Walburgis Brenner

Loss of tumor suppressor protein PTEN during renal carcinogenesis

The tumor suppressor gene PTEN (phosphatase and tensin homologue deleted from chromosome 10) encodes a dual specific protein and phospholipid phosphatase that affects cell proliferation, apoptosis and migration. In our study, we examined protein expression of PTEN in renal carcinogenesis. PTEN protein levels were examined in 42 clear cell renal cell carcinomas (ccRCC) and oncocytomas as well as in the corresponding normal renal tissue of the same patients using Western blot analysis. Cellular localization was analyzed by immunohistochemistry. PTEN was highly expressed in all investigated normal renal tissue specimens. Immunohistochemical analysis showed an almost exclusive staining of proximal tubulus epithelial cells known to be precursor cells of ccRCC. Within the proximal tubulus cells, PTEN exhibited a membrane predominant immunostaining pattern. In ccRCCs PTEN expression was markedly reduced to an average of less than 10% compared with normal tissue as evidenced by Western blot analysis (p < 0.001). The degree of reduction was similar in highly differentiated (G1) carcinomas and in less differentiated (G2–G4) carcinomas. These observations were reproduced by immunohistochemical studies, which revealed a loss of the characteristic membrane predominant immunostaining pattern in ccRCC. In contrast to the PTEN positive proximal tubulus epithelial cells, the distal tubulus epithelial cells, which are precursor cells of the benign oncocytomas, exhibited only a very weak PTEN expression. Compared with the distal tubulus epithelial cells, no downregulation of PTEN was seen in oncocytomas. We conclude that PTEN expression and PTEN membrane localization are lost during early renal cell carcinogenesis and may therefore be a valuable RCC tumor marker.

Identification of metabolic enzymes in renal cell carcinoma utilizing PROTEOMEX analyses.

PROTEOMEX, an approach which combines conventional proteome analysis with serological screening, is a powerful tool to separate proteins and identify immunogenic components in malignant diseases. By applying this approach, we characterized nine metabolic enzymes which were differentially expressed in renal cell carcinoma (RCC) cell lines and compared their expression profiles to that of normal kidney epithelium cells. Four of these proteins, superoxide dismutase (SODC), triosephosphatase isomerase (TPIS), thioredoxin (THIO) and ubiquitin carboxyl-terminal hydrolase (UBL1) were further analysed for both their constitutive and interferon (IFN)-gamma inducible protein expression pattern in cell lines or tissue specimens derived from RCC or normal kidney epithelium using Western blot analysis and immunohistochemistry, respectively. With the exception of the RCC cell line MZ1940RC, which completely lacks the expression of UBL1, a heterogeneous and variable expression pattern of the different metabolic enzymes was detected in RCC and normal renal epithelium. The highest differences in the expression levels were found for THIO in the RCC cell lines, which was 2-fold upregulated when compared to autologous normal kidney epithelium. Moreover, IFN-gamma treatment did not influence the constitutive expression of these metabolic enzymes. Thus, PROTEOMEX represents a valuable approach for the identification of metabolic enzymes which might be used as markers for the diagnosis of RCC.

Targeting of tumor associated antigens in renal cell carcinoma using proteome‐based analysis and their clinical significance

The suitability of proteome‐based strategies for the targeting of tumor‐associated markers along with further analysis regarding their clinical significance were investigated in human renal cell carcinoma (RCC). The immunogenic protein expression profile of normal kidney and RCC cell lines was studied by proteome analysis combined with immunoblotting using sera from healthy donors and RCC patients, also termed PROTEOMEX. Employing this approach, a series of proteins reactive with either RCC patient sera and/or reactive with control sera were identified by microanalysis of tryptic peptides. Some of these candidate antigens represent members of the cytoskeletal family, such as cytokeratins, in particular cytokeratin 8, cytoskeletal tropomyosin, F‐actin capping protein, γ‐actin, stathmin, tubulin‐α, tubulin‐β and vimentin. The expression pattern and clinical significance of three of these antigens, namely cytokeratin 8, stathmin and vimentin, were further analyzed in a large series of surgically removed RCC lesions of distinct subtypes. A heterogeneous expression pattern of cytokeratin 8, stathmin and vimentin was demonstrated in the different RCC subtypes. All epithelial cells of the autologous normal kidney showed a strong cytokeratin 8 staining pattern, whereas they totally lack vimentin expression. Stathmin was expressed in 10% of tubule cells. In conclusion, PROTEOMEX could be employed for the identification of tumor‐associated antigens of the cytoskeleton which are differentially expressed in RCC of distinct subtypes as well as in normal renal epithelium.

Short-Term Functional and Oncologic Outcomes of Nephron-Sparing Surgery for Renal Tumours ≥7 cm

BACKGROUND Nephron-sparing surgery (NSS) for renal tumours preserves renal function and has become the standard approach for small renal tumours. Little is known about perioperative and oncologic outcomes of patients following NSS in renal tumours ≥ 7 cm in the presence of a healthy contralateral kidney. OBJECTIVE To analyse oncologic outcomes and perioperative morbidity in patients treated by NSS for renal tumours ≥ 7 cm. DESIGN, SETTING, AND PARTICIPANTS In total, 5767 patients were treated for renal tumours at two institutions from 1984 to 2009. In 91 patients, elective NSS was performed for renal tumours ≥ 7 cm. MEASUREMENTS Complication rates were assessed in detail and stratified using the Clavien-Dindo score (CDS). Oncologic outcomes for overall survival (OS), cancer-specific survival (CSS), and progression-free survival (PFS) were estimated using the Kaplan-Meier method. Logistic regression analysis was used to identify clinical risk factors for complications and prognosticators that have an oncologic impact on OS. RESULTS AND LIMITATIONS The median follow-up was 28 mo (range: 1-247 mo). Twenty-seven patients (29.6%) had perioperative complications and, of these, 89.1% had CDS grade 1 and 2. Twenty-seven percent of the 91 patients had benign lesions. Seven patients (10.6%) died from cancer-related causes. The 5- and 10-yr rates for OS, CSS, and PFS were 88% and 64%, 97% and 83%, and 91% and 78%, respectively. None of the analysed parameters had an impact on morbidity or OS in the univariate analysis. Limitations of this study were its retrospective nature and the relatively short follow-up period for oncologic outcome. CONCLUSIONS NSS for renal tumours ≥ 7 cm can be performed with acceptable complication rates and with oncologic outcomes comparable to radical nephrectomy studies. Our findings support NSS whenever technically feasible to reduce the loss of renal function.

Heat shock protein expression and anti-heat shock protein reactivity in renal cell carcinoma.

Heat shock proteins (HSP) are families of highly conserved proteins which are induced in cells and tissues upon exposure to extreme conditions causing acute or chronic stress. They exhibit distinct functions and have been implicated in the pathogenesis of a number of diseases, including cancer. A causal relationship between HSP expression and immunogenicity has been demonstrated in murine and human tumors and is also associated with the immune response. In order to investigate the correlation of HSP expression and their immunogenic potential in renal cell carcinoma (RCC), we here analyzed (i) the protein expression profile of various members of the HSP family in untreated and interferon (IFN)‐γ treated RCC cell lines as well as normal kidney epithelium, and (ii) the anti‐heat shock protein reactivity in sera derived from RCC patients and healthy controls using proteomics‐based techniques. A heterogeneous expression pattern of members of the HSP families was demonstrated in RCC cell lines and in cells representing normal renal epithelium. In some cases the expression rate is moderately altered by IFN‐γ treatment. In addition, a distinct anti‐heat shock protein reactivity could be detected in autologous and allogeneic sera from RCC patients and healthy controls. These data suggest that HSP play a role in the immunogenicity of RCC and thus might be used for the design of immunization strategies to induce a potent antitumor response in this disease.

ERBB2-Mediated Transcriptional Up-regulation of the α5β1 Integrin Fibronectin Receptor Promotes Tumor Cell Survival Under Adverse Conditions

Oncogenic activation of the receptor tyrosine kinase ERBB2 is a key event in the development of a number of epithelial malignancies. In these tumors, high levels of ERBB2 are strongly associated with metastatic disease and poor prognosis. Paradoxically, an inherent cellular response to hypermitogenic signaling by ERBB2 and other oncogenes seems to be growth arrest, rather than proliferation. Molecular characterization of this yet undefined antiproliferative state in independent cell lines overexpressing either wild-type ERBB2 or the mutationally activated receptor unveiled a dramatic induction of the α5β1 integrin fibronectin receptor. α5 Integrin up-regulation is mainly a transcriptional response mediated by the hypoxia-inducible transcription factors (HIF), leading to a massive increase in membrane-resident receptor molecules and enhanced fibronectin adhesiveness of the respective cells. Functionally, ERBB2-dependent ligation of fibronectin results in improved survival of mammary adenocarcinoma cells under adverse conditions, like serum withdrawal, hypoxia, and chemotherapy. HIF-1α is an independent predictor of poor overall survival in patients with breast cancer. In particular, HIF-1α overexpression correlates significantly with early local relapse and distant metastasis, a phenotype also highly characteristic of ERBB2-positive tumors. As HIF-1α is known to be stabilized by ERBB2 signaling under normoxic conditions, we propose that α5 integrin is a major effector in this regulatory circuit and may represent the molecular basis for the HIF-1α-dependent aggressiveness observed in ERBB2-overexpressing breast carcinomas. Hypermitogenic ERBB2 signaling and tumor hypoxia may act synergistically to favor the establishment of chemoresistant dormant micrometastatic cells frequently observed in patients with breast cancer. This new insight could be the basis for additional approaches complementing current cancer therapy. (Cancer Res 2006; 66(7): 3715-25)

Genotyping NAT2 with only two SNPs (rs1041983 and rs1801280) outperforms the tagging SNP rs1495741 and is equivalent to the conventional 7-SNP NAT2 genotype.

Genotyping N-acetyltransferase 2 (NAT2) is of high relevance for individualized dosing of antituberculosis drugs and bladder cancer epidemiology. In this study we compared a recently published tagging single nucleotide polymorphism (SNP) (rs1495741) to the conventional 7-SNP genotype (G191A, C282T, T341C, C481T, G590A, A803G and G857A haplotype pairs) and systematically analysed if novel SNP combinations outperform the latter. For this purpose, we studied 3177 individuals by PCR and phenotyped 344 individuals by the caffeine test. Although the tagSNP and the 7-SNP genotype showed a high degree of correlation (R=0.933, P<0.0001) the 7-SNP genotype nevertheless outperformed the tagging SNP with respect to specificity (1.0 vs. 0.9444, P=0.0065). Considering all possible SNP combinations in a receiver operating characteristic analysis we identified a 2-SNP genotype (C282T, T341C) that outperformed the tagging SNP and was equivalent to the 7-SNP genotype. The 2-SNP genotype predicted the correct phenotype with a sensitivity of 0.8643 and a specificity of 1.0. In addition, it predicted the 7-SNP genotype with sensitivity and specificity of 0.9993 and 0.9880, respectively. The prediction of the NAT2 genotype by the 2-SNP genotype performed similar in populations of Caucasian, Venezuelan and Pakistani background. A 2-SNP genotype predicts NAT2 phenotypes with similar sensitivity and specificity as the conventional 7-SNP genotype. This procedure represents a facilitation in individualized dosing of NAT2 substrates without losing sensitivity or specificity.

Single Center Comparison of Anastomotic Strictures After Radical Perineal and Radical Retropubic Prostatectomy

OBJECTIVES To analyze the incidence and management of anastomotic strictures (ASs) after radical perineal prostatectomy (RPP) and retropubic prostatectomy (RRP) and to identify possible predisposing factors. METHODS Between 1997 and 2007, we performed 866 RPP and 2052 RRP for localized prostate cancer. Median follow-up was 52 months (12-136). We analyzed preoperative serum prostate-specific antigen, prostate size, clinical and pathologic tumor stage, neoadjuvant hormone deprivation, previous transurethral resection of the prostate, transfusion requirement, anastomotic insufficiency, and acute urinary retention (AUR) and its subsequent management to identify possible predisposing factors for AS formation. RESULTS The rate of AS after RPP and RRP was 3.8% (33/863) and 5.5% (113/2048), respectively (P = .067). In multivariate analysis, RRP was a statistically significant risk factor for AS (P = .0002). On survival analysis, the incidence of AS was lower for RPP as compared with RRP at median follow-up (P = .0229). Primary response to endoscopic AS incision or resection was 94% (31/33) and 72.6% (82/113) after RPP and RRP, respectively. On multivariate logistic regression analysis biopsy Gleason score, previous transurethral resection of the prostate, prostate volume, pathologic tumor stage and grade, transfusion requirement, AUR, and surgical technique were independent risk factors for the development of AS. An AS developed in 45.4% (20/44) and 10.9% (5/46) of the postoperative AUR cases treated with a suprapubic cystostomy tube and a transurethral Foley catheter, respectively (P <.05). CONCLUSIONS ASs occur more frequently after RRP in comparison with RPP. Primary endoscopic AS incision or resection are both highly successful. Treating postoperative AUR with a suprapubic cystostomy poses a high risk for AS formation and should be avoided.

Oncologic Long-term Outcome of Elective Nephron-sparing Surgery Versus Radical Nephrectomy in Patients With Renal Cell Carcinoma Stage pT1b or Greater in a Matched-pair Cohort

OBJECTIVES To analyze the oncologic outcome and overall survival (OS) for patients with renal cell carcinoma (RCC) >4 cm undergoing radical nephrectomy (RN) or elective nephron-sparing surgery (NSS) in a matched-pair cohort. METHODS From 1988 to 2007, we identified 829 patients in our clinic treated with either RN (n = 641) or open NSS (n = 188) for renal masses >4 cm. After matching the cohort for age, time of surgery, grade, TNM stage, tumor size, and sex and excluding patients with metastases, benign lesions with an imperative indication, and those with missing records, 173 remained for oncologic analysis. OS, cancer-specific survival, and progression-free survival were estimated using the Kaplan-Meier method. The association with death was evaluated with Cox proportional hazards regression analysis. RESULTS At the last follow-up visit, 39 patients had died of any cause and 134 were alive at a median of 7.0 years. RN and elective NSS had been performed in 100 and 73 patients, respectively. The OS (P = .357), progression-free survival (P = .558), and cancer-specific survival (P = .239) were not significantly different between the elective NSS and RN groups using the Kaplan-Meier method. On univariate and multivariate Cox regression analysis, the type of surgery did not have an effect on OS (hazard ratio 1.35, 95% confidence interval 0.71-2.54, P = .359). CONCLUSIONS Our results suggest that it is oncologically safe to perform NSS for renal tumors >4 cm, for which the surgical feasibility according to the tumor location, rather than the tumor size, seemed to be the limiting factor.

Perioperative morbidity and renal function in young and elderly patients undergoing elective nephron‐sparing surgery or radical nephrectomy for renal tumours larger than 4 cm

Study Type – Therapy (case series) Level of Evidence 4