Combined single nucleotide polymorphism-based genomic mapping and global gene expression profiling identifies novel chromosomal imbalances, mechanisms and candidate genes important in the pathogenesis of T-cell prolymphocytic leukemia with inv(14)(q11q32).

T-cell prolymphocytic leukemia (T-PLL) is a rare aggressive lymphoma derived from mature T cells, which is, in most cases, characterized by the presence of an inv(14)(q11q32)/t(14;14)(q11;q32) and a characteristic pattern of secondary chromosomal aberrations. DNA microarray technology was employed to compare the transcriptomes of eight immunomagnetically purified CD3+ normal donor-derived peripheral blood cell samples, with five highly purified inv(14)/t(14;14)-positive T-PLL blood samples. Between the two experimental groups, 734 genes were identified as differentially expressed, including functionally important genes involved in lymphomagenesis, cell cycle regulation, apoptosis and DNA repair. Notably, the differentially expressed genes were found to be significantly enriched in genomic regions affected by recurrent chromosomal imbalances. Upregulated genes clustered on chromosome arms 6p and 8q, and downregulated genes on 6q, 8p, 10p, 11q and 18p. High-resolution copy-number determination using single nucleotide polymorphism chip technology in 12 inv(14)/t(14;14)-positive T-PLL including those analyzed for gene expression, refined chromosomal breakpoints as well as regions of imbalances. In conclusion, combined transcriptional and molecular cytogenetic profiling identified novel specific chromosomal loci and genes that are likely to be involved in disease progression and suggests a gene dosage effect as a pathogenic mechanism in T-PLL.

Pseudo-precision in gene expression values can reduce efficiency.

OBJECTIVES: When estimating the expression of genes based on the scanned images from microarrays various algorithms are applied in a so-called low-level analysis which can calculate expression values with an arbitrary number of digits beyond the decimal point. However, too many digits (decimal places) are usually not justified because they do not represent the precision of the measured expression. Thus, there is pseudo-precision and, as a result, there are no tied values. METHODS: We suggest avoiding, or omitting, the pseudo-precision: ties can remain, or be created by rounding the computed expression values. Then, average ranks can be used in order to apply nonparametric tests when ties occur. We use two actual data sets and the Wilcoxon rank sum test. RESULTS: We demonstrate that rounding gives a more efficient test, i.e. the average p-value is decreased and the number of p-values smaller than 0.05 is increased. CONCLUSIONS: The random noise of pseudo-precision can reduce the efficiency of statistical tests applied to detect differentially expressed genes. This result is, obviously, relevant in many other areas of our digitalized world.

[Radiofrequency ablation of malignant liver tumors: use of a volumetric necrosis-tumor ratio for local control]. / Radiofrequenzablation von malignen Lebertumoren: Erlaubt ein volumetrischer Nekrose-/Tumor-Quotient eine Vorhersage zur lokalen Tumorkontrolle?

PURPOSE: Sufficient safety margins are essential for preventing local tumor recurrence after radiofrequency ablation RFA of malignant liver tumors. The aim was to determine the initial tumor volume, ablation necrosis volume, and the necrosis-tumor quotient in order to compare these parameters with the rate of local control during follow-up. MATERIALS AND METHODS: 35 patients with 53 tumor nodules (29 colorectal metastases and 24 HCC nodules) were enrolled. RFA procedures were performed under CT guidance with intravenous conscious sedation. Tumor volumes were measured based on CT data sets and the necrosis volume was assessed using the sum-of-area method. A volumetric necrosis/tumor quotient (NTQ) was calculated for all lesions. Follow-up examinations were performed after 3, 6, and 12 months and then on a yearly basis to identify local recurrent tumors. RESULTS: The CRC metastases and HCC nodules had a median tumor volume of 8.3 ml and 7.4 ml, respectively. The mean ablation volumes were 37.6 ml and 29.5 ml, respectively. This resulted in a median NTQ of 3.9 for metastases and 3.4 for HCC. The follow-up (mean time 18 months) revealed local tumor recurrence in 16 of 29 (55 %) metastases and 10 of 24 (42 %) HCC nodules. In lesions with local recurrence, the initial tumor volume was significantly greater and the NTQ was significantly smaller. A threshold value of 3.4 for NTQ has the highest predictive value for local tumor recurrence. CONCLUSION: The volumetric necrosis/tumor quotient NTQ makes it possible to predict the local outcome and can be used for the planning of additional therapy.

Normalization for Affymetrix GeneChips.

OBJECTIVES: The high density oligonucleotide microarrays from Affymetrix (Affymetrix GeneChips) are very popular in biomedical research. They enable to study the expression of thousands of genes simultaneously. In experiments with multiple arrays, normalization techniques are used to reduce the so-called obscuring variation, i.e. the technical variation that is of non-biological origin. Several different normalization methods have been proposed during the last years. METHODS: We review published results about the comparison of normalization methods proposed for Affymetrix GeneChips. RESULTS: The quantile normalization seems to perform favorably regarding precision (low variance), accuracy (low bias), and practicability (low computing time). However, according to very recent results, this normalization method can have an impact on the biological variability and, therefore, appears to be less than optimal from this point of view. CONCLUSION: Although the quantile normalization may be recommendable, more investigations based on more data sets are needed so that the different normalization methods can be evaluated on widely differing data.

Software packages for quantitative microarray-based gene expression analysis.

Microarray technology enables researchers to investigate the expression of several thousand genes simultaneously. The whole transcriptional response of these genes in normal cells or tissue, in disease condition, as an response to biological, genetical or chemical stimuli or during normal biological processes such as cell cycle or embryonic development can be investigated. This leads to a huge amount of data, from which the relevant information has to be extracted by statistical and computational methods. Several software packages for the analysis of gene expression data are available, both commercially and freely. They differ particularly with regard to the implemented analytical methods, the graphical display and the manageability. In this paper the commercial software packages arraySCOUT, GeneSpring and Spotfire DecisionSite for Functional Genomics are compared and their applicability for analysis of gene expression data is studied. Small artificial and application test datasets are used to compare the computational results of the software packages. As far as possible results are verified with standard statistical software package SAS.

Detection of inflammation in patients with acute aortic syndrome: comparison of FDG-PET/CT imaging and serological markers of inflammation.

OBJECTIVE: A substantial number of patients with acute aortic syndrome (AAS) require invasive therapy because of disease progression. Our study aimed to assess the impact of positron emission tomography (PET)/computed tomography (CT) and serological markers of inflammation to identify patients at high risk for disease progression. METHODS: 33 patients with AAS (thoracic aortic aneurysm 5, thoracic aortic dissection 14, penetrating aortic ulcer 8, intramural haematoma 6) were included. After intravenous administration of [18F] fluorodeoxyglucose a non-contrast enhanced PET/CT of the body trunk and CT angiography of the entire aorta was performed. Serological levels of D-dimers and C-reactive protein (CRP) were measured in all patients. Follow-up imaging was performed to detect disease progression. RESULTS: 11 (33%) of 33 patients showed elevated tracer uptake within the aortic pathology, whereas 22 patients were PET-negative. In 23 patients a CRP level exceeding 1.0 mg/dl or a D-dimer level larger than 250 microg/l was found. The follow-up time was 224 (195) days. Nine of 11 PET-positive patients (82%) showed progression of AAS. In contrast, 55% of PET-negative patients showed stable disease or regression during the follow-up period. Kaplan-Meier analysis showed a clear, but not yet significant trend to longer survival in PET-negative patients, whereas elevated CRP and D-dimers did not allow for distinguishing of high-risk patients. CONCLUSIONS: Vessel wall inflammation was found in one-third of the patients with AAS and this patient group seems to have a high risk for disease progression. These initial results needs further investigation.

Differential expression of drug-resistance-related genes between sensitive and resistant blasts in acute myeloid leukemia.

Drug resistance constitutes a considerable problem in the therapy of acute myeloid leukemia (AML). In order to identify genes which might be related to drug resistance, we retrospectively studied gene expression patterns in blast populations of 14 patients with de novo AML, focusing on known or potential resistance mechanisms against cytosine arabinoside and anthracyclines. Following induction and postremission chemotherapy, 7 patients achieved a complete remission (CR) for more than 1 year, while 7 patients showed blast persistence (BP) after induction and salvage chemotherapy. Gene expression analysis was performed using RNA extracted from archived guanidine extracts and Affymetrix HGU133A gene chips. We utilized the Gene Ontology category Biological Process to select genes implicated in DNA metabolism, nucleoside and nucleotide metabolism and transport, reactive oxygen species metabolism, apoptosis and response to drugs and identified 32 differentially expressed genes. From this functional perspective, we found differences between the CR and BP groups with regard to nucleotide metabolism (PBEF1, G6PD; p = 0.048), apoptosis (TNFAIP3, TNFAIP8, MPO, BCL2A1, BAX, SON, BNIP3L; p = 0.039) and reactive oxygen species metabolism (SOD2, KIAA0179; p = 0.048). However, the attempt to construct a predictive model of chemoresistance failed. BP samples had a 2-fold higher expression of CD34 than CR samples. Thus, our findings are in line with reports describing differences in apoptosis resistance between CD34+ and CD34- blast populations. Taken together, our results suggest that drug resistance in AML is a heterogenous phenomenon that might be better defined by means of disturbed biological processes than by focusing on the alteration of the expression of distinct genes.