**1. Introduction**

Pancreatic ductal adenocarcinoma (PDAC) is an extremely aggressive cancer [1, 2], as derived from the respective 5-year survival rate of about 6% [3, 4]. This poor prognosis results, at least in part, from a delayed diagnosis of the disease. All therapeutic efforts during the past 50 years,

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

such as surgery, radiation therapy, chemotherapy, or combinations thereof, have shown little impact on the course of this aggressive neoplastic disease. It is hoped, however, that a full understanding of the molecular biology of pancreatic cancer will help in diagnosing, prevent‐ ing, and treating this cancer.

A most recent study on pancreatic cancer patients describes genetic details, which have been detected through full genome sequencing and copy number analyses in 100 PDACs. Genetic drivers known for pancreatic cancer, such as K-ras, TP53, SMAD4, and mutations in CDKN2A, were confirmed, whereas newly identified mutations, such as in the KDM6A gene, highlighted the role of chromatin modifications [5]. Based on familial clustering studies, it was estimated that 10% of PDAC cases are linked to an inherited predisposition [4, 6, 7]. Germ line mutations were especially found in tumor suppressor genes, such as INK4A, BRCA2, and LKB1, the mismatch repair gene MLH1, and the cationic trypsinogen gene PRSS1 [8, 9]. These germ line mutations are responsible for onset and penetrance of PDAC, but they likely contribute more to the malignant progression of precursor lesions than to cancer initiation [10, 11]. Of course, exogenous factors, such as cigarette smoking, constitute a measurable risk, as well [12].

In the early stage of PDAC, the genetic mutations of oncogenes like K-ras and suppressor genes like TP53—or both—lead to constitutive activation of transcription factors, but in the late stage stress factors such as acidosis and hypoxia, which are frequently encountered in the tumor microenvironment, further deregulate the expression of metastasis-related proteins. PDAC metastasis is a progressive, debilitating disease that is characterized by pain, asthenia, anorexia, and cachexia. The formation of metastasis is a complex and progressive process, including four basic steps: dissociation of cells from the primary tumor, existence and survival in the circulatory system, break down and degrading of endothelium and basement mem‐ branes in target organs, and (d) establishment of a colony of metastatic cells [13].

Currently, it is hoped that new findings related to K-ras, the tumor's unique metabolic needs, and how the stroma and immune system affect the PDAC, will change the overall situation for the better [14].

In order to better understand the PDAC metastatic process, a liver metastasis model was developed by our group [15]. This model focused on the final steps of metastasis, which are related to organ colonization. To that purpose, ASML rat PDAC cells, which had been marked by luciferase (ASMLluc [15]) were implanted intraportally into isogenic BDX rats for mimicking the final phase of liver metastasis. This process of liver colonization was related to the size of the liver and the remaining life span of a given animal. Thus, for reisolation of tumor cells from rat liver, four periods were selected and classified as early (days 1, 3), intermediate (day 6), advanced (day 15), and terminal (day 21) stages. Also, tumor cells were recultivated in vitro after their reisolation at 21 days for three additional periods (3, 6, and 9 days) to define the microenvironment effect on gene expression after residing 21 days in rat liver. Reisolated ASML tumor cells were used for total RNA isolation and then investigated by Illumina chip array for genes and miRNAs, which show modulation of their expression during liver colonization. These modulations were evaluated by Ingenuity Pathway Analysis (IPA) as described in the study of Al-Taee et al. [16].

The aim of this study was to establish a rat model for identifying and understanding patho‐ physiological processes during metastatic liver colonization, as well as for finding new and specific tumor markers as tools for diagnostic and therapeutic approaches. Here, we report on the modulation of gene expression during stages of liver colonization by ASML pancreatic cancer cells in rat liver.
