Panel of candidate biomarkers for renal cell carcinoma.

The timely diagnosis and therapeutic monitoring of human renal cell carcinoma (RCC) is limited by the lack of specific biomarkers. To identify candidate RCC biomarkers, we used 2-DE gel electrophoresis with mass spectrometry and 2-DE spot intensity-based ROC analysis to analyze 18 sets of paired normal and RCC tumor tissue including conventional, papillary, and chromophobe subtypes. Validation was performed with RCC patient plasma samples and confirmed by clustergram, shRNA, and immunohistochemistry assays. Cardinal candidates were evaluated by ELISA. The leading candidate biomarker that was upregulated in RCC samples according to the clustergram and validation analysis was nicotinamide N-methyltransferase (NNMT) (13/15, P < 0.0001). Other upregulated candidate biomarkers that were identified by this method include ferritin, hNSE, NM23, secretagogin, and L-plastin. The upregulation of NNMT in RCC was confirmed by immunoblotting and immunohistochemistry. Analysis of fractionated membrane-associated proteins identified CAP-G, mitofillin, tubulin alpha, RBBP7, and HSP27. Of these, RBBP7 and HSP27 were highly expressed in the chromophobe subtype of RCC (3/3) but were absent from conventional RCC (0/3). The triple combination of the NNMT, FTL, and hNSE biomarkers had the highest predictive capacity of 0.993, while NNMT was the single, most powerful candidate diagnostic biomarker for all types of RCC.

Concentration of cytokines in age-related macular degeneration after consecutive intravitreal bevacizumab injection.

BACKGROUND: To evaluate the changes in aqueous humor cytokine levels following consecutive intravitreal bevacizumab (Avastin, Genentech Inc., San Francisco, CA, USA) injections in eyes with choroidal neovascularization (CNV) due to age-related macular degeneration (AMD). METHOD: Aqueous humor samples were collected at the time of intravitreal injection of 1.25 mg of bevacizumab every 7.0 (+/-2.0) weeks from ten eyes with AMD for the AMD group and during cataract surgery in nine eyes for the control group. Visual acuity with Early Treatment of Diabetic Retinopathy Study (ETDRS) letters and central macular thickness (CMT) using optical coherence tomography were measured before each injection in the AMD group. Aqueous cytokine levels were determined by immunoassay using multi-analyte biochip array technology (Evidence investigator cytokine and growth factor biochip array, RANDOX laboratories Ltd., Crumlin, UK). RESULT: In the AMD group, mean +/- standard deviation(SD) aqueous VEGF levels decreased from 68.0 +/- 32.1 pg/ml at baseline to 26.3 +/- 19.0 pg/ml after the first injection (p = 0.028) and to 25.2 +/- 12.8 pg/ml after the second injection (p = 0.005). While CMT decreased from 307.7 +/- 102.0 mum to 206.8 +/- 141.5 microm (p = 0.037), ETDRS visual acuity increased from 17.6 +/- 11.7 letters to 22.0 +/- 15.6 letters after three consecutive injections (p = 0.017). CONCLUSION: Significantly decreased VEGF levels were noted after the first injection of bevacizumab. These levels were maintained after the second injection, which paralleled the change in visual acuity and CMT.

Effect of intravitreal bevacizumab injection on aqueous humor cytokine levels in clinically significant macular edema.

PURPOSE: To determine the effect of intravitreal bevacizumab (Avastin, Genentech, Inc., San Francisco, CA) injection on aqueous humor cytokine levels in clinically significant macular edema (CSME). DESIGN: Retrospective, comparative study. PARTICIPANTS: Seventeen eyes undergoing intravitreal injection for CSME and 11 eyes undergoing cataract surgery as negative controls. Nine patients with CSME were naïve to intravitreal bevacizumab injection (CSME group 1), and 8 patients previously received 1 intravitreal bevacizumab injection at a mean of 9.6+/-1.4 weeks earlier (CSME group 2). METHODS: Aqueous humor samples were collected before performing intravitreal injection in the CSME group and during cataract surgery in the control group. Aqueous cytokine levels were determined by immunoassay using multianalyte biochip array technology. MAIN OUTCOME MEASURES: The concentration of cytokines in the aqueous humor was recorded as a mean (+/- standard deviation) in CSME groups 1 and 2 and the control group. Correlations of aqueous humor cytokine levels were evaluated. RESULTS: Significantly higher levels of interleukin (IL)-6, IL-8, and monocyte chemoattractant protein (MCP)-1 were noted in patients with CSME compared with controls. Although higher vascular endothelial growth factor (VEGF) levels were noted in CSME group 1 (P = 0.001), a lower level of VEGF was noted after intravitreal bevacizumab injection in CSME group 2 (P = 0.004) compared with the control group. VEGF levels were also lower in CSME group 2 compared with CSME group 1 (P = 0.001). In CSME group 2, a significant negative correlation was observed between VEGF and IL-6 (rho = -0.833, P = 0.01) and VEGF and MCP-1 (rho = -0.736, P = 0.037). CONCLUSIONS: Significantly higher levels of IL-6, IL-8, VEGF, and MCP-1 were noted in the aqueous humor of CSME. However, recurrence of macular edema was noted with significantly lower concentrations of VEGF after a single intravitreal injection of bevacizumab. This suggests the potential importance of IL-6 and MCP-1 in the pathogenesis of CSME. FINANCIAL DISCLOSURE(S): The author(s) have no proprietary or commercial interest in any materials discussed in this article.

Concentration of cytokines in the aqueous humor of patients with naive, recurrent and regressed CNV associated with amd after bevacizumab treatment.

PURPOSE: : To evaluate the concentration of various cytokines in the aqueous humor of patients with naive, recurrent, and regressed choroidal neovascularization (CNV) of age-related macular degeneration after bevacizumab treatment. METHODS: : Aqueous humor samples were collected from 36 eyes with age-related macular degeneration and 10 controls during cataract surgery. Of 36 patients with age-related macular degeneration, 5 eyes were naïve to bevacizumab injection, 14 eyes had recurrent CNV after bevacizumab treatment, and 17 eyes had regressed CNV after bevacizumab treatment. Cytokines were measured by an immunoassay using multianalyte biochip array technology (Evidence investigator cytokine and growth factor biochip array, RANDOX laboratories Ltd., Crumlin, UK). RESULTS: : No significant difference in the cytokine levels was noted between the control group and the naïve CNV group (all P > 0.05). Vascular endothelial growth factor in both naive (66.8 +/- 35.1 pg/mL) and recurrent CNV groups (55.7 +/- 63.0 pg/mL) was significantly higher compared with regressed CNV group (9.8 +/- 12.8 pg/mL, P = 0.025 and P = 0.004, respectively) but was not statistically different from the control group (81.8 +/- 43.7 pg/mL, P = 0.310 and P = 0.212, respectively). The aqueous humor level of tumor necrosis factor-alpha and interleukin (IL)-2 was significantly lower in recurrent CNV group (P = 0.036 and P = 0.019) compared with the control group. In the active CNV patients (recurrent and naïve CNV groups), the aqueous humor levels of IL-6 and IL-8 significantly correlated with the size of CNV (rho = 0.692, P = 0.001 and rho = 0.745, P < 0.001, respectively). CONCLUSION: : Levels of Vascular endothelial growth factor measured in the aqueous humor were significantly related to the disease activity of CNV in age-related macular degeneration. Moreover, IL-2, IL-6, IL-8, and tumor necrosis factor-alpha may be related to the activity of CNV.