AUTHORS: D. Delitto*, S.J. Hughes*, K.E. Behrns*, J.G. Trevino*
*Department of Surgery, University of Florida, Gainesville, Florida, United States.
BACKGROUND: Despite previously demonstrated stimulatory effects of nicotine on the growth and metastasis of
cancer cells, nicotine replacement strategies are increasingly employed as a safe alternative to tobacco use.
Additionally, the concept of the tumor microenvironment, which incorporates interactions between cancer cells
and the stromal support network, is essential to pancreatic cancer investigations. Here, we show that
upregulation of Id1, previously implicated as an essential mediator of nicotine-stimulated growth, metastasis and
chemoresistance in pancreatic cancer, is dependent on intact c-Met signaling. Further, we correlate
microenvironmental c-Met signaling, a known contributor to cancer metastasis, to a novel metastatic phenotype
following nicotine administration in a patient-derived xenograft model.
METHODS: Patient-derived subcutaneous xenograft models incorporating viable human stromal elements were
established from both primary and metastatic pancreatic cancer specimens. Mice were randomized to receive
nicotine vs. saline via intraperitoneal injections. Tumor volumes were measured thrice weekly using calipers. At
the conclusion of the study, tumors and lungs were resected and examined by H&E and IHC. Additionally,
following surgical resection of human pancreatic cancer specimens, tumor associated stroma (TAS) was cultured
using the outgrowth method. Primary cancer cells and TAS were exposed to nicotine in physiologic doses. HGF, cMet and Id1 expression was analyzed using a combination of western blot, ELISA and RT-PCR. Additionally, c-Met
and Id1 expression was examined by western blot in cancer cells exposed to supernatants from TAS cultures.
RESULTS: In contrast to xenografts obtained from cell lines, patient-derived pancreatic cancer xenografts retain
morphologic characteristics of the original tumor microenvironment after multiple passages. Nicotine exposure to
mice bearing patient-derived xenografts resulted in dramatically increased tumor growth and pulmonary
metastasis (Nicotine vs. Control: 4/7 vs. 0/8 and 6/8 vs. 1/9 pulmonary metastatic events in tumors of primary and
metastatic origin, respectively). Nicotine exposure resulted in a time-dependent increase in hepatocyte growth
factor (HGF) secretion in TAS. The HGF receptor, c-Met, was upregulated in response to nicotine exposure in
pancreatic cancer cells. Addition of TAS supernatants resulted in the overexpression of c-Met and Id1 in cancer
cells. Epigenetic silencing of c-Met in multiple pancreatic cancer cell lines abrogated TAS-mediated Id1 induction.
CONCLUSION: These results provide insight into a critical response of the tumor microenvironment to nicotine
administration. The activation of HGF/c-Met signaling between TAS and cancer cells is demonstrated as a required
step in Id1-mediated chemoresistance. Remarkably, nicotine administration to mice bearing patient-derived
xenografts, which conserve desmoplastic stromal elements present in the original tumor, results in dramatically
increased tumor growth and metastasis. To our knowledge, this is the first demonstration of a subcutaneous
patient-derived pancreatic cancer xenograft model exhibiting reliable metastatic behavior. Given this significant
augmentation of cancer progression, nicotine replacement therapies should be reconsidered as a safe alternative
to tobacco use. Furthermore, the c-Met signaling axis warrants further investigation in the context of novel
pancreatic cancer therapies.
Figure 1: Dual Mechanism of Nicotine Stimulation. Nicotine reacts with receptors in both the epithelial (top) and stromal
(bottom) elements of the pancreatic cancer microenvironment.
Figure 2: Nicotine Stimulates the Growth and Metastasis of Patient-Derived Xenografts. Nicotine administered to NOD-SCID
mice in a patient-derived xenograft model. Tumors originally derived from a liver metastasis (left) and primary pancreatic
adenocarcinoma (right) exhibited significant augmentation of metastatic behavior.