ABSTRACT
Clear cell renal cell carcinoma (CC-RCC) is the most lethal of all genitourinary cancers. The functional loss of the von Hippel-Lindau (VHL) gene occurs in 90% of CC-RCC, driving cancer progression. The objective of this study was to identify chemical compounds that are synthetically lethal with VHL deficiency in CC-RCC. An annotated chemical library, the library of pharmacologically active compounds (LOPAC), was screened in parallel on VHL-deficient RCC4 cells and RCC4VHL cells with re-introduced VHL. The ROCK inhibitor, Y-27632, was identified and validated for selective targeting of VHL-deficient CC-RCC in multiple genetic backgrounds by clonogenic assays. Downregulation of ROCK1 by small interfering RNA (siRNA) selectively reduced the colony-forming ability of VHL-deficient CC-RCC, thus mimicking the effect of Y-27632 treatment, whereas downregulation of ROCK2 had no effect. In addition, two other ROCK inhibitors, RKI 1447 and GSK 429286, selectively targeted VHL-deficient CC-RCC. CC-RCC treatment with ROCK inhibitors is cytotoxic and cytostatic based on bromodeoxyuridine (BrdU) assay, propidium iodide (PI) staining and growth curves, and blocks cell migration based on transwell assay. On the one hand, knockdown of hypoxia-inducible factor (HIF) ß in the VHL-deficient CC-RCC had a protective effect against Y-27632 treatment, mimicking VHL reintroduction. On the other hand, CC-RCCVHL cells were sensitized to Y-27632 treatment in hypoxia (2% O2). These results suggest that synthetic lethality between ROCK inhibition and VHL deficiency is dependent on HIF activation. Moreover, HIF1α or HIF2α overexpression in CC-RCCVHL cells is sufficient to sensitize them to ROCK inhibition. Finally, Y-27632 treatment inhibited growth of subcutaneous 786-OT1 CC-RCC tumors in mice. Thus, ROCK inhibitors represent potential therapeutics for VHL-deficient CC-RCC.
Subject(s)
Carcinoma, Renal Cell/pathology , Enzyme Inhibitors/pharmacology , Gene Expression Regulation, Neoplastic/drug effects , Kidney Neoplasms/pathology , Von Hippel-Lindau Tumor Suppressor Protein/metabolism , rho-Associated Kinases/antagonists & inhibitors , Amides/pharmacology , Animals , Apoptosis , Biomarkers, Tumor , Carcinoma, Renal Cell/genetics , Carcinoma, Renal Cell/metabolism , Cell Cycle , Cell Proliferation , Humans , Hypoxia-Inducible Factor 1, alpha Subunit/genetics , Hypoxia-Inducible Factor 1, alpha Subunit/metabolism , Kidney Neoplasms/genetics , Kidney Neoplasms/metabolism , Mice , Pyridines/pharmacology , RNA, Small Interfering/genetics , Tumor Cells, Cultured , Von Hippel-Lindau Tumor Suppressor Protein/genetics , Xenograft Model Antitumor Assays , rho-Associated Kinases/geneticsABSTRACT
Cancer screening is a major healthcare issue. Screening modalities are constantly changing due to improvements in technology. Whole body positron emission tomography (PET) with 18F-fluoro-2 deoxy-D-glucose (FDG) and the additional help of the serum levels of tumor markers have been considered as non-invasive methods for cancer screening in asymptomatic subjects. A total of 1283 subjects underwent whole-body FDG PET studies with the additional help of the serum levels of tumor markers in our center for cancer screening. The final diagnoses were confirmed by other imaging modalities or pathological findings in subjects with positive FDG-PET findings, and follow-up for at least 6 months were held in subjects with negative FDG-PET findings. Among a total of 18 (1.4%) subjects with cancers, FDG-PET detected cancers in 15 (1.2%) subjects but with false negative studies in 3 subjects with hepatoma (AFP = 129.6 ng/ml), prostate cancer (PSA = 25.1 ng/ml), and breast cancer (CEA and CA-153 were normal). False-positive FDG-PET studies were found in 24 (1.9%) subjects. However, none had abnormal serum levels of tumor markers. Whole body FDG-PET with the additional help of tumor markers could reduce the false negative and false positive results of FDG-PET only.
Subject(s)
Biomarkers, Tumor , CA-125 Antigen/blood , Fluorodeoxyglucose F18 , Neoplasms/diagnostic imaging , Radiopharmaceuticals , Adult , Aged , Aged, 80 and over , False Negative Reactions , False Positive Reactions , Female , Humans , Male , Mass Screening , Middle Aged , Neoplasms/diagnosis , Neoplasms/pathology , Tomography, Emission-ComputedABSTRACT
In osteosarcoma, some studies have suggested P-glycoprotein expression is a prognostic factor. The clearance of (99m)technetium hexakis-2-methoxyisobutylisonitrile ((99m)Tc-MIBI) has been used in some tumor systems as an in vivo measure of P-glycoprotein-mediated efflux. In this study we explored the correlation between (99m)Tc-MIBI clearance and histological necrosis following induction chemotherapy and P-glycoprotein expression in osteosarcoma. The primary tumors of 20 patients with high-grade osteosarcoma were imaged at diagnosis with (99m)Tc-MIBI, and the uptake ratios and biological half-lives were calculated. P-Glycoprotein expression in the tumor tissue was determined immunohistochemically and by measuring mRNA expression of the multidrug resistance-1 gene. The histological necrosis following induction chemotherapy was assessed by the Huvos grading system. The biological half-life of (99m)Tc-MIBI ranged from 1.4 to 52.5 h. Seven of the 20 tumor samples had a favorable extent of necrosis following induction chemotherapy. The (99m)Tc-MIBI half-life and uptake ratio showed no correlation with histological necrosis following induction chemotherapy. The (99m)Tc-MIBI half-life and uptake ratio did not correlate with either measure of P-glycoprotein expression. The results of this pilot study indicate that (99m)Tc-MIBI imaging is not an effective predictor of histological necrosis following induction chemotherapy in high-grade osteosarcoma. (99m)Tc-MIBI imaging did not correlate with measures of P-glycoprotein expression in the tumor tissue.