The clinical outcome for patients with lung cancer has not changed significantly for over two decades. New approaches to therapy are urgently needed, and these should be based on new understanding of fundamental growth-promoting pathways in the lung and possible sex differences in the natural history and therapeutic responses of NSCLC [1
]. The incidence of NSCLC, the predominant form of lung cancer, has now reached epidemic proportions particularly in women [1
]. Emerging data suggest that female sex hormones have a role in lung cancer development and progression. Endogenous and exogenous estrogens appear to be key contributors in stimulating NSCLC growth and progression. Aromatase, a key enzyme for estrogen biosynthesis, is expressed and active in NSCLC [5
]. The present and previous studies also reveal significant expression and activity of ERα and ERβ in both extranuclear and nuclear sites in most NSCLC. We now report on the occurrence of PR transcripts and protein in NSCLC. Using immunohistochemistry, expression of PR protein was observed in the nucleus and/or extranuclear compartment in the majority of human tumor specimens and NSCLC cell lines examined. These findings are consistent with independent reports on the expression of PR protein in NSCLC [25
]. Although some initial studies indicate that tumor PR levels correlate with clinical outcome [33
], others suggest no apparent correlation between tumor PR and clinicopathologic characteristics [25
]. Further investigation of the potential interplay between ER, PR and growth factor signaling pathways may be required to fully understand their contribution to clinical outcome [35
Previous work confirms that estrogen signaling in human lung interacts with other signaling pathways, such as EGFR, and contributes to tumor growth and clinical outcome [4
]. Additional studies indicate that overexpression of VEGF and EGF family ligands and their receptors also correlate with clinical outcome for NSCLC patients [45
]. New findings show that pulmonary vascularity is altered by sex steroids, particularly estradiol, with a greater number of lung microvessels in ovariectomized female mice receiving estradiol as compared to placebo [19
]. Similarly, cancer progression is dependent on development of a rich vascular network that supplies vital nutrients to the growing tumor (43
). Angiogenesis, the process of new blood vessel formation, is regulated by a number of potent growth factors, one of the most effective of which is VEGF [43
]. It is well-established that VEGF is produced by many tumor cells, including lung cancer cells [45
], and VEGF produced by tumor cells is essential for malignant expansion, presumably by increasing proliferation of endothelial cells from neighboring blood vessels through interactions with VEGF receptors present on these cells. Our findings further demonstrate that combinations of estradiol and progestins administered in vitro
cooperate in promoting NSCLC cell secretion of VEGF and, consequently, enhancing vascular endothelial cell proliferation, an event important for tumor-associated angiogenesis. Independent experiments suggest that progestins do not directly enhance the proliferation of vascular endothelial cells [68
]. The results of recent clinical trials indicate that hormone replacement therapy with estrogens and progestins is associated with higher risk of lung cancer in postmenopausal women than treatment with estrogens alone or placebo [29
]. This observation is consistent with the current studies showing that progestins as well as estrogens [7
] stimulate the secretion of VEGF by NSCLC cells which in turn stimulates local angiogenesis. These observations suggest that tumor-induced VEGF secretion may function in a paracrine manner to sustain tumor growth. Although endogenous progesterone levels are very low in postmenopausal women and men, in situ
production of progesterone is reported in NSCLC specimens [33
]. Further, it is possible that progesterone receptor, as in breast [69
], may also signal through ligand-independent mechanisms due to phosphorylation by kinases in the lung [33
]. Thus, it will be important to determine if PR activation by ligand-independent pathways can induce VEGF secretion from lung tumor cells to promote angiogenesis.
In view of recent evidence suggesting that sex steroids may influence the numbers of stem/progenitor cells in adult mammary tissues [41
], we investigated the role of estradiol and progestins in promoting expansion of stem/progenitor cells in NSCLC. Dual treatment with estrogen and progestin in vitro
increased the numbers of putative tumor stem/progenitor cells as evidenced by enrichment of CSPC markers and the formation of tumor spheres with self-renewal capability in vitro
and enhanced tumorigenicity in vivo
. It is reported that the number of tumor spheres (composed of tumor stem/progenitor cell populations) generated upon serial passage provides an indirect measure of cancer stem cell self-renewal [55
]. Independent experiments have also shown that ALDH-positive and CD133-positive lung tumor cell subsets [58
] can generate tumors in vivo
that recapitulate the heterogeneity of the parental cancer cells and maintain tumor progression. Immunohistochemical analysis of clinical specimens from lung cancer patients and controls further show that high tumor expression of ALDH and CD133 is correlated with a poor prognosis for patients with lung cancer. Although stem cells in other tissues, such as breast, are reported to lack expression of sex steroid receptors [41
], estrogen and progesterone may impact the function of stem/progenitor cells indirectly by mediating the supply of regulatory factors from neighboring cells (such as Wnt or receptor activator of NF-κB ligand [RANKL] signaling) or by regulating the activity of more differentiated progenitor cells [41
]. One recent report also shows that vascular endothelial cell-secreted factors can enhance the survival and self-renewal of neighboring head and neck cancer stem/progenitor cells [71
]. Of note, lung cancer stem/progenitor cells are also reported to produce substantially higher levels of VEGF than that of bulk tumor cells [58
]. Thus, treatments aimed to modulate the activity of ERα, ERβ and/or PR may modulate tumor progression by diverse molecular and cellular pathways and thereby offer new approaches to treat non-small cell lung cancer. Further studies are needed to pursue these alternatives for specific targets of sex steroid action in NSCLC.
Although progestins alone may not directly induce the proliferation of bulk NSCLC cells, the current and earlier findings suggest that these steroids regulate the activity of other tissue factors (such as VEGF, EGF) that indirectly determine the final proliferative or antiproliferative state. Similar actions of progesterone have been reported in breast cancers [72
]. Further studies will also be required to understand the significance of extranuclear and nuclear expression patterns of steroid receptors in malignant cells. In endometrial cancers, the PR-A isoform is localized in the nucleus, and PR-B is largely cytoplasmic in the absence of ligand [73
]. Similar investigations in lung cancer remain to be done [35
]. The distribution of steroid receptors in different subcellular compartments may well modulate signaling by genomic, nongenomic and/or ligand-independent pathways that contribute to tumor progression [15
As in studies of nuclear receptor expression in breast cancer [75
], a standardized and validated approach to the immunohistochemical detection and interpretation of ERα, ERβ, PR and possibly GPR30 expression is needed for potential screening of human lung tumor specimens in the clinic [35
]. A systematic examination of hormonal biomarkers could potentially be prognostic and may be useful to identify patients with NSCLC who might benefit from the administration of antihormone therapeutics.
- PR, ERα and ERβ are expressed in both extranuclear and nuclear sites in NSCLC.
- PR transcripts were lower in cancerous as compared to non-malignant tissue.
- Estrogen and progestins promote tumor VEGF secretion and angiogenesis.
- Estrogen and progestins stimulate putative tumor stem/progenitor cell expansion.
- ER- and/or PR-targeted therapies may offer new approaches to manage NSCLC.