysis of all the studies assessing OS (A) and DFS (B).
The funnel plots presented no evidence of publication bias in the studies of either outcome (Figure 5). No evidence for significant publication bias was found in OS (Egger's test, P = 0.851) and DFS (Egger's test, P = 0.458) studies.
Figure 5Begg's funnel plot estimated the publication bias of the included literature for OS (A) and DFS (B).
]. Many groups have investigated the relationship between ALDH1 expression and the clinicopathological features of lung cancer patients because of the important function of lung CSCs in tumourigenesis, development, and therapeutic outcomes [
]. However, discrepancies among the studies that attempted to assess the association necessitated a quantitative aggregation of the survival results. To the best of our knowledge, the present meta-analysis is the first to estimate the association between ALDH1 and lung cancer survival systematically.
The present results indicate that high ALDH1 expression is positively associated with tumour TNM stage and lymph node metastasis, as well as poor prognoses for patients with lung cancer. This trend suggests that ALDH1 can function as a prognostic factor for predicting the outcomes of lung cancer patients. Therefore, our data imply that elevated ALDH1 expression can contribute to lung cancer development and progression, and the detection of the ALDH1 aberrations may be useful for identifying poor prognoses in patients with lung cancer. ALDH1 may also provide a therapeutic target for the development of specific agents to eradicate lung CSCs and can potentially yield efficient therapeutic approaches for curing human lung cancer.
The ALDH1 subfamily comprises three isoforms (ALDH1A1, ALDH1A2, and ALDH1A3), which synthesise RA from the retina and are crucial regulators of the RA signalling pathway [
]. These enzymes have a high affinity for the oxidation of both all-trans- and 9-cis-retinal and regulate the self-renewal and differentiation of normal stem cells and CSCs [
]. Thus, considerable attention has been focussed on the relationship between the expression of this isoform and clinicopathological parameters, including the prognosis of lung cancer patients. In the subgroup analysis of the ALDH family members, a significant association was detected between the increased ALDH1a1 expression and poor OS and DFS of lung cancer patients. However, contrasting results were also reported. For instance, Kahlert et al. demonstrated that low ALDH1A1 expression is an independent prognostic marker for shortened DFS and OS in ductal adenocarcinoma of the pancreas [
]. Thus, more prospective studies are needed to draw a definite conclusion.
This meta-analysis has some limitations. First, the number of included studies, as well as the included lung cancer patients in each study, is relatively small. Thus, these factors might have reduced the power and accuracy of subcategory analysis. Second, the OS and DFS outcomes were based on individual unadjusted RRs. Thus, a more precise assessment should be adjusted using other prognostic factors. Third, no clear guidelines are available as regards the methods used for the evaluation of the levels of stem cell markers in lung cancer patients. Such evaluation differs among all the studies. In the assessment of biomarkers, the use of a standard threshold has great importance. Although immunohistochemistry was the most commonly applied method, differences in the cut-off values for the positive ALDH1 expression may have contributed to the observed heterogeneity. Subgroup analyses showed that the positive correlation between ALDH1 expression and poor OS and DFS could only be found in the cut-off > 10% subgroups. Thus, standardised methods and cut points that classify ALDH1 expression levels as “positive” or “negative” are urgently needed.
In summary, this study supports the CSC hypothesis by showing a significant correlation between stem cells and common clinical parameters, such as tumour TNM stage and lymph node metastasis. Based on the obtained data, ALDH1 expression has a significant association with poor survival. This result is consistent with findings in other cancers [
Aldehyde dehydrogenase 1/epidermal growth factor receptor coexpression is characteristic of a highly aggressive, poor-prognosis subgroup of high-grade serous ovarian carcinoma.
] and indicates that assessing ALDH1 expression could provide better prognostic information for patients with lung cancer and the development of more effective therapies for lung cancer requires effective targeting of this important cancer stem cell population. In addition, co-detection of ALDH1 with other CSC markers including CD133 and OCT4 may be more valuable and helpful in clinical application in lung cancer patients. However, further studies are required, with larger patient samples, unified methods and cut-off levels to detect ALDH1 expression, classified by tumour stage, therapeutic schedule, follow-up time and survival events, to confirm the findings of the present meta-analysis.
High expression of octamer-binding transcription factor 4A, prominin-1 and aldehyde dehydrogenase strongly indicates involvement in the initiation of lung adenocarcinoma resulting in shorter disease-free intervals.
European Journal of Cardio-Thoracic Surgery.2012; : ezs170
Genetic evidence that retinaldehyde dehydrogenase Raldh1 (Aldh1a1) functions downstream of alcohol dehydrogenase Adh1 in metabolism of retinol to retinoic acid.
Journal of Biological Chemistry.2003; 278: 36085-36090
Identification of human aldehyde dehydrogenase 1 family member A1 as a novel CD8+ T-cell–defined tumor antigen in squamous cell carcinoma of the head and neck.
Expression of the potential cancer stem cell markers, CD133, CD44, ALDH1, and β-catenin, in primary lung adenocarcinoma – their prognostic significance.
Aldehyde dehydrogenase 1/epidermal growth factor receptor coexpression is characteristic of a highly aggressive, poor-prognosis subgroup of high-grade serous ovarian carcinoma.
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