BCL-2 expression aids in the immunohistochemical prediction of the Oncotype DX breast cancer recurrence score.

BACKGROUND: The development of molecular techniques to estimate the risk of breast cancer recurrence has been a significant addition to the suite of tools available to pathologists and breast oncologists. It has previously been shown that immunohistochemistry can provide a surrogate measure of tumor recurrence risk, effectively providing a less expensive and more rapid estimate of risk without the need for send-out. However, concordance between gene expression-based and immunohistochemistry-based approaches has been modest, making it difficult to determine when one approach can serve as an adequate substitute for the other. We investigated whether immunohistochemistry-based methods can be augmented to provide a useful therapeutic indicator of risk. METHODS: We studied whether the Oncotype DX breast cancer recurrence score can be predicted from routinely acquired immunohistochemistry of breast tumor histology. We examined the effects of two modifications to conventional scoring measures based on ER, PR, Ki-67, and Her2 expression. First, we tested a mathematical transformation that produces a more diagnostic-relevant representation of the staining attributes of these markers. Second, we considered the expression of BCL-2, a complex involved in regulating apoptosis, as an additional prognostic marker. RESULTS: We found that the mathematical transformation improved concordance rates over the conventional scoring model. By establishing a measure of prediction certainty, we discovered that the difference in concordance between methods was even greater among the most certain cases in the sample, demonstrating the utility of an accompanying measure of prediction certainty. Including BCL-2 expression in the scoring model increased the number of breast cancer cases in the cohort that were considered high certainty, effectively expanding the applicability of this technique to a greater proportion of patients. CONCLUSIONS: Our results demonstrate an improvement in concordance between immunohistochemistry-based and gene expression-based methods to predict breast cancer recurrence risk following two simple modifications to the conventional scoring model.

Invasive breast carcinomas with ATM gene variants of uncertain significance share distinct histopathologic features.

The increasing availability of next-generation sequencing for clinical research dramatically improved our understanding of breast cancer genetics and resulted in detection of new mutation variants. Cancer risk data relating to some of these variants are insufficient, prompting the designation of variants of uncertain significance (VUS). The histopathologic characteristics of these variants have not been previously described. We propose to depict these characteristics and determine if invasive carcinomas with similar VUS genes share similar histomorphologic features. In total, 28 invasive breast cancers with VUS were retrospectively identified. Tumor sections were reviewed and a predefined set of histopathologic characteristics were documented and compared. Nine of the 28 cases were variants in the ATM gene and were found to share similar histologic characteristics; all had tumor cells with low nuclear grade, absent tumor infiltrating lymphocytes, as well as a marked desmoplastic response. A subset of the above findings were identified in variants of other genes but none had all findings collectively. Furthermore, variants of ATM gene had smaller tumor size, lower pathologic T stage at presentation, and more favorable surrogate molecular subtype compared to variants of other genes. These findings could potentially be used to reclassify VUS and predict which patients may harbor ATM mutations, and hence could have implications in triaging toward ATM variant identification for potential future targeted therapy.

Comparison of Tissue Molecular Biomarker Testing Turnaround Times and Concordance Between Standard of Care and the Biocartis Idylla Platform in Patients With Colorectal Cancer.

OBJECTIVES: Management of colorectal cancer warrants mutational analysis of KRAS/NRAS when considering anti-epidermal growth factor receptor therapy and BRAF testing for prognostic stratification. In this multicenter study, we compared a fully integrated, cartridge-based system to standard-of-care assays used by participating laboratories. METHODS: Twenty laboratories enrolled 874 colorectal cancer cases between November 2017 and December 2018. Testing was performed on the Idylla automated system (Biocartis) using the KRAS and NRAS-BRAF cartridges (research use only) and results compared with in-house standard-of-care testing methods. RESULTS: There were sufficient data on 780 cases to measure turnaround time compared with standard assays. In-house polymerase chain reaction (PCR) had an average testing turnaround time of 5.6 days, send-out PCR of 22.5 days, in-house Sanger sequencing of 14.7 days, send-out Sanger of 17.8 days, in-house next-generation sequencing (NGS) of 12.5 days, and send-out NGS of 20.0 days. Standard testing had an average turnaround time of 11 days. Idylla average time to results was 4.9 days with a range of 0.4 to 13.5 days. CONCLUSIONS: The described cartridge-based system offers rapid and reliable testing of clinically actionable mutation in colorectal cancer specimens directly from formalin-fixed, paraffin-embedded tissue sections. Its simplicity and ease of use compared with other molecular techniques make it suitable for routine clinical laboratory testing.

Insulin-like growth factor-I produced by seminal vesicles: relationship to intraepithelial basal cell hyperplasia in the prostate.

PURPOSE: This study examined the seminal vesicle fluid (SVF) as a potential local source of insulin-like growth factor-I (IGF-I) in the peripheral zone of the prostate. EXPERIMENTAL DESIGN: IGF-I levels in seminal fluid were measured. The levels of the IGF-I receptor (IGF-IR) in its active, phosphorylated form as well as direct downstream targets were examined in the peripheral zone of the prostate. RESULTS: In situ, we find that the IGF-IR is activated in the peripheral zone in areas of atrophy, prostatic intraepithelial hyperplasia, and cancer. In addition, immunostaining reveals preferential activation of the IGF-IR in p63-positive cells in areas of intermediate basal cell hyperplasia in the peripheral zone, indicating that prostate progenitor cells are highly sensitive to increases in local IGF-I levels. These areas of basal cell hyperplasia occur at high incidence in the peripheral zone of the prostate. Relatively high levels of IGF-I were identified in SVF. In addition, we find that SVF can stimulate the proliferation of both normal and cancer-derived prostate cells. CONCLUSIONS: These results suggest that SVF is a local source of IGF-I that provides chronic stimulation of prostate cells. This chronic stimulation could contribute to the development of prostate cancer in older men.

The ABCA5 protein: a urine diagnostic marker for prostatic intraepithelial neoplasia.

PURPOSE: To develop a urine diagnostic test for preneoplastic intraepithelial neoplasia of the prostate. EXPERIMENTAL DESIGN: We have used a DNA-binding assay and electrophoretic mobility shift assays (EMSA) to screen for novel duplexed DNA-binding sequences, which bind protein(s) overexpressed in crude protein extracts from high-grade prostatic intraepithelial neoplasia (HGPIN). EMSAs, immunohistochemistry, and ELISAs were used to measure expression of the ABCA5 protein identified as a specific marker in prostate tissue and patient urine. RESULTS: Following screening of 4,096 sequences, an 8-bp dsDNA sequence (i.e., TCCAGCGA) was identified, which binds the ABCA5 protein, a member of the ATP-binding cassette multidrug resistant family. EMSAs showed that ABCA5 was overexpressed in HGPIN tissue (n=11/11) and in the urine of patients with HGPIN (n=18/18) but was not expressed in prostate cancer, benign prostatic hyperplasia, or stroma. Immunohistochemistry indicated that ABCA5 was overexpressed in foci of intermediate basal cells in normal glands and in HGPIN. ABCA5 was faintly expressed in prostate cancer glands. ELISAs showed in 'blinded studies' that ABCA5 was a highly sensitive (>98% sensitivity) urine diagnostic marker for HGPIN in biopsy-positive patients (n=107) at a 'cutoff' of 25 ng/mL. ABCA5 was present at very low levels (i.e., <25 ng/mL) in the urine of patients diagnosed with benign prostatic hyperplasia (n=79) or prostatitis or kidney and bladder cancer (>86% specificity). CONCLUSIONS: The data indicate that ABCA5 might be a specific urine marker for diagnosis of patients with HGPIN.

Estimation of Fine-Scale Histologic Features at Low Magnification.

CONTEXT.­: Whole-slide imaging has ushered in a new era of technology that has fostered the use of computational image analysis for diagnostic support and has begun to transfer the act of analyzing a slide to computer monitors. Due to the overwhelming amount of detail available in whole-slide images, analytic procedures-whether computational or visual-often operate at magnifications lower than the magnification at which the image was acquired. As a result, a corresponding reduction in image resolution occurs. It is unclear how much information is lost when magnification is reduced, and whether the rich color attributes of histologic slides can aid in reconstructing some of that information. OBJECTIVE.­: To examine the correspondence between the color and spatial properties of whole-slide images to elucidate the impact of resolution reduction on the histologic attributes of the slide. DESIGN.­: We simulated image resolution reduction and modeled its effect on classification of the underlying histologic structure. By harnessing measured histologic features and the intrinsic spatial relationships between histologic structures, we developed a predictive model to estimate the histologic composition of tissue in a manner that exceeds the resolution of the image. RESULTS.­: Reduction in resolution resulted in a significant loss of the ability to accurately characterize histologic components at magnifications less than ×10. By utilizing pixel color, this ability was improved at all magnifications. CONCLUSIONS.­: Multiscale analysis of histologic images requires an adequate understanding of the limitations imposed by image resolution. Our findings suggest that some of these limitations may be overcome with computational modeling.

Heterogeneous breast tumoroids: An in vitro assay for investigating cellular heterogeneity and drug delivery.

Breast tumors are typically heterogeneous and contain diverse subpopulations of tumor cells with differing phenotypic properties. Planar cultures of cancer cell lines are not viable models of investigation of cell-cell and cell-matrix interactions during tumor development. This article presents an in vitro coculture-based 3-dimensional heterogeneous breast tumor model that can be used in drug resistance and drug delivery investigations. Breast cancer cell lines of different phenotypes (MDAMB231, MCF7, and ZR751) were cocultured in a rotating wall vessel bioreactor to form a large number of heterogeneous tumoroids in a single cell culture experiment. Cells in the rotating vessels were labeled with Cell Tracker fluorescent probes to allow for time course fluorescence microscopy to monitor cell aggregation. Histological sections of tumoroids were stained with hematoxylin and eosin, progesterone receptor, E-cadherin (E-cad), and proliferation marker ki67. In vitro tumoroids developed in this study recapture important features of the temporal-spatial organization of solid tumors, including the presence of necrotic areas at the center and higher levels of cell division at the tumor periphery. E-cad-positive MCF7 cells form larger tumoroids than E-cad-negative MDAMB231 cells. In heterogeneous tumors, the irregular surface roughness was mainly due to the presence of MDAMB231 cells, whereas MCF7 cells formed smooth surfaces. Moreover, when heterogeneous tumoroids were placed onto collagen gels, highly invasive MDAMB231 cell-rich surface regions produced extensions into the matrix, whereas poorly invasive MCF7 cells did not. The fact that one can form a large number of 1-mm tumoroids in 1 coculture attests to the potential use of this system at high-throughput investigations of cancer drug development and drug delivery into the tumor.

Increased sensitivity of anal cytology in evaluation of internal compared with external lesions.

OBJECTIVE: To investigate the impact of keratin on the accuracy of internal and external anal brush sampling of known lesions. STUDY DESIGN: A group of 46 human immunodeficiency virus (HIV)-seropositive patients underwent external and internal anal brush sampling before biopsy of known lesions. RESULTS; A total of 92 ThinPrep (46 external, 46 internal) an 211 biopsies were examined. The sensitivity and specificity for internal lesions positive and negative for anal squa mous intraepithelial lesion (ASIL) was 91.1% and 42.8%, respectively; and for external lesions was 79.4% and 100%, respectively. Low cellularity on cytology and markedly thickened keratin on biopsy were significantly more common in external compared with internal lesions (p < 0.0001). CONCLUSION: We conclude that hyperkeratosis interferes with adequate sampling and accurate grading of external anal lesions by brush sampling.

An alternative reference space for H&E color normalization.

Digital imaging of H&E stained slides has enabled the application of image processing to support pathology workflows. Potential applications include computer-aided diagnostics, advanced quantification tools, and innovative visualization platforms. However, the intrinsic variability of biological tissue and the vast differences in tissue preparation protocols often lead to significant image variability that can hamper the effectiveness of these computational tools. We developed an alternative representation for H&E images that operates within a space that is more amenable to many of these image processing tools. The algorithm to derive this representation operates by exploiting the correlation between color and the spatial properties of the biological structures present in most H&E images. In this way, images are transformed into a structure-centric space in which images are segregated into tissue structure channels. We demonstrate that this framework can be extended to achieve color normalization, effectively reducing inter-slide variability.

Impaired homologous recombination DNA repair and enhanced sensitivity to DNA damage in prostate cancer cells exposed to anchorage-independence.

During metastases, cancer cells are temporarily exposed to the condition in which interactions with extracellular environment can be restricted (anchorage-independence). We demonstrate that the sensitivity of prostate cancer cell lines, DU145 and PC-3, to genotoxic treatment (cisplatin and gamma-irradiation) increased several folds when cells were forced to grow in anchorage-independence. This enhanced drug sensitivity was associated with a severe impairment of homologous recombination-directed DNA repair (HRR). The mechanism involves Rad51, which is the major enzymatic component of HRR. The protein level of Rad51 and its recruitment to DNA double-strand breaks (DSBs) were both attenuated. Rad51 deficiency in anchorage-independence was not associated with Rad51 promoter activity, and was not compensated by a constitutive overexpression of Rad51 cDNA. Instead, Rad51 protein level and its ability to colocalize with DSBs were restored in the presence of proteosome inhibitors, or when cells from the suspension cultures were allowed reattachment. Presented results indicate that anchorage-independence sensitizes prostate cancer cells to genotoxic agents; however, it also attenuates faithful component of DNA repair by targeting stability of Rad51. This temporal attenuation of HRR may contribute to the accumulation mutations after DNA damage, and possibly the selection of new adaptations in cells, which survived genotoxic treatment.

Intraprostatic spermatozoa: zonal distribution and association with atrophy.

Intraprostatic spermatozoa (IS) have been demonstrated in only 2 articles in the literature reporting on postmortem prostates. Intraprostatic spermatozoa have not been previously described in radical prostatectomies. This is the first study that describes the presence of IS in radical prostatectomies with prostatic carcinoma (PC) and its association with atrophy. We examined whole mount sagittal sections from 69 consecutive radical prostatectomy cases for PC. A central section including the seminal vesicle ejaculatory duct urethra complex (SVEDU) from each case was stained with Berg's stain to identify spermatozoa and their location. The extent and the type of atrophy were assessed on the entire prostate by using a grid method. Eighteen cases (26.1%) revealed spermatozoa both in the SVEDU and in the prostate (IS) (group 1). Twenty-two cases (31.9%) showed spermatozoa exclusively in the SVEDU but not in the prostate (group 2). The remaining 29 cases (42.0%) had no spermatozoa in either site. Location of IS was 72.2% peripheral zone, 22.2% central zone, and 5.6% transitional zone. Intraprostatic spermatozoa were frequently seen accompanied by inflammatory infiltrate in the periglandular stroma. The extent of atrophy was greater in group 1 than in group 2 (25.7% versus 15.3%; P = .006). Postatrophic hyperplasia was seen more frequently in group 1 than in group 2 (72.2% versus 40.9%; P = .025). In conclusion, the frequency of IS is 26.1% when including all prostates and 45.0% when including prostates with evidence of residual ejaculate (spermatozoa in the SVEDU), more than previously reported. Intraprostatic spermatozoa are predominantly located in the peripheral zone similar to atrophy and PC. Prostates with IS have larger atrophic areas and increased frequency of postatrophic hyperplasia. The role of IS in the pathogenesis of prostate inflammation and atrophy should be investigated.

Use of a new method in reaching consensus on the cause of cytologic-histologic correlation discrepancy.

Pathologists exhibit very poor agreement in adjudicating the cause of cytologic-histologic correlation discrepancies, which contributes to problems in designing interventions to reduce discrepancy frequency. In this observational study, we developed a visual method of adjudicating discrepancy cause, termed the No-Blame Box method, which consisted of initially assessing specimen interpretability by separately evaluating specimen quality and the presence of tumor. Five pathologists blindly adjudicated the cause of discrepancy in pulmonary specimens from 40 patients. The kappa statistic of all pathologist pairs in adjudicating discrepancy cause using the No-Blame Box method ranged from 0.400 to 0.796, indicating acceptable to excellent agreement. Pathologists ranged in their assessment of specimen interpretability from 13% to 20%, and in no case did all 5 pathologists concur that a specimen was interpretable. Most discrepancies resulted from pathologists diagnosing noninterpretable samples. Pathologists who used the No-Blame Box showed significant agreement in the adjudication of discrepancy cause.

Large-scale computations on histology images reveal grade-differentiating parameters for breast cancer.

BACKGROUND: Tumor classification is inexact and largely dependent on the qualitative pathological examination of the images of the tumor tissue slides. In this study, our aim was to develop an automated computational method to classify Hematoxylin and Eosin (H&E) stained tissue sections based on cancer tissue texture features. METHODS: Image processing of histology slide images was used to detect and identify adipose tissue, extracellular matrix, morphologically distinct cell nuclei types, and the tubular architecture. The texture parameters derived from image analysis were then applied to classify images in a supervised classification scheme using histologic grade of a testing set as guidance. RESULTS: The histologic grade assigned by pathologists to invasive breast carcinoma images strongly correlated with both the presence and extent of cell nuclei with dispersed chromatin and the architecture, specifically the extent of presence of tubular cross sections. The two parameters that differentiated tumor grade found in this study were (1) the number density of cell nuclei with dispersed chromatin and (2) the number density of tubular cross sections identified through image processing as white blobs that were surrounded by a continuous string of cell nuclei. Classification based on subdivisions of a whole slide image containing a high concentration of cancer cell nuclei consistently agreed with the grade classification of the entire slide. CONCLUSION: The automated image analysis and classification presented in this study demonstrate the feasibility of developing clinically relevant classification of histology images based on micro- texture. This method provides pathologists an invaluable quantitative tool for evaluation of the components of the Nottingham system for breast tumor grading and avoid intra-observer variability thus increasing the consistency of the decision-making process.

Novel Small-Molecule CX3CR1 Antagonist Impairs Metastatic Seeding and Colonization of Breast Cancer Cells.

UNLABELLED: Recent evidence indicates that cancer cells, even in the absence of a primary tumor, recirculate from established secondary lesions to further seed and colonize skeleton and soft tissues, thus expanding metastatic dissemination and precipitating the clinical progression to terminal disease. Recently, we reported that breast cancer cells utilize the chemokine receptor CX3CR1 to exit the blood circulation and lodge to the skeleton of experimental animals. Now, we show that CX3CR1 is overexpressed in human breast tumors and skeletal metastases. To assess the clinical potential of targeting CX3CR1 in breast cancer, a functional role of CX3CR1 in metastatic seeding and progression was first validated using a neutralizing antibody for this receptor and transcriptional suppression by CRISPR interference (CRISPRi). Successively, we synthesized and characterized JMS-17-2, a potent and selective small-molecule antagonist of CX3CR1, which was used in preclinical animal models of seeding and established metastasis. Importantly, counteracting CX3CR1 activation impairs the lodging of circulating tumor cells to the skeleton and soft-tissue organs and also negatively affects further growth of established metastases. Furthermore, nine genes were identified that were similarly altered by JMS-17-2 and CRISPRi and could sustain CX3CR1 prometastatic activity. In conclusion, these data support the drug development of CX3CR1 antagonists, and promoting their clinical use will provide novel and effective tools to prevent or contain the progression of metastatic disease in breast cancer patients. IMPLICATIONS: This work conclusively validates the instrumental role of CX3CR1 in the seeding of circulating cancer cells and is expected to pave the way for pairing novel inhibitors of this receptor with current standards of care for the treatment of breast cancer patients. Mol Cancer Res; 14(6); 518-27. ©2016 AACR.

Interleukin 10 blocks matrix metalloproteinase-2 and membrane type 1-matrix metalloproteinase synthesis in primary human prostate tumor lines.

Insulin-like growth factor (IGF) I has been shown previously to up-regulate matrix metalloproteinase-2 (MMP-2) production, whereas the interleukin (IL) 10/IL-10 receptor axis has been found to down-regulate MMP-2 synthesis in tumor cells. In this paper, we showed that IL-10 activation of the IL-10 receptor blocked MMP-2 and membrane type 1 (MT1) -MMP transcription and protein synthesis in nonimmortalized primary human prostate cell strains (i.e., HPCA-10a and HPCA-10c) derived from high-grade cancer. Northern blots, Western blots, and ELISAs showed that IL-10 suppressed IGF-I induction of MMP-2 and MT1-MMP mRNA synthesis in these cell strains (P < 0.001). Inhibition studies with IL-10 and IGF-I receptor antibodies plus transfections experiments with IL-10 sense, and IGF-I receptor antisense constructs confirmed these results. Finally, transient transfection experiments and chloramphenicol acetyltransferase assays with different regions of the 5' promoter region of the MMP-2 gene (-1659 to -555 bp) additionally showed that IGF-I stimulated p53-dependent plasmid catecholamine acetyltransferase activity and that IL-10 blocked IGF-I-induced plasmid catecholamine acetyltransferase activity. Electrophoretic mobility shift assays revealed that IL-10 induced protein(s) binding to a putative "silencer element" (-1309 to -555 fragment) downstream of the p53 binding site (-1649 to -1640). The data show that IL-10 blocks IGF-I activation of MMP-2 and MT1-MMP mRNA expression and protein synthesis in primary prostate cell strains.

Evidence for prostate cancer-associated diagnostic marker-1: immunohistochemistry and in situ hybridization studies.

PURPOSE: The purpose of this study was to characterize a novel gene/protein associated with prostate cancer, termed prostate cancer-associated diagnostic marker-1 [PCADM-1 (Hu Y, Wang M, Garcia FU, Aoyaki K, Stearns ME. Identification of PCADM-1 as a novel diagnostic marker for prostate cancer, submitted for publication)]. EXPERIMENTAL DESIGN AND RESULTS: Immunological studies revealed that rabbit polyclonal antibodies generated against recombinant PCADM-1 specifically recognize the protein in crude protein extracts from a variety of prostate cancer cell lines (i.e., PC-3 ML, LNCaP, DU145, and CPTX-1532) and prostate cancer tissue. Combined immunolabeling and in situ hybridization studies demonstrated that PCADM-1 mRNA was expressed by the luminal epithelial cells of prostate cancer glands and was not expressed by high-grade prostatic intraepithelial neoplasia or HPV-MLC7 cells. Immunolabeling studies of tissue arrays from biopsies of archival material (n = 200 samples) confirmed that PCADM-1 was expressed by the luminal epithelial cells of prostate cancer. CONCLUSIONS: Taken together, the data suggest that PCADM-1 is a specific marker for human prostate cancer.

Expression of cell-cycle-regulated proteins pRb2/p130, p107, p27(kip1), p53, mdm-2, and Ki-67 (MIB-1) in prostatic gland adenocarcinoma.

PURPOSE: The quest for prognostic molecular markers in prostatic carcinoma is still in progress. Many proteins have already been screened by immunohistochemistry with the aim to find the most reliable indicator of progressive disease. In this study, we evaluated the expression of pRb2/p130, p107, p27(kip1), p53, mdm-2, and Ki-67 (MIB-1) by immunohistochemistry in 24 prostate carcinomas compared with the paired expression of normal prostates. EXPERIMENTAL DESIGN: Expression of the different proteins in normal and pathological specimens was evaluated by the Wilcoxon test. A matrix of correlation (Spearman coefficient) was used to evaluate the possible association in expression among the different proteins. Logistic regression analysis was used to test the multivariable prognostic value of the levels of protein expression for the probability of disease development. RESULTS: p53 and Ki-67 (MIB-1) showed a higher expression in cancer than in normal tissue (P = 0.006 and <0.001, respectively). pRb2/p130, p107, and p27(kip1) showed an overall lower expression in cancer, but the difference between cytoplasmic and nuclear expression was always higher for cancer (Ps, from <0.001 to 0.016). mdm-2 expression was lower in cancer, but the difference between cytoplasmic and nuclear expression was not significant (P = 0.571) when compared with that in normal tissue. A positive correlation between p27 and pRb2/p130 levels expressed, in normal and cancer counterparts in the same sample, as the difference between cytoplasmic and nuclear protein concentrations (P = 0.045) was found. Additionally, p107 expression showed an inverse correlation with Ki-67 (MIB-1) expression in the most aggressive tumors (P = 0.046). Logistic regression output showed that Ki-67 (MIB-1) and pRb2/p130 (expressed as differences between cytoplasmic and nuclear concentrations) were the variables associated with a higher risk of cancer. The highest value was reported for Ki-67 (MIB-1) (odds ratio, 2.11), followed by pRb2/p130 (odds ratio, 1.01). pRb2/p130 alone was associated with a sensitivity (rate of cases having a posterior probability of disease >/=0.5) of 61% with a false positive rate of 22%. Ki-67 (MIB-1) alone yielded a sensitivity of 69% and a false positive rate of 14%. The combined model (Ki-67 + pRb2/p130) yielded a sensitivity of 83% with a false positive rate of 17%. Interestingly, one specimen in which we also found a high-grade prostatic intraepithelial neoplasia showed the progressive loss of pRb2/p130 from normal prostatic cells to prostatic intraepithelial neoplasia cells, suggesting that in prostatic cancer, lack of expression of the tumor suppressor gene pRb2/p130 could be involved in the progression of the disease, from an early stage. CONCLUSIONS: This study showed that all of the proteins but mdm-2 were expressed at a different rate in normal and pathological prostate specimens. Multivariate analysis showed that pRb2/p130 and p107 may be involved in the pathogenesis and progression of prostate cancers, and that the expression of the retinoblastoma-related protein pRb2/p130 along with Ki-67 (MIB-1), expressed as differences between cytoplasmic and nuclear concentrations, could be considered new parameters to be evaluated in discriminating patients at a higher risk for prostate cancer.

Gastrointestinal neurofibromatosis: an unusual cause of gastric outlet obstruction.

Neurofibromatosis type-1 (NF-1), also known as von Recklinghausen disease, is a common autosomal dominant condition occurring in approximately 1/3000 births. NF-1 is known to be associated with gastrointestinal neoplasms in 2-25 per cent of patients. We report the first case of gastric outlet obstruction with perforation caused by neurofibroma in a patient with NF-1. The literature is reviewed, examining 61 previously reported cases of noncarcinoid gastrointestinal (GI) neoplasms in patients with NF-1 for symptoms, location, and types of neoplasms. Neoplasms were located most often in the small intestine (72%). Neurofibromas, found in 52 per cent of patients, were the most frequently diagnosed benign neoplasms followed by leiomyomas (13%), ganglioneurofibromas (9.8%), and gastrointestinal stomal tumor (GIST) (6.5%). Adenocarcinoma was present in 23 per cent of patients. Patients with NF-1 and GI symptoms are at risk for gastrointestinal neoplasms from which symptomatic patients are likely to experience significant morbidity.

An optimized color transformation for the analysis of digital images of hematoxylin & eosin stained slides.

Hematoxylin and eosin (H&E) staining is ubiquitous in pathology practice and research. As digital pathology has evolved, the reliance of quantitative methods that make use of H&E images has similarly expanded. For example, cell counting and nuclear morphometry rely on the accurate demarcation of nuclei from other structures and each other. One of the major obstacles to quantitative analysis of H&E images is the high degree of variability observed between different samples and different laboratories. In an effort to characterize this variability, as well as to provide a substrate that can potentially mitigate this factor in quantitative image analysis, we developed a technique to project H&E images into an optimized space more appropriate for many image analysis procedures. We used a decision tree-based support vector machine learning algorithm to classify 44 H&E stained whole slide images of resected breast tumors according to the histological structures that are present. This procedure takes an H&E image as an input and produces a classification map of the image that predicts the likelihood of a pixel belonging to any one of a set of user-defined structures (e.g., cytoplasm, stroma). By reducing these maps into their constituent pixels in color space, an optimal reference vector is obtained for each structure, which identifies the color attributes that maximally distinguish one structure from other elements in the image. We show that tissue structures can be identified using this semi-automated technique. By comparing structure centroids across different images, we obtained a quantitative depiction of H&E variability for each structure. This measurement can potentially be utilized in the laboratory to help calibrate daily staining or identify troublesome slides. Moreover, by aligning reference vectors derived from this technique, images can be transformed in a way that standardizes their color properties and makes them more amenable to image processing.