Expression of Stathmin in Localized Upper Urinary Tract Urothelial Carcinoma: Correlations With Prognosis

Oncology Expression of Stathmin in Localized Upper Urinary Tract Urothelial Carcinoma: Correlations With Prognosis Wei-Chou Lin, Shyh-Chyan Chen, Fu-Chang Hu, Shih-Chieh Chueh, Yeong-Shiau Pu, Hong-Jeng Yu, and Kuo-How Huang OBJECTIVES

METHODS

RESULTS

CONCLUSIONS

To investigate the expression of stathmin in upper urinary tract (UUT) urothelial carcinoma (UC) in human tissues and to determine whether the level of stathmin expression was correlated with prognosis because overexpression of stathmin has been observed in various malignancies. We first analyzed stathmin mRNA level in 5 UUT-UC paired fresh specimens (tumor and nontumoral urothelium) by RT-PCR. Besides, a total of 58 patients with localized UUT-UC (pT1-3N0M0) treated by nephroureterectomy were enrolled. The stathmin expression in UUT-UC specimens was analyzed by immunohistochemical (IHC) staining. Stathmin IHC score was defined as the proportion of positive staining tumor cells from each patient’s specimen. The stathmin IHC score ⱖ 0.5 was defined as strong (⫹) immunoreactivity and ⬍ 0.5 as weak (⫺) immunoreactivity. Significant differences in stathmin mRNA between UUT-UC and paired normal urothelium were noted in 5 of the patients. Of the 58 UUT-UC specimens, stathmin immunoreactivity (strong [⫹] vs weak [⫺]) was significantly associated with pT stage (P ⫽ .006) as well as with recurrence-free and cancer-specific survival. In multivariate analysis, stathmin IHC score was a significant predictor for both recurrence-free survival (hazard rates: 22.4; P ⫽ .001) and cancer-specific survival (hazard rates: 39.8; P ⫽ .0012). The stathmin immunostaining is a novel prognosticator for patients with localized UUT-UC stathmin may be a help identify such patients with poor outcomes to benefit from receiving close follow-up and early adjuvant therapy. UROLOGY 74: 1264 –1270, 2009. © 2009 Elsevier Inc.

U

pper urinary tract (UUT) urothelial carcinoma (UC), which might involve the renal pelvis and/or ureter, accounted for ⬍ 5% of all UCs in published studies.1 Standard treatment for patients with UUT-UC is ipsilateral radical nephroureterectomy (NU) and bladder cuff resection.2,3 The tumor-necrosis-metastasis stage and histologic grade are well-known prognostic factors and have the most significant impact on patient survival.4-6 The prognosis in locally advanced or lymph node-metastatic disease was worse than localized disease. However, there are still no ideal prognostic marker in patients with localized disease. Some factors including tumor site and lymphovascular invasion have been investigated and the results was controversial.7-9 Some molecular markers have been investigated to improve the This study was supported by the grant from the National Science Council of Taiwan (NSC 94WFA0102741). From the Departments of Pathology and Urology; and National Center of Excellence for General Clinical Trial and Research, National Taiwan University Hospital, Taipei, Taiwan Reprint requests: Kuo-How Huang, M.D., Department of Urology, National Taiwan University Hospital, No 7 Chung-Shan South Road, Taipei, Taiwan, 100. E-mail: [email protected] Submitted: December 23, 2008, accepted (with revisions): April 22, 2009

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© 2009 Elsevier Inc. All Rights Reserved

accuracy of predicting outcomes.10-12 A novel prognosticator for clinical practice is important to identify patients with localized UUT-UC at higher risk of recurrence to receive early adjuvant therapy. Stathmin (oncoprotein 18, Op 18) is a ubiquitous cytosolic protein, which is involved in diverse cell function such as cell cycle regulation, proliferation, and differentiation. Stathmin is encoded by the human STMN1 gene located at chromosome 1p35-36.13 Stathmin is a major microtubule-destabilizing protein and interacts directly with soluble tubulin to form a complex that sequesters free tubulin and impedes the polymerization of microtubules.14 The unphosphorylated form of stathmin, which is predominant in the interphase of the cell cycle, promotes the depolymerization of microtubules by increasing the catastrophe frequency by sequestering tubulin.12 Phosphorylated stathmin, which increases during mitosis, results in a reduced affinity of stathmin to tubulin and increased microtubule stabilization, thus allowing the formation of mitotic spindle.15 Overexpression of stathmin has been observed in acute leukemia, lymphomas, neuroblastoma, prostatic adenocarcinoma, lung cancer, ovarian cancer, hepatocellular 0090-4295/09/$34.00 doi:10.1016/j.urology.2009.04.088

carcinoma, oral squamous cell carcinoma, and breast cancer.16-21 Most of these studies have even shown that expression of stathmin was associated with tumor behavior and prognosis. Associations between stathmin expression and clinicopathologic features of UC have not been studied yet. We have found high expression level of stathmin in 4 UC cell lines (3 bladder UCs: NTUB1, T24, TSGH8301; 1 UUT-UC: BFTCC905) by Western blot technique (see supplements). We herein investigate the expression of stathmin in UUT-UC tissue samples and clarify the role of stathmin in prediction of prognosis.

MATERIAL AND METHODS Study Population We retrospectively analyzed the medical records of 85 patients with the clinical diagnosis of UUT-UC initially treated by ipsilateral radical Nu and bladder cuff resection during 19982004. We excluded 10 patients with distant or lymph node metastasis, 7 patients with previous bladder UC, and 10 patients with the presence of concomitant bladder UC. The remaining 58 patients met the criteria of localized UUT-UC (pT1-3N0M0) were followed up postoperatively with cystoscopy and urine cytology every 3 months for 2 years, and thereafter every 6 months. CT and/or MRI and/or excretory urography were performed every 6 months for 2 years and annually thereafter. Adjuvant cisplatin-based chemotherapy was not routinely administered until evidence of recurrence was documented. Pathologic tumor (pT) stage of the primary UC was judged according to the 2002 American Joint of Cancer Committee classification system. Tumor grade was assigned according to the 1998 World Health Organization/International Society of Urological Pathology Consensus classification. The tumor grading and staging were re-evaluated by an experienced pathologist (W.C.L.). Disease recurrence included documented local recurrence and metastasis.

IHC Analysis Stathmin antibodies (rabbit polyclonal antibody) were commercially obtained (ab11269, 1:500 dilution, Abcam Inc, Cambridge, MA). Stathmin immunostaining was performed on 5-␮m sections of formalin-fixed, paraffin-embedded specimens using the labeled-strepatavidin-biotin method after retrieving antigen and applying chromogen of diaminobenzidine (DAB), as previously described.22 To assess the immunoreactivity quantitatively, the mean percentage of positive staining tumor cells was determined by counting at least 10 random fields at both 40⫻ and 400⫻ magnification in each section. The stathmin IHC score was defined as the proportion of positive staining tumor cells from each patient’s specimen for stathmin immunostaining. The stathmin IHC score ⱖ 0.5 was defined as strong (⫹) immunoreactivity and ⬍ 0.5 as weak (⫺) immunoreactivity. As a negative control, duplicate sections were immunostained without exposure to primary antibodies. A specimen from human tonsil tissue was used as a positive control. The results were analyzed by a pathologist who was unaware of the clinical data.

Reverse Transcription-Polymerase Chain Reaction Reverse transcription-polymerase chain reaction was used to determine the mRNA levels of stathmin in paired UUT-UC UROLOGY 74 (6), 2009

and nontumoral mucosa of the renal pelvis in another 5 patients with UUT-UC. Total RNA was extracted using Trizol reagent from the fresh specimens obtained after nephroureterectomy. S-26 ribosomal protein mRNA, a housekeeping gene, was used as the internal control. Polymerase chain reaction (PCR) was stopped at the exponential phase of amplification for each gene: 30 cycles for stathmin and 25 cycles for S-26. PCR was performed in an automatic DNA thermal cycler 480 (PerkinElmer, Co, Wellesley, MA), with initial heating at 94°C for 2 minutes followed by 30 cycles of 94°C for 30 seconds, 60°C for 1 minute, 72°C for 1 minute, and, finally, 72°C for 10 minutes for stathmin. Primers for stathmin were stathmin-F (5=-AAGGATTTTTCCCTGGAGGA-3=) and stathmin-R (5=-TTTCTCCCCTTTAGCCCCTA-3=). The primers for S-26 were S-26-F (5=-CCGTGCCTCCAAGATGACAAAG3=) and S-26-R (5=-GTTCGGTCCTTGCGGGCTTCAC-3=). After PCR, 5 ␮L of reaction products were electrophoresed on a 2% agarose gel. The stathmin mRNA expression level was determined by the ratio of signal intensity of stathmin to that of S-26 measured by 1-D image analysis Software (Kodak Digital Science, Rochester, NY).

Statistical Analysis Statistical analyses were performed with the SAS software (Version 9.1.3, SAS Institute, Cary, NC). In statistical testing, two-sided P ⱕ.05 was considered statistically significant. Continuous data were expressed as mean ⫾ standard deviation (SD) unless otherwise specified. Percentage was calculated for categorical variables. Two-sample t test or Wilcoxon rank-sum test was used to compare the means or medians of continuous data between 2 groups, whereas ␹2 test or Fisher exact test was used to analyze categorical proportions between 2 groups. The Kaplan–Meier estimate of survival curve and log-rank test was used to explore the association between stathmin immunoreactivity and patient’s survival. In addition to univariate analyses, multivariate analyses of age, sex, tumor site, pathologic stage, smoking, hemodialysis, residency in the BFD endemic area, histologic grading, status of lymphovascular invasion, stathmin IHC score, and strong (⫹) or weak (⫺) stathmin immunoreactivity were conducted by fitting multiple Cox proportional hazards model to predict a patient’s recurrence-free survival and cancer-specific survival. Basic model-fitting techniques for (1) variable selection, (2) goodness-of-fit assessment, and (3) regression diagnostics were used in regression analyses to ensure the quality of analysis results. In stepwise variable selection procedure, the univariate significant and nonsignificant covariates were all considered and both the significance levels for entry and significance levels for stay were set to 0.15 or greater.

RESULTS The median follow-up period was 58 months (interquartile range: 36-80). The association between clinicopathologic characteristics and the status of stathmin expression are listed in Table 1. The mean age was 64.6 ⫾ 12.7 years. There were 30 male and 28 female patients. The tumors were located in the kidney in 40 patients, ureter in 10, or both in 8 patients. Thirty-three patients had pT1 disease, 20 patients had pT2 disease, and the remaining 5 patients had T3 disease. The histologic grading 1265

Table 1. Correlations between stathmin immunoreactivity and clinicopathologic data Immunoreactivity of Stathmin Weak Strong (⫺) (⫹) No. Patients (%) Age (y) ⱖ 65 ⬍ 65 Sex M F Site Kidney Ureter Kidney ⫹ ureter Laterality Right Left pT stage T1 T2 T3-4 Grading High grade Low grade Lymphovascular invasion (–) (⫹)

58

P

36 (62.1) 22 (37.9)

.84 — — .34 30 (51.7) 19 (63.3) 11 (36.7) — 28 (48.3) 17 (60.7) 11 (39.3) — .38 40 (68.9) 24 (60) 16 (40) — 10 (17.2) 8 (80) 2 (20) — 8 (13.9) 4 (50) 4 (50) — 27 (46.6) 15 (55.6) 12 (44.4) 31 (53.4) 21 (67.8) 10 (32.2)

.20 — — .006 26 (78.8) 7 (21.2) — 9 (45) 11 (55) — 1 (20) 4 (80) — .09 21 (48.9) 19 (44.1) — 12 (80) 3 (20) — .41

30 (51.7) 21 (70) 9 (30) 28 (48.3) 15 (53.6) 13 (46.4) 33 (56.9) 20 (34.5) 5 (8.6) 43 (74.1) 15 (25.9)

46 (79.3) 32 (69.6) 14 (30.4) .01 12 (20.7) 4 (33.3) 8 (66.7) —

P ⬍.05 was defined as significant.

showed high grade in 43 patients and low grade in 15 patients. Of the 58 UUT-UC tumor specimens, 36 (62%) showed strong (⫹) and 22 (38%) were weak (⫺) immunoreactivity. Stathmin immunoreactivity was significantly associated with pT stage and histologic grading (P ⫽ .006 and .05, respectively). The positive immunoreactivity was localized in the cytoplasm (Fig. 1A and B). The median IHC score for UUT-UC specimen was 0.4 (interquartile range: 0.25-0.61). The median IHC score for nontumoral urothelium was 0 (interquartile range: 0-0.05). Significant difference was noted in the IHC score between the UUT-UC and nontumoral urothelium (P ⬍.001). The stathmin IHC score in advanced stages (pT2-pT3) was significantly higher than that in early stages (pT1) (P ⬍.05). The median follow-up period was 58 months (interquartile range: 36-80). The association between clinicopathologic characteristics and the status of stathmin expression are listed in Table 1. The mean age was 64.6 ⫾ 12.7 years. There were 30 male and 28 female patients. The tumors were located in the kidney in 40 patients, ureter in 10, or both in 8 patients. Thirtythree patients had pT1 disease, 20 patients had pT2 disease, and the remaining 5 patients had T3 disease. The histologic grading showed high grade in 43 patients and low grade in 15 patients. Of the 58 UUT-UC tumor specimens, 36 (62%) showed strong (⫹) and 22 (38%) showed weak (⫺) immunoreactivity. Stathmin immuno1266

reactivity was significantly associated with pT stage and histologic grading (P ⫽ .006 and .05, respectively). The positive immunoreactivity was localized in the cytoplasm (Fig. 1A and B). The median IHC score for UUT-UC specimen was 0.4 (interquartile range: 0.25-0.61). The median IHC score for nontumoral urothelium was 0 (interquartile range: 0-0.05). Significant difference was noted in the IHC score between the UUT-UC and nontumoral urothelium (P ⬍.001). The stathmin IHC score in advanced stages (pT2-pT3) was significantly higher than that in early stages (pT1) (P ⬍.05). The levels of stathmin mRNA in tumors and paired nontumoral mucosa of the renal pelvis are shown in Fig. 1C. The findings were matched to protein expression levels. The quantitative data are represented in Fig. 1D, with a significant difference (P ⫽ .003). Disease recurred in 17 patients (29.3%) at a median of 24 months (range: 2-106). The recurrence sites at initial relapse were in the tumor bed or lymph nodes in 10 patients and at distant sites (lung, bone, or liver) in 7 patients. Twenty patients (34.5%) developed bladder recurrence. Fourteen (24.1%) patients died of UUT-UC. The recurrence-free survival and cancer-specific survival curves in relation to stathmin immunoreactivity are demonstrated in Fig. 1E and F. Stathmin expression was significantly correlated with recurrence-free and cancerspecific survival by the log-rank test (P ⫽ .007 and .043, respectively). An unfavorable 5-year cancer-specific survival rate (63.3%) was found in patients with strong (⫹) immunoreactivity compared with the rate (86.8%) for the patients with weak (⫺) immunoreactivity. The recurrence-free survival rate between patients with strong (⫹) immunoreactivity and those with weak (⫺) immunoreactivity was 53% vs 90.4%, respectively (Table 2). The results of multivariate analyses using Cox proportional hazards model are listed in Table 3. Stathmin IHC score, pT stage, and ureteral involvement were significantly associated with the recurrence-free survival. Moreover, stathmin IHC score and pT stage were significantly associated with the cancer-specific survival, whereas ureteral involvement had a borderline significance (P ⫽ .0711). After adjusting for the effects of the other covariates, 1 U increase in stathmin IHC score (between 0 and 1) could increase the hazard rates to 22.4 times high for recurrence-free survival (95% CI: 3.53-141.53; P ⫽ .001) and to 39.8 times high for cancer-specific survival (95% CI: 4.29-368.91; P ⫽ .0012), respectively.

COMMENT In this study, IHC staining demonstrated significant differences in IHC scores between UUT-UC specimens and nontumoral urothelium. The stathmin IHC score was significantly associated with increasing pT stage and histologic grading. Moreover, stathmin immunoreactivity was statistically correlated with unfavorable cancer-specific and recurrence-free survival. In UROLOGY 74 (6), 2009

Figure 1. Immunohistochemical staining of stathmin in normal urothelium and UUT-UC (A) Strong (⫹) immunoreactivity (ⱖ 50% of the tumor cells were positive for stathmin; IHC score ⫽ 0.85) with a sclerotic glomerulus in the renal parenchyma, ⫻100. (B) Weak (⫺) immunoreactivity (⬍ 50% of the tumor cells were positive for stathmin; IHC score ⫽ 0.4) and the regional urothelial cells were negative, ⫻40. Stathmin intracellular localization was typically diffuse in the cytoplasm of cancer cells. (C) Quantitative level of stathmin mRNA (expressed in %: Stathmin/S-26) in UUT-UC and normal urothelial mucosa of the renal pelvis. The relative expression showed significant difference between UUT-UC and normal urothelial mucosa (P ⫽ .003). (D) RT-PCR in the linear range of amplification showed stathmin mRNA overexpression in UUT-UC compared with normal urothelial mucosa of the renal pelvis in patients with UUT-UC. Kaplan–Meier analysis for (E). Recurrence-free survival (F). Cancer-specific survival in patients with UUT-UC related to stathmin immunoreactivity [strong (⫹) vs weak (⫺) (log-rank test: P ⫽ .007 and .043, respectively].

multivariate analysis, stathmin IHC score was an independent factor to predict recurrence-free and cancer-specific survival. All the above findings suggest that stathmin is a novel prognostic marker in localized UUT-UC. In the present study, we observed positive immunoreactivity of stathmin in all specimens. The IHC score of UUT-UC was significantly higher than that of nontumoral urothelium. The IHC score is also significantly UROLOGY 74 (6), 2009

higher in advanced pathologic stage (pT2-pT3). The stathmin IHC score was proportional to the probability of disease recurrence. These results are compatible with univariate and multivariate results. The IHC score was graded by cut-off point of 0.5 to compare the differences in oncological outcomes between strong (⫹) and weak (⫺) stathmin staining. The sensitivity for predicting disease recurrence was 45% and specificity was 89% in the cut-off point. 1267

Table 2. Univariate analysis of multiple prognostic variables for 5-year recurrence-free and cancer-specific survival

Variables

5-Year Recurrence-Free Survival Survival Rate (%) Log-Rank P

Age (y) ⬍ 65 ⱖ 65 Gender M F Ureteral involvement No Yes Laterality Right Left pT stage T1 T2 T3 Grading Low grade High grade Stathmin immunoreactivity Strong (⫹) Weak (⫺)

5-year Cancer-Specific Survival Survival Rate (%) Log-Rank P

.99

.39

79.0 72.9

80.5 63.2 .12

.11

62.2 84.4

66.5 85.3 .08

.09

77.3 68.2

75.1 71.6 .11

.29

67.4 81

65.9 81.6 .03

.04

86.9 61.8 30.0

82.8 66.1 30.0 .18

.26

80.8 72.0

74.0 71.2 .004

.011

89.5 53

86.3 57.6

Table 3. Multivariate analyses of the risk factors associated with recurrence-free survival and cancer-specific survival in patients with localized upper tract urothelial carcinoma using Cox proportional hazards model (n ⫽ 58) Variable Recurrence-free survival Stathmin IHC score pT stage Ureteral involvement Cancer-specific survival Stathmin IHC score pT stage Ureteral involvement

Estimate

Standard Error

␹2

P

Hazard Ratio

95% CI

3.1075 1.3002 1.2479

0.9414 0.6001 0.6203

10.8968 4.6941 4.0472

.0010 .0303 .0442

22.364 3.670 3.483

3.53-141.53 1.13-11.90 1.03-11.75

3.6839 1.8383 1.3067

1.1361 0.7687 0.7241

10.5149 5.7192 3.2567

.0012 .0168 .0711

39.802 6.286 3.694

4.29-368.91 1.39-28.36 0.89-15.27

Stathmin IHC score ⫽ the proportion of positive staining tumor cells from each patient’s specimen for stathmin immunostaining. The adjusted generalized R 2 was 0.279 for recurrence-free survival and 0.316 for cancer-specific survival, which indicated fair fits.

In IHC, most of the specimens showed homogenous staining. Nuclear stain is noted in some tumor cells with cytosolic stain but almost no pure nuclear staining could be seen. Staining of normal urothelium was mainly at intermediate and basal cell layers and less at superficial cells. For cases of multifocal tumors, we selected the more invasive part for staining. Recent studies have shown that inhibition of stathmin expression in malignant cells interferes with cell cycle progression and abrogates their transformed phenotype.23,24 Inhibition of stathmin has also been reported to have synergistic effects on chemotherapeutic agents including vinca alkaloids and paclitaxel.25 The main chemotherapeutic agents in metastatic UC are cisplatin, paclitaxel, gemcitabine, and vinca alkaloids.26 The combination effect between chemotherapy and anti-stathmin deserve further investigation. Stathmin may not only serve as a promising marker for predicting prognosis but provide an attractive molecular target for disrupting tumor proliferation. 1268

CONCLUSIONS This is the first systematic and comprehensive analysis of stathmin expression in UCs. Our results demonstrated that stathmin is highly expressed in UUT-UC. The level of stathmin expression was a significant factor to predict tumor recurrence and survival in patients with localized UUT-UC. The mechanism and clinical applications must be elucidated by further studies. Acknowledgments. The authors thank Ling-Chu Wu for her assistance in statistical computing.

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4. Hall MC, Womack S, Sagalowsky AI, et al. Prognostic factors, recurrence, and survival in transitional cell carcinoma of the upper urinary tract: a 30-year experience in 252 patients. Urology. 1998; 52:594-601. 5. Novara G, De Marco V, Gottardo F, et al. Independent predictors of cancer-specific survival in transitional cell carcinoma of the upper urinary tract: multi-institutional dataset from 3 European centers. Cancer. 2007;110:1715-1722. 6. Olgac S, Mazumdar M, Dalbagni G, et al. Urothelial carcinoma of the renal pelvis: a clinicopathologic study of 130 cases. Am J Surg Pathol. 2004;28:1545-1552. 7. Lin WC, Hu FC, Chung SD, et al. The role of lymphovascular invasion in predicting the prognosis of clinically localized upper tract urothelial carcinoma (pT1-3cN0M0). J Urol. 2008;180:879884. 8. Park S, Hong B, Kim CS, et al. The impact of tumor location on prognosis of transitional cell carcinoma of the upper urinary tract. J Urol. 2004;171(2 Pt. 1):621-625. 9. Saito K, Kawakami S, Fujii Y, et al. Lymphovascular invasion is independently associated with poor prognosis in patients with localized upper urinary tract urothelial carcinoma treated surgically. J Urol. 2007;178:2291-2296. 10. Fromont G, Roupret M, Amira N, et al. Tissue microarray analysis of the prognostic value of E-cadherin, Ki67, p53, p27, survivin and MSH2 expression in upper urinary tract transitional cell carcinoma. Eur Urol. 2005;48:764-770. 11. Langner C, Rupar G, Leibl S, et al. Alpha-methylacyl-CoA racemase (AMACR/P504S) protein expression in urothelial carcinoma of the upper urinary tract correlates with tumour progression. Virchows Arch. 2006;448:325-330. 12. Leibl S, Zigeuner R, Hutterer G, et al. EGFR expression in urothelial carcinoma of the upper urinary tract is associated with disease progression and metaplastic morphology. APMIS. 2008;116:27-32. 13. Ferrari AC, Seuanez HN, Hanash SM, et al. A gene that encodes for a leukemia-associated phosphoprotein (p18) maps to chromosome bands 1p35-36.1. Genes Chromosomes Cancer. 1990;2:125129. 14. Rubin CI, Atweh GF. The role of stathmin in the regulation of the cell cycle. J Cell Biochem. 2004;93:242-250. 15. Cassimeris L. The oncoprotein 18/stathmin family of microtubule destabilizers. Curr Opin Cell Biol. 2002;14:18-24. 16. Yuan RH, Jeng YM, Chen HL, et al. Stathmin overexpression cooperates with p53 mutation and osteopontin overexpression, and is associated with tumour progression, early recurrence, and poor prognosis in hepatocellular carcinoma. J Pathol. 2006;209:549-558. 17. Chen G, Wang H, Gharib TG, et al. Overexpression of oncoprotein 18 correlates with poor differentiation in lung adenocarcinomas. Mol Cell Proteomics. 2003;2:107-116. 18. Kouzu Y, Uzawa K, Koike H, et al. Overexpression of stathmin in oral squamous-cell carcinoma: correlation with tumour progression and poor prognosis. Br J Cancer. 2006;94:717-723. 19. Saal LH, Johansson P, Holm K, et al. Poor prognosis in carcinoma is associated with a gene expression signature of aberrant PTEN tumor suppressor pathway activity. Proc Natl Acad Sci USA. 2007; 104:7564-7569. 20. Ghosh R, Gu G, Tillman E, et al. Increased expression and differential phosphorylation of stathmin may promote prostate cancer progression. Prostate. 2007;67:1038-1052. 21. Wei SH, Lin F, Wang X, et al. Prognostic significance of stathmin expression in correlation with metastasis and clinicopathological characteristics in human ovarian carcinoma. Acta Histochem. 2008; 110:59-65. 22. Hsu HC, Jeng YM, Mao TL, et al. Beta-catenin mutations are associated with a subset of low-stage hepatocellular carcinoma negative for hepatitis B virus and with favorable prognosis. Am J Pathol. 2000;157:763-770. 23. Nishio K, Nakamura T, Koh Y, et al. Oncoprotein 18 overexpression increases the sensitivity to vindesine in the human lung carcinoma cells. Cancer. 2001;91:1494-1499.

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24. Mistry SJ, Bank A, Atweh GF. Synergistic antiangiogenic effects of stathmin inhibition and taxol exposure. Mol Cancer Res. 2007;5: 773-782. 25. Iancu C, Mistry SJ, Arkin S, et al. Taxol and anti-stathmin therapy: a synergistic combination that targets the mitotic spindle. Cancer Res. 2000;60:3537-3541. 26. Gallagher DJ, Milowsky MI, Bajorin DF. Advanced bladder cancer: status of first-line chemotherapy and the search for active agents in the second-line setting. Cancer. 2008;113:1284-1293.

APPENDIX SUPPLEMENTARY

DATA

Supplementary data associated with this article can be found, in the online version, at doi:10.1016/j.urology.2009.04.088.

EDITORIAL COMMENT Tumor stage and grade have been established as significant prognostic factors in the upper urinary tract urothelial carcinoma (UUTUC).1 Stathmin 1 (also known as oncoprotein 18) is the first discovered member of a family of phylogenetically related microtubule-destabilizing phosphoproteins critically involved in the construction and function of the mitotic spindle. It is overexpressed across a broad range of human malignancies (leukemia; lymphoma; neuroblastoma; ovarian, prostatic, breast and lung cancers; and mesothelioma).2 Authors of the present study assessed the immunohistochemical expression of stathmin in UUTUC samples from 58 patients who had undergone nephroureterectomy and determined whether it had any correlation with prognosis. They found that the stathmin expression was correlated with pathological stage, recurrence-free survival, and cancer-specific survival. These findings were also confirmed at multivariate analysis. Thus, stathmin immunostaining is advocated by the authors as a novel prognostic factor for patients with localized UUTUC.3 We note some methodological drawbacks related to this study: the retrospective design, the limited sample including patients with variable features (both pT1 and pT2-pT3 disease, both high- and low-risk tumors), the use of nontumoral urothelium specimens obtained from the same patients as controls for RT-PCR assessment, the slightly arbitrary definition of strong/ weak immunoreactivity, low sensitivity (45%) of the selected cut-off (0.5) for immunostaining score. Besides its potential role in the definition of candidates for any adjuvant therapy, it would be also interesting to see whether stathmin expression might be helpful in identifying patients amenable to conservative treatment. Of course, before addressing these further questions, external validation of the findings from the current study, on a larger sample, is awaited. Overall, the search for reliable prognosticators in the field UUTUC remains a challenging task for current clinical research. Riccardo Autorino, M.D., Ph.D., F.E.B.U., Urology Clinic, Second University of Naples, Naples, Italy Gianluca Giannarini, M.D., Ph.D., Department of Urology, University of Pisa, Pisa, Italy

References 1. Zigeuner R, Pummer K. Urothelial carcinoma of the upper urinary tract: surgical approach and prognostic factors. Eur Urol. 2008;53: 720-731.

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