Clinical Significance Of S100P Expression In Human Colorectal Carcinomas And The Effects Of S100P On The Biological Behaviors Of Colon Cancer Cell Lines

Background: Colorectal carcinoma is one of the most common types of malignant tumors worldwide. In China, the incidence of colorectal cancer ranks fourth and, colorectal cancer is the 4th leading cause of cancer-related mortality. Although great advances have been made in the diagnosis of this cancer as well as in the variety of tumor therapies, the survival rate for the patients with colorectal cancer is not improved profoundly. Insights into the molecular pathogenesis of colorectal carcinoma will be important for the early diagnosis, the improvement of cure rate and life quality of patients. Recently, it has been shown that S100P, a new member of the S100 calcium binding family, may contribute to the initiation and progression in a number of tumors. It is not clear, however, whether S100P plays a role in the tumorigenesis of colorectal carcinoma.Objective: To investigate the expression for S100P mRNA and protein in human colorectal carcinoma specimen and their paired normal colorectal tissue. If the expression of S100P was determined in the tumors, its clinical significance will be studied. Moreover, the expression for S100P mRNA and protein is detected in a number of colorectal cancer cell lines, and the effects of S100P on the biological behaviors of cancer cell lines are assessed.Methods: Sixty-two fresh frozen tumor tissue samples and their paired normal colorectal tissue from 62 patients with sporadic colorectal cancer were obtained postoperatively at our hospital between May 2005 and March 2006. The clinical data of patients and tumoral features, including age, gender, family history, history of radio-and/or chemotherapy prior to surgery, primary tumor location, tumor size and staging, histological pattern, degree of differentiation, fasting preoperative serum levels of tumor markers, were analyzed and correlated with the expression of S100P.The expression for S100P mRNA and protein as well as its receptor "receptor for advanced glycation end products" were detected both in colorectal cancer tissue and their paired normal tissue and in colon cancer cell lines by using Western blot and RT-PCR. The data from Western blot and RT-PCR were correlated with clinical data and tumoral features.The mammalian expression vector of S100P-pcDNA3.1(-) was constructed and transfected into SW480 cell line which did not express S100P. The effects of S100P gene on the proliferation of transfected SW480 cell line and control cell line (vector transfected) as well as wild type of SW480 were evaluated using MTT assay and cell growth curve. In addition, the cell cycle was assessed by flow cytometry; the ability of cell migration was evaluated by wound healing assay. To test signal transduction pathway of S100P, the intracellular level of phosphorylated extracellular signal regulated protein kinase 1/2 (ERK1/2) was detected by Western blot.Results: Of 62 patients with colorectal cancer, 38 were male patients. The average age of the patients was 60.8±13.6 years. Primary tumor site from 24 samples located in proximal colon, 14 tumors located in distal colon and 24 tumors in recta. The rate of expressing for S100P protein and mRNA in colorectal carcinomas was 58.1% (36/62) and 67.9% (38/56), respectively. S100P protein was expressed in only one normal tissue sample (1/62, 1.6%), while S100P mRNA was shown in 11 of 56 (19.6%) paired normal tissue. Of 10 human colorectal cancer cell lines, 7 were shown the expression for S100P mRNA whereas S100P protein was detected in 5 cell lines. Receptor for advanced glycation end products protein could be detected in all of the colorectal carcinomas tissue samples, paired normal tissue and 10 cell lines. The expressing rate of S100P protein and mRNA in proximal, distal and rectal cancer was 37.5%, 64.3%, 75% (P=0.027) and 59.1%, 69.2%, 76.2% (P=0.483), respectively. In tumors stagingⅠ,Ⅱ,ⅢandⅣ, the expressing rate of S100P protein and mRNA is 20%, 47.1%, 54.2%, 87.5% and 40%, 56.3%, 71.4%, 85.7%, respectively. This result demonstrated that the expression of S100P protein is significantly associated with advanced tumor stage (Spearman correlation, P=0.004). Similarly, expression of mRNA for S100P was also correlated with advanced stage (P=0.047).MTT assay and cell growth curve analysis indicated that SW480-S100P cells, which were stably transfected by S100P gene, proliferated more rapidly than wide type SW480 and the SW480-vector cells. Average doubling time of cell population was reduced from about 20h in SW480 and SW480-vector cells to 15h in SW480-S100P cells. Flow cytometry analysis showed a higher proportion of S phase in SW480-S100P cells compared with SW480-vector cells as well as wild type SW480 cell line [(30.6±3.8) % vs. (21.9±0.8) % and (21.4±1.8) %, respectively, P=0.007]. Twenty-four hours after wounding, the cell number of SW480-S100P migration were 11.3±3.1, much more than the number of migrated wild type SW480 and SW480-vector cells (10.7±3.7 and 19.3±3.5, respectively. P=0.035). After SW480 cells were transfected with S100P gene, the intracellular levels of phospho-ERK1/2 were upregulated, being 4.2 or 3.7 times more than that in SW480 or SW480-vector cells, respectively (P=0.001).Conclusions: S100P protein and mRNA were expressed in the most of human colorectal cancer tissue and cell lines, much higher than that in paired normal tissue. The S100P expression in colorectal cancer is significantly correlated with primary tumor location and staging, implying that S100P might play a role in the invasion and metastasis of colorectal carcinomas. S100P could promote proliferation and migration of colorectal cancer cells and regulate cell cycle. One of the mechanisms underlying these processes was that S100P could increase the level of phospho-ERK1/2.