Meta-analysis assessing the sensitivity and specificity of 18F-FDG PET/CT for the diagnosis of prosthetic valve endocarditis (PVE) using individual patient data (IPD).

IMPORTANCE: The use of 18F-FDG PET/CT in diagnostic algorithms for PVE has increased since publication of studies and guidelines advocating its use. The assessment of test accuracy has been limited by small study sizes. We undertook a systematic review using individual patient data (IPD) meta-analysis techniques. OBJECTIVE: To estimate the summary sensitivity and specificity of 18F-FDG PET/CT in diagnosing PVE. We also assessed the effect of patient factors on test accuracy as defined by changes in the odds ratios associated with each factor. The effect of the PET/CT study on the final diagnosis was also assessed when compared to the preliminary Duke classification to determine in which patient group 18F-FDG PET/CT had the greatest utility. STUDY SELECTION: Studies were included if PET/CT was performed for suspicion of PVE and IPD of both the PET/CT result and final diagnosis defined by a gold-standard assessment was available. There were 3 possible final diagnoses ("definite PVE," "possible PVE," and "rejected PVE"). RESULTS: Seventeen studies were included with IPD available for 537 patients (from 538 scans). The summary sensitivity and specificity were 85% (95% CI 74.2%-91.8%) and 86.5% (95% CI 75.8%-92.9%) respectively when patients with final diagnosis of "possible PVE" were classified as positive for PVE. When this group was classified as negative for PVE, sensitivity was 87.4% (95% CI 80.4%-92.1%) and specificity was 84.9% (95% CI 71.5%-92.6%). Patients with a known pathogen (especially coagulase negative staphylococcal species), elevated CRP, a biological or aortic valve infection appeared more likely to have an accurate PET/CT diagnosis. Those with a mechanical valve, prior antibiotic treatment or a transcatheter aortic valve replacement valve were less likely to have an accurate test. Time since valve implantation and the presence of surgical adhesive did not appear to affect test accuracy. Of the patients with a preliminary Duke classification of "possible PVE," 84% received a more conclusive final diagnosis of "definite" or "rejected" PVE after the PET/CT study. CONCLUSIONS AND RELEVANCE: 18F-FDG PET/CT has high sensitivity and specificity in diagnosing PVE and the diagnostic utility is greatest in patients with a preliminary Duke classification of "possible PVE." Some patient factors appear to affect test accuracy, though these results should be interpreted with caution given low patient numbers for subgroup analyses.

Effects of freshwater release on oyster reef density, reproduction, and disease in a highly modified estuary.

Few estuaries remain unaffected by water management and altered freshwater deliveries. The Caloosahatchee River Estuary is a perfect case study for assessing the impact of altered hydrology on natural oyster reef (Crassostrea virginica) populations. The watershed has been highly modified and greatly enlarged by an artificial connection to Lake Okeechobee. Accordingly, to generate data to support water management recommendations, this study monitored various oyster biometrics over 15 years along the primary salinity gradient. Oyster reef densities were significantly affected by both prolonged high volume freshwater releases creating hyposaline conditions at upstream sites and by a lack of freshwater input creating hypersaline conditions at downstream sites. Low freshwater input led to an increase in disease caused by Perkinsus marinus and predation. Moderate (< 2000 cfs) and properly timed (winter/spring) freshets benefited oysters with increased gametogenesis, good larval mixing, and a reprieve from disease. If high volume freshets occurred in the late summer, extensive mortality occurred at the upstream site due to low salinity. These findings suggest freshwater releases in the late summer, when reproductive stress is at its peak and pelagic larvae are most vulnerable, should be limited to < 2000 cfs, but that longer freshets (1-3 weeks) in the winter and early spring (e.g., December-April) benefit oysters by reducing salinity and lessening disease intensity. Similar strategies can be employed in other managed systems, and patterns regarding the timing of high volume flows are applicable to all estuaries where the management of healthy oyster reefs is a priority.

Fixation Method for Hip Arthroplasty Stem Following Hip Fracture: A Population-Level Cost-Effectiveness Analysis.

BACKGROUND: Hip arthroplasty is increasing in Australia. The number of procedures for fractured neck of femur was 7500 in 2017. Best practices for fixation method and procedure type require scrutiny. This paper is about the costs and health outcomes of cemented and uncemented hemiarthroplasty and total hip arthroplasty at a national level. METHODS: We created a Markov model for patients <75, aged 75-85, and over 85. Expected costs and health outcomes over 5 years from a decision to change from existing practice to a best practice policy in which all patients with fractured neck of femur received the same fixation method based on age and type of arthroplasty are estimated. The model was populated using prevalence and incidence data from the Australian Orthopedic Association National Joint Replacement Registry, costs from Metro North Hospital and Health Service in Queensland, and probabilities and utilities from the literature. We simulated the uncertainties in outcomes with probabilistic sensitivity analysis. RESULTS: We found that uncemented stem procedures were more costly and provided worse health outcomes compared to cemented stem fixation for hemiarthroplasty and total hip arthroplasty for all age groups. Moving from existing practice to cemented stem arthroplasty could save the Australian health system $2.0 million over 5 years with a gain of 203 quality-adjusted life years. CONCLUSION: We suggest that consideration be given to cemented fixation of the femoral stem for patients receiving both hemiarthroplasty and total hip arthroplasty for fractured neck of femur. Best practice guidelines focused on cost-effectiveness should recommend cemented stem fixation to both save costs and improve patient quality of life.

Rotational Malalignment of the Knee Extensor Mechanism: Defining Rotation of the Quadriceps and Its Role in the Spectrum of Patellofemoral Joint Instability.

Osseous rotational malalignment of the lower limb is widely accepted as a factor contributing to patellofemoral instability, particularly in pediatric patients. Patellar instability occurs when the lateral force vector generated by the quadriceps exceeds the restraints provided by osseous and soft-tissue anatomy. The anatomy and activation of the quadriceps are responsible for the force applied across the patellofemoral joint, which has previously been measured using the quadriceps (Q)-angle. To our knowledge, the contribution of the quadriceps anatomy in generating a force vector in the axial plane has not previously been assessed. The primary aim of this study was to introduce the quadriceps torsion angle, a measure of quadriceps rotational alignment in the juvenile population. The secondary aims of this study were to determine the inter-assessor and intra-assessor reliability of the quadriceps torsion angle in the juvenile population and to investigate whether a large quadriceps torsion angle is a classifier of patellar dislocator group membership in a mixed cohort of patellar dislocators and typically developing controls. METHODS: Participants between the ages of 8 and 19 years were recruited as either controls or recurrent patellar dislocators. A total of 58 knees in both groups were assessed from magnetic resonance imaging scans of the entire lower limbs. Axial cuts midway between the superior aspect of the femoral head and the articular surface of the medial femoral condyle were used to calculate the proximal reference for the quadriceps torsion angle. The quadriceps torsion angle was defined as the angle between the line connecting the anterior aspect of the sartorius and the junction of the anterior and posterior compartments at the lateral intermuscular septum and the posterior condylar axis line. Inter-assessor reliability was calculated using the intraclass correlation coefficient. The relationship between the quadriceps torsion angle and the femoral torsion was assessed in the entire cohort. These values were compared between the control group and the dislocator group to determine if the raw values or an interplay between the 2 factors played a role in the pathoanatomy of recurrent patellofemoral dislocation. RESULTS: The quadriceps torsion angle was a reproducible assessment in both inter-assessor and intra-assessor reliability analyses. A moderate positive correlation (r = 0.624; p < 0.01) was found between the femoral torsion and the quadriceps torsion angle. Although the quadriceps torsion angle was a fair classifier of patellar dislocation group membership, femoral torsion was not. CONCLUSIONS: This study has quantified the rotational alignment of the extensor mechanism using the quadriceps torsion angle. The measurement is shown to be reliable and reproducible and a fair classifier of patellofemoral instability. CLINICAL RELEVANCE: This article introduces an objective measure of soft-tissue rotational malalignment in the pathogenesis of recurrent patellar dislocation.

FDG PET/CT of Metabolic Myopathy With Posttreatment Follow-up.

A 38-year-old woman presented with extreme fatigue and multiple lung nodules. She was referred for a PET/CT, which demonstrated multiple FDG-avid pulmonary nodules and lymph nodes with intense uptake within multiple muscle groups predominantly involving the paraspinal muscles and muscles of mastication. Histopathology of a paraspinal muscle biopsy revealed increased skeletal muscle lipid stores and increased mitochondria with normal morphology. This abnormality is seen in metabolic myopathy due to a disorder of fatty acid oxidation. Transbronchial biopsy showed no evidence of sarcoidosis. The patient was commenced on carnitine and riboflavin supplementation, and a follow-up PET/CT was performed.

Genetic predisposition to ductal carcinoma in situ of the breast.

BACKGROUND: Ductal carcinoma in situ (DCIS) is a non-invasive form of breast cancer. It is often associated with invasive ductal carcinoma (IDC), and is considered to be a non-obligate precursor of IDC. It is not clear to what extent these two forms of cancer share low-risk susceptibility loci, or whether there are differences in the strength of association for shared loci. METHODS: To identify genetic polymorphisms that predispose to DCIS, we pooled data from 38 studies comprising 5,067 cases of DCIS, 24,584 cases of IDC and 37,467 controls, all genotyped using the iCOGS chip. RESULTS: Most (67 %) of the 76 known breast cancer predisposition loci showed an association with DCIS in the same direction as previously reported for invasive breast cancer. Case-only analysis showed no evidence for differences between associations for IDC and DCIS after considering multiple testing. Analysis by estrogen receptor (ER) status confirmed that loci associated with ER positive IDC were also associated with ER positive DCIS. Analysis of DCIS by grade suggested that two independent SNPs at 11q13.3 near CCND1 were specific to low/intermediate grade DCIS (rs75915166, rs554219). These associations with grade remained after adjusting for ER status and were also found in IDC. We found no novel DCIS-specific loci at a genome wide significance level of P < 5.0x10(-8). CONCLUSION: In conclusion, this study provides the strongest evidence to date of a shared genetic susceptibility for IDC and DCIS. Studies with larger numbers of DCIS are needed to determine if IDC or DCIS specific loci exist.

Expression of regulators of mitotic fidelity are associated with intercellular heterogeneity and chromosomal instability in primary breast cancer.

Regulators of transition through mitosis such as SURVIVIN and Aurora kinase A (AURKA) have been previously implicated in the initiation of chromosomal instability (CIN), a driver of intratumour heterogeneity. We investigate the relationship between protein expression of these genes and directly quantified CIN, and their prognostic utility in breast cancer. The expression of SURVIVIN and AURKA was determined by immunohistochemistry in a cohort of 426 patients with primary breast cancer. The association between protein expression and histopathological characteristics, clinical outcome and CIN status, as determined by centromeric FISH and defined by modal centromere deviation, was analysed. Significantly poorer clinical outcome was observed in patients with high AURKA expression levels. Expression of SURVIVIN was elevated in ER-negative relative to ER-positive breast cancer. Both AURKA and SURVIVIN increased expression were significantly associated with breast cancer grade. There was a significant association between increased CIN and both increased AURKA and SURVIVIN expression. AURKA gene amplification was also associated with increased CIN. To our knowledge this is the largest study assessing CIN status in parallel with the expression of the mitotic regulators AURKA and SURVIVIN. These data suggest that elevated expression of AURKA and SURVIVIN, together with AURKA gene amplification, are associated with increased CIN in breast cancer, and may be used as a proxy for CIN in breast cancer samples in the absence of more advanced molecular measurements.

Genetic predisposition to in situ and invasive lobular carcinoma of the breast.

Invasive lobular breast cancer (ILC) accounts for 10-15% of all invasive breast carcinomas. It is generally ER positive (ER+) and often associated with lobular carcinoma in situ (LCIS). Genome-wide association studies have identified more than 70 common polymorphisms that predispose to breast cancer, but these studies included predominantly ductal (IDC) carcinomas. To identify novel common polymorphisms that predispose to ILC and LCIS, we pooled data from 6,023 cases (5,622 ILC, 401 pure LCIS) and 34,271 controls from 36 studies genotyped using the iCOGS chip. Six novel SNPs most strongly associated with ILC/LCIS in the pooled analysis were genotyped in a further 516 lobular cases (482 ILC, 36 LCIS) and 1,467 controls. These analyses identified a lobular-specific SNP at 7q34 (rs11977670, OR (95%CI) for ILC = 1.13 (1.09-1.18), P = 6.0 × 10(-10); P-het for ILC vs IDC ER+ tumors = 1.8 × 10(-4)). Of the 75 known breast cancer polymorphisms that were genotyped, 56 were associated with ILC and 15 with LCIS at P<0.05. Two SNPs showed significantly stronger associations for ILC than LCIS (rs2981579/10q26/FGFR2, P-het = 0.04 and rs889312/5q11/MAP3K1, P-het = 0.03); and two showed stronger associations for LCIS than ILC (rs6678914/1q32/LGR6, P-het = 0.001 and rs1752911/6q14, P-het = 0.04). In addition, seven of the 75 known loci showed significant differences between ER+ tumors with IDC and ILC histology, three of these showing stronger associations for ILC (rs11249433/1p11, rs2981579/10q26/FGFR2 and rs10995190/10q21/ZNF365) and four associated only with IDC (5p12/rs10941679; rs2588809/14q24/RAD51L1, rs6472903/8q21 and rs1550623/2q31/CDCA7). In conclusion, we have identified one novel lobular breast cancer specific predisposition polymorphism at 7q34, and shown for the first time that common breast cancer polymorphisms predispose to LCIS. We have shown that many of the ER+ breast cancer predisposition loci also predispose to ILC, although there is some heterogeneity between ER+ lobular and ER+ IDC tumors. These data provide evidence for overlapping, but distinct etiological pathways within ER+ breast cancer between morphological subtypes.

Simulation of potential oyster density with variable freshwater inflow (1965-2000) to the Caloosahatchee River Estuary, southwest Florida, USA.

Oyster beds are disappearing worldwide through a combination of over-harvesting, diseases, and salinity alterations in the coastal zone. Sensitivity of oysters to variable discharge and salinity is particularly acute in small sub-tropical estuaries subject to regulated freshwater releases. South Florida has sub-tropical estuaries where watershed flood control sometimes results in excessive freshwater inflow to estuaries during the wet season (May-Oct) and reduced discharge and increased salinities in the dry season (Nov-Apr). The potential to reserve freshwater accumulated during the wet season could offer the capacity to regulate freshwater at different temporal scales, thus optimizing salinity conditions for estuarine biota. The goal of this study was to use simulation modeling to explore the effects of freshwater inflows and salinity on adult oyster survival in the Caloosahatchee River Estuary (CRE) in southwest Florida. Water managers derived three different freshwater inflow scenarios for the CRE based on historical and modified watershed attributes for the time period of 1965-2000. Three different salinity time series were generated from the inflow scenarios at each of three sites in the lower CRE and used to conduct nine different oyster simulations. Overall, the predicted densities of adult oysters in the upstream site were 3-4 times greater in seasons that experienced reduced freshwater inflow (e.g., increased salinity) with oyster density in the lower estuary much less influenced by the inflows. Potential storage of freshwater reduced the frequency of extreme flows in the wet season and helped to maintain minimum inflow in the dry season near the estuarine mouth. Analyses of inflows indicated that discharges ranging from 0 to 1,500 cfs could promote favorable salinities of 10-25 in the lower CRE depending on wet versus dry season climatic conditions. This range of inflows is similar to that derived in other studies of the CRE and emphasizes the value of simulation models to help prescribe freshwater releases which benefit estuarine biota.

CERT depletion predicts chemotherapy benefit and mediates cytotoxic and polyploid-specific cancer cell death through autophagy induction.

Chromosomal instability (CIN) has been implicated in multidrug resistance and the silencing of the ceramide transporter, CERT, promotes sensitization to diverse cytotoxics. An improved understanding of mechanisms governing multidrug sensitization might provide insight into pathways contributing to the death of CIN cancer cells. Using an integrative functional genomics approach, we find that CERT-specific multidrug sensitization is associated with enhanced autophagosome-lysosome flux, resulting from the expression of LAMP2 following CERT silencing in colorectal and HER2(+) breast cancer cell lines. Live cell microscopy analysis revealed that CERT depletion induces LAMP2-dependent death of polyploid cells following exit from mitosis in the presence of paclitaxel. We find that CERT is relatively over-expressed in HER2(+) breast cancer and CERT protein expression acts as an independent prognostic variable and predictor of outcome in adjuvant chemotherapy-treated patients with primary breast cancer. These data suggest that the induction of LAMP2-dependent autophagic flux through CERT targeting may provide a rational approach to enhance multidrug sensitization and potentiate the death of polyploid cells following paclitaxel exposure to limit the acquisition of CIN and intra-tumour heterogeneity.

Relationship of extreme chromosomal instability with long-term survival in a retrospective analysis of primary breast cancer.

BACKGROUND: Chromosomal instability (CIN) is thought to be associated with poor prognosis in solid tumors; however, evidence from preclinical and mouse tumor models suggest that CIN may paradoxically enhance or impair cancer cell fitness. Breast cancer prognostic expression signature sets, which reflect tumor CIN status, efficiently delineate outcome in estrogen receptor ER-positive breast cancer in contrast to ER-negative breast cancer, suggesting that the relationship of CIN with prognosis differs in these two breast cancer subtypes. METHODS: Direct assessment of CIN requires single-cell analysis methods, such as centromeric FISH, aimed at determining the variation around the modal number of two or more chromosomes within individual tumor nuclei. Here, we document the frequency of tumor CIN by dual centromeric FISH analysis in a retrospective primary breast cancer cohort of 246 patients with survival outcome. RESULTS: There was increased CIN and clonal heterogeneity in ER-negative compared with ER-positive breast cancer. Consistent with a negative impact of CIN on cellular fitness, extreme CIN in ER-negative breast cancer was an independent variable associated with improved long-term survival in multivariate analysis. In contrast, a linear relationship of increasing CIN with poorer prognosis in ER-positive breast cancer was observed, using three independent measures of CIN. CONCLUSIONS: The paradoxical relationship between extreme CIN and cancer outcome in the ER-negative cohorts may explain why prognostic expression signatures, reflecting tumor CIN status, fail to predict outcome in this subgroup. IMPACT: Assessment of tumor CIN status may support risk stratification in ER-negative breast cancer and requires prospective validation.

Cyclin D-1 protein over-expression is not associated with gene amplification in benign and atypical apocrine lesions of the breast.

Cyclin D-1 protein over-expression and/or gene amplification have been shown to be frequent events in subsets of breast carcinomas. Cyclin D-1 is generally considered as a weak oncogene, and its over-expression has been shown to occur in occasional benign breast lesions. In a previous series, we have shown that cyclin D-1 was over-expressed in subsets of apocrine metaplasia (APM) and apocrine adenosis (AA) of the breast and that such over-expression was mostly associated with a significant increase in the proliferative capacity of these lesions. We examined the mechanisms involved in cyclin D-1 over-expression in apocrine lesions. A total of 41 cases were analysed in this study. The cases were divided into: 18 cases of APM and 23 cases of AA. All cases analysed had been previously analysed by immunohistochemistry, and all showed over-expression of the cyclin D-1 protein. Fluorescence in situ hybridisation (FISH) was performed using a dual cyclin D-1 (spectrum orange)/chromosome 11 centromere (spectrum green) DNA probe. The results showed that none of the cases studied had concomitant gene amplification. These results suggest that other post-transcriptional mechanisms might be responsible for cyclin D-1 protein over-expression in benign apocrine lesions. Further studies are needed to understand the mechanisms involved in abnormal cyclin D-1 expression in these lesions.

Homozygous PMS2 deletion causes a severe colorectal cancer and multiple adenoma phenotype without extraintestinal cancer.

BACKGROUND & AIMS: We report a patient of Indian descent with parental consanguinity, who developed 10 carcinomas and 35 adenomatous polyps at age 23 and duodenal adenocarcinoma at age 25. He also had dysmorphic features, mental retardation, and café-au-lait spots but no brain tumor. We aimed to establish his molecular diagnosis. METHODS: Germ-line screening for APC and MYH/MUTYH mutations was normal as was immunohistochemistry for MLH1 and MSH2 proteins. Investigation by array-comparative genomic hybridization revealed deletion of a small region on chromosome 7. Using polymerase chain reaction, this region was refined to a 400-kilobase deletion, which included exons 9-15 of the PMS2 gene, and all coding regions of oncomodulin, TRIAD3, and FSCN1. RESULTS: The deletion was confirmed as homozygous, and both parents were carriers. Immunohistochemistry showed absent PMS2 expression in all tumors and normal tissue. Most tumors showed microsatellite instability, more marked at dinucleotide than mononucleotide repeats. The tumors harbored no somatic mutations in APC, BRAF, AXIN2, or beta-catenin, but KRAS2 and TGFBR2 mutations were found. CONCLUSIONS: Our patient represents a novel phenotype for homozygous PMS2 mutation and perhaps the most severe colorectal cancer phenotype-in terms of numbers of malignancies at an early age-described to date. PMS2 mutations-and perhaps other homozygous mismatch repair mutations-should be considered in any patient presenting with multiple gastrointestinal tumors, since our patient could not be distinguished clinically from cases with attenuated familial adenomatous polyposis or MUTYH-associated polyposis.

Genomic profiling identifies discrete deletions associated with translocations in glioblastoma multiforme.

Glioblastoma multiforme is the most common tumor arising in the central nervous system. Patients with these tumors have limited treatment options and their disease is invariably fatal. Molecularly targeted agents offer the potential to improve patient treatment, however the use of these will require a fuller understanding of the genetic changes in these complex tumors. In this study, we identify copy number changes in a series of glioblastoma multiforme tumors and cell lines by applying high-resolution microarray comparative genomic hybridization. Molecular cytogenetic characterization of the cell lines revealed that copy number changes define translocation breakpoints. We focused on chromosome 6 and further characterized three regions of copy number change associated with translocations including a discrete deletion involving IGF2R, PARK2, PACRG and QKI and an unbalanced translocation involving POLH, GTPBP2 and PTPRZ1.

Combined array-comparative genomic hybridization and single-nucleotide polymorphism-loss of heterozygosity analysis reveals complex changes and multiple forms of chromosomal instability in colorectal cancers.

Cancers with chromosomal instability (CIN) are held to be aneuploid/polyploid with multiple large-scale gains/deletions, but the processes underlying CIN are unclear and different types of CIN might exist. We investigated colorectal cancer cell lines using array-comparative genomic hybridization (CGH) for copy number changes and single-copy number polymorphism (SNP) microarrays for allelic loss (LOH). Many array-based CGH changes were not found by LOH because they did not cause true reduction-to-homozygosity. Conversely, many regions of SNP-LOH occurred in the absence of copy number change, comprising an average per cell line of 2 chromosomes with complete LOH; 1-2 terminal regions of LOH (mitotic recombination); and 1 interstitial region of LOH. SNP-LOH detected many novel changes, representing possible locations of uncharacterized tumor suppressor loci. Microsatellite unstable (MSI+) lines infrequently showed gains/deletions or whole-chromosome LOH, but their near-diploid karyotypes concealed mitotic recombination frequencies similar to those of MSI- lines. We analyzed p53 and chromosome 18q (SMAD4) in detail, including mutation screening. Almost all MSI- lines showed LOH and/or deletion of p53 and 18q; some near-triploid lines had acquired three independent changes at these loci. We found consistent results in primary colorectal cancers. Overall, the distributions of mitotic recombination and whole-chromosome LOH in the MSI- cell lines differed significantly from random, with some lines having much higher than expected levels of these changes. Moreover, lines with more LOH changes had significantly fewer copy number changes. These data suggest that CIN is not synonymous with copy number change and some cancers have a specific tendency to whole-chromosome deletion and regain or to mitotic recombination.

Fluorescence in situ hybridization and comparative genomic hybridization.

This chapter discusses the complementary methodologies of fluorescence in situ hybridization and comparative genomic hybridization. Fluorescence in situ hybridization uses fluorescently labeled DNA probes (whole chromosomes, centromere, or locus-specific sequences) to visualize complementary DNA sequences in the target DNA (metaphase chromosomes or interphase nuclei). Comparative genomic hybridization is essentially a modified in situ hybridization, whereby the whole genome can be screened for gains and losses in a single experiment. Tumor and normal DNA are differentially fluorescently labeled and competitively co-hybridized to normal metaphase spreads. Both techniques require multiple steps in their preparation. These can be divided into preparation of the target, preparation of the probes, hybridization, post-hybridization washes, and image acquisition and analysis, each of which will be described in detail. Success of each experiment is dependent on all of these steps, but where steps are of critical importance to particular techniques, they will be highlighted, together with notes on how to optimize the process.

Array-CGH analysis of microsatellite-stable, near-diploid bowel cancers and comparison with other types of colorectal carcinoma.

Microsatellite-stable, near-diploid (MSI-CIN-) colorectal carcinomas have been reported, but it is not clear as to whether these tumours form a discrete group or represent one end of the distribution of MSI-CIN+ cancers. In order to address this question, we screened 23 MSI-CIN- colorectal cancers for gains and losses using array-based comparative genomic hybridization (aCGH) based on large-insert clones at about 1 Mb density. We compared our findings with those from a small set of MSI+CIN+ cancers, and with our reported data from MSI-CIN+ and MSI+CIN- cancers. We found no evidence of any form of genomic instability in MSI-CIN- cancers. At the level of the chromosome arm, the MSI-CIN- cancers had significantly fewer gains and losses than MSI-CIN+ tumours, but more than the MSI+CIN- and MSI+CIN+ lesions. The chromosomal-scale changes found in MSI-CIN- cancers generally involved the same sites as those in MSI-CIN+ tumours, and in both cancer groups, the best predictor of a specific change was the total number of such changes in that tumour. A few chromosomal-scale changes did, however, differ between the MSI-CIN- and MSI-CIN+ pathways. MSI-CIN- cancers showed: low frequencies of gain of 9p and 19p; infrequent loss of 5q and a high frequency of 20p gain. Overall, our data suggested that the MSI-CIN- group is heterogeneous, one type of MSI-CIN- cancer having few (< or =6) chromosomal-scale changes and the other with more (> or =10) changes resembling MSI-CIN+ cancers. At the level of individual clones, frequent and/or discrete gains or losses were generally located within regions of chromosomal-scale changes in both MSI-CIN- and MSI-CIN+ cancers, and fewer losses and gains were present in MSI-CIN- than MSI-CIN+ tumours. No changes by clone, which were specific to the MSI-CIN- cancers, were found. In addition to indicating differences among the cancer groups, our results also detected over 50 sites (amplifications, potential homozygous deletion and gains or losses which extended over only a few megabases) which might harbour uncharacterized oncogenes or tumour suppressor loci. In conclusion, our data support the suggestion that some MSI-CIN- carcinomas form a qualitatively different group from the other cancer types, and also suggest that the MSI-CIN- group is itself heterogeneous.

Analysis of ovarian cancer cell lines using array-based comparative genomic hybridization.

In this study, 23 ovarian cancer cell lines were screened using array-comparative genomic hybridization (aCGH) based on large-insert clones at about 1 Mb density from throughout the genome. The most frequent recurrent changes at the level of the chromosome arm were loss of chromosome 4 or 4q, loss of 18q and gain of 20 or 20q; other recurrent changes included losses of 6q, 8p, 9p, 11p, 15q, 16q, 17p, and 22q, and gain of 7q. Losses of 4q and 18q occurred together more often than expected. Evidence was found for two types of ovarian cancer, one typically near-triploid and characterized by a generally higher frequency of chromosomal changes (especially losses of 4p, 4q, 13q, 15q, 16p, 16q, 18p and 18q), and the other typically near-diploid/tetraploid and with fewer changes overall, but with relatively high frequencies of 9p loss, 9q gain, and 20p gain. Multiple novel changes (amplifications, homozygous deletions, discrete regions of gain or loss, small overlapping regions of change and frequently changed clones) were also detected, each of which might indicate the locations of oncogenes or tumour suppressor loci. For example, at least two regions of amplification on chromosome 11q13, one including cyclin D1 and the other the candidate oncogene PAK1, were found. Amplification on 11q22 near the progesterone receptor gene and a cluster of matrix metalloproteinase loci was also detected. Other potential oncogenes, which mapped to regions found by this study, included cyclin E and PIK3C2G. Candidate tumour suppressor genes in regions of loss included CDKN2C, SMAD4-interacting protein and RASSF2.

MSI-low, a real phenomenon which varies in frequency among cancer types.

This study assessed whether low-level microsatellite instability (MSI-L) is a phenomenon specific to colorectal cancers or is also present in other tumour types. Breast (grade III ductal and lobular), endometrial and ovarian carcinomas, as well as colorectal cancers, were analysed for MSI-L using eight microsatellite markers. The markers were selected from a panel that had previously been shown to be sensitive for the detection of MSI-L in colorectal cancers. It was found that MSI-L was present in 30 of 87 (35%) colorectal cancers, 2 of 59 (3%) grade III breast carcinomas, 1 of 35 (3%) lobular breast cancers, 16 of 50 (32%) endometrial cancers, and 9 of 34 (26%) ovarian cancers. These results suggest that MSI-L is a very rare occurrence in breast carcinomas, but does occur as a real phenomenon in colorectal, endometrial, and ovarian carcinomas, which are all part of the hereditary non-polyposis colon cancer (HNPCC) syndrome. PCR artefact was also found to masquerade as MSI-L; criteria for the assessment of MSI-L are suggested to eliminate this problem.