A reliable transcriptomic risk-score applicable to formalin-fixed paraffin-embedded biopsies improves outcome prediction in localized prostate cancer.

BACKGROUND: Clinical manifestation of prostate cancer (PCa) is highly variable. Aggressive tumors require radical treatment while clinically non-significant ones may be suitable for active surveillance. We previously developed the prognostic ProstaTrend RNA signature based on transcriptome-wide microarray and RNA-sequencing (RNA-Seq) analyses, primarily of prostatectomy specimens. An RNA-Seq study of formalin-fixed paraffin-embedded (FFPE) tumor biopsies has now allowed us to use this test as a basis for the development of a novel test that is applicable to FFPE biopsies as a tool for early routine PCa diagnostics. METHODS: All patients of the FFPE biopsy cohort were treated by radical prostatectomy and median follow-up for biochemical recurrence (BCR) was 9 years. Based on the transcriptome data of 176 FFPE biopsies, we filtered ProstaTrend for genes susceptible to FFPE-associated degradation via regression analysis. ProstaTrend was additionally restricted to genes with concordant prognostic effects in the RNA-Seq TCGA prostate adenocarcinoma (PRAD) cohort to ensure robust and broad applicability. The prognostic relevance of the refined Transcriptomic Risk Score (TRS) was analyzed by Kaplan-Meier curves and Cox-regression models in our FFPE-biopsy cohort and 9 other public datasets from PCa patients with BCR as primary endpoint. In addition, we developed a prostate single-cell atlas of 41 PCa patients from 5 publicly available studies to analyze gene expression of ProstaTrend genes in different cell compartments. RESULTS: Validation of the TRS using the original ProstaTrend signature in the cohort of FFPE biopsies revealed a relevant impact of FFPE-associated degradation on gene expression and consequently no significant association with prognosis (Cox-regression, p-value > 0.05) in FFPE tissue. However, the TRS based on the new version of the ProstaTrend-ffpe signature, which included 204 genes (of originally 1396 genes), was significantly associated with BCR in the FFPE biopsy cohort (Cox-regression p-value < 0.001) and retained prognostic relevance when adjusted for Gleason Grade Groups. We confirmed a significant association with BCR in 9 independent cohorts including 1109 patients. Comparison of the prognostic performance of the TRS with 17 other prognostically relevant PCa panels revealed that ProstaTrend-ffpe was among the best-ranked panels. We generated a PCa cell atlas to associate ProstaTrend genes with cell lineages or cell types. Tumor-specific luminal cells have a significantly higher TRS than normal luminal cells in all analyzed datasets. In addition, TRS of epithelial and luminal cells was correlated with increased Gleason score in 3 studies. CONCLUSIONS: We developed a prognostic gene-expression signature for PCa that can be applied to FFPE biopsies and may be suitable to support clinical decision-making.

Focal structural variants revealed by whole genome sequencing disrupt the histone demethylase KDM4C in B-cell lymphomas.

Histone methylation-modifiers, such as EZH2 and KMT2D, are recurrently altered in B-cell lymphomas. To comprehensively describe the landscape of alterations affecting genes encoding histone methylation-modifiers in lymphomagenesis we investigated whole genome and transcriptome data of 186 mature B-cell lymphomas sequenced in the ICGC MMML-Seq project. Besides confirming common alterations of KMT2D (47% of cases), EZH2 (17%), SETD1B (5%), PRDM9 (4%), KMT2C (4%), and SETD2 (4%), also identified by prior exome or RNA-sequencing studies, we here found recurrent alterations to KDM4C in chromosome 9p24, encoding a histone demethylase. Focal structural variation was the main mechanism of KDM4C alterations, and was independent from 9p24 amplification. We also identified KDM4C alterations in lymphoma cell lines including a focal homozygous deletion in a classical Hodgkin lymphoma cell line. By integrating RNA-sequencing and genome sequencing data we predict that KDM4C structural variants result in loss-offunction. By functional reconstitution studies in cell lines, we provide evidence that KDM4C can act as a tumor suppressor. Thus, we show that identification of structural variants in whole genome sequencing data adds to the comprehensive description of the mutational landscape of lymphomas and, moreover, establish KDM4C as a putative tumor suppressive gene recurrently altered in subsets of B-cell derived lymphomas.

Characterization and evaluation of gene fusions as a measure of genetic instability and disease prognosis in prostate cancer.

BACKGROUND: Prostate cancer (PCa) is one of the most prevalent cancers worldwide. The clinical manifestations and molecular characteristics of PCa are highly variable. Aggressive types require radical treatment, whereas indolent ones may be suitable for active surveillance or organ-preserving focal therapies. Patient stratification by clinical or pathological risk categories still lacks sufficient precision. Incorporating molecular biomarkers, such as transcriptome-wide expression signatures, improves patient stratification but so far excludes chromosomal rearrangements. In this study, we investigated gene fusions in PCa, characterized potential novel candidates, and explored their role as prognostic markers for PCa progression. METHODS: We analyzed 630 patients in four cohorts with varying traits regarding sequencing protocols, sample conservation, and PCa risk group. The datasets included transcriptome-wide expression and matched clinical follow-up data to detect and characterize gene fusions in PCa. With the fusion calling software Arriba, we computationally predicted gene fusions. Following detection, we annotated the gene fusions using published databases for gene fusions in cancer. To relate the occurrence of gene fusions to Gleason Grading Groups and disease prognosis, we performed survival analyses using the Kaplan-Meier estimator, log-rank test, and Cox regression. RESULTS: Our analyses identified two potential novel gene fusions, MBTTPS2,L0XNC01::SMS and AMACR::AMACR. These fusions were detected in all four studied cohorts, providing compelling evidence for the validity of these fusions and their relevance in PCa. We also found that the number of gene fusions detected in a patient sample was significantly associated with the time to biochemical recurrence in two of the four cohorts (log-rank test, p-value < 0.05 for both cohorts). This was also confirmed after adjusting the prognostic model for Gleason Grading Groups (Cox regression, p-values < 0.05). CONCLUSIONS: Our gene fusion characterization workflow revealed two potential novel fusions specific for PCa. We found evidence that the number of gene fusions was associated with the prognosis of PCa. However, as the quantitative correlations were only moderately strong, further validation and assessment of clinical value is required before potential application.

Distinct Immune Signatures Indicative of Treatment Response and Immune-Related Adverse Events in Melanoma Patients under Immune Checkpoint Inhibitor Therapy.

To identify potential early biomarkers of treatment response and immune-related adverse events (irAE), a pilot immune monitoring study was performed in stage IV melanoma patients by flow cytometric analysis of peripheral blood mononuclear cells (PBMC). Overall, 17 patients were treated with either nivolumab or pembrolizumab alone, or with a combination of nivolumab and ipilimumab every three weeks. Of 15 patients for which complete response assessment was available, treatment responders (n = 10) as compared to non-responders (n = 5) were characterized by enhanced PD-1 expression on CD8+ T cells immediately before treatment (median ± median absolute deviation/MAD 26.7 ± 10.4% vs. 17.2 ± 5.3%). Responders showed a higher T cell responsiveness after T cell receptor ex vivo stimulation as determined by measurement of programmed cell death 1 (PD-1) expression on CD3+ T cells before the second cycle of treatment. The percentage of CD8+ effector memory (CD8+CD45RA-CD45RO+CCR7-) T cells was higher in responders compared to non-responders before and immediately after the first cycle of treatment (median ± MAD 39.2 ± 7.3% vs. 30.5 ± 4.1% and 37.7 ± 4.6 vs. 24.0 ± 6.4). Immune-related adverse events (irAE) were accompanied by a higher percentage of activated CD4+ (CD4+CD38+HLADR+) T cells before the second treatment cycle (median ± MAD 14.9 ± 3.9% vs. 5.3 ± 0.4%). In summary, PBMC immune monitoring of immune-checkpoint inhibition (ICI) treatment in melanoma appears to be a promising approach to identify early markers of treatment response and irAEs.

Lupus erythematosus: correlation of clinical and histological findings and proposal for a modified disease classification.

BACKGROUND: Classification of lupus erythematosus (LE) is conflicting as it is carried out from different starting points. Whereas dermatological classifications categorize LE morphologically based on specific cutaneous lesions, rheumatologic classifications are based on symptomatic aspects. Indeed, LE is a systemic autoimmune disease with variable acuity and organ involvement. All cutaneous disease patterns may occur in both limited-cutaneous and systemic LE. PATIENTS AND METHODS: 76 LE-patients with complete clinical data, clinical photographs and biopsy of cutaneous manifestations as well as paraclinical findings were retrospectively analyzed. Based on a published two-dimensional classification system that considers disease-specific skin manifestations and final disease diagnosis separately, patients' diagnoses were revised and compared with those in medical records. In addition, the extent to which patients could be clustered by diagnosis based on their LE-specific skin manifestations, corresponding histopathological changes, and paraclinical data was investigated. RESULTS: After re-evaluation, the proportion of patients with limited-cutaneous LE decreased from 82% previously to 24%. More than two-thirds of patients indeed showed intermediate or systemic LE. Disease-specific skin manifestations, histologic characteristics and paraclinical data did not cluster with final diagnoses. CONCLUSIONS: First, the work underlines the systemic character of the disease. Second, a two-dimensional approach can help overcome classification difficulties in LE, as skin-morphologic and symptomatic aspects can be considered separately.

The impact of SOCS1 mutations in diffuse large B-cell lymphoma.

Mutations in SOCS1 are frequent in primary mediastinal B-cell lymphoma and classical Hodgkin lymphoma. In the latter, SOCS1 mutations affect the length of the encoded protein (major mutations) and are associated with shorter patient survival. Two independent studies examined the prognostic impact of SOCS1 mutations in diffuse large B-cell lymphoma (DLBCL) and showed differing results. This may be due to the small number of included patients, the heterogeneity of patients' demographics and the distinct treatment schemes in these studies. To overcome the size limitations of these previous studies, we assessed SOCS1 mutations in the RICOVER-60 cohort. The cohort uniformly consists of elderly patients (aged 61-80 years) treated with the CHOP-14 scheme (cyclophosphamide, hydroxydaunorubicin, vincristine, prednisolone at 14-day intervals) with or without an additional rituximab treatment. Patient outcomes were analysed with regard to overall SOCS1 mutation frequency, major and minor mutations and a novel impact-based classifier - against the treatment modalities. Patients harbouring putative pathogenic SOCS1 mutations showed significant reduced overall survival within the CHOP plus rituximab group. Hence, putative pathogenic SOCS1 mutations seem to efface the beneficial effect of the therapeutic CD20 antibody. Comparing published data of whole exome and transcriptome sequencing of a large DLBCL cohort confirmed that predicted deleterious SOCS1 mutations forecast pre-eminent survival in early onset DLBCL.

Stress-related hormones in association with periodontal condition in adolescents-results of the epidemiologic LIFE Child study.

OBJECTIVES: The aim of this study was to investigate the associations between blood levels of stress-related hormones and early signs of periodontal disease in children and adolescents. MATERIALS AND METHODS: Within the LIFE (Leipzig research center for civilization diseases) Child study, 498 adolescents (10 to 18 years) were included. Early signs of periodontal inflammation were measured by probing depth (PD) at six index teeth (16, 11, 26, 36, 31, 46). Blood levels of stress-related hormones (cortisol, dehydroepiandosterone-sulfate [DHEA-S]) and, additionally interleukine-6 (IL-6) were measured. Socioeconomic status, oral hygiene, orthodontic appliances, and nutritional status, recorded by body-mass-index-standard-deviation-score (BMI-SDS), were considered as confounding factors. Additionally, in 98 participants, an oral chairside active matrix metalloproteinase-8 (aMMP-8) test was performed. Statistical tests are the Mann-Whitney U tests, chi-squared tests and multivariate logistic regression model. RESULTS: IL-6, BMI-SDS as well as positive aMMP-8 test result were significantly associated with maximum PD > 3 mm (p < 0.05). However, no statistically significant associations between stress-related hormones (cortisol and DHEA-S) and presence of maximum PD > 3 mm were found (p > 0.05). Higher DHEA-S and BMI were associated with positive aMMP-8 result, even after adjusting for age and gender (p = 0.027, padj = 0.026). CONCLUSION: The results reveal no associations between PD and stress-related hormones cortisol and DHEA-S. aMMP-8 test result might be associated with DHEA-S level. Nutritional status seems to influence periodontal disease in adolescents. CLINICAL RELEVANCE: DHEA-S and BMI-SDS show associations with early signs of periodontal disease in adolescents aged 10 to 18 years. This association should be confirmed by the investigation of high-risk groups.

Gene expression profiling reveals a close relationship between follicular lymphoma grade 3A and 3B, but distinct profiles of follicular lymphoma grade 1 and 2.

A linear progression model of follicular lymphomas (FL) FL1, FL2 and FL3A has been favored, since FL3A often co-exist with an FL1/2 component. FL3B, in contrast, is thought to be more closely related to diffuse large B-cell lymphoma (DLBCL), and both are often simultaneously present in one tumor (DLBCL/FL3B). To obtain more detailed insights into follicular lymphoma progression, a comprehensive analysis of a well-defined set of FL1/2 (n=22), FL3A (n=16), FL3B (n=6), DLBCL/FL3B (n=9), and germinal center B-cell-type diffuse large B-cell lymphoma (n=45) was undertaken using gene expression profiling, immunohistochemical stainings and genetic analyses by fluorescence in situ hybridization. While immunohistochemical (CD10, IRF4/MUM1, Ki67, BCL2, BCL6) and genetic profiles (translocations of BCL2, BCL6 and MYC) delineate FL1-3A from FL3B and DLBCL/FL3B, significant differences were observed between FL1/2 and FL3A upon gene expression profiling. Interestingly, FL3B turned out to be closely related to FL3A, not categorizing within a separate gene expression cluster, and both FL3A and FL3B showed overlapping profiles in between FL1/2 and diffuse large B-cell lymphoma. Finally, based upon their gene expression pattern, DLBCL/FL3B represent a composite form of FL3B and DLBCL, with the majority of samples more closely resembling the latter. The fact that gene expression profiling clearly separated FL1/2 from both FL3A and FL3B suggests a closer biological relationship between the latter. This notion, however, is in contrast to immunohistochemical and genetic profiles of the different histological FL subtypes that point to a closer relationship between FL1/2 and FL3A, and separates them from FL3B.

MINCR is a MYC-induced lncRNA able to modulate MYC's transcriptional network in Burkitt lymphoma cells.

Despite the established role of the transcription factor MYC in cancer, little is known about the impact of a new class of transcriptional regulators, the long noncoding RNAs (lncRNAs), on MYC ability to influence the cellular transcriptome. Here, we have intersected RNA-sequencing data from two MYC-inducible cell lines and a cohort of 91 B-cell lymphomas with or without genetic variants resulting in MYC overexpression. We identified 13 lncRNAs differentially expressed in IG-MYC-positive Burkitt lymphoma and regulated in the same direction by MYC in the model cell lines. Among them, we focused on a lncRNA that we named MYC-induced long noncoding RNA (MINCR), showing a strong correlation with MYC expression in MYC-positive lymphomas. To understand its cellular role, we performed RNAi and found that MINCR knockdown is associated with an impairment in cell cycle progression. Differential gene expression analysis after RNAi showed a significant enrichment of cell cycle genes among the genes down-regulated after MINCR knockdown. Interestingly, these genes are enriched in MYC binding sites in their promoters, suggesting that MINCR acts as a modulator of the MYC transcriptional program. Accordingly, MINCR knockdown was associated with a reduction in MYC binding to the promoters of selected cell cycle genes. Finally, we show that down-regulation of Aurora kinases A and B and chromatin licensing and DNA replication factor 1 may explain the reduction in cellular proliferation observed on MINCR knockdown. We, therefore, suggest that MINCR is a newly identified player in the MYC transcriptional network able to control the expression of cell cycle genes.

aMMP-8 in correlation to caries and periodontal condition in adolescents-results of the epidemiologic LIFE child study.

OBJECTIVES: The suitability of a chairside aMMP-8 test in determination of periodontal inflammation and caries in adolescents was assessed. Secondly, the influence of orthodontic treatment on aMMP-8 test result was analyzed. MATERIALS AND METHODS: Within the LIFE Child study, 434 adolescents (10 to 18 years) were included. Clinical dental examinations comprised caries experience (DMF/T-Index), signs of periodontal inflammation (probing pocket depth, PPD; community periodontal index of treatment needs; CPITN) at six index teeth and oral hygiene (OH). Information about orthodontic appliances (OA) and socioeconomic status (SES) were obtained by validated questionnaires. Test's sensitivity and specificity to detect periodontal inflammation and carious lesions were evaluated. The influence of OA on the test result was analyzed (multivariate model). RESULTS: No associations between age, gender, SES or OH, and test outcome were found (p > 0.05). Positive test results correlated to periodontal findings (CPITN, mean PPD; p < 0.001). However, for the detection of ≥ 1 site(s) with PPD ≥ 4 mm, the test's sensitivity and specificity were found to be 61 and 69%, respectively. Multivariate analysis revealed a higher probability for a positive test result in cases of fixed OA (odds ratio 5.02, 95% confidence interval 1.90-13.19). The test had no diagnostic value considering carious lesions. CONCLUSIONS: The chairside aMMP-8 test does not reliably identify adolescents with periodontal inflammation. Positive test results were more frequent in case of OA. CLINICAL RELEVANCE: The chairside aMMP-8 test is no appropriate tool to screen children and adolescents neither for periodontal inflammation nor for carious lesions.

TP53 mutation and survival in aggressive B cell lymphoma.

TP53 is mutated in 20-25% of aggressive B-cell lymphoma (B-NHL). To date, no studies have addressed the impact of TP53 mutations in prospective clinical trial cohorts. To evaluate the impact of TP53 mutation to current risk models in aggressive B-NHL, we investigated TP53 gene mutations within the RICOVER-60 trial. Of 1,222 elderly patients (aged 61-80 years) enrolled in the study and randomized to six or eight cycles of CHOP-14 with or without Rituximab (NCT00052936), 265 patients were analyzed for TP53 mutations. TP53 mutations were demonstrated in 63 of 265 patients (23.8%). TP53 mutation was associated with higher LDH (65% vs. 37%; p < 0.001), higher international prognostic index-Scores (IPI 4/5 27% vs. 12%; p = 0.025) and B-symptoms (41% vs. 24%; p = 0.011). Patients with TP53 mutation were less likely to obtain a complete remission CR/CRu (CR unconfirmed) 61.9% (mut) vs. 79.7% (wt) (p = 0.007). TP53 mutations were associated with decreased event-free (EFS), progression-free (PFS) and overall survival (OS) (median observation time of 40.2 months): the 3 year EFS, PFS and OS were 42% (vs. 60%; p = 0.012), 42% (vs. 67.5%; p < 0.001) and 50% (vs. 76%; p < 0.001) for the TP53 mutation group. In a Cox proportional hazard analysis adjusting for IPI-factors and treatment arms, TP53 mutation was shown to be an independent predictor of EFS (HR 1.5), PFS (HR 2.0) and OS (HR 2.3; p < 0.001). TP53 mutations are independent predictors of survival in untreated patients with aggressive CD20+ lymphoma. TP53 mutations should be considered for risk models in DLBCL and strategies to improve outcome for patients with mutant TP53 must be developed.

Novel IGH and MYC Translocation Partners in Diffuse Large B-Cell Lymphomas.

Chromosomal translocations involving an immunoglobulin (IG) locus and a proto-oncogene play a major role in diffuse large B-cell lymphoma (DLBCL) pathogenesis. Recurrent IG translocation partners in DLBCL are the BCL6, BCL2, and MYC genes, but other rare translocation partners are also known. We studied 20 DLBCL with fluorescence in situ hybridization-based evidence for IG heavy chain (IGH) locus-associated translocations not involving BCL6, BCL2, MALT1, or MYC by long distance inverse PCR to identify the translocation partners. Moreover, we studied eight DLBCL with MYC translocations not involving IG or known non-IG loci as translocation partner to search for novel MYC translocations. We identified three novel IGH-associated translocations. Chromosomal breakpoints involved the IMMP2L gene in 7q31, the BCAS2 gene in 1p13, and the PVRL2 gene in 19q13. The latter gene, which is recurrently translocated in T-cell lymphomas, is significantly higher expressed in the biopsy with the translocation compared to cases without this genetic aberration, indicating a pathogenetic role of PVRL2 also in DLBCL. In one case with a MYC break we obtained a novel MYC-SOCS1 translocation representing an unusual translocation of a proto-oncogene with a tumor suppressor gene. Indeed, we demonstrate that the oncogene was deregulated and the tumor suppressor gene inactivated. As both genes undergo aberrant somatic hypermutation in the region of the chromosomal breakpoints, this translocation likely happened as a byproduct of the hypermutation process. Overall, our study suggests that chromosomal translocations in DLBCL are more heterogeneous than previously known. © 2016 Wiley Periodicals, Inc.

A recurrent 11q aberration pattern characterizes a subset of MYC-negative high-grade B-cell lymphomas resembling Burkitt lymphoma.

The genetic hallmark of Burkitt lymphoma (BL) is the t(8;14)(q24;q32) and its variants leading to activation of the MYC oncogene. It is a matter of debate whether true BL without MYC translocation exists. Here, we identified 59 lymphomas concordantly called BL by 2 gene expression classifiers among 753 B-cell lymphomas. Only 2 (3%) of these 59 molecular BL lacked a MYC translocation, which both shared a peculiar pattern of chromosome 11q aberration characterized by interstitial gains including 11q23.2-q23.3 and telomeric losses of 11q24.1-qter. We extended our analysis to 17 MYC-negative high-grade B-cell lymphomas with a similar 11q aberration and showed this aberration to be recurrently associated with morphologic and clinical features of BL. The minimal region of gain was defined by high-level amplifications in 11q23.3 and associated with overexpression of genes including PAFAH1B2 on a transcriptional and protein level. The recurrent region of loss contained a focal homozygous deletion in 11q24.2-q24.3 including the ETS1 gene, which was shown to be mutated in 4 of 16 investigated cases. These findings indicate the existence of a molecularly distinct subset of B-cell lymphomas reminiscent of BL, which is characterized by deregulation of genes in 11q.

Alterations of microRNA and microRNA-regulated messenger RNA expression in germinal center B-cell lymphomas determined by integrative sequencing analysis.

MicroRNA are well-established players in post-transcriptional gene regulation. However, information on the effects of microRNA deregulation mainly relies on bioinformatic prediction of potential targets, whereas proof of the direct physical microRNA/target messenger RNA interaction is mostly lacking. Within the International Cancer Genome Consortium Project "Determining Molecular Mechanisms in Malignant Lymphoma by Sequencing", we performed miRnome sequencing from 16 Burkitt lymphomas, 19 diffuse large B-cell lymphomas, and 21 follicular lymphomas. Twenty-two miRNA separated Burkitt lymphomas from diffuse large B-cell lymphomas/follicular lymphomas, of which 13 have shown regulation by MYC. Moreover, we found expression of three hitherto unreported microRNA. Additionally, we detected recurrent mutations of hsa-miR-142 in diffuse large B-cell lymphomas and follicular lymphomas, and editing of the hsa-miR-376 cluster, providing evidence for microRNA editing in lymphomagenesis. To interrogate the direct physical interactions of microRNA with messenger RNA, we performed Argonaute-2 photoactivatable ribonucleoside-enhanced cross-linking and immunoprecipitation experiments. MicroRNA directly targeted 208 messsenger RNA in the Burkitt lymphomas and 328 messenger RNA in the non-Burkitt lymphoma models. This integrative analysis discovered several regulatory pathways of relevance in lymphomagenesis including Ras, PI3K-Akt and MAPK signaling pathways, also recurrently deregulated in lymphomas by mutations. Our dataset reveals that messenger RNA deregulation through microRNA is a highly relevant mechanism in lymphomagenesis.

The PCBP1 gene encoding poly(rC) binding protein I is recurrently mutated in Burkitt lymphoma.

The genetic hallmark of Burkitt lymphoma is the translocation t(8;14)(q24;q32), or one of its light chain variants, resulting in IG-MYC juxtaposition. However, these translocations alone are insufficient to drive lymphomagenesis, which requires additional genetic changes for malignant transformation. Recent studies of Burkitt lymphoma using next generation sequencing approaches have identified various recurrently mutated genes including ID3, TCF3, CCND3, and TP53. Here, by using similar approaches, we show that PCBP1 is a recurrently mutated gene in Burkitt lymphoma. By whole-genome sequencing, we identified somatic mutations in PCBP1 in 3/17 (18%) Burkitt lymphomas. We confirmed the recurrence of PCBP1 mutations by Sanger sequencing in an independent validation cohort, finding mutations in 3/28 (11%) Burkitt lymphomas and in 6/16 (38%) Burkitt lymphoma cell lines. PCBP1 is an intron-less gene encoding the 356 amino acid poly(rC) binding protein 1, which contains three K-Homology (KH) domains and two nuclear localization signals. The mutations predominantly (10/12, 83%) affect the KH III domain, either by complete domain loss or amino acid changes. Thus, these changes are predicted to alter the various functions of PCBP1, including nuclear trafficking and pre-mRNA splicing. Remarkably, all six primary Burkitt lymphomas with a PCBP1 mutation expressed MUM1/IRF4, which is otherwise detected in around 20-40% of Burkitt lymphomas. We conclude that PCBP1 mutations are recurrent in Burkitt lymphomas and might contribute, in cooperation with other mutations, to its pathogenesis.

Characterization of genomic imbalances in diffuse large B-cell lymphoma by detailed SNP-chip analysis.

The pathogenesis of diffuse large B-cell lymphomas (DLBCL) is only partly understood. We analyzed 148 DLBCL by single nucleotide polymorphism (SNP)-chips to characterize genomic imbalances. Seventy-nine cases were of the germinal center B-cell like (GCB) type of DLBCL, 49 of the activated B-cell like (ABC) subtype and 20 were unclassified DLBCL. Twenty-four regions of recurrent genomic gains and 38 regions of recurrent genomic losses were identified over the whole cohort, with a median of 25 imbalances per case for ABC-DLBCL and 19 per case for GCB-DLBCL. Several recurrent copy number changes showed differential frequencies in the GCB- and ABC-DLBCL subgroups, including gains of HDAC7A predominantly in GCB-DLBCL (38% of cases) and losses of BACH2 and CASP8AP2 predominantly in ABC-DLBCL (35%), hinting at disparate pathogenetic mechanisms in these entities. Correlating gene expression and copy number revealed a strong gene dosage effect in all tumors, with 34% of probesets showing a concordant expression change in affected regions. Two new potential tumor suppressor genes emerging from the analysis, CASP3 and IL5RA, were sequenced in ten and 16 candidate cases, respectively. However, no mutations were found, pointing to a potential haploinsufficiency effect of these genes, considering their reduced expression in cases with deletions. Our study thus describes differences and similarities in the landscape of genomic aberrations in the DLBCL subgroups in a large collection of cases, confirming already known targets, but also discovering novel copy number changes with possible pathogenetic relevance.

The role of HPV RNA transcription, immune response-related gene expression and disruptive TP53 mutations in diagnostic and prognostic profiling of head and neck cancer.

Stratification of head and neck squamous cell carcinomas (HNSCC) based on HPV16 DNA and RNA status, gene expression patterns, and mutated candidate genes may facilitate patient treatment decision. We characterize head and neck squamous cell carcinomas (HNSCC) with different HPV16 DNA and RNA (E6*I) status from 290 consecutively recruited patients by gene expression profiling and targeted sequencing of 50 genes. We show that tumors with transcriptionally inactive HPV16 (DNA+ RNA-) are similar to HPV-negative (DNA-) tumors regarding gene expression and frequency of TP53 mutations (47%, 8/17 and 43%, 72/167, respectively). We also find that an immune response-related gene expression cluster is associated with lymph node metastasis, independent of HPV16 status and that disruptive TP53 mutations are associated with lymph node metastasis in HPV16 DNA- tumors. We validate each of these associations in another large data set. Four gene expression clusters which we identify differ moderately but significantly in overall survival. Our findings underscore the importance of measuring the HPV16 RNA (E6*I) and TP53-mutation status for patient stratification and identify associations of an immune response-related gene expression cluster and TP53 mutations with lymph node metastasis in HNSCC.

Molecular classification of diffuse cerebral WHO grade II/III gliomas using genome- and transcriptome-wide profiling improves stratification of prognostically distinct patient groups.

Cerebral gliomas of World Health Organization (WHO) grade II and III represent a major challenge in terms of histological classification and clinical management. Here, we asked whether large-scale genomic and transcriptomic profiling improves the definition of prognostically distinct entities. We performed microarray-based genome- and transcriptome-wide analyses of primary tumor samples from a prospective German Glioma Network cohort of 137 patients with cerebral gliomas, including 61 WHO grade II and 76 WHO grade III tumors. Integrative bioinformatic analyses were employed to define molecular subgroups, which were then related to histology, molecular biomarkers, including isocitrate dehydrogenase 1 or 2 (IDH1/2) mutation, 1p/19q co-deletion and telomerase reverse transcriptase (TERT) promoter mutations, and patient outcome. Genomic profiling identified five distinct glioma groups, including three IDH1/2 mutant and two IDH1/2 wild-type groups. Expression profiling revealed evidence for eight transcriptionally different groups (five IDH1/2 mutant, three IDH1/2 wild type), which were only partially linked to the genomic groups. Correlation of DNA-based molecular stratification with clinical outcome allowed to define three major prognostic groups with characteristic genomic aberrations. The best prognosis was found in patients with IDH1/2 mutant and 1p/19q co-deleted tumors. Patients with IDH1/2 wild-type gliomas and glioblastoma-like genomic alterations, including gain on chromosome arm 7q (+7q), loss on chromosome arm 10q (-10q), TERT promoter mutation and oncogene amplification, displayed the worst outcome. Intermediate survival was seen in patients with IDH1/2 mutant, but 1p/19q intact, mostly astrocytic gliomas, and in patients with IDH1/2 wild-type gliomas lacking the +7q/-10q genotype and TERT promoter mutation. This molecular subgrouping stratified patients into prognostically distinct groups better than histological classification. Addition of gene expression data to this genomic classifier did not further improve prognostic stratification. In summary, DNA-based molecular profiling of WHO grade II and III gliomas distinguishes biologically distinct tumor groups and provides prognostically relevant information beyond histological classification as well as IDH1/2 mutation and 1p/19q co-deletion status.

Histopathology of rhinophyma - a clinical-histopathologic correlation.

BACKGROUND: Whereas early rhinophyma is histopathologically thought to resemble fully developed rosacea, a fibromatous variant has previously been described for severe rhinophyma. In terms of clinical characteristics, recently a new Rhinophyma Severity Index (RHISI) was introduced. METHODS: We studied 24 patients who had been treated with wide shave excisions for rhinophyma. Specimens were stained with hematoxylin-eosin, periodic acid-Schiff reaction and a panel of immunohistochemical stains and observed for any correlation between clinical severity and histopathologic features as well as for predictive markers of clinical recurrence. RESULTS: There were no significant histopathologic differences between the groups reflecting the different clinical expressions. From a histopathologic perspective, clinically severe forms did not show with exclusive fibrotic changes. Further, there was no histopathologic marker predicting the clinical course or possible recurrence of the disease after surgical treatment. Only the clinical pre-operative RHISI score correlated with the postoperative outcome, with a high pre-operative RHISI being a risk factor for recurrence. CONCLUSION: Histopathologic features do not correlate with the clinical expression of rhinophyma. An exclusively 'fibrotic' rhinophyma form does not appear to exist and could possibly be the result of sampling error based on small biopsies studied.