Differential expression of metabolic genes in tumor and stromal components of primary and metastatic loci in pancreatic adenocarcinoma

Abstract

Background: Pancreatic cancer is the fourth leading cause of cancer related deaths in the United States with a five-year survival rate of 6%. It is characterized by extremely aggressive tumor growth rate and high incidence of metastasis. One of the most common and profound biochemical phenotypes of animal and human cancer cells is their ability to metabolize glucose at high rates, even under aerobic conditions. However, the contribution of metabolic interrelationships between tumor cells and cells of the surrounding microenvironment to the progression of cancer is not well understood. We evaluated differential expression of metabolic genes and, hence, metabolic pathways in primary tumor and metastases of patients with pancreatic adenocarcinoma.

Methods and findings: We analyzed the metabolic gene (those involved in glycolysis, tri-carboxylic acid pathway, pentose-phosphate pathway and fatty acid metabolism) expression profiles of primary and metastatic lesions from pancreatic cancer patients by gene expression arrays. We observed two principal results: genes that were upregulated in primary and most of the metastatic lesions; and genes that were upregulated only in specific metastatic lesions in a site-specific manner. Immunohistochemical (IHC) analyses of several metabolic gene products confirmed the gene expression patterns at the protein level. The IHC analyses also revealed differential tumor and stromal expression patterns of metabolic enzymes that were correlated with the metastasis sites.

Conclusions: Here, we present the first comprehensive studies that establish differential metabolic status of tumor and stromal components and elevation of aerobic glycolysis gene expression in pancreatic cancer.

Figure 1. mRNA expression profiles of metabolic genes in primary and metastatic lesions.

The mRNA expression profiles from pancreatic cancer patients were analyzed on 80 Agilent human whole genome 4×44 K DNA microarrays. The heatmap was created by Genesis . The heatmap display the mean log2 fold change of the gene intensity between the samples from the metastasis site or the primary pancreatic cancer site and the reference mRNA samples. The genes examined were functionally involved in glycolysis (A), Krebs tricarboxylic acid cycle (B) or pentose phosphate pathways (C).

Figure 2. Metabolic protein expression in primary and metastatic lesions of pancreatic adenocarcinomas.

Immunohistochemical examination of GLUT1 (A) HK2 (B), ALDOB (C), ENO3 (D), PKM2 (E), LDHA (F) IDH3A (G) expression in uninvolved pancreas, pancreatic tumor, and liver, lung, lymph node and diaphragm metastatic lesions. The staining was performed with Fast-Red stain. The images were captured at 200× magnification. Tumor cell area (T), stromal cell area (S) and uninvolved region (U) are indicated in the IHC images.

Figure 3. Semiquantitative evaulation of protein expression levels based on immunohistochemical findings.

Tumor cell (left) and stromal (right) expression levels of GLUT1, HK2, ALDOB, ENO3, PKM2, LDHA and IDH3A in pancreatic tumor (A), and liver (B), lung (C), lymph node (D) and diaphragm (E) metastatic lesions were graded on a scale of 0–3. The staining intensities are tabularized and indicated with gradations of red color.

Figure 4. Differential protein expression levels by immunohistochemistry.

Average tumor cell versus stromal cell expression intensities of GLUT1, HK2, ALDOB, ENO3, PKM2, LDHA and IDH3A in pancreatic tumor, and liver, lung, lymph node and diaphragm metastatic lesions are tabularized and indicated with gradations of red color.