Protein Expression of STAT3, pSTAT3, MMP-7 and VEGF in Colorectal Adenocarcinoma: An Immunohistochemical Study

Rakesh Naidu^1*, Lim Vin Nee², Mak Joon Wah², Kevin Moissinac³, A. Rahman A. Jamal⁴, Isa Mohd Rose⁵, Yunus Gul Alif Gul⁶, Patricia Lim Kim Chooi², Gregory Tan Jin San⁷

¹School of Medicine and Health Sciences, Monash University Malaysia, Selangor, Malaysia.
²International Medical University, Kuala Lumpur, Malaysia.
³Department of Surgery, Penang Medical College, Pulau Pinang, Malaysia.
⁴Department of Paediatrics, Faculty of Medicine, Universiti Kebangsaan Malaysia, Kuala Lumpur, Malaysia.
⁵Department of Pathology, Faculty of Medicine, Universiti Kebangsaan Malaysia, Kuala Lumpur, Malaysia.
⁶Department of Surgery, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia, Selangor, Malaysia.
⁷School of Health Sciences, The University of Notre Dame Australia, Fremantle, Australia.

DOI: 10.4236/jct.2014.513119                

Abstract

Background: The purpose of the present study is to investigate the expression levels of STAT3, pSTAT3, MMP-7 and VEGF in colorectal adenocarcinoma, and also to determine association with the clinico-pathological parameters and co-expression of these genes. Methods: An immunohistochemical method was used to evaluate the expression of MMP-7 and VEGF genes in 93 archival tissues whereas STAT3 and pSTAT3 expression was determined in 75 cases. Results: Overexpression of STAT3 was detected in 26.7% (20/75), pSTAT3 in 13.4% (10/75), MMP-7 in 38.8% (36/93) and VEGF in 59.2% (55/93) of the colorectal carcinomas. STAT3, MMP-7 and VEGF immunopositivity were significantly correlated with poorly-differentiated tumors (P = 0.004; P = 0.03; P = 0.002, respectively) but not with other parameters. However, pSTAT3 immunostaining was not significantly associated with the clinico-pathological characteristics. Significant relationship was noted between overexpression of pSTAT3 and STAT3 (P < 0.001), pSTAT3 and VEGF (P = 0.044), pSTAT3 and MMP-7 (P = 0.003), and STAT3 and VEGF (P = 0.037) but marginal association was detected between STAT3 and MMP-7 (P = 0.057), and MMP-7 and VEGF (P = 0.052). Conclusion: Our data suggest that expression of these genes may have an important role in tumor dedifferentiation and may be useful as indicators of biologic aggressiveness. Co-expression of the biomarkers by cancer cells may have important implications in colorectal cancer biology and could be useful biological markers of the malignant phenotype.

Keywords

STAT3, pSTAT3, MMP-7, VEGF, Colorectal Adenocarcinoma

Conflicts of Interest

The authors declare no conflicts of interest.

References

[1]	Levy, D.E. and Darnell Jr., J.E. (2002) Stats: Transcriptional Control and Biological Impact. Nature Reviews Molecular Cell Biology, 3, 651-662. http://dx.doi.org/10.1038/nrm909
[2]	Bromberg, J. (2002) Stat Proteins and Oncogenesis. Journal of Clinical Investigation, 109, 1139-1142. http://dx.doi.org/10.1172/JCI0215617
[3]	Tsareva, S.A., Moriggl, R., Corvinus, F.M., Wiederanders, B., Schütz, A., Kovacic, B. and Friedrich, K. (2007) Signal Transducer and Activator of Transcription 3 Activation Promotes Invasive Growth of Colon Carcinomas through Matrix Metalloproteinase Induction. Neoplasia, 9, 279-291. http://dx.doi.org/10.1593/neo.06820
[4]	Ma, X.T., Wang, S., Ye, Y.J., Du, R.Y., Cui, Z.R. and Somsouk, M. (2004) Constitutive Activation of Stat3 Signaling Pathway in Human Colorectal Carcinoma. World Journal of Gastroenterology, 10, 1569-1573.
[5]	Bowman, T., Garcia, R., Turkson, J. and Jove, R. (2000) STATs in Oncogenesis. Oncogene, 19, 2474-2488. http://dx.doi.org/10.1038/sj.onc.1203527
[6]	Niu, G., Wright, K.L., Huang, M., Song, L., Haura, E., Turkson, J., Zhang, S., Wang, T., Sinibaldi, D., Loppola, D., Heller, R., Ellis, L.M., Karras, J., Bromberg, J., Pardoll, D., Jove, R. and Yu, H. (2002) Constitutive Stat3 Activity Up-Regulates VEGF Expression and Tumor Angiogenesis. Oncogene, 21, 2000-2008. http://dx.doi.org/10.1038/sj.onc.1205260
[7]	Real, P.J., Sierra, A., De Juan, A., Segovia, J.C., Lopez-Vega, J.M. and Fernandez-Luna, J.L. (2002) Resistance to Chemotherapy via Stat3-Dependent Overexpression of Bcl-2 in Metastatic Breast Cancer Cells. Oncogene, 21, 7611-7668. http://dx.doi.org/10.1038/sj.onc.1206004
[8]	Kusaba, T., Nakayama, T., Yamazumi, K., Yakata, Y., Yoshizaki, A., Inoue, K., Nagayasu, T. and Sekine, I. (2006) Activation of STAT3 Is a Marker of Poor Prognosis in Human Colorectal Cancer. Oncology Reports, 15, 1445-1451.
[9]	Park, J.K., Hong, R., Kim, K.J., Lee, T.B. and Lim, S.C. (2008) Significance of p-STAT3 Expression in Human Colorectal Adenocarcinoma. Oncology Reports, 20, 597-604.
[10]	Curran, S. and Murray, G.I. (2000) Matrix Metalloproteinases: Molecular Aspects of Their Roles in Tumour Invasion and Metastasis. European Journal of Cancer, 36, 1621-1630. http://dx.doi.org/10.1016/S0959-8049(00)00156-8
[11]	Stetler-Stevenson, W.G., Hewitt, R. and Corcoran, M. (1996) Matrix Metalloproteinases and Tumour Invasion: From Correlation and Causality to the Clinic. Seminar Cancer Biology, 7, 147-154. http://dx.doi.org/10.1006/scbi.1996.0020
[12]	Wilson, C.L. and Matrisian, L.M. (1996) Matrilysin: An Epithelial Matrix Metalloproteinase with Potentially Novel Functions. International Journal of Biochemistry and Cell Biology, 28, 123-136. http://dx.doi.org/10.1016/1357-2725(95)00121-2
[13]	Newell, K.J., Witty, J.P., Rodgers, W.H. and Matrisian, L.M. (1994) Expression and Localization of Matrix-Degrading Metalloproteinases during Colorectal Tumorigenesis. Molecular Carcinogenesis, 10, 199-206. http://dx.doi.org/10.1002/mc.2940100404
[14]	Ishikawa, T., Ichikawa, Y., Mitsuhashi, M., Momiyama, N., Chishima, T., Tanaka, K., Yamaoka, H., Miyazakic, K., Nagashima, Y., Akitaya, T. and Shimada, H. (1996) Matrilysin Is Associated with Progression of Colorectal Tumor. Cancer Letters, 107, 5-10. http://dx.doi.org/10.1016/0304-3835(96)04336-4
[15]	Adachi, Y., Yamamoto, H., Itoh, F., Arimura, Y., Nishi, M., Endo, T. and Imai, K. (2001) Clinicopathologic and Prognostic Significance of Matrilysin Expression at the Invasive Front in Human Colorectal Cancers. International Journal of Cancer, 95, 290-294.
[16]	Masaki, T., Sugiyama, M., Matsuoka, H., Abe, N., Izumisato, Y., Sakamoto, A. and Atomi, Y. (2003) Matrix Metalloproteinases May Contribute Compensationally to Tumor Invasion in T1 Colorectal Carcinomas. Anticancer Research, 23, 4169-4173.
[17]	Masaki, T., Matsuoka, H., Sugiyama, M., Abe, N., Goto, A., Sakamoto, A. and Atomi, Y. (2001) Matrilysin (MMP-7) as a Significant Determinant of Malignant Potential of Early Invasive Colorectal Carcinomas. British Journal of Cancer, 84, 1317-1321. http://dx.doi.org/10.1054/bjoc.2001.1790
[18]	Hanahan, D. and Folkman, J. (1996) Patterns and Emerging Mechanisms of the Angiogenic Switch during Tumorigenesis. Cell, 86, 353-364. http://dx.doi.org/10.1016/S0092-8674(00)80108-7
[19]	Dvorak, H.F., Detmar, M., Claffey, K.P., Nagy, J.A., van de Water, L. and Senger, D.R. (1995) Vascular Permeability Factor/Vascular Endothelial Growth Factor: An Important Mediator of Angiogenesis in Malignancy and Inflammation. International Archives of Allergy and Immunology, 107, 233-235. http://dx.doi.org/10.1159/000236988
[20]	Guba, M., Seeliger, H., Kleespies, A., Jauch, K.W. and Bruns, C. (2004) Vascular Endothelial Growth Factor in Colorectal Cancer. International Journal of Colorectal Disease, 19, 510-517. http://dx.doi.org/10.1007/s00384-003-0576-y
[21]	Zafirellis, K., Agrogiannis, G., Zachaki, A., Gravani, K., Karameris, A. and Kombouras, C. (2008) Prognostic Significance of VEGF Expression Evaluated by Quantitative Immunohistochemical Analysis in Colorectal Cancer. Journal of Surgical Research, 147, 99-107. http://dx.doi.org/10.1016/j.jss.2007.05.041
[22]	Altomare, D.F., Rotelli, M.T., Pentimone, A., Rossiello, M.R., Martinelli, E., Guglielmi, A., De Fazio, M., Marino, F., Memeo, V., Colucci, M. and Semeraro, N. (2007) Tissue Factor and Vascular Endothelial Growth Factor Expression in Colorectal Cancer: Relation with Cancer Recurrence. Colorectal Disease, 9, 133-138. http://dx.doi.org/10.1111/j.1463-1318.2006.01158.x
[23]	Lim, G.C.C. and Halimah, Y. (2004) Second Report of the National Cancer Registry. Cancer Incidence in Malaysia 2003. National Cancer Registry, Lumpur.
[24]	Madbouly, K.M., Senagore, A.J., Mukerjee, A., Delaney, C.P., Connor, J. and Fazio, V.W. (2007) Does Immunostaining Effectively Upstage Colorectal Cancer by Identifying Micrometastatic Nodal Disease? International Journal of Colorectal Disease, 22, 39-48. http://dx.doi.org/10.1007/s00384-006-0098-5
[25]	Hbibi, A.T., Lagorce, C., Wind, P., Spano, J.P., Guetz, G., Milano, G., Benamouzig, R., Rixe, O., Morere, J.F., Breau, J.L., Martin, A. and Fagard, R. (2008) Identification of a Functional EGF-R/p60c-src/STAT3 Pathway in Colorectal Carcinoma: Analysis of Its Long-Term Prognostic Value. Cancer Biomarkers, 4, 83-91.
[26]	Kawada, M., Seno, H., Uenoyama, Y., Sawabu, T., Kanda, N., Fukui, H., Shimahara, Y. and Chiba, T. (2006) Signal Transducers and Activators of Transcription 3 Activation Is Involved in Nuclear Accumulation of Beta-Catenin in Colorectal Cancer. Cancer Research, 66, 2913-2917. http://dx.doi.org/10.1158/0008-5472.CAN-05-3460
[27]	Kusaba, T., Nakayama, T., Yamazumi, K., Yakata, Y., Yoshizaki, A., Nagayasu, T. and Sekine, I. (2005) Expression of p-STAT3 in Human Colorectal Adenocarcinoma and Adenoma; Correlation with Clinicopathological Factors. Journal of Clinical Pathology, 58, 833-838. http://dx.doi.org/10.1136/jcp.2004.023416
[28]	Xiong, H., Zhang, Z.G., Tian, X.Q., Sun, D.F., Liang, Q.C., Zhang, Y.J., Lu, R., Chen, Y.X. and Fang, J.Y. (2008) Inhibition of JAK1, 2/STAT3 Signaling Induces Apoptosis, Cell Cycle Arrest, and Reduces Tumor Cell Invasion in Colorectal Cancer Cells. Neoplasia, 10, 287-297.
[29]	Lassmann, S., Schuster, I., Walch, A., Gobel, H., Jütting, U., Makowiec, F., Hopt, U. and Werner, M. (2007) STAT3 mRNA and Protein Expression in Colorectal Cancer: Effects on STAT3-Inducible Targets Linked to Cell Survival and Proliferation. Journal of Clinical Pathology, 60, 173-179. http://dx.doi.org/10.1136/jcp.2005.035113
[30]	Adachi, Y., Yamamoto, H., Itoh, F., Hinoda, Y., Okada, Y. and Imai, K. (1999) Contribution of Matrilysin (MMP-7) to the Metastatic Pathway of Human Colorectal Cancers. Gut, 45, 252-258. http://dx.doi.org/10.1136/gut.45.2.252
[31]	Remy, L., Trespeuch, C., Bachy, S., Scoazec, J.Y. and Rousselle, P. (2006) Matrilysin 1 Influences Colon Carcinoma Cell Migration by Cleavage of the Laminin-5 β3 Chain. Cancer Research, 66, 11228-11237. http://dx.doi.org/10.1158/0008-5472.CAN-06-1187
[32]	Jeffery, N., McLean, M.H., El-Omar, E.M. and Murray, G.I. (2009) The Matrix Metalloproteinase/Tissue Inhibitor of Matrix Metalloproteinase Profile in Colorectal Polyp Cancers. Histopathology, 4, 820-828. http://dx.doi.org/10.1111/j.1365-2559.2009.03301.x
[33]	Fang, Y.J., Lu, Z.H., Wang, G.Q., Pan, Z.Z., Zhou, Z.W., Yun, J.P., Zhang, M.F. and Wan, D.S. (2009) Elevated Expressions of MMP7, TROP2, and Survivin Are Associated with Survival, Disease Recurrence, and Liver Metastasis of Colon Cancer. International Journal of Colorectal Disease, 24, 875-884. http://dx.doi.org/10.1007/s00384-009-0725-z
[34]	Roca, F., Mauro, L.V., Morandi, A., Bonadeo, F., Vaccaro, C., Quintana, G.O., Specterman, S., Joffé, E.B.K., Pallotta, M.Q., Puricelli, L.I. and Lastiri, J. (2006) Prognostic Value of E-Cadherin, β-Catenin, MMPs (7 and 9), and TIMPs (1 and 2) in Patients with Colorectal Carcinoma. Journal of Surgical Oncology, 93, 151-160. http://dx.doi.org/10.1002/jso.20413
[35]	Barresi, V., Di Gregorio, C., Regiani-Bonetti, L., Leon, M., Barresi, G. and Vitarelli, E. (2010) Stage I Colorectal Carcinoma: VEGF Immunohistochemical Expression, Microvessel Density, and Their Correlation with Clinical Outcome. Virchows Archiv, 457, 11-19. http://dx.doi.org/10.1007/s00428-010-0933-5
[36]	Yin, Y., Cao, L.Y., Wu, W.Q., Li, H., Jiang, Y. and Zhang, H.F. (2010) Blocking Effects of siRNA on VEGF Expression in Human Colorectal Cancer Cells. World Journal of Gastroenterology, 16, 1086-1092. http://dx.doi.org/10.3748/wjg.v16.i9.1086
[37]	Kwon, H.C., Kim, S.H., Oh, S.Y., Lee, S., Kwon, K.A., Lee, J.H., Choi, H.J., Park, K.J., Lee, H.S., Roh, M.S. and Kim, H.J. (2010) Clinicopathological Significance of Nuclear Factor-κB, HIF-1α, and Vascular Endothelial Growth Factor Expression in Stage III Colorectal Cancer. Cancer Science, 10, 1557-1561. http://dx.doi.org/10.1111/j.1349-7006.2010.01553.x
[38]	Ljujic, B., Radosavljevic, G., Jovanovic, I., Pavlovic, S., Zdravkovic, N., Milovanovic, M., Acimovic, L., Knezevic, M., Bankovic, D., Zdravkovic, D. and Arsenijevic, N. (2010) Elevated Serum Level of IL-23 Correlates with Expression of VEGF in Human Colorectal Carcinoma. Archive of Medical Research, 41, 182-189. http://dx.doi.org/10.1016/j.arcmed.2010.02.009
[39]	Cao, D., Hou, M., Guan, Y.S., Jiang, M., Yang, Y. and Gou, H.F. (2009) Expression of HIF-1α and VEGF in Colorectal Cancer: Association with Clinical Outcomes and Prognostic Implications. BMC Cancer, 9, 432. http://dx.doi.org/10.1186/1471-2407-9-432
[40]	Doger, F.K., Meteoglu, I., Tuncyurek, P., Okyay, P. and Cevikel, H. (2006) Does the EGFR and VEGF Expression Predict the Prognosis in Colon Cancer? European Surgical Research, 38, 540-544. http://dx.doi.org/10.1159/000096774
[41]	Noike, T., Miwa, S., Soeda, J., Kobayashi, A. and Miyagawa, S. (2008) Increased Expression of Thioredoxin-1, Vascular Endothelial Growth Factor, and Redox Factor-1 Is Associated with Poor Prognosis in Patients with Liver Metastasis from Colorectal Cancer. Human Pathology, 39, 201-208. http://dx.doi.org/10.1016/j.humpath.2007.04.024
[42]	Zhang, C., Hao, L., Wang, L., Xiao, Y., Ge, H., Zhu, Z., Luo, Y. and Zhang, Y. (2010) Elevated IGFIR Expression Regulating VEGF and VEGF-C Predicts Lymph Node Metastasis in Human Colorectal Cancer. BMC Cancer, 10, 184. http://dx.doi.org/10.1186/1471-2407-10-184
[43]	Khorana, A.A., Ryan, C.K., Cox, C., Eberly, S. and Sahasrabudhe, D.M. (2003) Vascular Endothelial Growth Factor, CD68, and Epidermal Growth Factor Receptor Expression and Survival in Patients with Stage II and Stage III Colon Carcinoma: A Role for the Host Response in Prognosis. Cancer, 97, 960-968. http://dx.doi.org/10.1002/cncr.11152
[44]	Uner, A., Ebinc, F.A., Akyurek, N., Unsal, D., Mentes, B.B. and Dursun, A. (2005) Vascular Endothelial Growth Factor, c-erbB-2 and c-erbB-3 Expression in Colorectal Adenoma and Adenocarcinoma. Experimental Oncology, 27, 225-228.
[45]	Ottaiano, A., Franco, R., Talamanca, A., Liguori, G., Tatangelo, F., Delrio, P., Nasti, G., Barletta, E., Facchini, G., Daniele, B., Di Blasi, A., Napolitano, M., Ieranò, C., Calemma, R., Leonardi, E., Albino, V., De Angelis, V., Falanga, M., Boccia, V., Capuozzo, M., Parisi, V., Botti, G., Castello, G., Iaffaioli, V.R. and Scala, S. (2006) Overexpression of both CXC Chemokine Receptor 4 and Vascular Endothelial Growth Factor Proteins Predicts Early Distant Relapse in Stage II-III Colorectal Cancer Patients. Clinical Cancer Research, 12, 2795-2803. http://dx.doi.org/10.1158/1078-0432.CCR-05-2142
[46]	Dassoulas, K., Gazouli, M., Theodoropoulos, G., Christoni, Z., Rizos, S., Zisi-Serbetzoglou, A., et al. (2010) Vascular Endothelial Growth Factor and Endoglin Expression in Colorectal Cancer. Journal of Cancer Research Clinical Oncology, 136, 703-708. http://dx.doi.org/10.1007/s00432-009-0709-y
[47]	Cascio, S., Ferla, R., D’Andrea, A., Gerbino, A., Bazan, V., Surmacz, E. and Russo, A. (2009) Expression of Angiogenic Regulators, VEGF and Leptin, Is Regulated by the EGF/PI3K/STAT3 Pathway in Colorectal Cancer Cells. Journal of Cell Physiology, 221, 189-194. http://dx.doi.org/10.1002/jcp.21843
[48]	Overall, C.M. and Kleifeld, O. (2006) Validating Matrix Metalloproteinases as Drug Targets and Anti-Targets for Cancer Therapy. Nature Review Cancer, 6, 227-239. http://dx.doi.org/10.1038/nrc1821
[49]	Roy, R., Zhan, B. and Moses, M.A. (2006) Making the Cut: Protease-Mediated Regulation of Angiogenesis. Experimental Cell Research, 312, 608-622. http://dx.doi.org/10.1016/j.yexcr.2005.11.022
[50]	Ito, T.K., Ishii, G., Chiba, H. and Ochiai, A. (2007) The VEGF Angiogenic Switch of Fibroblasts Is Regulated by MMP-7 from Cancer Cells. Oncogene, 26, 7194-7203. http://dx.doi.org/10.1038/sj.onc.1210535