Evaluation of polymorphisms in angiogenesis-related genes as predictive and prognostic markers for sunitinib-treated metastatic renal cell carcinoma patients.

PURPOSE: Single nucleotide polymorphisms (SNPs) in angiogenesis-associated genes might play an important role in activity of the tyrosine kinase inhibitor sunitinib and could affect survival of cancer patients treated with this drug. The aim of this retrospective study was to elucidate the role of 10 known SNPs in VEGFA, VEGFR1, VEGFR2 and VEGFR3 as potential prognostic and predictive markers in an independent cohort of patients with metastatic renal cell carcinoma (mRCC). METHODS: DNA from 121 mRCC patients treated with sunitinib was used to analyze SNPs by TaqMan genotyping assays. Disease control rate was evaluated according to RECIST. Adverse effects of sunitinib were registered from medical records. The results of Cox and logistic regression were verified by correction for multiple testing. RESULTS: Kaplan-Meier analysis revealed a reduced progression-free survival in patients with the wild-type (WT) allele of the VEGFA SNP rs699947 compared to variant alleles. Patients with the AA/AC-alleles of the VEGFR1 SNP rs9582036 had an improved median overall survival compared to those with the CC-WT allele what could be confirmed by multivariable Cox proportional hazard regression analyses. No statistically significant associations between the analyzed SNPs and higher risk for adverse effects were observed. CONCLUSIONS: The results of this study suggest that most of the selected SNPs in angiogenesis-related genes are not associated with survival of mRCC patients after sunitinib therapy or with adverse effects. Only the VEGFR1 SNP rs9582036 showed a statistically significant association with overall survival. The potential of SNPs as prognostic and predictive markers for sunitinib-treated mRCC patients should be finally assessed by prospective studies.

Urine protein profiling identified alpha-1-microglobulin and haptoglobin as biomarkers for early diagnosis of acute allograft rejection following kidney transplantation.

PURPOSE: Early diagnosis of acute rejection and effective immunosuppressive therapy lead to improvement in graft survival following kidney transplantation. In this study, we aimed to establish a urinary protein profile suitable to distinguish between patients with rejection and stable graft function and to predict acute rejection based on postoperatively collected urine samples. A further objective was to identify candidate proteins for the use as biomarkers in clinical practice. METHODS: Urine samples of 116 kidney recipients were included. Rejection was proven by biopsy (n = 58), and stable transplant function was monitored for at least 2 years (n = 58). Postoperative urine samples were collected between 3rd and 10th day following transplantation. Urinary protein profiles were obtained by surface-enhanced laser desorption/ionization time-of-flight mass spectrometry. Protein identification and validation were performed using multiplex fluorescence 2DE, peptide mass fingerprinting and enzyme-linked immunosorbent assay. RESULTS: A protein profile including four mass peaks differentiated acute rejection from stable transplants at the time point of rejection and at the postoperative state with 73 % sensitivity and 88 % specificity. Alpha-1-microglobulin (A1MG) and Haptoglobin (Hp) were identified as putative rejection biomarkers. Protein levels were significantly higher in postoperative urine from patients with rejection (A1MG 29.13 vs. 22.06 µg/ml, p = 0.001; Hp 628.34 vs. 248.57 ng/ml, p = 0.003). The combination of both proteins enabled the diagnosis of early rejection with 85 % sensitivity and 80 % specificity. CONCLUSION: Protein profiling using mass spectrometry is suitable for noninvasive detection of rejection-specific changes following kidney transplantation. A specific protein profile enables the prediction of early acute allograft rejection in the immediate postoperative period. A1MG and Hp appear to be reliable rejection biomarkers.

Specific protein and miRNA patterns characterise tumour-associated fibroblasts in bladder cancer.

PURPOSE: Tumour development and progression are strongly affected by interaction of tumour cells and tumour stroma. Different tumour models demonstrate a supportive effect of tumour-associated fibroblasts (TAF) on the tumour genesis. Aims of the present study are the isolation of TAF from primary urinary bladder tumour specimens and the proteomic and epigenetic characterisation. METHODS: TAF were isolated from cultured urinary bladder tumour specimens. Therefore, primary tumour material was treated with EDTA followed by two separated detachment steps. Non-tumour fibroblasts were isolated from foreskin and normal bladder tissues. Proteins and total RNA were isolated from cultured fibroblasts. Protein pattern analyses were carried out by SELDI-TOF-MS. The miRNA expression profile was analysed by miRNA microarray. RESULTS: By optimising cell culture routines, we achieved to isolate and subsequently cultivate TAF from primary tumour material of the urinary bladder. SELDI-TOF-MS measurements reveal distinct differences in the proteomic patterns of TAF and non-tumour fibroblasts. Microarray analyses indicate specific expression of several miRNAs in TAF and non-tumour fibroblasts. CONCLUSION: In summary, we determined proteomic and epigenetic differences between non-tumour fibroblasts and TAF of urinary bladder carcinoma and identified specific protein expression patterns as well as miRNA profiles of TAF in comparison with non-tumour fibroblasts. These findings provide more insights into the complex tumour network and a good starting point for the identification of markers for the prediction of tumour development and progression based on specific TAF expression patterns.

Immunochemotherapy-associated protein patterns in tumour tissue and serum of patients with metastatic renal cell carcinoma.

CONTEXT: Systemic treatment of metastatic renal cell carcinoma (mRCC) with targeted therapies became widely accepted; however, there are few patients who greatly benefit from immunochemotherapy (ICT). It is crucial to recognize these patients for individual treatment. OBJECTIVES: Definition of protein patterns in tissue and serum from mRCC-patients to predict benefit from ICT. MATERIALS AND METHODS: Twenty-five tissue samples and 59 sera were analysed by surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF MS). Protein peaks of interest were identified by 2D-PAGE and peptide mass fingerprinting. Validation was carried out by Western Blot and ELISA. RESULTS: Protein patterns associated with therapy response were determined. Caveolin-1 (CAV-1) and plasminogen activator inhibitor 1 (PAI-1) were identified in tissue; serum amyloid A (SAA) and transthyretin (TTR) were found in serum. CONCLUSION: Individual prediction of therapy benefit and selecting patients for ICT based on molecular biological profiles appear to be feasible in the future.

HIV-1, HCV and HBV seronegative window reduction by the new Roche cobas TaqScreen MPX test in seroconverting donors.

BACKGROUND: By shortening the pre-seroconversion window in the viral screening of donated blood, nucleic acid amplification testing greatly improves safety and efficiency, particularly when combined with multiple target detection and maximal automation. OBJECTIVES: Evaluation of seronegative window reduction during HIV-1, HCV and HBV infection by the novel cobas TaqScreen MPX test for simultaneous nucleic acid detection of HIV-1 (groups M and O), HIV-2, HCV and HBV using the cobas s 201 system. STUDY DESIGN: Testing of HIV-1, HCV, and HBV seroconversion panels (20 each) using the cobas TaqScreen MPX test versus reference immuno- and nucleic acid technology assays. RESULTS: The cobas TaqScreen MPX test detected HIV-1 and HCV infection earlier than immunoassays in 20/20 and 19/20 panels, and HBV DNA earlier than or on the same day as HBsAg in 19/20 and 18/20 panels, and later in 1 and 2 panels on neat samples and 1:6 dilutions. Pre-seroconversion sensitivity exceeded that of COBAS AmpliScreen testing in pools of 24. CONCLUSION: The cobas TaqScreen MPX test shortens the pre-seroconversion window in minipools of six, evidencing high sensitivity, and significantly enhances blood-screening efficiency by the simultaneous automated detection of multiple viruses in a single test.