Increased Expression of Matrix Metalloproteinases 2 and 9 and Tissue Inhibitors of Metalloproteinases 1 and 2 Correlate With

Vol. 7, 3113–3119, October 2001 Clinical Cancer Research 3113

Increased Expression of Matrix Metalloproteinases 2 and 9 and Tissue Inhibitors of Metalloproteinases 1 and 2 Correlate with Poor Prognostic Variables in Renal Cell Carcinoma

Bhaskar V. S. Kallakury,1 Shridevi Karikehalli, pression of each of these markers correlated with shortened Aptullah Haholu, Christine E. Sheehan, survival (P range, 0.004–0.05). On multivariate analysis, Norio Azumi, and Jeffrey S. Ross including tumor grade, stage, and all four markers, only and increased TIMP1 expression (0.047 ؍ advanced stage (P .independently predicted shortened survival (0.007 ؍ Department of Pathology and Laboratory Medicine, Albany Medical (P College, Albany, New York 12208 [B. V. S. K., S. K., C. E. S., J. S. R.], and Georgetown University Medical Center, Washington, Conclusion: Increased expression of MMP2, MMP9, DC 20007 [A. H., N. A.] TIMP1, and TIMP2 proteins in RCCs correlate with poor prognostic variables including shortened patient survival. The paradoxical poor prognostic implication of TIMP over- ABSTRACT expression complements the recently documented dual func- Purpose: Matrix metalloproteinases (MMPs) degrade tion of TIMPs and warrants further investigation. components of the extracellular matrix and are implicated in tissue remodeling and tumor infiltration. Tissue inhibitor INTRODUCTION of metalloproteinases (TIMPs) inhibit enzymes of the MMP 2 Proteolytic degradation of the ECM is a fundamental family and preserve stromal integrity, thus inhibiting tumor aspect of cancer development and a key event in the regulation migration. Although numerous studies on several human of tumor proliferation and metastasis. MMPs are a family of carcinomas have demonstrated a role for MMPs in tumor zinc-dependent endopeptidases that are collectively capable of metastasis and patient survival, their prognostic role in degrading most components of the basement membrane and patients with renal cell carcinoma (RCC) has not been well ECM, facilitating cell migration (1). Given their ubiquitous defined. More importantly, the recently documented para- presence, MMPs play an important role in several physiological doxical functions of TIMPs have not been characterized in and benign pathological processes such as in embryogenesis and these neoplasms. inflammatory diseases (2, 3). MMPs are secreted as inactive Experimental Design: Five-␮m, formalin-fixed, paraf- preenzymes and are transformed into active forms after cleavage fin-embedded tissue sections from 153 RCCs were immuno- of a propeptide domain of the molecule (4). On the basis of their stained using specific antibodies against MMP2, MMP9, structure, cell localization, and substrate specificity, the Ͼ20 (Novocastra, Burlingame, CA) TIMP1, and TIMP2 (Neo- human MMPs are divided into several groups such as collag- Markers, Fremont, CA) proteins. Immunostaining was enases, gelatinases, stromelysins, and membrane-type MMPs semiquantitatively scored based on intensity and distribu- (MT-MMPs; Ref. 5). TIMPs are the major endogenous regulation, and results were correlated with histological and prog- tors of MMPs and consist of four homologous members nostic variables. (TIMP1–4) identified to date (6). TIMPs are multifunctional Results: The rates of increased expression of MMPs and proteins that inhibit cell invasion in vitro and tumorigenesis and TIMPs in RCC were as follows: MMP2, 67%; MMP9, 43%; metastasis in vivo (6). Although each TIMP appears capable of TIMP1, 46%; and TIMP2, 73%. Each of these four markers inhibiting several MMPs, these proteins exhibit preferential individually correlated with histological tumor type with a inhibitory capacity; e.g.,TIMP1 and -2 selectively inhibit MMP9 vast majority of papillary and sarcomatoid RCCs expressing and -2, respectively (7). these proteins as compared with clear cell tumors (P range, Numerous investigators used various techniques and re- 0.0001–0.003). Significant coexpression of MMPs and ported the prognostic significance of increased MMP expression -Increased immunoreac .(0.0001 ؍ TIMPs was observed (P in several human malignancies. A significant correlation be- tivity for each of these proteins correlated with high tumor tween increased MMPs and poor prognosis, including shortened grade (P range, 0.0001–0.01). On univariate analysis, ex- patient survival, has been documented in carcinomas of the esophagus (8), stomach (9), colon (10), breast (11), pancreas (12), prostate (13), lung (14), and ovary (15). Although early studies have shown TIMPs to have antitumor or antimetastatic Received 4/9/01; revised 6/21/01; accepted 6/26/01. The costs of publication of this article were defrayed in part by the effects, more recent reports indicate a dual function with posi- payment of page charges. This article must therefore be hereby marked tive correlation between increased TIMP levels and poor out- advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 1 To whom requests for reprints should be addressed, at Department of Pathology and Laboratory Medicine, Georgetown University Medical Center, 121, Basic Science Building, 3900 Reservoir Road, NW, Wash- 2 The abbreviations used are: ECM, extracellular matrix; MMP, matrix ington, DC 20007. Phone: (202) 687-3444; Fax: (202) 687-8934; E-mail: metalloproteinase; TIMP, tissue inhibitor of metalloproteinase; DAB, [email protected]. 3,3Ј-diaminobenzidine.

Table 1 Antibodies and immunohistochemical procedure Citrate antigen Antibody Primary antibody Antibody Manufacturer Clone retrieval (min) dilution incubation Positive controls MMP2 Novocastra 4D3 60 1:10 32 min at 41°C Colon carcinoma MMP9 Novocastra 2C3 None 1:15 Overnight at 4°C Colon carcinoma TIMP1 Neomarkers 102D1 None 1:10 32 min at 41°C Breast carcinoma TIMP2 Neomarkers 2TMP04 60 1:10 Overnight at 4°C Breast carcinoma

come in some human malignancies (16–18). With regard to parameters by two observers (B. V. S. K., S. K.) using a con- MMP expression in RCC, there are only a few studies in the sensus method. For all markers, both the intensity of staining literature, all of which used small numbers of cases and reported and the approximate percentage of positive tumor cells were variable findings. Kugler et al. (19) isolated RNA from 17 fresh considered in the semiquantitative assessment as published pre- tumor specimens and, using a reverse transcription-PCR tech- viously (24–25). Briefly, the distribution of positive staining in nique, demonstrated a correlation between increased MMP: the tumors was graded as focal (Յ10%), regional (11–50%), or TIMP ratio and aggressive tumor behavior. Subsequently, Lein diffuse (Ͼ50%). The staining intensity was subjectively scored et al. (20) reported increased MMP9 levels in both tumor tissue as weak, moderate, or intense. Staining patterns of moderate and sera of patients with RCC but found no correlation with diffuse, intense regional, and intense diffuse were considered as prognostic variables such as tumor grade and stage. increased expression of each protein. The aim of the current study was to evaluate the immuno- Statistical Analysis. Statistical comparisons were per- histochemical expression of MMP2, MMP9, TIMP1, and formed using Stata software (Stata Corp, College Station, TX). TIMP2 proteins in RCC and to determine whether the expres- Correlation between protein expression and pathological vari- sion of these markers correlates with prognostic variables in- ables was performed using the ␹2 univariate analysis. Survival cluding survival. curves for all univariate analyses were assessed using the Kaplan-Meier method. Overall survival was defined as the in- MATERIALS AND METHODS terval between surgery and death, the end point being either Patients and Specimens. One hundred fifty-three ran- death or the closing date of this study. Multivariate analyses of domly selected surgically resected RCC specimens obtained clinicopathological parameters including survival were per- from the files of the pathology departments at Albany Medical formed using the Cox proportional hazards model. The level of Center Hospital and Georgetown University Medical Center significance was set at 0.05. between 1991 and 1999 were included. All of the H&E-stained slides from each case were reviewed and tumors were histolog- ically classified as clear cell, papillary, chromophobe, and sar- RESULTS comatoid RCC. Tumors were graded according to Fuhrman’s Of the 153 RCCS, there were 118 clear cell, 22 papillary, system (21) and staged according to TNM criteria (22). For 8 chromophobe, and 5 sarcomatoid carcinomas. The tumors statistical evaluations, grade 1 and 2 tumors were considered as were categorized as grade 1 and 2 in 114 (75%) cases and grade low grade and grade 3 and 4 as high grade (23). Similarly, stage 3 and 4 in 39 (25%) cases; and stage 1 and 2 in 104 (68%) cases 3 and 4 tumors were considered in the advanced stage category. and stage 3 and 4 in 49 (32%) cases. Follow-up information was Follow-up information was obtained from review of the pa- available for 139 (91%) patients. Whereas 34 (24%) patients tients’ medical records. died of disease, the remaining 105 (76%) were alive at the time Immunohistochemistry. To analyze for the expression of the last follow-up (mean, 40 months; range, 2–95 months). of MMP2, MMP9, TIMP1, and TIMP2 proteins, contiguous Immunohistochemistry. The frequency of increased ex- 5-␮m sections were cut from a representative block of formalin- pression of each protein and correlation with histological tumor fixed, paraffin-embedded (FFPE) tissue and placed on charged type, grade, and survival are summarized in Table 2. Immuno- slides. After deparaffinization, primary antibody incubation was staining pattern for each protein was cytoplasmic, with tumor performed manually for MMP9 and TIMP2 and by an auto- cells showing moderate-to-intense positivity as opposed to rel- mated system (Ventana Medical Systems, Tucson, AZ) for atively weaker expression in the benign elements. There was an MMP2 and TIMP1. Pertinent details regarding antibodies and overall significant coexpression of MMPs and TIMPs in all of staining procedure are summarized in Table 1. The remainder of the RCCs (P ϭ 0.0001). Of particular interest was the similarity the staining procedure included incubation with a biotintylated in the frequency of expression of MMP2 and TIMP2, and antimouse secondary, DAB substrate and hematoxylin counter- MMP9 and TIMP1, supporting their preferential association. stain and was performed on the Ventana ES automated immu- The non-clear cell RCCs (Fig. 1, A–D) showed a significant nohistochemistry system (Ventana Medical Systems, Tucson, predilection to overexpress each of these proteins as compared AZ). Negative control slides were incubated with isotype- with clear cell tumors. matched immunoglobulin in parallel with each batch of staining Increased expression of each protein correlated with high to confirm the specificity of the antibodies. tumor grade (Figs. 2, A–D and 3, A–D). However, no correlation Staining Interpretation. Staining results were inter- was found between increased expression of these proteins and preted without prior knowledge of clinical and pathological advanced tumor stage.

Table 2 Increased expression of MMPs and TIMPs by tumor type, grade, and survival Tumor type (%) Grade (%) Survival (%) Increased Protein expression (%) Clear cell Non-clear cell High Low Dead Alive MMP2 67 59 91 85 60 75 52 P ϭ 0.003 P ϭ 0.005 P ϭ 0.05 MMP9 43 38 63 79 32 64 33 P ϭ 0.001 P ϭ 0.0001 P ϭ 0.004 TIMP1 46 38 77 74 38 63 40 P ϭ 0.0001 P ϭ 0.0001 P ϭ 0.02 TIMP2 73 67 91 89 67 85 63 P ϭ 0.05 P ϭ 0.01 P ϭ 0.04

Fig. 1 An example of the same case of papillary RCC showing moderate-to-intense, diffuse, cytoplasmic immunore- activity within tumor cells for MMP2 (A), MMP9 (B), TIMP1 (C), and TIMP2 (D) proteins; ϫ200; staining: DAB and hematoxylin.

Survival Analysis. On both ␹2 and Cox univariate anal- in this model may reflect the proliferative capacity of tumor ysis, high tumor grade (P ϭ 0.02 and P ϭ 0.01, respectively) cells rather than their ability to spread. and advanced stage (P ϭ 0.003 and P ϭ 0.04, respectively) correlated with death (Fig. 4, A and B). On ␹2 analysis, increased expression of each of the four proteins correlated with DISCUSSION survival (Table 2). However, on Cox univariate analysis, which Tumor growth, invasion, and metastasis is a complex, takes into account the duration of survival, only the increased multistep process that is facilitated by the proteolytic degrada- expression of MMP9 (P ϭ 0.01) and TIMP1 (P ϭ 0.006) tion of components of the ECM and basement membrane. The correlated with a shortened patient survival (Fig. 5, A and B). By role of MMPs in this process has been firmly established based the latter analysis, increased expression of MMP2 (P ϭ 0.07) on numerous previously published experimental and clinical and TIMP2 (P ϭ 0.1) showed a trend toward shortened survival studies. Increased expression of various MMPs has correlated that did not reach statistical significance. On multivariate anal- with enhanced metastatic properties in rat embryo fibroblasts ysis of tumor grade, stage, and all four proteins using Cox (26) and cancer cell lines of rat bladder, mouse lung, and human proportional hazards model (Table 3), only tumor stage (P ϭ prostate (27–29). In human tumors, MMP expression has been 0.047) and increased TIMP1 expression (P ϭ 0.007) independ- reported to be low or undetectable in most benign elements and ently predicted shortened survival. Although the remaining vari- substantially increased in a majority of malignancies, including ables were not significant indicators of patient survival, this carcinomas of colon, breast, lung, pancreas, and prostate (10– model (Table 3) suggests a relatively higher magnitude of death 14). Analysis of both primary and metastatic tumors demon- hazard for MMP9 and TIMP2 expression over tumor grade, strated increased MMPs at the metastatic site, which supports which supports the role of these proteins in regulating matrix their role in tumor migration and spread (30). Additionally, degradation. The relatively lower death hazard for nuclear grade increased MMP levels have been reported in the plasma and

Fig. 2 Grade 3 clear cell RCCs with increased expression of MMP2 and MMP9 in comparison with their absence in grade 2 tumors. A and B, MMP2 in high-grade versus low-grade tumors. C and D, MMP9 in high-grade versus low-grade tumors. ϫ200; staining: DAB and hematoxylin.

Fig. 3 High-grade clear cell RCCs overexpressing TIMP1 and TIMP2 proteins in comparison with their absence in low- grade tumors. A and B, TIMP1 in high-grade versus low-grade tumors. C and D, TIMP2 in high-grade versus low-grade tumors. ϫ200; staining: DAB and hematoxylin.

urine of patients with cancers of colon, breast, prostate, and MMP9, TIMP1, and TIMP2 in 17 RCCs by PCR and demon- bladder compared with levels in healthy subjects (31). Further- strated a strong correlation between increased gene expression more, there is ample evidence correlating increased expression and tumor stage. Additionally, these authors also reported that with poor prognosis (8, 10, 32). With regard to genitourinary the MMP:TIMP ratio significantly increased in a graded fashion tumors, Gohji et al. (33) described increased serum levels of from normal tissues to organ-confined tumors and metastatic MMP2 and MMP3 in patients with urothelial cancer that cor- carcinomas. In contrast, Lein et al. (20) reported that only related with disease progression and poor outcome. Studies on MMP9 was increased in RCCs and found no correlation with MMP/TIMP expression in RCCs have been limited and have tumor type, grade, or stage. Our results are more in keeping with reported variable findings. Kugler et al. (19) analyzed MMP2, Kugler’s data, in that an increased expression of MMP2, MMP9,

Fig. 4 Kaplan-Meier survival estimates showing correlation of high Fig. 5 Kaplan-Meier survival estimates showing correlation of in- tumor grade (A; P ϭ 0.01) and advanced stage (B; P ϭ 0.04) with death. creased expression of MMP9 (A; P ϭ 0.01) and TIMP1 (B; P ϭ 0.006) proteins with shortened patient survival.

TIMP1, and TIMP2 portend a poor prognosis in patients with Table 3 Cox multivariate analysis of prognostic variables RCC. Furthermore, we also demonstrated that increased TIMP1 with survival protein expression independently predicted shortened patient Variable Coefficient SE P survival. MMP2 Ϫ0.0959459 0.6312752 0.879 In the context of tumor invasion, the original concept of Grade 0.2526737 0.4672274 0.589 TIMPs was that of an inhibition of MMPs, thus serving as TIMP2 Ϫ0.3059951 0.5548827 0.581 anti-invasive/anti-metastatic agents (as evidenced by data from MMP9 0.5454763 0.4471249 0.222 both experimental and clinical studies). Recombinant TIMP2 Stage 0.7109947 0.3576072 0.047 TIMP1 1.000858 0.3685423 0.007 was shown to inhibit the invasion of HT 1080 fibrosarcoma cells in vitro (34). Increased TIMP expression has been associated with decreased tumor growth, invasiveness, and metastasis in cancer cell lines of the stomach (35), pancreas (36), lung (14), and breast (37). However, the results of the current study, which of TIMP in tumor progression are not completely understood demonstrate a poor prognostic significance for increased TIMP1 and are the subject of intense investigation. This tumor-promot- and TIMP2 expression, are contrary to the previous notion and ing activity may be attributable to either proteolytic degradation more in line with recent evidence documenting a multifunc- of ECM or direct influence on cell survival and growth. TIMP2, tional complex role for TIMPs. Nemeth et al. (38) described the acting through a membrane-type MMP (MT1-MMP; 40, 41), is growth-promoting abilities of TIMP2 in several human cell reported to regulate matrix degradation. MT1-MMP is a key types including fibroblasts, keratocytes, lymphocytes, and stem enzyme in tumor angiogenesis and metastasis, hydrolyzes a cells. With regard to human cancers, increased TIMP1/TIMP2 variety of ECM components, and is a physiological activator of mRNA levels correlated with tumor stage, lymph node metas- pro-MMP2 (42). TIMP2 forms a complex with MT1-MMP and tasis, and shortened survival in patients with carcinomas of the pro-MMP2 on the cell surface, promoting hydrolysis of pro- colon (16), breast (17), and bladder (39). Our findings of the MMP2 to its active form (MMP2) and resulting in degradation poor prognostic role of increased TIMP expression concur with of ECM. Also, formation of this complex decreases the autoca- the data of Kugler (2), in which the author demonstrated a talysis of MT1-MMP, which results in increased levels of its correlation of increased TIMP2 levels with aggressive pheno- active form. It has also been reported that some TIMPs can type in RCC. directly affect cell growth/survival independent of their actions The mechanisms supporting the paradoxical positive effect on MMPs. Stimulation of cell growth by TIMPs is thought to be

mediated by c-AMP-dependent activation of protein kinase A 11. Talvensaari-Mattila, A., Paakko, P., Hoyhtya, M., Blanco-Sequei- (43) and increased tyrosine phosphorylation (44). Cell survival ros, G., and Turpeenniemi-Hujanen, T. Matrix metalloproteinase-2 im- is prolonged by the TIMP1-mediated up-regulation of antiapo- munoreactive protein: a marker of aggressiveness in breast carcinoma. Cancer (Phila.), 83: 1153–1162, 1998. ptotic protein bcl-X and decreased nuclear factor ␬B activity L 12. Bramhall, S. R., Neoptolemos, J. R., Stamp, G. W., and Lemoine, (45). What are the factors that regulate TIMP activity toward N. R. Imbalance of expression of matrix metalloproteinases (MMPs) tumor suppression or tumor promotion? Several factors that are and tissue inhibitors of matrix metalloproteinases (TIMPs) in human likely to play a key role include local TIMP concentration, pancreatic carcinoma. J. Pathol., 182: 347–355, 1997. cellular distribution, association with pro-MMPs, and presence 13. Stearns, M., and Stearns, M. E. Evidence for increased activated of a putative “TIMP receptor” (46, 47). metalloproteinase 2 (MMP-2a) expression associated with human pros- In view of the important role in tumor invasion and me- tate cancer progression. Oncol. Res., 8: 69–75, 1996. tastasis, inhibitors of MMP activity have been investigated as a 14. Kawano, N., Osawa, H., Ito, T., Nagashima, Y., Hirahara, F., method of preventing/decreasing tumor spread. Several pharma- Inayama, Y., Nakatani, Y., Kimura, S., Kitajima, H., Koshikawa, N., Miyazaki, K., and Kitamura, H. Expression of gelatinase A, tissue ceutical companies are currently developing low-molecular- inhibitor of metalloproteinases-2, matrilysin, and trypsin (ogen) in lung weight MMP inhibitors for clinical use. Clinical trials involving neoplasms: an immunohistochemical study. Hum. Pathol., 28: 613–622, batimastat (British Biotech), a potent broad based inhibitor of 1997. MMPs 1, 2, 3, and 9 (48) and marimastat (British Biotech), a 15. Young, T. N., Rodriguez, G. C., Rinehart, A. R., Bast, R. C., Jr., second-generation water-soluble synthetic MMP inhibitor, have Pizzo, S. V., and Stack, M. S. Characterization of gelatinases linked to been evaluated in patients with pancreatic, pulmonary, ovarian, extracellular matrix invasion in ovarian adenocarcinoma: purification of matrix metalloproteinase 2. Gynecol. Oncol., 62: 89–99, 1996. and mammary carcinomas. In conclusion, MMPs and their inhibitors are key enzymes 16. Murashige, M. Miyahara, M., Shiraishi, N., Saito, T., Kohno, K., and Kobayashi, M. Enhanced expression of tissue inhibitors of metal- for tumor progression, and increased expression of these pro- loproteinases in human colorectal tumors. Jpn. J. Clin. Oncol., 26: teins is associated with poor outcome in patients with RCC, 303–309, 1996. which supports a potential role for synthetic MMP inhibitors in 17. Ree, A. H., Florenes, V. A., Berg, J. P., Malandsmo, G. M., the management of this disease. The paradoxical poor prognos- Nesland, J. M., and Fodstad, O. High levels of messenger RNAs for tic significance of TIMP overexpression, as demonstrated in this tissue inhibitors of metalloproteinase (TIMP-1 and TIMP-2) in primary study and in recent literature, warrants further investigation to breast carcinomas are associated with development of distant metastasis. Clin. Cancer Res., 3: 1623–1628, 1997. fully understand the complex MMP/TIMP interactions and to 18. Fong, K. M., Kida, Y., Zimmerman, P. V., and Smith, P. J. TIMP-1 evaluate the potential efficacy of TIMP modulation in cancer and adverse prognosis in non-small cell cancer. Clin. Cancer Res., 2: therapy. 1369–1372, 1996. 19. Kugler, A., Hemmerlein, B., Thelen, P., Kallerhoff, M., Radzun, H-J., and Ringert, R-H. Expression of metalloproteinase 2 and 9 and REFERENCES their inhibitors in renal cell carcinoma. J. Urol., 160: 1914–1918, 1998. 1. Nagase, H., and Woessner, J. F. Matrix metalloproteinases. J. Biol. 20. Lein, M., Jung, K., Laube, C., Hubner, T., Winkelmann, B., Chem., 274: 21491–21494, 1999. Stephan, C., Hauptmann, S., Rudolph, B., Schnorr, D., and Loening, 2. Kugler, A. Matrix metalloproteinases and their inhibitors. Anticancer S. A. Matrix-metalloproteinases and their inhibitors in plasma and tumor Res., 19: 1589–1592, 1999. tissue in patients with renal cell carcinoma. Int. J. Cancer, 85: 801–804, 3. Manicourt, D. H., Fujimoto, N., Obata, K., and Thonar, E. J. M. 2000. Levels of circulating collagenase, stromelysin-1, and tissue inhibitor of 21. Fuhrman, S. A., Lasky, L. C., and Limas, C. Prognostic significance matrix metalloproteinase 1 in patients with rheumatoid arthritis. Arthri- of morphologic parameters in renal cell carcinoma. Am. J. Surg. Pathol., tis Rheum., 38: 1031–1039, 1995. 6: 655–663, 1982. 4. Carmeliet, P., Moons, L., Lijnen, R., Baes, M., Lemaitre, V., Tip- 22. Guinan, P., Sobin, L. H., Algaba, F., Badellino, F., Kameyama, S., ping, P., Drew, A., Eeckhout, Y., Shapiro, S., Lupu, F., and Collen, D. Mac Lennan, G., and Novick, A. TNM staging of renal cell carcinoma: Urokinase-generated plasmin activates matrix metalloproteinases during Workgroup No. 3. Union International Contre le Cancer (UICC) and the aneurysm formation. Nat. Genet., 17: 439–444, 1997. American Joint Committee on Cancer (AJCC). Cancer (Phila.), 80: 5. Curran, S., and Murray, G. I. Matrix metalloproteinase in tumour 992–993, 1997. invasion and metastasis. J. Pathol., 189: 300–308, 1999. 23. Medeiros, L. J., Gelb, A. B., and Weiss, L. M. Low-grade renal cell 6. Gomez, D. E., Alonso, D. F., Yoshiji, H., and Thorgeirsson, U. P. carcinoma. A clinicopathologic study of 53 cases. Am. J. Surg. Pathol., Tissue inhibitors of metalloproteinases: structure, regulation and bio- 11: 633–642, 1987. logical functions. Eur. J. Cell Biol., 74: 111–122, 1997. 24. Kallakury, B. V. S., Yang, F., Figge, J., Smith, K. E., Kausik, S. J., 7. Baker, A. H., Ahonen, M., and Kahari, V-M. Potential applications Tacy, N. J., Fisher, H. A. G., Kaufman, R., Figge, H., and Ross, J. S. of tissue inhibitor of metalloproteinase (TIMP) overexpression for can- Decreased levels of CD44 protein and mRNA in prostate cancer. Cor- cer gene therapy. Adv. Exp. Med. Biol., 465: 469–483, 2000. relation of tumor grade and ploidy. Cancer (Phila.), 78: 1461–1469, 8. Murray, G. I., Duncan, M. E., O’Neil, P., McKay, J. A., Melvin, 1996. W. T., and Fothergill, J. E. Matrix metalloproteinase-1 is associated with 25. Kallakury, B. V. S., Sheehan, C. E., Ambros, R. A., Fisher, poor prognosis in esophageal cancer. J. Pathol., 185: 256–261, 1998. H. A. G., Kaufman, R. P., Jr., Muraca, P.J., and Ross, J. S. Correlation cdc2 9. Sier, C. F. M., Kubben, F. J. G. M., Ganesh, S., Heerding, M. M., of p34 cyclin-dependent kinase overexpression, CD44s downregu- Griffioen, G., Hanemaaijer, R., van Krieken, J. H., Lamers, C. B., and lation, and HER-2/neu oncogene amplification with recurrence in pros- Verspaget, H. W. Tissue levels of matrix metalloproteinase MMP-2 and tatic adenocarcinomas. J. Clin. Oncol., 16: 1302–1309, 1998. MMP-9 are related to the overall survival of patients with gastric 26. Hua, J., and Muschel, R. J. Inhibition of matrix metalloproteinase 9 carcinoma. Br. J. Cancer, 74: 413–417, 1996. expression by a ribozyme blocks metastasis in a rat sarcoma model 10. Murray, G. I., Duncan, M. E., O’Neil, P., Melvin, W. T., and system. Cancer Res., 56: 5279–5284, 1996. Fothergill, J. E. Matrix metalloproteinase-1 is associated with poor 27. Kawamata, H., Kameyama, S., Kawai, K., Tenaka, Y., Nan, L., prognosis in colorectal cancer. Nat. Med., 2: 461–462, 1996. Barch, D., Stetler-Stevenson, W. G., and Oyasu, R. Marked acceleration

of the metastatic phenotype of a rat bladder carcinoma cell line by the 38. Nemeth, J. A., Rafe, A., Steiner, M., and Goolsby, C. L. TIMP-2 expression of human gelatinase A. Int. J. Cancer, 63: 568–575, 1995. growth stimulatory activity: a concentration and cell-type specific re- 28. Tsunezuka, Y., Kinoh, H., Takino, T., Watanabe, Y., Okada, Y., sponse in the presence of insulin. Exp. Cell Res., 224: 110–115, 1996. Shinagawa, A., Sato, H., and Seiki, M. Expression of membrane-type 39. Grignon, D. J., Sakr, W., Toth, M., Ravery, V., Angulo, J., Shamsa, matrix metalloproteinase 1 (MT1-MMP) in tumor cells enhances pul- F., Pontes, J. E., Crissman, J. C., and Fridman, R. High levels of tissue monary metastasis in an experimental metastasis assay. Cancer Res., 56: inhibitor of metalloproteinase-2 (TIMP-2) expression are associated 5678–5683, 1996. with poor outcome in invasive bladder cancer. Cancer Res., 56: 1654– 29. Powell, W. C., Knoz, J. D., Navre, M., Grogan, T. M., Kittelson, J., 1659, 1996. and Nagle, R. B. Expression of the metalloproteinase matrilysin in 40. Shofuda, K., Moriyama, K., Nishihashi, A., Higashi, S., Mizushima, DU-145 cells increases their invasive potential in severe combined H., Yasumitsu, H., Miki, K., Sato, H., Seiki, M., and Miyazaki, K. Role immunodeficient mice. Cancer Res., 53: 417–422, 1993. of tissue inhibitor of metalloproteinase-2 (TIMP-2) in regulation of pro-gelatinase A activation catalyzed by membrane-type matrix metal- 30. Sutinen, M. Kainulainen, T., Hurskainen, T., Vesterlund, E., Alex- loproteinase-1 (MT1-MMP) in human cancer cells. J. Biochem., 124: ander, J. P., Overall, C. M. Sorsa, T., and Salo, T. Expression of matrix 462–470, 1998. metalloproteinases (MMP-1 and -2) and their inhibitors (TIMP-1, -2, and -3) in oral lichen planus, dysplasia, squamous cell carcinoma and 41. Butler, G. S., Butler, M. J., Atkinson, S. J., Will, H., Tamura, T., lymph node metastasis. Br. J. Cancer, 77: 2239–2245, 1998. VanWestrum, S. S., Crabbe, T., Clements, J., D’Ortho, M. P., and Murphy, G. The TIMP2 membrane type 1 metalloproteinase “receptor” 31. Zucker, S., Hymowitz, M., Conner, C., Zarrabi, H. M., Hurewitz, regulates the concentration and efficient activation of pro-gelatinase A. N., Matrisian, L., Boyd, D., Nicolson, G., and Montana, S. Meas- A—a kinetic study. J. Biol. Chem., 273: 871–880, 1998. urement of matrix metalloproteinases and tissue inhibitors of metallo- 42. Toth, M., Bernardo, M. M., Gervasi, D. C., Soloway, P. D., Wang, proteinases in blood and tissues. Clinical and experimental applications. Z., Bigg, H. F., Overall, C. M., DeClerck, Y. A., Tschesche, H., Cher, Ann. NY Acad. Sci., 878: 212–227, 1999. M. L., Brown, S., Mobashery, S., and Fridman, R. Tissue inhibitor of 32. Yamamoto, H., Adachi, Y., Itoh, F., Iku, S., Matsuno, K., Kusano, metalloproteinase (TIMP)-2 acts synergistically with synthetic matrix M., Arimura, Y., Endo, T., Hinoda, Y., Hosokawa, M., and Imai, K. metalloproteinase (MMP) inhibitors but not with TIMP-4 to enhance the Association of matrilysin expression with recurrence and poor prognosis (membrane type 1)-MMP-dependent activation of pro-MMP-2. J. Biol. in human esophageal squamous cell carcinoma. Cancer Res., 59: 3313– Chem., 275: 41415–41423, 2000. 3316, 1999. 43. Corcoran, M. L., and Stetler-Stevenson, W. G. Tissue inhibitor of 33. Gohji, K., Fujimoto, N., Komiyama, T., Fujii, A., Ohkawa, J., metalloproteinase-2 stimulates fibroblast proliferation via c-AMP- Kamidono, S., and Nikajima, M. Evaluation of serum levels of matrix dependent mechanism. J. Biol. Chem., 270: 13458–13459, 1995. metalloproteinase-2 and -3 as new predictors of recurrence in patients 44. Yamashita, K., Suzuki, M., Iwata, H., Koike, T., Hamaguchi, M., with urothelial carcinoma. Cancer (Phila.), 78: 2379–2387, 1996. Shinagawa, A., Noguchi, T., and Hayakawa, T. Tyrosine phosphoryla- 34. Albini, A., Melchiori, A., Santi, L., Liotta, L. A., Brown, P. D., and tion is crucial for growth signaling by tissue inhibitors of metallopro- Stetler-Stevenson, W. G. Tumor cell invasion inhibited by TIMP-2. teinases (TIMP-1 and TIMP-2). FEBS Lett., 396: 103–107, 1996. J. Natl. Cancer Inst. (Bethesda), 83: 775–779, 1991. 45. Guedez, L., Stetler-Stevenson, W. G., Wolff, L., Wang, J., Fuku- 35. Watanabe, M., Takahashi, Y., Ohta, T., Mai, M., Sasaki, T., and shima, P., Mansoor, A., and Stetler-Stevenson, M. In vitro suppression Seiki, M. Inhibition of metastasis in human gastric cancer cells trans- of programmed cell death of B cells by tissue inhibitor of metallopro- fected with tissue inhibitor metalloproteinase1 gene in nude mice. teinases-1. J. Clin. Investig., 102: 2002–2010, 1998. Cancer (Phila.), 77: 1676–1680, 1996. 46. Hayakawa, T., Yamashita, K., Ohuchi, E., and Shinagawa, A. Cell 36. Bramhall, S. R., Stamp, G. W., Dunn, J., Lemoine, N. R., and growth promoting activity of tissue inhibitor of metalloproteinases-2 Neoptolemos, J. P. Expression of collagenase (MMP2), stromelysin (TIMP-2). J. Cell Sci., 107: 2373–2379, 1994. (MMP3) and tissue inhibitor of the metalloproteinases (TIMP1) in 47. DeClerck, Y. A., Yean, T. D., Chan, D., Shimada, H., and Langley, pancreatic and ampullary disease. Br. J. Cancer, 73: 972–978, 1996. K. E. Inhibition of tumor invasion of smooth muscle cell layers by 37. Alonso, D. F., Skilton, G., DeLorenzo, M. S., Scursoni, A. M., recombinant human metalloproteinase inhibitor. Cancer Res., 51: 2151– Yoshiji, H., and Gomez, D. E. Histopathologic findings in a highly 2157, 1991. invasive mouse mammary carcinoma transfected with human tissue 48. Brown, P. D., and Giavazzi, R. Matrix metalloproteinase inhibition: inhibitor of metalloproteinases-1. Oncol. Rep., 5: 1083–1087, 1998. a review of anti-tumour activity. Ann. Oncol., 6: 967–974, 1995.

Downloaded from clincancerres.aacrjournals.org on September 25, 2021. © 2001 American Association for Cancer Research. Increased Expression of Matrix Metalloproteinases 2 and 9 and Tissue Inhibitors of Metalloproteinases 1 and 2 Correlate with Poor Prognostic Variables in Renal Cell Carcinoma

Bhaskar V. S. Kallakury, Shridevi Karikehalli, Aptullah Haholu, et al.

Clin Cancer Res 2001;7:3113-3119.

Updated version Access the most recent version of this article at: http://clincancerres.aacrjournals.org/content/7/10/3113

Cited articles This article cites 48 articles, 13 of which you can access for free at: http://clincancerres.aacrjournals.org/content/7/10/3113.full#ref-list-1

Citing articles This article has been cited by 22 HighWire-hosted articles. Access the articles at: http://clincancerres.aacrjournals.org/content/7/10/3113.full#related-urls

E-mail alerts Sign up to receive free email-alerts related to this article or journal.

Reprints and To order reprints of this article or to subscribe to the journal, contact the AACR Publications Subscriptions Department at [email protected].

Permissions To request permission to re-use all or part of this article, use this link http://clincancerres.aacrjournals.org/content/7/10/3113. Click on "Request Permissions" which will take you to the Copyright Clearance Center's (CCC) Rightslink site.