Involvement of aryl hydrocarbon receptor signaling in the development of small cell lung cancer induced by HPV E6/E7 oncoproteins

Abstract

Background: Lung cancers consist of four major types that and for clinical-pathological reasons are often divided into two broad categories: small cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC). All major histological types of lung cancer are associated with smoking, although the association is stronger for SCLC and squamous cell carcinoma than adenocarcinoma. To date, epidemiological studies have identified several environmental, genetic, hormonal and viral factors associated with lung cancer risk. It has been estimated that 15-25% of human cancers may have a viral etiology. The human papillomavirus (HPV) is a proven cause of most human cervical cancers, and might have a role in other malignancies including vulva, skin, oesophagus, head and neck cancer. HPV has also been speculated to have a role in the pathogenesis of lung cancer. To validate the hypothesis of HPV involvement in small cell lung cancer pathogenesis we performed a gene expression profile of transgenic mouse model of SCLC induced by HPV-16 E6/E7 oncoproteins.

Methods: Gene expression profile of SCLC has been performed using Agilent whole mouse genome (4 × 44k) representing ~ 41000 genes and mouse transcripts. Samples were obtained from two HPV16-E6/E7 transgenic mouse models and from littermate's normal lung. Data analyses were performed using GeneSpring 10 and the functional classification of deregulated genes was performed using Ingenuity Pathway Analysis (Ingenuity® Systems, http://www.ingenuity.com).

Results: Analysis of deregulated genes induced by the expression of E6/E7 oncoproteins supports the hypothesis of a linkage between HPV infection and SCLC development. As a matter of fact, comparison of deregulated genes in our system and those in human SCLC showed that many of them are located in the Aryl Hydrocarbon Receptor Signal transduction pathway.

Conclusions: In this study, the global gene expression of transgenic mouse model of SCLC induced by HPV-16 E6/E7 oncoproteins led us to identification of several genes involved in SCLC tumor development. Furthermore, our study revealed that the Aryl Hydrocarbon Receptor Signaling is the primarily affected pathway by the E6/E7 oncoproteins expression and that this pathway is also deregulated in human SCLC. Our results provide the basis for the development of new therapeutic approaches against human SCLC.

Figure 1

Confirmation of microarray data. RT-PCR was done using total RNA from wild-type mouse lung (line 1), transgenic mouse lung (line 2) and PPAP9 cells (line 3). Scg2: secretogranin 2, Chga: chromogranin, Cav-1: caveolin1, Cav-2: caveolin 2, Ascl1: achete-scute complex homologue 1, Igf-2: insulin-like growth factor 2, FoxA2: forkhead box A2, A, 18S: 18 S ribosomal RNA M: marker.

Figure 2

Normal lung tissue and transgenic lung tumor histology. (A) Normal lung; original magnification 150×; (B) Transgenic SCLC; original magnification 150×; (C) Transgenic SCLC; original magnification 400×.

Figure 3

Human SCLC hierarchical clustering of the significantly deregulated genes. Analysis show human normal control lung tissues and SCLC samples. Up-regulated genes are shown in red, down-regulated genes are shown in green and black bars indicate not significantly changed genes.

Figure 4

Expansion of the significantly up-regulated genes human SCLC hierarchical clustering. The expansion highlights the common up-regulated genes in human SCLC and in SCLC transgenic mouse induced by E6/E7 oncoproteins.

Figure 5

Expansion of the significantly down-regulated genes human SCLC hierarchical clustering. The expansion highlights the common down-regulated genes in human SCLC and in SCLC transgenic mouse induced by E6/E7 oncoproteins.

Figure 6

IPA pathway graphical representation of Aryl Hydrocarbon Receptor Signaling. 51 deregulated genes are represented out of 154. Gene products are positioned according to sub cellular localization. Only direct connections (i.e., direct physical contact between two molecules) among the individual gene products are shown for clarity of presentation; lines indicate protein-protein binding interactions, and arrows refer to "acts on" interactions such as proteolysis, expression, and protein-protein interactions. Genes up regulated are shown in red, down-regulated genes are shown in green.

Figure 7

Comparison analysis of most significant pathways in human SCLC and in SCLC induced by E6/E7. The comparison of top ten canonical pathways identified by IPA in human SCLC and in transgenic mouse emphasizes the common differential regulation in the tumor development.