Pancreatic adenocarcinoma is the fourth leading cause of cancer-related death in the Western industrialized world, owing to rapid primary tumor growth and ensuing metastasis (36
). The metastatic cascade is dependent upon angiogenesis and cell migration, two processes that are regulated acutely by the local microenvironment (37
). Therefore, understanding the interaction of pancreatic tumor cells with stromal components is critical to developing improved therapeutic options for patients. We sought to determine if two proteins linked to the desmoplastic response of pancreatic cancer interact at a functional level in the progression of this disease. SPARC is a matricellular protein implicated in tumor growth (11
) with an in vivo
function in the desmoplastic response, characteristic of pancreatic cancer. MMP9 on the other hand is associated with ECM turnover and cell migration through the ECM.
Our results show for the first time that orthotopic pancreatic tumors grow larger and more aggressively in the absence of host SPARC. Furthermore, forced expression of MMP9 by tumor cells increases tumor size but does not increase metastasis in WT and SPARC-/- animals. We also identified that SPARC regulates MMP9 expression in vivo and that SPARC and MMP9 both impact ECM deposition and angiogenesis in the tumor microenvironment. We found that, in general, tumor size correlated inversely with collagen deposition and that the lack of SPARC or the increased expression of MMP9 resulted in reduced collagen.
The expression of SPARC in tumor tissue from patients with pancreatic cancer has been shown to correlate with a worse prognosis (40
). In particular, patients whose tumor had fibroblasts that expressed SPARC by immunohistochemistry had a median survival of 15 months whereas patients whose tumor stroma did not express SPARC had a median survival of 30 months. The authors conclude that, after controlling for other prognostic factors, the relative hazard for patients with stromal expression of SPARC was 1.89 (40
). These studies support the idea that SPARC contributes to the progression of pancreatic cancer in humans, which is contrary to our results in SPARC-deficient animals. The reason(s) underlying this difference is unclear but might be due to the fact that the ECM of tumors from SPARC-/-
animals is likely quite different from the ECM of human tumors from patients that have low or hard to detect SPARC protein. It is possible that defects inherent to the ECM of tumors grown in SPARC-/-
mice are not replicated effectively by low or absent SPARC expression by stromal cells present in human tumors.
There are conflicting reports on whether SPARC is a substrate for MMP9. Sasaki et al. (41
) found that MMP9 could cleave SPARC while Sage et al. (21
) found that MMP3 but not MMP9 was the relevant MMP that mediated cleavage of SPARC. We did not determine if there was an increase in SPARC cleavage in PAN02-MMP9 tumors. It is possible however, to evaluate if SPARC is cleaved at L197 or L196 using neo-epitope specific antibodies (20
), these studies are now in progress. Our results are however, consistent with previous studies (42
) that have demonstrated that SPARC can increase the expression of MMPs.
Metastasis is the leading cause of death in pancreatic cancer patients. We provide evidence here that metastatic events occur more frequently in SPARC-/-
animals. Since the initial steps in the metastatic cascade occur in the context of the ECM, we hypothesize that the compromised ECM formed in the absence of host SPARC favors invasion and metastasis. SPARC expression by pancreatic tumors cells has been implicated in tumor cell migration in vitro
and as a possible factor contributing to metastasis in vivo
). Our results suggest that an ECM formed in the absence of host SPARC can also facilitate metastasis. It is important to note that PAN02 cells produce SPARC. Therefore, we cannot exclude the possibility that tumor cell-derived SPARC contributes to metastatic spread in WT
MMP9 expression is frequently associated with metastasis and is thought to facilitate tumor cell invasion (39
). However, results from this study supported by previous compelling reports suggest that the underlying mechanisms and the function of MMP9 in metastasis are more complex (31
). Our study demonstrates that orthotopic implantation of tumor cells forced to express MMP9 results in a decrease in metastatic burden compared to parental PAN02 tumor cells. Altogether, these results are in striking contrast to our own in vitro
cell migration data and the generally perceived function of MMP9 as a promoter of metastasis but consistent with, Deryugina et al. (31
) who showed that down-regulation of MMP9 in HT1080 cells resulted in increased intravasation and metastasis in the CAM.
These findings highlight the complex nature of MMP9 activity in the tumor microenvironment and might be partly explained by the paradigm that excess MMP9 expression inhibits tumor progression by generating endothelial cell inhibitors from both ECM and non-ECM sources, such as angiostatin and tumstatin (46
). Thus, the outcome of MMP activity in the tumor microenvironment may be dependent on a variety of factors including ECM deposition, the presence of other proteases and cytokines, the time, level, and site of MMP production, and the level and activity of adaptor proteins such as SPARC.