The N-terminal transactivation domain of the glucocorticoid receptor mediates apoptosis of human small cell lung cancer cells.

Small cell lung cancer (SCLC) is an aggressive disease with a poor prognosis. These cancers are deficient in glucocorticoid receptor (GR) expression, and therefore, resistant to glucocorticoids. Overexpression of the GR both in vivo and in vitro leads to apoptotic cell death suggesting that loss of GR is favorable for cancer growth. Indeed, the GR promoter is silenced in SCLC cells by methylation. We now show that treatment of the SCLC cell line (DMS79) cells with the demethylating agent, 5-aza-2'-deoxycytidine (5-aza), results in significant endogenous re-expression of both GRα and the ligand-independent GR-P. The GR gene has a complex promoter region comprising nine alternative promoters, the proximal seven of which lie within a CpG island. The endogenous re-expression seen is attributed to the constitutive promoters 1B and 1C and 1J but predominantly 1F, which we show to be heavily methylated in SCLC cells. Flow cytometric analysis using the apoptotic marker, Annexin V, shows that this endogenous re-expression is sufficient to drive the SCLC cells to apoptosis. Apoptotic induction is specific to GR re-expression as cotreatment with 5-aza and the GR antagonist, RU486 prevented apoptosis. Of the three functional GR domains (the DNA binding domain, ligand binding domain, and transactivation domain), we identified that the transactivation domain is essential for apoptosis in SCLC. The discovery that endogenous re-expression of the GR in SCLC cells is sufficient to induce apoptotic cell death by reversing a cancer-driven DNA methylation effect may lead to the development of novel adjunct therapies.

The tobacco carcinogen NNK drives accumulation of DNMT1 at the GR promoter thereby reducing GR expression in untransformed lung fibroblasts.

Small cell lung cancer (SCLC) is a highly aggressive, predominantly cigarette smoke-induced tumour with poor prognosis. The glucocorticoid receptor (GR), a SCLC tumour suppressor gene, is typically reduced in SCLC. We now show that SCLC cells express high levels of DNA methyltransferase 1 (DNMT1) which accumulates at the GR promoter. DNMT1 expression is further increased by exposure to the tobacco carcinogen NNK. In the untransformed human lung fibroblast cell line, MRC-5, short term NNK treatment decreases GRα mRNA and protein expression due to accumulation of DNMT1 at the GR promoter. Long term NNK treatment results in persistently augmented DNMT1 levels with lowered GR levels. Long term exposure to NNK slows cell proliferation and induces DNA damage, while the GR antagonist RU486 stimulates proliferation and protects against DNA damage. Although both NNK and RU486 treatment increases methylation at the GR promoter, neither are sufficient to prevent senescence in this context. NNK exposure results in accumulation of DNMT1 at the GR promoter in untransformed lung cells mimicking SCLC cells, directly linking tobacco smoke exposure to silencing of the GR, an important step in SCLC carcinogenesis.

Glucocorticoid receptors in lung cancer: new perspectives.

Proper expression of the glucocorticoid receptor (GR) plays an essential role in the development of the lung. GR expression and signalling in the lung is manipulated by administration of synthetic glucocorticoids (Gcs) for the treatment of neonatal, childhood and adult lung diseases. In lung cancers, Gcs are also commonly used as co-treatment during chemotherapy. This review summarises the effect of Gc monotherapy and co-therapy on lung cancers in vitro, in mouse models of lung cancer, in xenograft, ex vivo and in vivo The disparity between the effects of pre-clinical and in vivo Gc therapy is commented on in light of the recent discovery of GR as a novel tumour suppressor gene.