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Item does not contain fulltext, OBJECTIVES: To explore whether urinary cytokine and chemokine (CK) levels differed between cold mitomycin-C (cold-MMC)-treated patients and chemohyperthermia (C-HT)-treated patients, to shed light on the possible molecular mechanisms that might explain the superior outcome of C-HT. Furthermore, CK-differences were explored between C-HT responders and C-HT non-responders. METHODS: Twelve NMIBC patients were included. Nine received six-weekly C-HT, and three received four-weekly cold-MMC instillations. Urine was collected on 8-12 time points before and after every treatment. MDC, IL-2, IL-6, IL-8, IP-10, MCP-1 and RANTES were determined by Luminex((R))-analysis. RESULTS: Elevated urinary CK levels were observed in both groups after treatment. In general, CK-peaks were lower in the cold-MMC group in comparison with levels in the C-HT group. Significant higher MCP-1 and IL-6 levels were observed in C-HT-treated patients. Additionally, significant cumulative effects were observed for IP-10 and IL-2. However, IP-10 and IL-2 levels did not significantly differ between treatments. MDC levels after the first week of treatment were significantly higher in the C-HT responders compared with the non-responders. CONCLUSION: MMC treatment leads to elevated urinary CK levels with significantly higher MCP-1 and IL-6 levels in C-HT-treated patients. Increased MDC levels after the first C-HT instillation appear to be related to good clinical outcome and might be of additional value to personalize treatment. Studies involving more patients and longer follow-up are needed to substantiate this observation.

Contains fulltext : 154027.pdf (publisher's version ) (Open Access), Anti-angiogenic treatment with tyrosine kinase inhibitors (TKI) has lead to an impressive increase in progression-free survival for patients with metastatic RCC (mRCC), but mRCC remains largely incurable. We combined sunitinib, targeting the endothelial cells with Girentuximab (monoclonal antibody cG250, recognizing carbonic anhydrase IX (CAIX) targeting the tumor cells to study the effect of sunitinib on the biodistribution of Girentuximab because combination of modalities targeting tumor vasculature and tumor cells might result in improved effect. Nude mice with human RCC xenografts (NU12, SK-RC-52) were treated orally with 0.8 mg/day sunitinib, or vehicle for 7 to 14 days. Three days before start or cessation of treatment mice were injected i.v. with 0.4 MBq/5 mug (111)In-Girentuximab followed by biodistribution studies. Immunohistochemical analyses were performed to study the tumor vasculature and CAIX expression and to confirm Girentuximab uptake. NU12 appeared to represent a sunitinib sensitive tumor: sunitinib treatment resulted in extensive necrosis and decreased microvessel density (MVD). Accumulation of Girentuximab was significantly decreased when sunitinib treatment preceded the antibody injection but remained unchanged when sunitinib followed Girentuximab injection. Cessation of therapy led to a rapid neovascularization, reminiscent of a tumor flare. SK-RC-52 appeared to represent a sunitinib-resistant tumor: (central) tumor necrosis was minimal and MVD was not affected. Sunitinib treatment resulted in increased Girentuximab uptake, regardless of the sequence of treatment. These data indicate that sunitinib can be combined with Girentuximab. Since these two modalities have different modes of action, this combination might lead to enhanced therapeutic efficacy.

Item does not contain fulltext, Tyrosine kinase inhibitors (TKIs) have revolutionized the treatment of metastatic clear cell renal cell carcinoma (RCC). Although TKIs have demonstrated good clinical efficacy, the lack of complete responses, the chronic nature of the treatment, and the side effects are clear disadvantages. An interesting new approach in the treatment of clear cell RCC is antibody-mediated therapy with the chimeric anti-carbonic anhydrase IX (CAIX) antibody girentuximab (cG250). As the results of several girentuximab trials become available, the question arises of whether TKI treatment can be combined with girentuximab-based therapy. In this study, we assessed the effect of the widely used TKI sorafenib on the tumor-targeting potential of (111)In-labeled girentuximab. METHODS: (111)In-girentuximab imaging was performed on 15 patients suspected of having a renal malignancy, with surgery being part of their treatment plan. Of these, 10 patients were treated in a neoadjuvant setting with sorafenib (400 mg orally twice daily). Five patients received treatment during 1 wk, and 5 patients received treatment during 4 wk. In both sorafenib-treated groups, baseline and posttreatment tumor targeting of (111)In-girentuximab were compared. Surgery was performed 3 d after the last image acquisition. Five additional patients were included as a control group and had only a single (111)In-girentuximab injection and scintigraphy without any treatment. Distribution of (111)In-girentuximab was determined scintigraphically ex vivo in a 1-cm lamella of the resected tumorous kidney. Expression of CAIX and of the vascular marker CD31 was determined immunohistochemically on specimens of both tumor and normal kidney tissue. RESULTS: Treatment with sorafenib resulted in a marked decrease of (111)In-girentuximab uptake in the tumor in clear cell RCC patients, especially in the group treated for 4 wk (mean change in both sorafenib-treated groups, -38.4%; range, +9.1% to -79.4%). Immunohistochemical analysis sh

Contains fulltext : 58051.pdf (publisher's version ) (Closed access), Renal cell carcinoma of the clear cell type (ccRCC) is associated with loss of functional von Hippel-Lindau (VHL) protein and high, homogeneous expression of the G250MN protein, an isoenzyme of the carbonic anhydrase family. High expression of G250MN is found in all ccRCCs, but not in most normal tissues, including normal human kidney. We specifically studied the mechanism of transcriptional regulation of the CAIXG250 gene in RCC. Previous studies identified Sp1 and hypoxia-inducible factor (HIF) as main regulatory transcription factors of G250MN in various non-RCC backgrounds. However, G250MN regulation in RCC has not been studied and may be differently regulated in view of the HIF accumulation under normoxic conditions due to VHL mutations. Transient transfection of different G250MN promoter constructs revealed strong promoter activity in G250MN -positive RCC cell lines, but no activity in G250MN -negative cell lines. DNase-I footprint and band-shift analysis demonstrated that Sp1 and HIF-1alpha proteins in nuclear extracts of RCC cells bind to the CAIX promoter and mutations in the most proximal Sp1 binding element or HIF binding element completely abolished CAIX promoter activity, indicating their critical importance for the activation of G250 expression in RCC. A close correlation between HIF-1alpha expression and G250MN expression was observed. In contrast, no relationship between HIF-2alpha expression and G250MN was seen. The participation of cofactor CBP/p300 in the regulation of G250 transcription was shown. In conclusion, HIF-1alpha and Sp1, in combination with CBP/p300, are crucial elements for G250MN expression in ccRCC, and CAIXG250 can be regarded as a unique HIF-1alpha target gene in ccRCC.

Item does not contain fulltext, OBJECTIVES: In renal cell carcinoma (RCC) cell lines, expression of the RCC-associated antigen G250 correlates with hypomethylation of the investigated CpG dinucleotides in the G250 promoter region, despite the absence of a CpG island. To gain insight into the molecular mechanism leading to G250 expression in vivo, we ascertained whether this correlation between G250 gene expression and the methylation status of the G250 gene also existed in primary RCC and normal kidney tissue. METHODS: G250 mRNA and protein expression was determined by reverse transcriptase-polymerase chain reaction, fluorescence activated cell sorting analysis, and immunohistochemistry in 15 RCC cell lines and 13 paired primary RCC/normal kidney tissue specimens. The methylation status of the G250 gene was determined by bisulfite genomic sequencing. RESULTS: RCC cell lines revealed a clear correlation between G250 expression and hypomethylation. In contrast, no hypomethylation was observed in primary RCC compared with normal kidney tissue. The CpG dinucleotides investigated were generally completely methylated in RCC, as well as in normal kidney tissue. Furthermore, a primary culture of RCC tissue revealed increasing hypomethylation of the G250 gene in successive passages, suggesting that the G250 hypomethylation observed in vitro is tissue culture induced. CONCLUSIONS: The methylation status of the G250 gene correlated with G250 expression in vitro but not in vivo.

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Contains fulltext : 25024___.PDF (publisher's version ) (Open Access)

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