Nck adaptor proteins modulate differentiation and effector function of T cells.

Understanding the molecular mechanisms regulating T cell reactivity is required for successful reprogramming of immune responses in medical conditions, characterized by dysfunctions of the immune system. Nck proteins are cytoplasmic adaptors mediating diverse cellular functions, including TCR signaling. By enhancing TCR signal strength, Nck proteins influence thymic selection and regulate the size and sensitivity of the peripheral T cell repertoire. Here, we investigated the contribution of Nck proteins to CD4(+) T cell differentiation and effector function using Nck.T(-/-) mice. Impaired GC formation and reduced Tfh were observed in Nck.T(-/-) mice after immunization with T cell-dependent antigens. Th2/Tfh-related cytokines, such as IL-4, IL-10, and IL-21, were decreased in Nck.T(-/-) mice T cells. Moreover, an increased susceptibility to cell death of Tfh cells in Nck.T(-/-) mice was associated with decreased levels of Akt phosphorylation. As a result of this dysregulation in Tfh cells of Nck.T(-/-) mice, we found impaired production and affinity maturation of antibodies against T cell-dependent antigens. Thus, Nck proteins not only participate in thymic selection and generation of the peripheral T cell repertoire but also are involved in the differentiation and effector functions of CD4(+) T cells.

Fractal and stochastic geometry inference for breast cancer: a case study with random fractal models and Quermass-interaction process.

Fractals are models of natural processes with many applications in medicine. The recent studies in medicine show that fractals can be applied for cancer detection and the description of pathological architecture of tumors. This fact is not surprising, as due to the irregular structure, cancerous cells can be interpreted as fractals. Inspired by Sierpinski carpet, we introduce a flexible parametric model of random carpets. Randomization is introduced by usage of binomial random variables. We provide an algorithm for estimation of parameters of the model and illustrate theoretical and practical issues in generation of Sierpinski gaskets and Hausdorff measure calculations. Stochastic geometry models can also serve as models for binary cancer images. Recently, a Boolean model was applied on the 200 images of mammary cancer tissue and 200 images of mastopathic tissue. Here, we describe the Quermass-interaction process, which can handle much more variations in the cancer data, and we apply it to the images. It was found out that mastopathic tissue deviates significantly stronger from Quermass-interaction process, which describes interactions among particles, than mammary cancer tissue does. The Quermass-interaction process serves as a model describing the tissue, which structure is broken to a certain level. However, random fractal model fits well for mastopathic tissue. We provide a novel discrimination method between mastopathic and mammary cancer tissue on the basis of complex wavelet-based self-similarity measure with classification rates more than 80%. Such similarity measure relates to Hurst exponent and fractional Brownian motions. The R package FractalParameterEstimation is developed and introduced in the paper.

High-resolution genomic profiling reveals clonal evolution and competition in gastrointestinal marginal zone B-cell lymphoma and its large cell variant.

We studied marginal zone B-cell lymphomas of the gastrointestinal tract including seven small cell lymphomas, eight large cell areas of composite lymphomas and 13 large cell variants using SNP array profiling. We found an increase of genomic complexity with lymphoma progression from small to large cytology, and identified gains of prominent (proto) oncogenes such as REL, BCL11A, ETS1, PTPN1, PTEN and KRAS which were found exclusively in the large cell variants. Copy numbers of ADAM3A, SCAPER and SIRPB1 were varying between the three different modes of presentation, hence suggestive for aberrations associated with progression from small to large cell lymphoma. The number of aberrations was slightly higher in the large cell part of composite lymphomas than in large cell lymphomas, suggesting that clonal selection takes place and that composite lymphomas are in a transition state. To further investigate this, we comparatively analyzed samples of two morphologically different regions of the same small cell tumor with a BIRC3-MALT1 translocation, as well as material acquired at two different time points from one composite lymphoma. We found genomic heterogeneity in both cases, supporting the theory of competing subclones in the evolution and progression of extranodal marginal zone B-cell lymphoma.

A brief introduction to computer-intensive methods, with a view towards applications in spatial statistics and stereology.

Computer-intensive methods may be defined as data analytical procedures involving a huge number of highly repetitive computations. We mention resampling methods with replacement (bootstrap methods), resampling methods without replacement (randomization tests) and simulation methods. The resampling methods are based on simple and robust principles and are largely free from distributional assumptions. Bootstrap methods may be used to compute confidence intervals for a scalar model parameter and for summary statistics from replicated planar point patterns, and for significance tests. For some simple models of planar point processes, point patterns can be simulated by elementary Monte Carlo methods. The simulation of models with more complex interaction properties usually requires more advanced computing methods. In this context, we mention simulation of Gibbs processes with Markov chain Monte Carlo methods using the Metropolis-Hastings algorithm. An alternative to simulations on the basis of a parametric model consists of stochastic reconstruction methods. The basic ideas behind the methods are briefly reviewed and illustrated by simple worked examples in order to encourage novices in the field to use computer-intensive methods.

BCL6 gene rearrangement and protein expression are associated with large cell presentation of extranodal marginal zone B-cell lymphoma of mucosa-associated lymphoid tissue.

Extranodal marginal zone B-cell lymphoma of mucosa-associated lymphoid tissue (MALT lymphoma) is an indolent B-cell lymphoma, which is often localized in the stomach. It is characterized by typical morphology, immunology, cytogenetics and expression profile. The coexistence of a large B-cell lymphoma and a MALT lymphoma in the gastrointestinal tract is defined as a composite lymphoma (ComL) and, as we have previously shown, is almost always the consequence of secondary transformation of MALT lymphoma. Here, we have analyzed a panel of seven MALT lymphomas, seven ComL and thirteen large cell variants of marginal zone B-cell lymphomas (MZBL) using FISH for the detection of rearrangements of IGH, MALT1, BCL6, BCL10 and FOXP1 and immunohistochemistry for Bcl6, Bcl10 and FoxP1. Translocations involving IGH were found in 10/27 lymphomas including two cases with IGH-BCL6 fusion and one with IGH-BCL10 fusion; in 7/10 cases, the translocation partner was not identified. Bcl10 and FoxP1 protein expression was heterogeneous throughout the series. Genetic rearrangements of BCL6 and Bcl6 protein expression were found almost exclusively in the large cell components of the ComL and the large cell extranodal MZBL (p = 0.2093 and p = 0.0261, respectively). These findings suggest Bcl6 as a marker for transformation of MALT lymphoma.

Sex-related differences in length and erosion dynamics of human telomeres favor females.

Telomeres are repetitive DNA sequences at chromosomal ends contributing to genomic integrity. In somatic cells, telomeres are shortened during DNA reduplication. Thus, telomere erosion has been regarded as a biological clock. Applying the telomere/centromere (T/C)-FISH technique to human peripheral blood lymphocytes, we showed that pangenomically, telomere shortening is linear in centenarians and that this attrition is delayed in females. Statistics reveal a greater skewness in telomere length distribution in females. As the morphological correlate, we find abnormally long telomeres distributed at random. This "erratic extensive elongation" (EEE) of telomeres is a hitherto unrecognized phenomenon in non-neoplastic cells, and females are more successful in this respect. As evidenced by endoreduplication, EEE is transmitted to the cells' progeny. The mechanism involved is likely to be the alternative pathway of telomere elongation (ALT), counteracting erosion and already known to operate in neoplastic cells.

Statistical modelling of the geometry of planar sections of prostatic capillaries on the basis of stationary Strauss hard-core processes.

In a recent study, the capillarization of normal prostatic tissue and prostatic carcinoma tissue was characterized by means of explorative methods of spatial statistics. In the present paper, an attempt was made to go beyond the explorative approach and to characterize the observed point patterns of the capillary profiles on sections by means of a parametric model. For this purpose, the flexible class of Gibbs processes was considered. Specifically, stationary Strauss hard-core processes were fitted to the observed point patterns. The goodness of fit achieved by the model was checked by simulations with the Markov chain Monte Carlo method using the Metropolis-Hastings algorithm. Model fitting and simulations were performed with the help of the spatstat package under R. The observed point patterns were in some cases compatible with realizations of stationary Strauss hard-core processes for all ranges of spatial interaction. However, deviations from the model were found for one or more domains of ranges in other cases. In the tumour tissue, a highly significant decrease of the interaction parameter of the Strauss hard-core process could be found as compared to the normal prostatic tissue. This finding is discussed in terms of a loss of the normal lobular architecture of the glands in the tumour tissue.

Early assessment of therapy response in malignant lymphoma with the thymidine analogue [18F]FLT.

PURPOSE: The aim of this study was to determine whether the thymidine analogue 3'-deoxy-3'-[(18)F]fluorothymidine ([(18)F]FLT) is adequate for early evaluation of the response of malignant lymphoma to antiproliferative treatment in a mouse xenotransplant model. METHODS: Immunodeficient mice bearing a follicular lymphoma xenotransplant were treated with high-dose chemotherapy (cyclophosphamide, n = 10), immunotherapy (CD20 mAb, ibritumomab-tiuxetan, n = 10) or radioimmunotherapy ([(90)Y]CD20 mAb, Zevalin, n = 10). Forty-eight hours after treatment, antiproliferative effects were assessed with [(18)F]FLT. Ninety minutes after i.v. injection of 5-10 MBq [(18)F]FLT, mice were sacrificed and radioactivity within the tumour and normal organs was measured using a gamma counter and calculated as % ID/g. The proliferation fraction in tissue samples derived from treated and untreated tumours was evaluated by Ki-67 immunohistochemistry, which served as the reference for proliferative activity. RESULTS: In untreated lymphoma, the mean proliferation fraction was 83.6%. After chemotherapy, the mean proliferation fraction decreased to 39.3% (p = 0.0001), after immunotherapy to 77.6% (p = 0.0078) and after radioimmunotherapy to 78.8% (p = 0.014). In none of the animals was a significant change in tumour size observed. In untreated lymphoma, tumoural [(18)F]FLT uptake was 5.4% ID/g, after chemotherapy it was 1.5% (p = 0.0005), after immunotherapy, 3.9% (non-significant), and after radioimmunotherapy, 5.8% (non-significant). CONCLUSION: In a lymphoma xenotransplant model, [(18)F]FLT detects early antiproliferative drug activity before changes in tumour size are visible. These findings further support the use of [(18)F]FLT-PET for imaging early response to treatment in malignant lymphoma.

Molecular imaging of proliferation in malignant lymphoma.

We have determined the ability of positron emission tomography (PET) with the thymidine analogue 3'-deoxy-3'-[(18)F]fluorothymidine (FLT) to detect manifestation sites of malignant lymphoma, to assess proliferative activity, and to differentiate aggressive from indolent tumors. In this prospective study, FLT-PET was done additionally to routine staging procedures in 34 patients with malignant lymphoma. Sixty minutes after i.v. injection of approximately 330 MBq FLT, emission and transmission scanning was done. Tracer uptake in lymphoma was evaluated semiquantitatively by calculation of standardized uptake values (SUV) and correlated to tumor grading and proliferation fraction as determined by Ki-67 immunohistochemistry. FLT-PET detected a total of 490 lesions compared with 420 lesions revealed by routine staging. In 11 patients with indolent lymphoma, mean FLT-SUV in biopsied lesions was 2.3 (range, 1.2-4.5). In 21 patients with aggressive lymphoma, a significantly higher FLT uptake was observed (mean FLT-SUV, 5.9; range, 3.2-9.2; P < 0.0001) and a cutoff value of SUV = 3 accurately discriminated between indolent and aggressive lymphoma. Linear regression analysis indicated significant correlation of FLT uptake in biopsied lesions and proliferation fraction (r = 0.84; P < 0.0001). In this clinical study, FLT-PET was suitable for imaging malignant lymphoma and noninvasive assessment of tumor grading. Due to specific imaging of proliferation, FLT may be a superior PET tracer for detection of malignant lymphoma in organs with high physiologic fluorodeoxyglucose uptake and early detection of progression to a more aggressive histology or potential transformation.

Prediction of the variance of stereological volume estimates from systematic sections using computer-intensive methods.

The Cavalieri method is an unbiased estimator of the total volume of a body from its transectional areas on systematic sections. The coefficient of error (CE) of the Cavalieri estimator was predicted by a computer-intensive method. The method is based on polynomial regression of area values on section number and simulation of systematic sectioning. The measurement function is modelled as a quadratic polynomial, with an error term superimposed. The relative influence of the trend and the error component is estimated by techniques of analysis of variance. This predictor was compared with two established short-cut estimators of the CE based on transitive theory. First, all predictors were applied to data sets from six deterministic models with analytically known CE. For these models, the CE was best predicted by the older short-cut estimator and by the computer-intensive approach, if the measurement function had finite jumps. The best prediction was provided by the newer short-cut estimator when the measurement function was continuous. The predictors were also applied to published empirical datasets. The first data set consisted of 10 series of areas of systematically sectioned rat hearts with 10-13 items, the second data set consisted of 13 series of systematically sampled transectional areas of various biological structures with 38-90 items. On the whole, similar mean values for the predicted CE were obtained with the older short-cut estimator and the computer-intensive method. These ranged in the same order of magnitude as resampling estimates of the CE from the empirical data sets, which were used as a cross-check. The mean values according to the newer short-cut CE estimator ranged distinctly lower than the resampling estimates. However, for individual data sets, it happened that the closest prediction as compared to the cross-check value could be provided by any of the three methods. This finding is discussed in terms of the statistical variability of the resampling estimate itself.

Clinical relevance of imaging proliferative activity in lung nodules.

PURPOSE: Recently, the thymidine analogue 3'-deoxy-3'[18F]fluorothymidine (FLT) has been introduced for imaging proliferation with positron emission tomography (PET). In this prospective study, we examined the accuracy of FLT for differentiation of benign from malignant lung lesions and for tumour staging. METHODS: A total of 47 patients with newly diagnosed pulmonary nodules on chest CT suspicious for malignancy were examined with FLT-PET in addition to routine staging procedures. A total of 43 patients also underwent 2-[18F]fluoro-2-deoxy-D-glucose (FDG) PET imaging. Within 2 weeks, patients underwent resective surgery or core biopsy of the pulmonary lesion. RESULTS: Histopathology revealed malignant lung tumours in 32 patients (20 non-small cell lung cancer, 1 small cell lung cancer, 1 pulmonary carcinoid, 1 non-Hodgkin's lymphoma, nine metastases from extrapulmonary tumours) and benign lesions in 15 patients. Increased FLT uptake was exclusively related to malignant tumours. FLT-PET was false negative in two patients with non-small cell lung cancer, in the patient with a pulmonary carcinoid and in three patients with lung metastases. The sensitivity of FLT-PET for detection of lung cancer was 90%, the specificity 100% and the accuracy 94%. Fifteen out of 21 patients with lung cancer had mediastinal lymph node metastases. FLT-PET was true positive in 7/15 patients, resulting in a sensitivity of 53% for N-staging (specificity 100%, accuracy 67%). Clinical TNM stage was correctly identified in 67% (20/30) patients, compared to 85% (23/27) with FDG-PET. CONCLUSION: FLT-PET has a high specificity for the detection of malignant lung tumours. Compared with FDG, FLT-PET is less accurate for N-staging in patients with lung cancer and for detection of lung metastases. FLT-PET therefore cannot be recommended for staging of lung cancer.

Classification of incidental carcinoma of the prostate using learning vector quantization and support vector machines.

The subclassification of incidental prostatic carcinoma into the categories T1a and T1b is of major prognostic and therapeutic relevance. In this paper an attempt was made to find out which properties mainly predispose to these two tumor categories, and whether it is possible to predict the category from a battery of clinical and histopathological variables using newer methods of multivariate data analysis. The incidental prostatic carcinomas of the decade 1990-99 diagnosed at our department were reexamined. Besides acquisition of routine clinical and pathological data, the tumours were scored by immunohistochemistry for proliferative activity and p53-overexpression. Tumour vascularization (angiogenesis) and epithelial texture were investigated by quantitative stereology. Learning vector quantization (LVQ) and support vector machines (SVM) were used for the purpose of prediction of tumour category from a set of 10 input variables (age, Gleason score, preoperative PSA value, immunohistochemical scores for proliferation and p53-overexpression, 3 stereological parameters of angiogenesis, 2 stereological parameters of epithelial texture). In a stepwise logistic regression analysis with the tumour categories T1a and T1b as dependent variables, only the Gleason score and the volume fraction of epithelial cells proved to be significant as independent predictor variables of the tumour category. Using LVQ and SVM with the information from all 10 input variables, more than 80 of the cases could be correctly predicted as T1a or T1b category with specificity, sensitivity, negative and positive predictive value from 74-92%. Using only the two significant input variables Gleason score and epithelial volume fraction, the accuracy of prediction was not worse. Thus, descriptive and quantitative texture parameters of tumour cells are of major importance for the extent of propagation in the prostate gland in incidental prostatic adenocarcinomas. Classical statistical tools and neuronal approaches led to consistent conclusions.

Biological characterisation of breast cancer by means of PET.

Breast cancer is associated with increased glucose consumption and can therefore be visualised with the glucose analogue [(18)F]2-deoxy-2-fluoro-D-glucose (FDG) and positron emission tomography (PET). FDG uptake in the primary tumour can vary substantially, and specific tumour characteristics have been demonstrated to determine the degree of glucose metabolism. Factors with a major influence on FDG uptake in breast cancer comprise expression of glucose transporter Glut-1 and hexokinase I, number of viable tumour cells per volume, histological subtype, tumour grading, microvessel density and proliferative activity. Recently, an association between high FDG uptake and a worse prognosis was suggested. Several studies have been performed correlating FDG uptake with a variety of prognostic and molecular biomarkers as well as parameters predicting tumour response to therapy. However, a correlation with important clinical prognostic markers such as axillary lymph node status and size of the primary tumour, expression of oestrogen and progesterone receptors, proto-oncogene c-erbB-2 or VEGF could not be demonstrated. The lack of correlation with important markers of prognosis does not suggest that FDG uptake might be used as a prognostic criterion in breast cancer. Innovative radiotracers for specific imaging of tumoural perfusion ([(15)O]H(2)O), hormone receptor expression ([(18)F]FES), protein synthesis ([(11)C]methionine), proliferation rate ([(18)F]FLT) or bone mineralisation ([(18)F]fluoride) may provide additional information compared with that provided by FDG PET.

Imaging proliferation in lung tumors with PET: 18F-FLT versus 18F-FDG.

UNLABELLED: Recently, the thymidine analog 3'-deoxy-3'-(18)F-fluorothymidine (FLT) was suggested for imaging tumoral proliferation. In this prospective study, we examined whether (18)F-FLT better determines proliferative activity in newly diagnosed lung nodules than does (18)F-FDG. METHODS: Twenty-six patients with pulmonary nodules on chest CT were examined with PET and the tracers (18)F-FDG and (18)F-FLT. Tumoral uptake was determined by calculation of standardized uptake value (SUV). Within 2 wk, patients underwent resective surgery or had core biopsy. Proliferative activity was estimated by counting nuclei stained with the Ki-67-specific monoclonal antibody MIB-1 per total number of nuclei in representative tissue specimens. The correlation between the percentage of proliferating cells and the SUVs for (18)F-FLT and (18)F-FDG was determined using linear regression analysis. RESULTS: Eighteen patients had malignant tumors (13 with non-small cell lung cancer [NSCLC], 1 with small cell lung cancer, and 4 with pulmonary metastases from extrapulmonary tumors); 8 had benign lesions. In all visible lesions, mean (18)F-FDG uptake was 4.1 (median, 4.4; SD, 3.0; range, 1.0-10.6), and mean (18)F-FLT uptake was 1.8 (median, 1.2; SD, 2.0; range, 0.8-6.4). Statistical analysis revealed a significantly higher uptake of (18)F-FDG than of (18)F-FLT (Mann-Whitney U test, P < 0.05). (18)F-FLT SUV correlated better with proliferation index (P < 0.0001; r = 0.92) than did (18)F-FDG SUV (P < 0.001; r = 0.59). With the exception of 1 carcinoma in situ, all malignant tumors showed increased (18)F-FDG PET uptake. (18)F-FLT PET was false-negative in the carcinoma in situ, in another NSCLC with a low proliferation index, and in a patient with lung metastases from colorectal cancer. Increased (18)F-FLT uptake was related exclusively to malignant tumors. By contrast, (18)F-FDG PET was false-positive in 4 of 8 patients with benign lesions. CONCLUSION: (18)F-FLT uptake correlates better with proliferation of lung tumors than does uptake of (18)F-FDG and might be more useful as a selective biomarker for tumor proliferation.

Statistical analysis of intramembranous particles using freeze fracture specimens.

We studied the point processes of intramembranous particles of mitochondrial membranes from HeLa cells using the freeze fracture technique. Three groups - under normal conditions, after exposition with rotenone, and after exposition with sodium acid - were compared. First, we used several summary statistics in order to study the two-dimensional point patterns of intramembranous particles within each group. Then, we compared the patterns in different groups by bootstrap tests using the K-function and the nearest neighbour distance function G(r). Estimation of the G-function provided significant results but no significant differences between groups were found using the classical K-function; estimation of G(r) should therefore not be omitted when studying observed planar point patterns.

Classification of prostatic carcinoma with artificial neural networks using comparative genomic hybridization and quantitative stereological data.

Staging of prostate cancer is a mainstay of treatment decisions and prognostication. In the present study, 50 pT2N0 and 28 pT3N0 prostatic adenocarcinomas were characterized by Gleason grading, comparative genomic hybridization (CGH), and histological texture analysis based on principles of stereology and stochastic geometry. The cases were classified by learning vector quantization and support vector machines. The quality of classification was tested by cross-validation. Correct prediction of stage from primary tumor data was possible with an accuracy of 74-80% from different data sets. The accuracy of prediction was similar when the Gleason score was used as input variable, when stereological data were used, or when a combination of CGH data and stereological data was used. The results of classification by learning vector quantization were slightly better than those by support vector machines. A method is briefly sketched by which training of neural networks can be adapted to unequal sample sizes per class. Progression from pT2 to pT3 prostate cancer is correlated with complex changes of the epithelial cells in terms of volume fraction, of surface area, and of second-order stereological properties. Genetically, this progression is accompanied by a significant global increase in losses and gains of DNA, and specifically by increased numerical aberrations on chromosome arms 1q, 7p, and 8p.

FDG uptake in breast cancer: correlation with biological and clinical prognostic parameters.

The aim of this study was to evaluate the possible correlation between preoperative FDG-PET results in human breast cancer and the prognostic markers Ki-67, c- erb B2, p53, oestrogen/progesterone receptor status, axillary lymph node status, tumour size and tumour grading. Seventy-five female patients with breast cancer were included in this prospective study. Patient selection was independent of tumour size and the suspected clinical stage of disease. A high-resolution full-ring scanner (Siemens ECAT HR+) was used for PET imaging. The FDG uptake of breast tumours was calculated as the tumour to background ratio (TBR). In resected cancer tissue specimens, the proliferative fraction was evaluated by Ki-67 immunostaining. Additionally, immunostaining of the prognostic markers c-erb B2, p53, and progesterone and oestrogen receptors was performed. Haematoxylin and eosin-stained sections were used for tumour grading. Correlations between FDG uptake and prognostic markers were assumed to be significant at P<0.05 using the Mann-Whitney U test. In ductal breast cancer, mean TBR was 17.3 (median 7.7, range 1.6-122.7), while in lobular cancer it was 6.5 (median 3.7, range 1.4-22.7). Mean proliferative fraction (% Ki-67 positive tumour cells) was 15%+/-13.8% (median 10%, range 0%-60%). Twenty-three carcinomas showed <5% Ki-67 positive tumour cells. Statistical analysis indicated a positive correlation between FDG uptake and proliferative index in ductal breast cancer ( P<0.0001, r=0.63). By contrast, there was no correlation between FDG uptake and c- erb B2 ( P=0.79), p53 ( P=0.92), tumour grading ( P=0.09), oestrogen receptor status ( P=0.41), progesterone receptor status ( P=0.34), axillary lymph node status ( P=0.90) and tumour size ( P=0.3). It is concluded that FDG uptake is significantly higher in ductal breast cancer than in lobular cancer ( P<0.05). FDG uptake correlates with proliferative activity assessed by Ki-67 immunostaining ( P<0.05). A significant correlation with the other prognostic markers, however, could not be demonstrated.

alpha-Methylacyl-CoA racemase: expression levels of this novel cancer biomarker depend on tumor differentiation.

alpha-Methylacyl-CoA racemase (AMACR) has previously been shown to be a highly sensitive marker for colorectal and clinically localized prostate cancer (PCa). However, AMACR expression was down-regulated at the transcript and protein level in hormone-refractory metastatic PCa, suggesting a hormone-dependent expression of AMACR. To further explore the hypothesis that AMACR is hormone regulated and plays a role in PCa progression AMACR protein expression was characterized in a broad range of PCa samples treated with variable amounts and lengths of exogenous anti-androgens. Analysis included standard slides and high-density tissue microarrays. AMACR protein expression was significantly increased in localized hormone-naive PCa as compared to benign (P < 0.001). Mean AMACR expression was lower in tissue samples from patients who had received neoadjuvant hormone treatment but still higher compared to hormone-refractory metastases. The hormone-sensitive tumor cell line, LNCaP, demonstrated stronger AMACR expression by Western blot analysis than the poorly differentiated cell lines DU-145 and PC-3. AMACR protein expression in cells after exposure to anti-androgen treatment was unchanged, whereas prostate-specific antigen, known to be androgen-regulated, demonstrated decreased protein expression. Surprisingly, this data suggests that AMACR expression is not regulated by androgens. Examination of colorectal cancer, which is not hormone regulated, demonstrated high levels of AMACR expression in well to moderately differentiated tumors and weak expression in anaplastic colorectal cancers. Taken together, these data suggest that AMACR expression is not hormone-dependent but may in fact be a marker of tumor differentiation.

Chromosomal changes in incidental prostatic carcinomas detected by comparative genomic hybridization.

OBJECTIVES: The genetic changes underlying the development and progression of prostate cancer are poorly understood. To identify chromosomal regions in incidental prostatic carcinoma (T1a and T1b) was the primary aim of this study. MATERIALS AND METHODS: We used comparative genomic hybridization (CGH) to search for DNA sequence copy number changes on a series of 48 T1 prostate cancer diagnosed by transurethral resection (TURP) and by adenomectomy. Incidental prostatic carcinomas have not been studied by CGH previously. RESULTS: CGH analysis indicated that 14 cases (29.2%) of incidental prostatic carcinoma showed chromosome alterations. The most frequent alterations were chromosomal losses of 8p (10.4%), 13q (6.3%), 5q (4.2%) and 18q (4.2%), and gains of 17p (10.4%), 17q (10.4%), 9q (6.3%) and 7q (4.2%). Minimal overlapping chromosomal regions of loss, indicative for the presence of tumor suppressor genes (TSGs), were mapped to 8p22 and 13q14.1-q21.3, and minimal overlapping regions of gain, indicative for the presence of oncogenes, were found at 9q34.2-qter, 17p12 and 17q24-qter. The statistical analysis displayed a significant association between chromosomal aberration detected by CGH and high Gleason score (P < 0.005) as well as between tumor categories T1a and T1b and chromosomal imbalance (P = 0.041). CONCLUSIONS: Studies directed at incidental prostatic carcinomas allow discovery of chromosomal changes in small and highly malignant tumors. Our results suggest that loss or gain of DNA in these regions are important in prostate cancer. This is the first study, which documents the spectrum of chromosomal changes in incidental prostatic carcinomas.

3-deoxy-3-[(18)F]fluorothymidine-positron emission tomography for noninvasive assessment of proliferation in pulmonary nodules.

We investigated whether uptake of the thymidine analogue 3-deoxy-3-[(18)F]fluorothymidine ([(18)F]FLT) reflects proliferation in solitary pulmonary nodules (SPNs). Thirty patients with SPNs were prospectively examined with positron emission tomography. Standardized uptake values were calculated for quantification of FLT uptake. Histopathology revealed 22 malignant and 8 benign lesions. Proliferation was evaluated by Ki-67 immunostaining and showed a mean proliferation fraction of 30.9% (range, 1-65%) in malignant SPNs and <5% in benign lesions. Linear regression analysis indicated a significant correlation between FLT-standardized uptake values and proliferative activity (P < 0.0001; r = 0.87). FLT uptake was specific for malignant lesions and may be used for differential diagnosis of SPNs, assessment of proliferation, and estimation of prognosis.