Stat3 downstream genes serve as biomarkers in human lung carcinomas and chronic obstructive pulmonary disease

Abstract

Smoking causes lung cancer and chronic obstructive pulmonary disease (COPD) that impose severe health problem to humans. Both diseases are related to each other and can be induced by chronic inflammation in the lung. To identify the molecular mechanism for lung cancer formation, a CCSP-rtTA/(teto)(7)Stat3C bitransgenic model was generated recently. In this model, persistent activation of the Stat3 signaling pathway induced pulmonary inflammation and adenocarcinoma formation in the lung. A group of Stat3 downstream genes were identified by Affymetrix GeneChip microarray analysis that can be used as biomarkers for lung cancer diagnosis and prognosis. To determine which human lung cancers are related to the Stat3 pathway, multiple Stat3 downstream genes were screened in human lung cancers (adenocarcinomas and squamous cell carcinomas) and lung tissue with COPD. In both cancer and COPD, the Stat3 gene was up-regulated. A panel of Stat3-up-regulated downstream genes in mice was up-regulated in human adenocarcinomas, but not in human squamous cell carcinomas. This panel of genes was also modestly up-regulated in lung tissue with COPD from patients with a history of smoking and not up-regulated in those without histories of smoking. Several Stat3-down-regulated downstream genes also showed differential expression patterns in carcinoma and COPD. These studies support a concept that Stat3 is a potent oncogenic molecule that plays a role in formation of lung adenocarcinomas in both mice and humans. The carcinogenesis of adenocarcinoma and squamous cell carcinoma is mediated by different molecular mechanisms and pathways in vivo. Stat3 and its downstream genes can serve as biomarkers for lung adenocarcinoma and COPD diagnosis and prognosis in mice and humans.

Figure 1. Stat3 mRNA expression in adenocarcinoma, squamous cell carcinoma and COPD

Expression of Stat3 mRNA was measured by quantitative Real-Time PCR and normalized by GAPDH mRNA expression. Numbers in each column represent average ΔCt. Average fold changes were determined by 2^(ΔΔCt), in which ΔΔCT= ΔCt(normal human samples) − ΔCt(cancer or COPD samples). Numbers on the bottom represent average fold changes compared to the normal human samples. Normal human samples were set up as one. Nor: normal samples. COPD: COPD samples from smokers. Benign: COPD samples from non-smokers. Adeno: adenocarcinoma. Squa: squamous cell carcinoma.

Figure 2. Expression of Stat3-up-regulated genes in adenocarcinoma

Expression of Stat3-up-regulated genes was measured by quantitative Real-Time PCR and normalized by GAPDH mRNA expression. Numbers in each column represent average ΔCt. Average fold changes were determined by 2^(ΔΔCt), in which ΔΔCT= ΔCt(normal human samples) − ΔCt(adenocarcinoma samples). Numbers on the bottom represent average fold changes compared to the normal human samples. Normal human samples were set up as one. Nor: normal samples. Adeno: adenocarcinoma samples.

Figure 3. Expression of Stat3-up-regulated genes in squamous cell carcinoma

Expression of Stat3 up-regulated genes was measured by quantitative Real-Time PCR and normalized by GAPDH mRNA expression. Numbers in each column represent average ΔCt. Average fold changes were determined by 2^(ΔΔCt), in which ΔΔCT= ΔCt(normal human samples) − ΔCt(squamous cell carcinoma samples). Numbers on the bottom represent average fold changes compared to the normal human samples. Normal human samples were set up as one. Nor: normal samples. Squa: squamous cell carcinoma samples.

Figure 4. Expression of Stat3-up-regulated genes in COPD from non-smokers

Expression of Stat3 up-regulated genes was measured by quantitative Real-Time PCR and normalized by GAPDH mRNA expression. Numbers in each column represent average ΔCt. Average fold changes were determined by 2^(ΔΔCt), in which ΔΔCT= ΔCt(normal human samples) − ΔCt(benign COPD samples). Numbers on the bottom represent average fold changes compared to the normal human samples. Normal human samples were set up as one. Nor: normal samples. Benign: COPD from non-smoker samples.

Figure 5. Expression of Stat3-up-regulated genes in COPD from smokers

Expression of Stat3 up-regulated genes was measured by quantitative Real-Time PCR and normalized by GAPDH mRNA expression. Numbers in each column represent average ΔCt. Average fold changes were determined by 2^(ΔΔCt), in which ΔΔCT= ΔCt(normal human samples) − ΔCt(COPD from smokers). Numbers on the bottom represent average fold changes compared to the normal human samples. Normal human samples were set up as one. Nor: normal samples. COPD: COPD samples from smokers.

Figure 6. Expression of Stat3-down-regulated genes in adenocarcinomas, squamous cell carcinoma and COPD

Expression of Stat3-down-regulated Krtdap (A), Lor (B) and Rdhe2 (C) genes was measured by quantitative Real-Time PCR and normalized by GAPDH mRNA expression. Numbers in each column represent average ΔCt. Average fold changes were determined by 2^(ΔΔCt), in which ΔΔCT= ΔCt(normal human samples) − ΔCt(cancer or COPD samples). Numbers on the bottom represent average fold changes compared to the normal human samples. Normal human samples were set up as one. Nor: normal samples. COPD: COPD samples from smokers. Benign: Benign COPD samples from non-smokers. Adeno: adenocarcinoma. Squa: squamous cell carcinoma.

Figure 6. Expression of Stat3-down-regulated genes in adenocarcinomas, squamous cell carcinoma and COPD

Expression of Stat3-down-regulated Krtdap (A), Lor (B) and Rdhe2 (C) genes was measured by quantitative Real-Time PCR and normalized by GAPDH mRNA expression. Numbers in each column represent average ΔCt. Average fold changes were determined by 2^(ΔΔCt), in which ΔΔCT= ΔCt(normal human samples) − ΔCt(cancer or COPD samples). Numbers on the bottom represent average fold changes compared to the normal human samples. Normal human samples were set up as one. Nor: normal samples. COPD: COPD samples from smokers. Benign: Benign COPD samples from non-smokers. Adeno: adenocarcinoma. Squa: squamous cell carcinoma.

Figure 6. Expression of Stat3-down-regulated genes in adenocarcinomas, squamous cell carcinoma and COPD

Expression of Stat3-down-regulated Krtdap (A), Lor (B) and Rdhe2 (C) genes was measured by quantitative Real-Time PCR and normalized by GAPDH mRNA expression. Numbers in each column represent average ΔCt. Average fold changes were determined by 2^(ΔΔCt), in which ΔΔCT= ΔCt(normal human samples) − ΔCt(cancer or COPD samples). Numbers on the bottom represent average fold changes compared to the normal human samples. Normal human samples were set up as one. Nor: normal samples. COPD: COPD samples from smokers. Benign: Benign COPD samples from non-smokers. Adeno: adenocarcinoma. Squa: squamous cell carcinoma.