We report an association between serum estrogen, tumor ER-α mRNA expression levels, estrogen-related SNPs and lung cancer survival in independent cohorts. Serum estrogen and estrogen-related SNPS were analyzed in three cohorts, whereas tumor ER-α and ER-β gene expression was analyzed in the two cohorts with available RNA. Higher serum estrogen levels, as well as tumor ER-α mRNA positive status, were associated with worse lung cancer prognosis. We also demonstrated that specific estrogen-related SNPs, which were associated with either higher serum estrogen levels or tumor ER-α mRNA levels, were also associated with poorer lung cancer prognosis. To our knowledge, this is the first study to demonstrate a significant association between serum estrogen and poorer lung cancer survival and to support functionality of estrogen-related SNPs in lung cancer prognosis.
The associations between serum estrogen and tumor ER-α mRNA expression and lung cancer survival were independent of gender and race. One previous study demonstrated that tumor ER expression in the lung was dependent on gender (14
); however, our studies were in accordance with another study reporting no difference in tumor ER expression between men and women (13
). Our data support the notion that the association we observe between serum estrogen levels and lung cancer survival is not due to estrogen therapy. First, we observe this association not only among females but also among males. Second, the removal of women who were taking estrogen therapy in each study did not alter our findings. Our studies also agree with previous research demonstrating an association between tumor ER-α expression and poor survival (13
). We also observed a stronger association of tumor ER-α expression with lung cancer survival among adenocarcinomas in the NCI-MD case–case cohort (supplementary Table VII
is available at Carcinogenesis
Online). Finally, much of the previous research examining the association between tumor ER-α expression and lung cancer focused on tumor ER-α protein expression measured by immunohistochemistry. Our studies are complementary with these past studies, because we examined ER-α mRNA levels via quantitative reverse transcription polymerase chain reaction, which is more quantitative and less subjective. Our studies also agree with a recent study in which ER-α and ER-β mRNA examined via quantitative reverse transcription polymerase chain reaction were higher in lung carcinoma cells compared with normal epithelium (30
). Although ER gene amplification is common in breast cancer (31
), there are no studies examining whether the same phenomenon exists in lung cancer.
Our study demonstrated a significant positive association between serum estrogen levels and lung cancer survival, independent of histological subtype (data not shown). Although some researchers have suggested that nicotine may have an anti-estrogenic effect (34
), neither serum estrogen nor ER-α levels differed by smoking status or pack-years (data not shown). In animal models, estrogen increases lung tumor growth (6
). Taioli and Wynder (35
) suggested that estrogen plays a role in the development of lung cancer among women, similar to its effect on breast cancer development, which is supported by our results. Their study demonstrated that an early age at menopause was associated with a significant decrease in risk of lung adenocarcinoma, whereas use of estrogen replacement therapy was associated with an increased risk (35
). Another study using Surveillance, Epidemiology and End Results data supported the detrimental effect of estrogen on lung cancer through its observation that premenopausal women were more probably to have poorly differentiated tumors and to present at a more advanced stage than postmenopausal women (36
). Other studies also observed associations between lung cancer risk and earlier age of menarche (37
), greater number of menstrual cycles (38
), higher parity (39
) and earlier age at menopause (35
). Adami et al.
) observed that women who took estrogen replacement therapy had a higher risk of developing lung cancer, however, they did not report on survival. In another study, median lung cancer survival was shorter in patients taking hormone replacement therapy (42
). Other studies also observed a higher risk of lung cancer among women taking hormone replacement therapy, although many findings were not significant (43
). Recently, data have demonstrated that women on hormone replacement therapy of combined estrogen and progesterone for >5 years have a higher risk of lung cancer risk and mortality (46
). We observed an association of serum progesterone with lung cancer survival among men in the NCI-MD case–case cohort; however, this was not validated by the other two cohorts. Furthermore, there was no association with serum progesterone and lung cancer survival among the women in any of the cohorts.
To our knowledge, this is the first study to report associations between estrogen-related SNPs and lung cancer prognosis. Specifically, the variant genotype (A/A) of the ESR1-07
SNP was associated with higher tumor ER-α mRNA levels and poorer lung cancer prognosis. This SNP is present in exon 8 of the ER-α
gene, which translates to the F domain of the ER-α protein (48
). The significance of this domain has been widely researched although its exact function is still unknown. The F domain is often combined with the E domain when investigating function, therefore, this E/F domain contains the ligand-binding domain, the dimerization domain and a second activation function (AF2) (20
). Transcriptional activity of ER-α is mediated, in part, by the AF2 domain and appears to be hormone dependent (20
). Our study is consistent with the above observations because we observe an association between the A/A variant genotype of ESR1-07
and increased tumor PR mRNA, an estrogen-regulated gene (52
), suggesting that the variant genotypes (G/A or A/A) of ESR1-07
has higher transcriptional activity than the referent genotype (G/G). It is possible that the variant genotype of ESR1-07
results in a longer half-life of the ER protein and in turn, higher levels of tumor PR mRNA. This increase in ER levels, along with an increase in transcriptional activity, may help explain why patients who are tumor ER-α positive have a poorer prognosis in our study.
Aromatase inhibitors have been shown to inhibit lung cancer epithelial cell proliferation in vitro
, lung tumor growth in vitro
and lung tumor xenograft growth in nude mice, supporting their role in lung carcinogenesis (7
). Our study illustrates that the variant genotype of an SNP in the aromatase gene, CYP19a1-08
, is associated with decreased levels of serum estrogen and a better prognosis among lung cancer patients. Previous research demonstrated that another SNP in the aromatase gene, CYP19a1-09
, which is similar to CYP19a1-08
, is also located in the 3′ untranslated region and is associated with lower serum estrogen levels in postmenopausal women (53
). We confirmed this result in our study. The variant genotype of CYP19a1-08
may impair aromatase function and thus reduce serum estrogens, which result in better lung cancer survival.
There are some limitations to this study. First, we do not have serum hormone, hormone receptor or SNP data on all the patients in the NCI-MD case–case cohort. This study began recruitment in 1984; therefore, many of the samples were depleted before our analyses. However, comparison of race, age, gender, smoking status and pack-years illustrated no significant difference between the study participants that underwent serum hormone, hormone receptor or SNP analysis and those that did not. Significantly, we were able to confirm the results of the test cohort with two independent validation cohorts. Second, although lung cancer diagnosis typically occurs at a later age and our study results are independent of gender, it is important to confirm whether these associations are present in premenopausal women. It is possible that premenopausal women may have a poorer survival than postmenopausal women due to the effect of serum estrogen. The one study that has examined this comparison did not report a difference in survival (36
), however, in this previous study, menopausal status was determined solely by age with women <50 categorized as premenopausal and therefore, the potential for misclassification was high. The associations between an increased lung cancer risk and earlier age at menarche, later age at menopause, increased number of menstrual cycles, and lower parity suggest that estrogen plays a role in lung carcinogenesis (35
In conclusion, we observed that serum estrogen and tumor ER-α levels were inversely associated with lung cancer prognosis in three independent cohorts. In addition, specific genotypes affecting serum estrogen and tumor ER-α expression were associated with prognosis. Our findings suggest that further research investigating the use of therapeutic agents to inhibit estrogen synthesis or ER activation in lung cancer patients may be warranted.